chapter
stringlengths 1.97k
1.53M
| path
stringlengths 47
241
|
|---|---|
Describe and give an example of an acute viral infection, a late complication following an acute infection, a latent viral infection, a chronic viral infection, and a slow viral infection.
Most viruses that infect humans, such as those that cause routine respiratory infections (e.g., cold viruses, influenza viruses) and gastrointestinal infections (e.g., Rotaviruses, Noroviruses), cause acute infections. Acute infections are of relatively short duration with rapid recovery.
In persistent infections, the viruses are continually present in the body. Some persistent infections are late complications following an acute infection and include subacute sclerosing panencephalitis (SSPE) that can follow an acute measles infection and progressive encephalitis that can follow rubella. Other persistent infections are known as latent viral infection. In a latent viral infection the virus remains in equilibrium with the host for long periods of time before symptoms again appear, but the actual viruses cannot be detected until reactivation of the disease occurs. Examples include infections caused by HSV-1 (fever blisters), HSV-2 (genital herpes), and VZV (chickenpox-shingles). In the case of chronic virus infections, the virus can be demonstrated in the body at all times and the disease may be present or absent for an extended period of time. Examples include hepatitis B (caused by HBV) and hepatitis C (caused by HCV). Slow infections are ones in which the infectious agents gradually increase in number over a very long period of time during which no significant symptoms are seen. Examples include AIDS (caused by HIV-1 and HIV-2) and certain lentiviruses that cause tumors in animals. Although not viruses, prions also cause slow infections.
Summary
1. Acute infections are of relatively short duration with rapid recovery.
2. Persistent infections are where the viruses are continually present in the body.
3. In a latent viral infection the virus remains in equilibrium with the host for long periods of time before symptoms again appear, but the actual viruses cannot be detected until reactivation of the disease occurs.
4. In a chronic virus infection, the virus can be demonstrated in the body at all times and the disease may be present or absent for an extended period of time.
5. Slow infections are ones in which the infectious agents gradually increase in number over a very long period of time during which no significant symptoms are seen.
10.E: Viruses (Exercises)
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied.
10.1: General Characteristics of Viruses
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. State 2 living characteristics of viruses.
2. State 2 nonliving characteristics of viruses.
3. List 3 criteria used to define a virus.
4. A virus that infects only bacteria is termed a ___________________. (ans)
5. State why viruses can't replicate on environmental surfaces or in synthetic laboratory medium. (ans)
10.2: Size and Shapes of Viruses
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Compare the size of most viruses to that of bacteria. (ans)
2. List 4 shapes of viruses.
10.3: Viral Structure
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the structure of most viruses that infect humans. (ans)
2. Define the following:
1. capsid (ans)
2. capsomeres (ans)
3. nucleocapsid (ans)
3. Describe how most animal viruses obtain their envelope. (ans)
4. State why some bacteriophages are more complex than typical polyhedral or helical viruses. (ans)
5. Multiple Choice (ans)
10.5: Other Acellular Infectious Agents: Viroids and Prions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Small, circular, single-stranded molecules of infectious that cause of a few plant diseases such as potato spindle-tuber disease,cucumber pale fruit, citrus exocortis disease, and cadang-cadang (coconuts) are called ____________. (ans)
2. Infectious protein particlesthought to be responsible for a group of transmissible and/or inherited neurodegenerative diseases including Creutzfeldt-Jakob disease, kuru, and Gerstmann-Straussler- syndrome in humans as well as scrapie in sheep and goats are called ______________. (ans)
3. Name 3 other neurological protein misfolding diseases that apprear to be initiated by prions. (ans)
10.7: Bacteriophage Life Cycles: An Overview
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Name the 2 types of bacteriophage life cycles and state what the bacteriophage capable of each is called.
10.7A: The Lytic Life Cycle of Bacteriophages
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describethe 5 steps involved in the lytic life cycle of bacteriophages.
2. Multiple Choice (ans)
10.7B: The Lysogenic Life Cycle of Bacteriophages
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describehow the lysogenic life cycle of temperate bacteriophages differs from the lytic life cycle of lytic bacteriophages. (ans)
2. What is spontaneous induction as it relates to the lysogenic life cycle? (ans)
3. When a bacteriophage inserts its DNA into the DNA of the host bacterium, this form of the virus is called a ________________. (ans)
4. The host bacterium for a bacteriophage is called a ________________. (ans)
5. A virus capable of the lysogenic life cycle is called a __________________. (ans)
6. Multiple Choice (ans)
10.8: Pathogenicity of Animal Viruses
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe 4 ways viruses can damage infected host cells.
2. Briefly describe 2 different ways viruses can evade host immune defenses and give an example of a virus that uses each mechanism.
3. Multiple Choice (ans)
10.9: Bacteriophage-Induced Alterations of Bacteria
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe how a bacteriophage may in some cases enable a bacterium to become virulent and state 2 examples. (ans)
10.10: Antiviral Agents
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Explain why the antibiotics we use to treat bacterial infections are not effective against viral infections. (ans)
2. Match the following drugs with the viral infections they are used against:
_____ amantadine, rimantidine, zanamivar, and oseltamivir (ans)
_____ acyclovir, famciclovir, penciclovir, and valacyclovir(ans)
_____ foscarnet, gancyclovir, cidofovir, valganciclovir, and fomivirsen(ans)
_____ AZT (ZDV), didanosine, zalcitabine, stavudine, nevirapine, delavirdine, saquinavir, and ritonavir (ans)
1. HIV infection and AIDS
2. influenza A
3. severe CMV infections such as retinitis
4. HSV and VZV infections
3. Match the following:
_____ These are drugs that bind to the active site of an HIV-encoded protease and prevent it from cleaving the long gag-pol polyprotein and the gag polyprotein into essential proteins essential to the structure of HIV and to RNA packaging within its nucleocapsid. As a result, viral maturation does not occur and noninfectious viral particles are produced. (ans)
_____ These drugs chemically resemble normal DNA nucleotides, the building block molecules for DNA synthesis. They bind to the active site of the reverse transcriptase which, in turn, inserts it into the growing DNA strand in place of a normal nucleotide. Once inserted, however, new DNA nucleotides are unable to attach to the drug and DNA synthesis is stopped. This results in an incomplete provirus. (ans)
1. nucleoside reverse transcriptase inhibitors
2. non-nucleoside reverse transcriptase inhibitors
3. protease inhibitors
4. entry inhibitors
4. Multiple Choice (ans)
10.11: General Categories of Viral Infections
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following:
_____ Viral infections in which the infectious agents gradually increase in number over a very long period of time during which no significant symptoms are seen. (ans)
_____ Viral infections of relatively short duration with rapid recovery. (ans)
_____ Viral infections where the virus can be demonstrated in the body at all times and the disease may be present or absent for an extended period of time. (ans)
_____ Viral infections where the virus remains in equilibrium with the host for long periods of time before symptoms again appear, but the actual viruses cannot be detected until reactivation of the disease occurs. (ans)
1. acute viral infection
2. chronic viral infection
3. latent viral infection
4. slow viral infection
2. Give an example of of a virus causing each of the following:
1. acute viral infection (ans)
2. chronic viral infection (ans)
3. latent viral infection (ans)
4. slow viral infection (ans)
3. Multiple Choice (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_4%3A_Eukaryotic_Microorganisms_and_Viruses/10%3A_Viruses/10.11%3A_General_Categories_of_Viral_Infections.txt |
Innate immunity is an antigen-nonspecific defence mechanisms that a host uses immediately or within several hours after exposure to almost any microbe. This is the immunity one is born with and is the initial response by the body to eliminate microbes and prevent infection. Innate immunity can be divided into immediate innate immunity and early induced innate immunity. In this section we will learn about immediate innate immunity.
• 11.1: The Innate Immune System: An Overview
The body has two immune systems: the innate immune system and the adaptive immune system. Innate immunity is an antigen-nonspecific defense mechanisms that a host uses immediately or within several hours after exposure to almost any microbe. Innate immunity is the immunity one is born with and is the initial response by the body to eliminate microbes. Immediate innate immunity begins 0 - 4 hours after exposure to an infectious agent. Early induced innate immunity begins 4 - 96 hours afterward.
• 11.2: Defense Cells in the Blood: The Leukocytes
The complete blood count (CBC) is a laboratory test that, among other things, determines the total number of both leukocytes and erythrocytes per ml of blood. In general, an elevated WBC count (leukocytosis ) is seen in infection, inflammation, leukemia, and parasitic infestations. Neutrophils are the most abundant of the leukocytes, normally accounting for 54-75% of the WBCs. Neutrophils are important phagocytes and also promote inflammation. Eosinophils normally comprise 1-4% of the WBCs.
• 11.3: Defense Cells in the Tissue - Dendritic Cells, Macrophages, and Mast Cells
Most dendritic cells are derived from monocytes and are referred to as myeloid dendritic cells and are located throughout the epithelium of the skin, the respiratory tract, and the gastrointestinal tract, as well as lymphoid tissues and organ parenchyma. Upon capturing antigens through pinocytosis and, the dendritic cells detach from their initial site, enter lymph vessels, and are carried to regional lymph nodes where they present antigens to the ever changing populations of naive T-lymphocytes
• 11.3: Immediate Innate Immunity
Immediate innate immunity begins 0-4 hours after exposure to an infectious agent and involves the action of soluble preformed antimicrobial molecules that circulate in the blood, our found in extracellular tissue fluids, and are secreted by epithelial cells. These include: antimicrobial enzymes and peptides, and complement system proteins. These preformed antimicrobial molecules are designed to immediately begin to remove infectious agents as soon as they enter the body.
• 11.4: Early Induced Innate Immunity
Early induced innate immunity begins 4 - 96 hours after exposure to an infectious agent and involves the recruitment of defense cells as a result of pathogen-associated molecular patterns or PAMPs binding to pattern-recognition receptors or PRRs. These recruited defense cells include phagocytic cells (leukocytes such as neutrophils, eosinophils, and monocytes; tissue phagocytic cells in the tissue such as macrophages), cells that release inflammatory mediators and natural killer cells (NK cells)
• 11.E: Innate Immunity (Exercises)
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied.
Thumbnail: A scanning electron microscope (SEM) image of a single human lymphocyte. (Public Domain; Dr. Triche National Cancer Institute).
Unit 5: Innate Immunity
Compare adaptive (acquired) immunity with innate immunity. Compare immediate innate immunity with early induced innate immunity. Define the following: pathogen-associated molecular patterns (PAMPs) pattern-recognition receptors (PRRs) antigen immunogen epitope.
In Units 1-4 we looked at microorganisms: how they replicate, why some are potentially more pathogenic than others, and how we can control them with antimicrobial agents. Units 4 and 5 are devoted to the ways in which the body defends itself against microbes and other potentially harmful cells and molecules. The body has two immune systems: the innate immune system and the adaptive immune system. Unit 5 deals with innate immunity while Unit 6 will cover adaptive immunity. Let's first briefly compare acquired and innate immunity.
Innate immunity
Innate immunity is an antigen-nonspecific defense mechanisms that a host uses immediately or within several hours after exposure to almost any microbe. This is the immunity one is born with and is the initial response by the body to eliminate microbes and prevent infection. Innate immunity can be divided into immediate innate immunity and early induced innate immunity.
Immediate innate immunity begins 0 - 4 hours after exposure to an infectious agent and involves the action of soluble preformed antimicrobial molecules that circulate in the blood, our found in extracellular tissue fluids, and are secreted by epithelial cells. These include:
• antimicrobial enzymes and peptides;
• complement system proteins; and
• anatomical barriers to infection, mechanical removal of microbes, and bacterial antagonism by normal body microbiota
These preformed innate defense molecules will be discussed in greater detail later in this unit.
Early induced innate immunity begins 4 - 96 hours after exposure to an infectious agent and involves the recruitment of defense cells as a result of pathogen-associated molecular patterns or PAMPS binding to pattern-recognition receptors or PRRs . These recruited defense cells include:
• phagocytic cells: leukocytes such as neutrophils, eosinophils, and monocytes; tissue phagocytic cells in the tissue such as macrophages ;
• cells that release inflammatory mediators: inflammatory cells in the tissue such as macrophages and mast cells ; leukocytes such as basophils and eosinophils; and
• natural killer cells (NK cells ).
Unlike adaptive immunity, innate immunity does not recognize every possible antigen. Instead, it is designed to recognize molecules shared by groups of related microbes that are essential for the survival of those organisms and are not found associated with mammalian cells. These unique microbial molecules are called pathogen-associated molecular patterns or PAMPS and include LPS from the gram-negative cell wall, peptidoglycan and lipotechoic acids from the gram-positive cell wall, the sugar mannose (a terminal sugar common in microbial glycolipids and glycoproteins but rare in those of humans), bacterial and viral unmethylated CpG DNA, bacterial flagellin, the amino acid N-formylmethionine found in bacterial proteins, double-stranded and single-stranded RNA from viruses, and glucans from fungal cell walls. In addition, unique molecules displayed on stressed, injured, infected, or transformed human cells also act as PAMPS. (Because all microbes, not just pathogenic microbes, possess PAMPs, pathogen-associated molecular patterns are sometimes referred to as microbe-associated molecular patterns or MAMPs.)
Most body defense cells have pattern-recognition receptors or PRRs for these common PAMPS (see Figure \(1\)) and so there is an immediate response against the invading microorganism. Pathogen-associated molecular patterns can also be recognized by a series of soluble pattern-recognition receptors in the blood that function as opsonins and initiate the complement pathways. In all, the innate immune system is thought to recognize approximately 103 of these microbial molecular patterns.
Examples of innate immunity include anatomical barriers, mechanical removal, bacterial antagonism, antigen-nonspecific defense chemicals, the complement pathways, phagocytosis, inflammation, fever, and the acute-phase response. In this current unit we will look at each of these in greater detail.
Adaptive (acquired) immunity
Adaptive (acquired) immunity refers to antigen-specific defense mechanisms that take several days to become protective and are designed to react with and remove a specific antigen . This is the immunity one develops throughout life. During adaptive immunity, antigens are transported to lymphoid organs where they are recognized by naive B-lymphocytes and T-lymphocytes. These activated B- and T-lymphocytes subsequently proliferate and differentiate into effector cells.
An antigen is defined as a substance that reacts with antibody molecules and antigen receptors on lymphocytes. An immunogen is an antigen that is recognized by the body as nonself and stimulates an adaptive immune response. For simplicity we will use the term antigen when referring to both antigens and immunogens. The actual portions or fragments of an antigen that react with antibodies and lymphocyte receptors are called epitopes .
As we will see later in Unit 5, the body recognizes an antigen as foreign when epitopes of that antigen bind to B-lymphocytes and T-lymphocytes by means of epitope-specific receptor molecules having a shape complementary to that of the epitope. The epitope receptor on the surface of a B-lymphocyte is called a B-cell receptor and is actually an antibody molecule . The receptor on a T-lymphocyte is called a T-cell receptor (TCR).
It is estimated that the human body has the ability to recognize 107 or more different epitopes and make up to 109 different antibodies, each with a unique specificity. In order to recognize this immense number of different epitopes, the body produces 107 or more distinct clones of both B-lymphocytes and T-lymphocytes, each with a unique B-cell receptor or T-cell receptor. Among this large variety of B-cell receptors and T-cell receptors there is bound to be at least one that has an epitope-binding site able to fit, at least to some degree, any antigen the immune system eventually encounters. With the adaptive immune responses, the body is able to recognize any conceivable antigen it may eventually encounter.
The downside to the specificity of adaptive immunity is that only a few B-cells and T-cells in the body recognize any one epitope. These few cells then must rapidly proliferate in order to produce enough cells to mount an effective immune response against that particular epitope, and that typically takes several days. During this time the pathogen could be causing considerable harm, and that is why innate immunity is also essential.
Adaptive immunity usually improves upon repeated exposure to a given infection and involves the following:
• antigen-presenting cells (APCs) such as macrophages and dendritic cells ;
• the activation and proliferation of antigen-specific B-lymphocytes ;
• the activation and proliferation of antigen-specific T-lymphocytes ; and
• the production of antibody molecules , cytotoxic T-lymphocytes (CTLs) , activated macrophages , and cytokines .
Acquired immunity includes both humoral immunity and cell-mediated immunity and will be the topic of Unit 6.
Compare and contrast how innate immunity and adaptive immunity are typically initiated in response to microbes.
We will now take a closer look at innate immunity.
Summary
1. The body has two immune systems: the innate immune system and the adaptive immune system.
2. Innate immunity is an antigen-nonspecific defense mechanisms that a host uses immediately or within several hours after exposure to almost any microbe.
3. Innate immunity is the immunity one is born with and is the initial response by the body to eliminate microbes and prevent infection.
4. Immediate innate immunity begins 0 - 4 hours after exposure to an infectious agent and involves the action of soluble preformed antimicrobial molecules that circulate in the blood and in extracellular tissue fluids.
5. Early induced innate immunity begins 4 - 96 hours after exposure to an infectious agent and involves the recruitment of defense cells as a result of pathogen-associated molecular patterns or PAMPS binding to pattern-recognition receptors or PRRs.
6. Adaptive (acquired) immunity refers to antigen-specific defense mechanisms that take several days to become protective and are designed to react with and remove a specific antigen.
7. Adaptive immunity is the immunity one develops throughout life.
8. An antigen is defined as a substance that reacts with antibody molecules and antigen receptors on lymphocytes.
9. The actual portions or fragments of an antigen that react with antibodies and lymphocyte receptors are called epitopes. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.1%3A_The_Innate_Immune_System%3A_An_Overview.txt |
State what each of the following determine: CBC and leukocyte differential count. State the significance of the following: an elevated white blood cell count a shift to the left (elevated bands) Describe and state the major functions of the following leukocytes: neutrophils basophils eosinophils monocytes B-lymphocytes T4-lymphocytes T8-lymphocytes NK cells State what type of cell monocytes differentiate into when they enter tissue. State 2 functions of platelets.
All leukocytes are critical to body defense. There are normally between 5,000-10,000 leukocytes per cubic millimeter (mm3) of blood and these can be divided into five major types: neutrophils, basophils, eosinophils, monocytes, and lymphocytes. The production of colonies of the different types of leukocytes is called leukopoiesis and is induced by various cytokines known as colony stimulating factors or CSFs .
The complete blood count (CBC) is a laboratory test which, among other things, determines the total number of both leukocytes and erythrocytes per ml of blood. In general, an elevated WBC count (leukocytosis ) is seen in infection, inflammation, leukemia, and parasitic infestations. A decreased WBC count (leukopenia ) is generally seen in bone marrow depression, severe infection, viral infections, autoimmune diseases, malignancies, and malnutrition. For example, infections may increase the total leukocyte count two to three times the normal level by dramatically increasing the number of neutrophils.
The differential white blood cell count (leukocyte differential count) determines the number of each type of leukocyte calculated as a percentage of the total number of leukocytes. This information can be useful diagnostically because different diseases or disorders can cause an increase or a decrease in the various types of WBCs. For example, when doing a differential WBC count, neutrophils are usually divided into segs (a mature neutrophile having a segmented nucleus) and bands (an immature neutrophil with an incompletely segmented or banded nucleus). During an active infection, people are generally producing large numbers of new neutrophils and therefore will have a higher percentage of the immature band forms. (An increase in band forms is sometimes referred to as a "shift to the left" because on laboratory slips used for differential WBC counts, the heading for bands is to the left of the heading for mature neutrophils or segs.)
The five types of leukocytes fall into one of two groups: the polymorphonuclear leukocytes and the mononuclear leukocytes.
Polymorphonuclear Leukocytes
Polymorphonuclear leukocytes (granulocytes) have irregular shaped nuclei with several lobes and their cytoplasm is filled with granules containing enzymes and antimicrobial chemicals. They include the following:
Neutrophils
Neutrophils are the most abundant of the leukocytes, normally accounting for 54-75% of the WBCs. An adult typically has 3,000-7,500 neutrophils/mm3 of blood but the number may increase two- to three-fold during active infections. They are called neutrophils because their granules stain poorly - they have a neutral color - with the mixture of dyes used in staining leukocytes. The nucleus of a neutrophil has multiple lobes.
Neutrophils are important phagocytes. Their granules contain various agents for killing microbes. Primary azurophil granules contain acid hydrolase, myeloperoxidase, defensins, cathepsin G, cationic proteins, and bactericidal permeability increasing protein (BPI ). Secondary specific granules contain such defense chemicals as lysozyme, lactoferrin, collagenase, and elastase. These agents kill microbes intracellularly during phagocytosis but are also often released extracellularly where they kill not only microbes but also surrounding cells and tissue, as will be discussed later under phagocytosis.
They release the enzyme kallikrein that catalyzes the generation of bradykinins. Bradykinins promote inflammation by causing vasodilation, increasing vascular permeability, and increasing mucous production. They are also chemotactic for leukocytes and stimulate pain. They produce enzymes that catalyze the synthesis of prostaglandins from arachidonic acid in cell membranes. Certain prostaglandins promote inflammation by causing vasodilation and increasing capillary permeability. They also cause constriction of smooth muscles, enhance pain, and induce fever.
They are short-lived, having a life span of a few hours to a few days, and do not multiply. They circulate in the blood for around 6 hours and if the are not recruited, they undergo apoptosis. In tissue, they function for several hours and die. However, the bone marrow makes about 80,000,000 new neutrophils per minute to replace these.
• To view an electron micrograph of a neutrophil, see the Web page for the University of Illinois College of Medicine.
• Scanning electron micrograph of a neutrophil engulfing Escherichia coli from sciencephotogallery.com.
• Transmission electron micrograph of a neutrophil engulfing Neisseria gonorrhoeae from sciencephotogallery.com.
Eosinophils
Eosinophils normally comprise 1-4% of the WBCs (50-400/mm3 of blood). They are called eosinophils because their granules stain red with the acidic dye eosin, one of the mixture of dyes used when staining leukocytes. The nucleus of an eosinophil typically appears lobed.
Their granules contain destructive enzymes for killing infectious organisms. These enzymes include acid phosphatase, peroxidases, major basic protein, RNase, DNases, lipase, and plasminogen. They are capable of phagocytosis but primarily they release their contents into the surrounding environment to kill microbes extracellularly. The substances they release defend primarily against fungi, protozoa, and parasitic worms (helminths), pathogens that are too big to be consumed by phagocytosis. They secrete leukotrienes, prostaglandins, chemicals that promotes inflammation by causing vasodilation and increasing capillary permeability. They also secrete various cytokines such as IL-1, IL-2, IL-4, IL-5, IL-6, IL-8, IL-13, and TNF alpha. Their life span is 8-12 days.
• To view an electron micrograph of an eosinophil, see the Web page for the University of Illinois College of Medicine.
• Transmission electron micrograph of an eosinophil from sciencephotogallery.com.
Basophils
Basophils normally make up 0-1% of the WBCs (25-100/mm3 of blood). They are called basophils because their granules stain a dark purplish blue with the basic dye methylene blue, one of the dyes that are used when staining leukocytes. Basophils have a lobed nucleus. Basophils release histamine, leukotrienes, and prostaglandins, chemicals that promotes inflammation by causing vasodilation, increasing capillary permeability, and increasing mucous production. Basophils also produce heparin, platelet-activating factor (PAF) and the cytokine IL-4. Their life span is probably a few hours to a few days.
Mononuclear Leukocytes
Mononuclear leukocytes (agranulocytes) have compact nuclei and have no visible cytoplasmic granules. The following are agranulocytes:
Monocytes
Monocytes normally make up 2-8% of the WBCs (100-500/mm3 of blood). Monocytes are important phagocytes. Monocytes differentiate into macrophages and dendritic cells when they leave the blood and enter the tissue. Macrophages and dendritic cells are very important in phagocytosis and serve as antigen-presenting cells in the adaptive immune responses (see below). They produce a variety of cytokines that play numerous roles in body defense. They are long-lived (life span of months) and can multiply.
• To view an electron micrograph of a monocyte, see the Web page for the University of Illinois College of Medicine.
• Transmission electron micrograph of a monocyte from sciencephotogallery.com.
Lymphocytes
Lymphocytes normally represent 25-40% of the WBCs (1,500-4,500/mm3 of blood). Lymphocytes mediate the adaptive immune responses (Unit 6). Only a small proportion of the body's lymphocytes are found in the blood. The majority are found in lymphoid tissue. In fact the collective mass of all the lymphocytes in the human body is about the same as the mass of the brain! Lymphocytes circulate back and forth between the blood and the lymphoid system of the body. They have a life span of days to years. There are 3 major populations of lymphocytes:
B-lymphocytes (B-cells) mediate humoral immunity, the production of antibody molecules against a specific antigen,and have B-cell receptors (BCR) on their surface for antigen recognition. Generally 10-20% of the lymphocytes are B-lymphocytes. Once activated, most B-lymphocytes differentiate into antibody-secreting plasma cells.
T-lymphocytes (T-cells) are responsible for cell-mediated immunity, the production of cytotoxic T-lymphocytes (CTLs), activated macrophages, activated NK cells, and cytokines against a specific antigen. They also regulate the adaptive immune responses. Generally 60-80% of the lymphocytes are T-lymphocytes. Based on biochemical markers on their surface, there are two major classes of T-lymphocytes:
• T4-lymphocytes (CD4+ T-lymphocytes) have CD4 molecules and T-cell receptors (TCRs) on their surface for protein antigen recognition. They function to regulate the adaptive immune responses through cytokine production. Once activated, they differentiate into effector T4-lymphocytes such as Th1 cells, Th2 cells, and Th17 cells.
• T8-lymphocytes (CD8+ T-lymphocytes) have CD8 molecules and T-cell receptors (TCRs) on their surface for protein antigen recognition. Once activated, they differentiate into cytotoxic T-lymphocytes (CTLs ).
Invariant natural killer T (iNKT) cells are a subset of lymphocytes that bridge the gap between innate and adaptive immunity. They have T-cell receptors (TCRs) on their surface for glycolipid antigen recognition. Through the cytokines they produce once activated,i NKT cells are essential in both innate and adaptive immune protection against pathogens and tumors. They also play a regulatory role in the development of autoimmune diseases and transplantation tolerance.
NK cells (natural killer cells ) are lymphocytes that lack B-cell receptors and T-cell receptors. They function to kill infected cells and tumor cells. NK cells are able to kill cells to which antibody molecules have attached through a process called antibody-dependent cellular cytotoxicity (ADCC). They also kill human cells lacking MHC-I molecules on their surface. Lymphocytes will be discussed in greater detail in Unit 6.
Although not white blood cells, platelets (thrombocytes) are another formed element in the blood. They promote clotting by sticking together after becoming activated and forming platelet plugs to close up damaged capillaries. They also secrete cytokines and chemokines to promote inflammation.
Summary
1. The complete blood count (CBC) is a laboratory test that, among other things, determines the total number of both leukocytes and erythrocytes per ml of blood.
2. In general, an elevated WBC count (leukocytosis ) is seen in infection, inflammation, leukemia, and parasitic infestations.
3. Neutrophils are the most abundant of the leukocytes, normally accounting for 54-75% of the WBCs. Neutrophils are important phagocytes and also promote inflammation.
4. Eosinophils normally comprise 1-4% of the WBCs. They are capable of phagocytosis but primarily they release their contents into the surrounding environment to kill microbes, especially parasitic worms, extracellularly. They also promote inflammation.
5. Basophils normally make up 0-1% of the WBCs and release histamine, leukotrienes, and prostaglandins, chemicals that promotes inflammation.
6. Monocytes normally make up 2-8% of the WBCs and differentiate into macrophages and dendritic cells when they leave the blood and enter the tissue.
7. Lymphocytes normally represent 25-40% of the WBCs and mediate the specific immune responses.
8. B-lymphocytes (B-cells) mediate humoral immunity, the production of antibody molecules against a specific antigen, and have B-cell receptors (BCR) on their surface for antigen recognition. Most B-lymphocytes differentiate into antibody-secreting plasma cells.
9. T-lymphocytes (T-cells) are responsible for cell-mediated immunity, the production of cytotoxic T-lymphocytes (CTLs), activated macrophages, activated NK cells, and cytokines against a specific antigen.
10. T4-lymphocytes have CD4 molecules and T-cell receptors on their surface for antigen recognition. They function to regulate the adaptive immune responses through cytokine production. Once activated, they differentiate into effector T4-lymphocytes.
11. T8-lymphocytes have CD8 molecules and T-cell receptors on their surface for antigen recognition. Once activated, they differentiate into T8-suppressor cells and cytotoxic T-lymphocytes (CTLs).
12. NK cells (natural killer cells) are lymphocytes that lack B-cell receptors and T-cell receptors. They function to kill infected cells and tumor cells. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.2%3A_Defense_Cells_in_the_Blood%3A_The_Leukocytes.txt |
Learning Objectives
1. State 3 different functions of macrophages in body defense.
2. State the primary function of dendritic cells in body defense.
3. Name the cells in the tissue whose primary function is to present antigen to naive T-lymphocytes.
4. Name the cells in the tissue whose primary function is to present antigen to effector T-lymphocytes.
5. State the primary function of mast cells in body defense.
Dendritic Cells
Most dendritic cells are derived from monocytes and are referred to as myeloid dendritic cells. They are located throughout the epithelium of the skin, the respiratory tract, and the gastrointestinal tract, as well as lymphoid tissues and organ parenchyma. In these locations, in their immature form, they are attached by long cytoplasmic processes. Upon capturing antigens through pinocytosis and phagocytosis and becoming activated by inflammatory cytokines, the dendritic cells detach from their initial site, enter lymph vessels, and are carried to regional lymph nodes. By the time they enter the lymph nodes, they have matured and are now able to present antigen to the ever changing populations of naive T-lymphocytes located in the cortex of the lymph nodes.
The primary function of dendritic cells is to capture and present protein antigens to naive T-lymphocytes. (Naive lymphocytes are those that have not yet encountered an antigen.) Dendritic cells engulf microorganisms and other materials and degrade them with their lysosomes. Peptides from microbial proteins are then bound to a groove of unique molecules called MHC-II molecules produced by macrophages, dendritic cells, and B-lymphocytes. The peptide epitopes bound to the MHC-II molecules are then put on the surface of the dendritic cell (Figure \(1\)) where they can be recognized by complementary shaped T-cell receptors (TCR) and CD4 molecules on naive T4-lymphocyte (see Figure \(2\)).
In addition, dendritic cells can bind peptide epitopes to MHC-I molecules and present them to naiveT8-lymphocytes. The MHC-I molecules with bound peptide on the dendritic cell are recognized by complementary shaped T-cell receptors (TCR) and CD8 molecules on naive T8-lymphocyte (Figure \(3\)).
A dendritic cell. (CC BY-SA 2.5; Judith Behnsen, Priyanka Narang, Mike Hasenberg, Frank Gunzer, Ursula Bilitewski, Nina Klippel, Manfred Rohde, Matthias Brock, Axel A. Brakhage, Matthias Gunzer - Source: PLoS Pathogens ).
These interactions enable the T4-lymphocytes or T8-lymphocytes to become activated, proliferate, and differentiate into effector cells. This will be discussed in detail in Unit 6. Myeloid dendritic cells also use pattern-recognition receptors called toll-like receptors (TLRs) to recognize pathogen-associated molecular patterns or PAMPs (Figure \(4\)). The interaction of the PAMP with its TLR stimulates the production of co-stimulatory molecules that are also required for T-lymphocyte activation. Dendritic cells produce many of the same inflammatory cytokines as macrophages, such as tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), interleukin-6 (IL-6), and interleukin-8 (IL-8). They also can produce interleukin-12 (IL-12), a cytokine that can activate natural killer T-lymphocytes (NKT cells).
Another type of dendritic cell, the plasmacytoid dendritic cell, uses its TLRs to recognize viral PAMPs. This interaction results in the production and secretion of type I interferons. Antigen-presenting cells or APCs will be discussed in greater detail in Unit 6.
Macrophages
When monocytes leave the blood and enter the tissue, they become activated and differentiate into macrophages. Those that have recently left the blood during inflammation and move to the site of infection through positive chemotaxis are sometimes referred to as wandering macrophages. In addition, the body has macrophages already stationed throughout all tissues and organs of the body. These are sometimes referred to as fixed macrophages.
Many fixed macrophages are part of the mononuclear phagocytic (reticuloendothelial) system. They, along with B-lymphocytes and T-lymphocytes, are found supported by reticular fibers in lymph nodules, lymph nodes, and the spleen where they filter out and phagocytose foreign matter such as microbes. Similar cells are also found in the liver (Kupffer cells), the kidneys (mesangial cells), the brain (microglia), the bones (osteoclasts), the lungs (alveolar macrophages), and the gastrointestinal tract (peritoneal macrophages). Macrophages actually have a number of very important functions in body defense including:
Function 1
Killing of microbes, infected cells, and tumor cells by phagocytosis. Macrophages that have engulfed microorganisms become activated by a subset of T-helper lymphocytes called Th1 cells (Figure \(6\)). Activated macrophages develop a ruffled cytoplasmic membrane and produce increased numbers of lysosomes.
Function 2
Processing antigens so they can be recognized by effector T-lymphocytes during the adaptive immune responses. Macrophages, as well as the dendritic cells mentioned below, process antigens through phagocytosis and present them to T-lymphocytes. Because of this function, they are often referred to as antigen-presenting cells or APCs.
Macrophages primarily capture and present protein antigens to effector T-lymphocytes. (Effector lymphocytes are lymphocytes that have encountered an antigen, proliferated, and matured into a form capable of actively carrying out immune defenses.) Macrophages engulf the microorganism and degrade it with their lysosomes. Peptides from microbial proteins are then bound to a groove of unique molecules called MHC-II molecules produced by macrophages, dendritic cells, and B-lymphocytes. The peptide epitopes bound to the MHC-II molecules are then put on the surface of the macrophage (Figure \(1\)) where they can be recognized by complementary shaped T-cell receptors (TCR) and CD4 molecules on an effector T4-lymphocyte (Figure \(2\)). This interaction leads to the activation of that macrophage.
Like dendritic cells discussed above, macrophages are also capable of capturing and presenting protein antigens to naive T-lymphocytes although they are not as important in this function.
Function 3
Secreting lipid mediators of inflammation such as leukotrienes, prostaglandins, and platelet-activating factor (PAF).
Function 4
Secreting proteins called cytokines that play a variety of roles in non-specific body defense. Macrophage-produced cytokines promote inflammation and induce fever, increase phagocytosis and energy output, promote sleep, activate resting T-lymphocytes , attract and activate neutrophils, and stimulate the replication of endothelial cells to form capillaries and fibroblasts to form connective scar tissue. Four important cytokines that macrophages produce (as mentioned in Unit 1 under endotoxin) are tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), interleukin-6 (IL-6), and interleukin-8 (IL-8).
There is growing evidence that monocytes and macrophages can be “trained” by an earlier infection to do better in future infections, that is, develop memory. It is thought that microbial pathogen-associated molecular patterns (PAMPs) binding to pattern-recognition (PRRs) on monocytes and macrophages triggers the cell’s epigenome to reprogram or train that cell to react better against new infections.
Macrophages show great functional diversity. In addition to the populations of macrophages involved in body defense and immunity, there are populations of macrophages that play important roles in:
1. The development of a variety of tissues and organs within the body, including the brain, blood cells, mammary gland, pancreas, and kidneys.
2. Modulating normal physiology and maintaining homeostasis in the body, including insulin resistance and sensitivity, long term nutrient storage, thermogenesis, and liver and pancreas function in response to caloric intake.
3. Tissue repair, including the formation of scar tissue and the growth of new capillaries into injured tissues.
Mast Cells
Mast cells are typically the immunological first responders to infection and carry out many of the same inflammatory-mediating functions as basophils. There are two types of mast cells in the body: mast cells found in the connective tissue and mast cells found throughout the mucous membranes. The granules of mast cells contain such mediators as histamine, eosinophil chemotactic factor, neutrophil chemotactic factor, platelet activating factor, and cytokines such as IL-3, IL-4, IL-5, IL-6, and TNF-alpha. They also possess pathways for synthesizing leukotrienes and prostaglandins, chemicals that promote inflammation by causing vasodilation, increasing capillary permeability, and increasing mucous production.
Photo of cultured mast cells at 100X using an oil immersion lens and an olympus digital camera. The cells are stained with Tol Blue, and might appear slightly degranulated as they were activated using an artificial antigen during the course of an experiment. Image use with permission (Kauczuk).
Mast cells have pattern-recognition receptors or PRRs on their surface that interact with pathogen-associated molecular patterns or PAMPs of microbes. After the PAMPs bind to their respective PRRs, they release the contents of their granules. These chemical mediators promote inflammation and attract neutrophils to the infected site.
Summary
1. Most dendritic cells are derived from monocytes and are referred to as myeloid dendritic cells and are located throughout the epithelium of the skin, the respiratory tract, and the gastrointestinal tract, as well as lymphoid tissues and organ parenchyma.
2. Upon capturing antigens through pinocytosis and, the dendritic cells detach from their initial site, enter lymph vessels, and are carried to regional lymph nodes where they present antigens to the ever changing populations of naive T-lymphocytes.
3. The primary function of dendritic cells is to capture and present protein antigens to naive T-lymphocytes.
4. When monocytes leave the blood and enter the tissue, many become activated and differentiate into macrophages. These macrophages that have recently left the blood during inflammation and move to the site of infection through positive chemotaxis are sometimes referred to as wandering macrophages.
5. The body has macrophages already stationed throughout the tissues and organs of the body and these are sometimes referred to as fixed macrophages.
6. Functions of macrophages include killing of microbes, infected cells, and tumor cells by phagocytosis, processing antigens so they can be recognized by effector T-lymphocytes during the adaptive immune responses, and secreting mediators of inflammation such as leukotrienes, prostaglandins, and platelet-activating factor, and cytokines.
7. Mast cells are typically the immunological first responders to infection and carry out many of the same inflammatory-mediating functions as basophils. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.3%3A_Defense_Cells_in_the_Tissue_-_Dendritic_Cells_Macrophages_and_Mast_Cells.txt |
Immediate innate immunity begins 0-4 hours after exposure to an infectious agent and involves the action of soluble preformed antimicrobial molecules that circulate in the blood, our found in extracellular tissue fluids, and are secreted by epithelial cells. These include: antimicrobial enzymes and peptides, and complement system proteins. These preformed antimicrobial molecules are designed to immediately begin to remove infectious agents as soon as they enter the body. In addition to preformed antimicrobial molecules, the following also play a role in immediate innate immunity: anatomical barriers to infection, mechanical removal of microbes, and bacterial antagonism by the body's normal microbiota
11.3: Immediate Innate Immunity
State how long it takes for immediate innate immunity to become activated and what it involves. State the function of the following antimicrobial enzymes and peptides: lysozyme phospholipase A2 defensins cathelicidins lactotransferrin and transferrin
Examples include:
a. Lysozyme , found in in tears, mucous, saliva, plasma , tissue fluid, etc., breaks down peptidoglycan in bacteria causing osmotic lysis. Specifically, it breaks the bond between the N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM), the two sugars that make up the backbone of peptidoglycan (see Figure \(1\)).
b. Phospholipase A2 is an enzyme that penetrates the bacterial cell wall and hydrolyzes the phospholipids in the bacterial cytoplasmic membrane.
c. Human defensins ) are short cationic peptides 30-40 amino acids long that are directly toxic by disrupting the cytoplasmic membrane of a variety of microorganisms causing leakage of cellular needs (see Figure \(2\)). They also activate cells for an inflammatory response. Defensins are produced by leukocytes, epithelial cells, and other cells. They are also found in blood plasma and mucus. Certain defensins also disrupt the envelopes of some viruses.
d. Cathelicidins are proteins produced by skin and mucosal epithelial cells. The two peptides produced upon cleavage of the cathelicidin are directly toxic to a variety of microorganisms. One pepitide also can bind to and neutralize LPS from Gram-negative cell walls to reduce inflammation.
e. Lactic and fatty acids, found in perspiration and sebaceous secretions , inhibit microbes on the skin.
f. Lactoferrin and transferrin , found in body secretions, plasma, and tissue fluid, trap iron for use by human cells while preventing its use by microorganisms.
g. Hydrochloric acid and enzymes found in gastric secretions destroy microbes that are swallowed.
Keep in mind that in Unit 3 under "Virulence Factors that Promote Bacterial Colonization of the Host" we learned several mechanisms that various bacteria use to resist the body's antibacterial peptides. By resisting these immediate innate immune defenses, some bacteria have a better chance of colonizing their host.
Summary
1. Immediate innate immunity begins 0-4 hours after exposure to an infectious agent and involves the action of soluble preformed antimicrobial molecules that circulate in the blood and are found in extracellular tissue fluids.
2. Lysozyme, found in in tears, mucous, saliva, plasma, tissue fluid, etc., breaks down peptidoglycan in bacteria causing osmotic lysis.
3. Phospholipase A2 is an enzyme that penetrates the bacterial cell wall and hydrolyzes the phospholipids in the bacterial cytoplasmic membrane.
4. Human defensins are short cationic peptides 30-40 amino acids long that are directly toxic by disrupting the cytoplasmic membrane of a variety of microorganisms causing leakage of cellular needs.
5. Cathelicidins are proteins produced by skin and mucosal epithelial cells that are directly toxic to a variety of microorganisms.
6. Lactoferrin and transferrin, found in body secretions, plasma, and tissue fluid, trap iron for use by human cells while preventing its use by microorganisms. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.3%3A_Immediate_Innate_Immunity/11.3A%3A_Antimicrobial_Enzymes_and_Antimicrobial_Peptides.txt |
Briefly describe how the classical complement pathway is activated. Briefly describe the beneficial effects of the following complement pathway products: C5a C3a C3b C4b C3d C5b6789n (MAC) Briefly describe how the lectin pathway is activated. Briefly describe how the alternative complement pathway is activated.
In this section we will look at how the body's complement system functions to remove infectious agents. The complement system refers to a series of more than 30 soluble, preformed proteins circulating in the blood and bathing the fluids surrounding tissues. The proteins circulate in an inactive form, but in response to the recognition of molecular components of microorganism, they become sequentially activated, working in a cascade where in the binding of one protein promotes the binding of the next protein in the cascade. There are 3 complement pathways that make up the complement system: the classical complement pathway, the lectin pathway, and the alternative complement pathway. The pathways differ in the manner in which they are initiated and ultimately produce a key enzyme called C3 convertase:
• The classical complement pathway is initiated by activation of C1. C1 is primarily activated by interacting with the Fc portion of the antibody molecules IgG or IgM after they have bound to their specific antigen. C1 is also able to directly bind to the surfaces of some pathogens as well as with the C-reactive protein (CRP) that is produced during the acute phase response of innate immunity.
• The lectin pathway is activated by the interaction of microbial carbohydrates (lectins) with mannose-binding lectin (MBL) or ficolins found in the plasma and tissue fluids.
• The alternative complement pathway is activated by C3b binding to microbial surfaces and to antibody molecules.
The end results and defense benefits of each pathway, however, are the same. All complement pathways carry out 6 beneficial innate defense functions. Proteins produced by the complement pathways:
1. Trigger inflammation,
2. Chemotactically attract phagocytes to the infection site,
3. Promote the attachment of antigens to phagocytes (enhanced attachment or opsonization),
4. Cause lysis of Gram-negative bacteria, human cells displaying foreign epitopes,and viral envelopes,
5. Play a role in the activation of naive B-lymphocytes during adaptive immunity, and
6. Remove harmful immune complexes from the body.
We will now look at each of these complement pathways and see how they function to protect the body.
The Classical Complement Pathway
The classical complement pathway is primarily activated when a complement protein complex called C1 interacts with the Fc portion of the antibody molecules IgG or IgM after they have bound to their specific antigen via their Fab portion. C1 is also able to directly bind to the surfaces of some pathogens as well as with the C-reactive protein (CRP) that is produced during the acute phase response of innate immunity. The C1 complex is composed of three complement proteins called C1q, C1r, and C1s.
C1 is also able to directly bind to the surfaces of some pathogens as well as with the C-reactive protein (CRP) that is produced during the acute phase response of innate immunity.
2. The binding of C1q activates the C1r portion of C1 which, in turn, activates C1s. This activation gives C1s enzymatic activity to cleave complement protein C4 into C4a and C4b (see Figure \(2\)A and Figure \(2\)B).
3. C2 then binds to C4b and is cleaved by C1 into C2a and C2b (see Figure \(3\)A and Figure \(3\)B).
4. C4b and C2a combine to form C4b2a, the C3 convertase. C3 convertase can now cleave hundreds of molecules of C3 into C3a and C3b (see Figure \(4\)).
5. Some molecules of C3b bind to C4b2a, the C3 convertase, to form C4b2a3b, a C5 convertase that cleaves C5 into C5a and C5b (see Figure \(5\)).
6. C5b binds to the surface of the target cell and subsequently binds C6, C7, C8, and a number of monomers of C9 to form C5b6789n, the Membrane Attack Complex (MAC) (see Figure \(6\) and Figure \(7\)).
As mentioned above, proteins of the complement pathways carry out 6 beneficial innate defense functions. These include:
1. Triggering inflammation: C5a is the most potent complement protein triggering inflammation. It reacts with blood vessels causing vasodilation. It also causes mast cells to release vasodilators such as histamine,increasing blood vessel permeability as well as increasing the expression of adhesion molecules on leukocytes and the vascular endothelium so that leukocytes can squeeze out of the blood vessels and enter the tissue (diapedesis). C5a also causes neutrophils to release toxic oxygen radicals for extracellular killing and induces fever. To a lesser extent C3a and C4a also promote inflammation. As we will see later in this unit, inflammation is a process in which blood vessels dilate and become more permeable, thus enabling body defense cells and defense chemicals to leave the blood and enter the tissues.
2. Chemotactically attracting phagocytes to the infection site: C5a also functions as a chemoattractant for phagocytes. Phagocytes will move towards increasing concentrations of C5a and subsequently attach, via their CR1 receptors to the C3b molecules attached to the antigen. This will be discussed in greater detail later in this unit under phagocytosis.
3. Promoting the attachment of antigens to phagocytes (enhanced attachment or opsonization): C3b and to a lesser extent, C4b can function as opsonins, that is, they can attach antigens to phagocytes. One portion of the C3b binds to proteins and polysaccharides on microbial surfaces; another portion attaches to CR1 receptors on phagocytes, B-lymphocytes, and dendritic cells for enhanced phagocytosis. (see Figure \(8\)). In actuality, C3b molecule can bind to pretty much any protein or polysaccharide. Human cells, however, produce Factor H that binds to C3b and allows Factor I to inactivate the C3b. On the other hand, substances such as LPS on bacterial cells facilitate the binding of Factor B to C3b and this protects the C3b from inactivation by Factor I. In this way, C3b does not interact with our own cells but is able to interact with microbial cells. C3a and C5a increase the expression of C3b receptors on phagocytes and increase their metabolic activity.
4. Causing lysis of Gram-negative bacteria, human cells displaying foreign epitopes,and viral envelopes: C5b6789n, functions as a Membrane Attack Complex (MAC). This helps to destroy gram-negative bacteria as well as human cells displaying foreign antigens (virus-infected cells, tumor cells, etc.) by causing their lysis; see Figure \(6\) and Figure \(7\). It can also damage the envelope of enveloped viruses.
5. Serving as a second signal for activating naive B-lymphocytes during adaptive immunity: Some C3b is converted to C3d. C3d binds to CR2 receptors on B-lymphocytes. This serves as a second signal for the activation of B-lymphocytes whose B-cell receptors have just interacted with their corresponding antigen.
6. Removing harmful immune complexes from the body: C3b and to a lesser extent, C4b help to remove harmful immune complexes from the body. The C3b and C4b attach the immune complexes to CR1 receptors on erythrocytes. The erythrocytes then deliver the complexes to fixed macrophages within the spleen and liver for destruction. Immune complexes can lead to a harmful Type III hypersensitivity, as will be discussed later in Unit 5 under Hypersensitivities.
The Lectin Pathway
The lectin pathway is activated by the interaction of microbial carbohydrates with mannose-binding lectin (MBL) or ficolins found in the plasma and tissue fluids. (Lectins are carbohydrate-binding proteins.) The lectin pathway is mediated by two groups of proteins found in the plasma of the blood and in tissue fluids:
1. Mannose-binding lectin (MBL) - also known as mannose-binding protein or MBP. MBL is a soluble pattern-recognition receptor that binds to various microbial carbohydrates such as those rich in mannose or fucose, and to N-acetylglucosamine (NAG). These glycans are common in microbial glycoproteins and glycolipids but rare in those of humans. MBL is synthesized by the liver and released into the bloodstream as part of the acute phase response that will be discussed later in this unit. The MBL is equivalent to C1q in the classical complement pathway.
Ficolins are similar in their structure to MBL and bind to microbial carbohydrates such as N-acetylglucosamine (NAG), lipoteichoic acids, and lipopolysaccharide (LPS). Ficolin is also equivalent to C1q in the classical complement pathway.
2. Both mannose-binding lectin (MBL) and ficolin form complexes with MBL-associated serine proteases called MASP1 and MASP2, which are equivalent to C1r and C1s of the classical pathway.
a. The binding of the MBL (or the ficolin) to the microbial carbohydrate activates the associated MASP2 giving it the enzymatic activity to split C4 into C4a and C4b (see Figure \(9\)A and Figure \(9\)B).
b. C2 then binds to C4b and is cleaved by MASP2 into C2a and C2b (see Figure \(10\)A and Figure \(10\)B).
c. C4b and C2a combine to form C4b2a, the C3 convertase. C3 convertase can now cleave hundreds of molecules of C3 into C3a and C3b (see Figure \(11\)).
d. Some molecules of C3b bind to C4b2a, the C3 convertase, to form C4b2a3b, a C5 convertase that cleaves C5 into C5a and C5b (see Figure \(12\)).
e. C5b binds to the surface of the target cell and subsequently binds C6, C7, C8, and a number of monomers of C9 to form C5b6789n, the Membrane Attack Complex (MAC) (see Figure \(6\) and Figure \(7\)).
The beneficial results of the activated complement proteins are the same as in the classical complement pathway above. The complement proteins:
1. Trigger inflammation : C5a>C3a>c4a;
2. Chemotactically attract phagocytes to the infection site: C5a;
3. Promote the attachment of antigens to phagocytes via enhanced attachment or opsonization : C3b>C4b;
4. Cause lysis of Gram-negative bacteria and human cells displaying foreign epitopes : MAC;
5. Serve as a second signal for the activation of naive B-lymphocytes ): C3d; and
6 Remove harmful immune complexes from the body: C3b>C4b.
The Alternative Complement Pathway
The alternative complement pathway is mediated by C3b, produced either by the classical or lectin pathways or from C3 hydrolysis by water. (Water can hydrolyze C3 and form C3i, a molecule that functions in a manner similar to C3b.)
Activation of the alternative complement pathway begins when C3b (or C3i) binds to the cell wall and other surface components of microbes. C3b can also bind to IgG antibodies. Alternative pathway protein Factor B then combines with the cell-bound C3b to form C3bB. Factor D then splits the bound Factor B into Bb and Ba, forming C3bBb. A serum protein called properdin then binds to the Bb to form C3bBbP that functions as a C3 convertase (see Figure \(13\)) capable of enzymatically splitting hundreds of molecules of C3 into C3a and C3b. The alternative complement pathway is now activated.
Some of the C3b subsequently binds to some of the C3bBb to form C3bBb3b, a C5 convertase capable of splitting molecules of C5 into C5a and C5b (see Figure \(14\)). From here, the alternative complement pathway is identical to the other complement pathways.
The beneficial results are the same as in the classical complement pathway above. The complement proteins:
1. Trigger inflammation : C5a>C3a>c4a;
2. Chemotactically attract phagocytes to the infection site: C5a;
3. Promote the attachment of antigens to phagocytes via enhanced attachment or opsonization : C3b>C4b;
4. Cause lysis of Gram-negative bacteria, human cells displaying foreign epitopes,and viral envelopes: MAC; and
5. Serve as a second signal for the activation of naive B-lymphocytes ): C3d;
6. Remove harmful immune complexes from the body: C3b>C4b.
Keep in mind that in Unit 3, we learned several mechanisms that various bacteria use to resist the body's complement pathways. By resisting these immediate innate immune defenses, some bacteria have a better chance of colonizing their host.
Summary
1. The proteins of the complement system circulate in an inactive form, but in response to the recognition of molecular components of microorganism, they become sequentially activated, working in a cascade where in the binding of one protein promotes the binding of the next protein in the cascade.
2. There are 3 complement pathways that make up the complement system: the classical complement pathway, the lectin pathway, and the alternative complement pathway.
3. The classical complement pathway is initiated by activation of C1. C1 is primarily activated by interacting with the Fc portion of the antibody molecules IgG or IgM after they have bound to their specific antigen. C1 is also able to directly bind to the surfaces of some pathogens as well as with the C-reactive protein (CRP) that is produced during the acute phase response of innate immunity.
4. The lectin pathway is activated by the interaction of microbial carbohydrates (lectins) with mannose-binding lectin (MBL) or ficolins found in the plasma and tissue fluids.
5. The alternative complement pathway is activated by C3b binding to microbial surfaces and to antibody molecules.
6. All complement pathways carry out the same 6 beneficial innate defense functions.
7. The complement proteins C5a and, to a lesser extent, C3a, and C4a trigger vasodilation and inflammation in order to deliver defense cells and defense chemicals to the infection site.
8. The complement protein C5a also functions as a chemoattractant for phagocytes.
9. The complement proteins C3b and to a lesser extent, C4b can function as opsonins, that is, they can attach antigens to phagocytes.
10. The complement proteins C5b6789n, functions as a Membrane Attack Complex (MAC) causing lysis of Gram-negative bacteria, human cells displaying foreign epitopes, and viral envelopes.
11. The complement protein C3d serves as a second signal for activating naive B-lymphocytes during adaptive immunity.
12. The complement proteins C3b and to a lesser extent, C4b help to remove harmful immune complexes from the body. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.3%3A_Immediate_Innate_Immunity/11.3B%3A_The_Complement_System.txt |
Describe what is meant by anatomical barriers to infection. List 4 ways in which the body can physically remove microorganisms or their products. Briefly describe how intraepithelial T-lymphocytes and B-1 cells play a role in innate immunity. Describe how bacterial antagonism by normal microbiota acts as a non-specific body defense mechanism and name 2 opportunistic microbes that may cause superinfection upon destruction of the normal microbiota. Briefly describe the process involved in the development of antibiotic-associated colitis.
Anatomical barriers are tough, intact barriers that prevent the entry and colonization of many microbes. Examples include the skin, the mucous membranes, and bony encasements.
The skin
The skin, consisting of the epidermis and the dermis, is dry, acidic, and has a temperature lower than 37 degrees Celsius (body temperature). These conditions are not favorable to bacterial growth. Resident normal microbiota of the skin also inhibits potentially harmful microbes. In addition, the dead, keratinized cells that make up the surface of the skin are continuously being sloughed off so that microbes that do colonize these cells are constantly being removed. Hair follicles and sweat glands produce lysozyme and toxic lipids that can kill bacteria. Epithelial cells also produce defensins and cathelicidins to kill microbes. Beneath the epidermis of the skin are Langerhans' cells - immature dendritic cells - that phagocytose and kill microbes, carry them to nearby lymph nodes, and present antigens of these microbes to T-lymphocytes to begin adaptive immune responses against them. Finally, intraepithelial T-lymphocytes and B-1 lymphocytes are associated with the epidermis and the mucosal epithelium. These cells recognize microbes common to the epidermis and mucous membranes and start immediate adaptive immune responses against these commonly encountered microbes.
The mucous membranes
Mucous membranes line body cavities that open to the exterior, such as the respiratory tract, the gastrointestinal tract, and the genitourinary tract. Mucous membranes are composed of an epithelial layer that secretes mucus, and a connective tissue layer. The mucus is a physical barrier that traps microbes. Mucus also contains lysozyme to degrade bacterial peptidoglycan, an antibody called secretory IgA that prevents microbes from attaching to mucosal cells and traps them in the mucous, lactoferrin to bind iron and keep it from from being used by microbes, and lactoperoxidase to generate toxic superoxide radicals that kill microbes. Resident normal microbiota of the mucosa also inhibits potentially harmful microbes. In addition, the mucous membrane, like the skin, is constantly sloughing cells to remove microbes that have attached to the mucous membranes. Beneath the mucosal membrane is mucosa-associated lymphoid tissue (MALT) that contains Langerhans' cells - immature dendritic cells - that phagocytose and kill microbes, carry them to nearby lymph nodes, and present antigens of these microbes to T-lymphocytes to begin adaptive immune responses against them. Intraepithelial T-lymphocytes and B-1 lymphocytes are associated with the epidermis and the mucosal epithelium. These cells recognize microbes common to the epidermis and mucous membranes and start immediate adaptive immune responses against these commonly encountered microbes.
Bony encasements
Bony encasements, such as the skull and the thoracic cage, protect vital organs from injury and entry of microbes.
Mechanical removal is the process of physically flushing microbes from the body. Methods include:
1. Mucus and cilia: Mucus traps microorganisms and prevents them from reaching and colonizing the mucosal epithelium. Mucus also contains lysozyme to degrade bacterial peptidoglycan, an antibody called secretory IgA that prevents microbes from attaching to mucosal cells and traps them in the mucus, lactoferrin to bind iron and keep it from from being used by microbes, and lactoperoxidase to generate toxic superoxide radicals that kill microbes. Cilia on the surface of the epithelial cells propel mucus and trapped microbes upwards towards the throat where it is swallowed and the microbes are killed in the stomach. This is sometimes called the tracheal toilet.
2. The cough and sneeze reflex: Coughing and sneezing removes mucus and trapped microbes.
3. Vomiting and diarrhea: These processes remove pathogens and toxins in the gastrointestinal tract.
4. he physical flushing action of body fluids: Fluids such as urine, tears, saliva, perspiration, and blood from injured blood vessels also flush microbes from the body.
Bacterial Antagonism by Normal Microbiota
Approximately 100 trillion bacteria and other microorganisms reside in or on the human body. The normal body microbiota keeps potentially harmful opportunistic pathogens in check and also inhibits the colonization of pathogens by:
1. Producing metabolic products (fatty acids, bacteriocins, etc.) that inhibit the growth of many pathogens;
2. Adhering to target host cells so as to cover them and preventing pathogens from colonizing;
3. Depleting nutrients essential for the growth of pathogens; and
4. Non-specifically stimulating the immune system.
Destruction of normal bacterial microbiota by the use of broad spectrum antibiotics may result in superinfections or overgrowth by antibiotic-resistant opportunistic microbiota. For example, the yeast Candida, that causes infections such as vaginitis and thrush, and the bacterium Clostridium difficile, that causes potentially severe antibiotic-associated colitis, are opportunistic microorganisms normally held in check by the normal microbiota.
In the case of Candida infections, the Candida resists the antibacterial antibiotics because being a yeast, it is eukaryotic, not prokaryotic like the bacteria. Once the bacteria are eliminated by the antibiotics, the Candida has no competition and can overgrow the area.
Clostridium difficile is an opportunistic Gram-positive, endospore-producing bacillus transmitted by the fecal-oral route that causes severe antibiotic-associated colitis. C. difficile is a common healthcare-associated infection (HAIs) and is the most frequent cause of health-care-associated diarrhea. C. difficile infection often recurs and can progress to sepsis and death. CDC has estimated that there are about 500,000 C. difficile infections (CDI) in health-care associated patients each year and is linked to 15,000 American deaths each year.
Antibiotic-associated colitis is especially common in older adults. It is thought that C. difficile survives the exposure to the antibiotic by sporulation. After the antibiotic is no longer in the body, the endospores germinate and C. difficile overgrows the intestinal tract and secretes toxin A and toxin B that have a cytotoxic effect on the epithelial cells of the colon. C. difficile has become increasingly resistant to antibiotics in recent years making treatment often difficult. There has been a great deal of success in treating the infection with fecal transplants, still primarily an experimental procedure. Polymerase chain reaction (PCRs) assays, which test for the bacterial gene encoding toxin B, are highly sensitive and specific for the presence of a toxin-producing Clostridium difficile organism. The most successful technique in restricting C. difficile infections has been the restriction of the use of antimicrobial agents.
Summary
Anatomical barriers such as the skin, the mucous membranes, and bony encasements are tough, intact barriers that prevent the entry and colonization of many microbes. Mechanical removal is the process of physically flushing microbes from the body. Examples include mucus and cilia, coughing and sneezing, vomiting and diarrhea, and the flushing action of bodily fluids. The normal microbiota keeps potentially harmful opportunistic pathogens in check and also inhibits the colonization of pathogens by producing metabolic products that inhibit the growth of many pathogens, adhering to target host cells so as to cover them and prevent pathogens from colonizing, depleting nutrients essential for the growth of pathogens, and non-specifically stimulating the immune system.
Destruction of normal bacterial microbiota by the use of broad spectrum antibiotics may result in superinfections or overgrowth by antibiotic-resistant opportunistic microbiota such as Candida and Clostridium difficile. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.3%3A_Immediate_Innate_Immunity/11.3C%3A_Anatomical_Barriers_to_Infection_Mechanical_Removal_of_Microbes_and_Bacterial_Antagonism_by_Normal_Body_Microbiota.txt |
Most body defense cells have pattern-recognition receptors or PRRs for these common PAMPs enabling an immediate response against the invading microorganism. Pathogen-associated molecular patterns can also be recognized by a series of soluble pattern-recognition receptors in the blood that function as opsonins and initiate the complement pathways. In all, the innate immune system is thought to recognize approximately 103 of these microbial molecular patterns.
11.4: Early Induced Innate Immunity
State how long it takes for early induced innate immunity to become activated and what it involves. State what is meant by pathogen-associated molecular patterns (PAMPs), and the role PAMPs play in inducing innate immunity. Name at least 5 PAMPS associated with bacteria. Name at least 2 PAMPS associated with viruses. Define DAMPs and give two examples.
Examples of microbial-associated PAMPs include:
1. lipopolysaccharide (LPS) from the outer membrane of the Gram-negative cell wall (see Figure \(1\)A);
2. bacterial lipoproteins and lipopeptides (see Figure \(1\)A);
3. porins in the outer membrane of the Gram-negative cell wall (see Figure \(1\)A);
4. peptidoglycan found abundantly in the Gram-positive cell wall and to a lesser degree in the gram-negative cell wall (see Figure \(1\)B);
5. lipoteichoic acids found in the Gram-positive cell wall (Figure \(1\)B);
6. lipoarabinomannan and mycolic acids found in acid-fast cell walls (Figure \(2\)B)
7. mannose-rich glycans (short carbohydrate chains with the sugar mannose or fructose as the terminal sugar). These are common in microbial glycoproteins and glycolipids but rare in those of humans (see Figure \(6\)).
8. flagellin found in bacterial flagella;
9. bacterial and viral nucleic acid. Bacterial and viral genomes contain a high frequency of unmethylated cytosine-guanine dinucleotide or CpG sequences (a cytosine lacking a methyl or CH3 group and located adjacent to a guanine). Mammalian DNA has a low frequency of CpG sequences and most are methylated which may mask recognition by pattern-recognition receptors . Also, human DNA and RNA does not normally enter cellular endosomes where the pattern-recognition receptors for microbial DNA and RNA are located;
10. N-formylmethionine , an amino acid common to bacterial proteins;
11. double-stranded viral RNA unique to many viruses in some stage of their replication;
12. single-stranded viral RNA from many` viruses having an RNA genome;
13. lipoteichoic acids, glycolipids, and zymosan from yeast cell walls; and
14. phosphorylcholine and other lipids common to microbial membranes.
Examples of DAMPs associated with stressed, injured, infected, or transformed host cells and not found on normal cells include:
1. heat-shock proteins;
2. altered membrane phospholipids; and
3. molecules normally located inside phagosomes and lysosomes that enter the cytosol only when these membrane-bound compartments are damaged as a result of infection, including antibodies bound to microbes from opsonization.
4. molecules normally found within cells, such as ATP, DNA, and RNA, that spill out of damaged cells.
To recognize PAMPs such as those listed above, various body cells have a variety of corresponding receptors called pattern-recognition receptors or PRRs capable of binding specifically to conserved portions of these molecules. Cells that typically have pattern recognition receptors include macrophages , dendritic cells , endothelial cells , mucosal epithelial cells, and lymphocytes .
Summary
1. Early induced innate immunity begins 4 - 96 hours after exposure to an infectious agent and involves the recruitment of defense cells as a result of pathogen-associated molecular patterns or PAMPS binding to pattern-recognition receptors or PRRs.
2. Pathogen-associated molecular patterns or PAMPs are molecules shared by groups of related microbes that are essential for the survival of those organisms and are not found associated with mammalian cells. Examples include LPS, porins, peptidoglycan, lipoteichoic acids, mannose-rich glycans, flagellin, bacterial and viral genomes, mycolic acid, and lipoarabinomannan.
3. Danger-associated molecular patterns or DAMPs are unique molecules displayed on stressed, injured, infected, or transformed human cells also be recognized as a part of innate immunity. Examples include heat-shock proteins and altered membrane phospholipids.
4. PAMPs and DAMPs bind to pattern-recognition receptors or PRRs associated with body cells to induce innate immunity. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.4%3A_Early_Induced_Innate_Immunity/11.3A%3A_Pathogen-Associated_Molecular_Patterns_%28PAMPs%29_and_Danger-Associated_Molecular_Patterns_%28DAMPs%29.txt |
State the function of the following as they relate to innate immunity. pattern recognition receptors (PRRs) endocytic pattern recognition receptors signaling pattern recognition receptors danger-associated molecular patterns danger recognition receptors inflammasome pyroptosis Name 2 endocytic PRRs. Name 2 signaling PRRs found on host cell surfaces. Name 2 signaling PRRs found in the endosomes of phagocytic cells. Name 2 signaling PRRs found on the host cell cytoplasm. Briefly describe the major difference between the effect of the cytokines produced in response to PAMPs that bind to cell surface signaling PRRs and endosomal PRRs.
In order to recognize PAMPs, various body cells have a variety of corresponding receptors called pattern-recognition receptors or PRRs (see Figure \(5\)) capable of binding specifically to conserved portions of these molecules. Cells that typically have pattern recognition receptors include macrophages, dendritic cells, endothelial cells, mucosal epithelial cells, and lymphocytes.
Many pattern-recognition receptors are located on the surface of these cells where they can interact with PAMPs on the surface of microbes. Others PRRs are found within the phagolysosomes of phagocytes where they can interact with PAMPs located within microbes that have been phagocytosed. Some PRRs are found in the cytosol of the cell.
There are two functionally different major classes of pattern-recognition receptors: endocytic pattern-recognition receptors and signaling pattern-recognition receptors.
Endocytic (Phagocytic) Pattern-Recognition Receptors
Endocytic pattern-recognition receptors, also called phagocytic pattern-recognition receptors, are found on the surface of phagocytes and promote the attachment of microorganisms to phagocytes leading to their subsequent engulfment and destruction. They include:
1. Mannose receptors
Mannose receptors on the surface of phagocytes bind to various microbial carbohydrates such as those rich in mannose or fucose, and to N-acetylglucosamine (NAG). Human glycoproteins and glycolipids typically have terminal N-acetylglucosamine and sialic acid groups. C-type lectins found on the surface of phagocytes are mannose receptors (see Figure \(6\)).
It is now thought that mannose receptors may be quite important in removing potentially harmful mannose-containing glycoproteins such as lysosomal hydrolases that are produced in increased amounts during inflammation.
2. Dectin-1
Dectin-1 recognizes beta-glucans (polymers of glucose) commonly found in fungal cell walls.
3. Scavenger receptors
Scavenger receptors found on the surface of phagocytic cells bind to bacterial cell wall components such as LPS, peptidoglycan and teichoic acids (see Figure \(7\)). There are also scavenger receptors for certain components of other types of microorganisms, as well as for stressed, infected, or injured cells. Scavenger receptors include CD-36, CD-68, and SRB-1.
4. Opsonin receptors
Opsonins are soluble molecules produced as a part of the body's immune defenses that bind microbes to phagocytes. One portion of the opsonin binds to a PAMP on the microbial surface and another portion binds to a specific receptor on the phagocytic cell.
• Acute phase proteins circulating in the plasma, such as:
• mannose-binding lectin (also called mannose-binding protein) that binds to various microbial carbohydrates such as those rich in mannose or fucose, and to N-acetylglucosamine (NAG); and
• C-reactive protein (CRP) that binds to phosphorylcholine portion of teichoic acids and lipopolysaccharides of bacterial and fungal cell walls. It also binds to the phosphocholine found on the surface of damaged or dead human cells.
• Complement pathway proteins, such as C3b (see Figure \(8\)) and C4b recognize a variety of PAMPS.
• Surfactant proteins in the alveoli of the lungs, such as SP-A and SP-D are opsonins.
• During adaptive immunity, the antibody molecule IgG can function as an opsonin (see Figure \(16\)).
5. N-formyl Met receptors
N-formyl methionine is the first amino acid produced in bacterial proteins since the f-met-tRNA in bacteria has an anticodon complementary to the AUG start codon (see Figure \(17\)). This form of the amino acid is not typically seen in mammalian proteins. FPR and FPRL1 are N-formyl receptors on neutrophils and macrophages. Binding of N-formyl Met to its receptor promotes the motility and the chemotaxis of these phagocytes. It also promotes phagocytosis.
Signaling Pattern-Recognition Receptors
Signaling pattern-recognition receptors bind a number of microbial molecules: LPS, peptidoglycan, teichoic acids, flagellin, pilin, unmethylated cytosine-guanine dinucleotide or CpG sequences from bacterial and viral genomes; lipoteichoic acid, glycolipids, and zymosan from fungi; double-stranded viral RNA, and certain single-stranded viral RNAs. Binding of microbial PAMPs to signaling PRRs promotes the production of:
• inflammatory cytokines, such as such as interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF-alpha), and interleukin-12 (IL-12);
• antiviral cytokines called type-1 interferons (IFN), such as IFN-alpha and IFN-beta;
• chemotactic factors, such as the chemokines interleukin-8 (IL-8), MCP-1, and RANTES; and
• antimicrobial peptides, such as human defensins ) and cathelicidins.
These molecules are crucial to initiating innate immunity and adaptive immunity.
1. Signaling PRRs found on cell surfaces (see Figure \(5\)):
A series of signaling pattern-recognition receptors known as toll-like receptors (TLRs) are found on the surface of a variety of defense cells and other cells. These TLRs play a major role in the induction of innate immunity and contribute to the induction of adaptive immunity.
Different combinations of TLRs appear in different cell types and may occur in pairs. Different TLRs directly or indirectly bind different microbial molecules. For example:
a. TLR-2 - recognizes peptidoglycan, bacterial lipoproteins, lipoteichoic acid (Gram-positive bacteria), and porins (gram-negative bacteria).
b. TLR-4 - recognizes lipopolysaccharide (Gram-negative bacteria), fungal mannans, viral envelope proteins, parasitic phospholipids, heat-shock proteins.
c. TLR-5 - recognizes bacterial flagellin;
d. TLR-1/TLR-2 pairs - binds to bacterial lipopeptides, lipomannans (mycobacteria) lipoteichoic acids (Gram-positive bacteria), cell wall beta gucans (bacteria and fungi), zymosan (fungi) and glycosylphosphatidylinositol (GPI)-anchored proteins (protozoa).
e. TLR-2/TL6 pairs - also binds to bacterial lipopeptides, lipomannans (mycobacteria) lipoteichoic acids (Gram-positive bacteria), cell wall beta gucans (bacteria and fungi), zymosan (fungi) and glycosylphosphatidylinositol (GPI)-anchored proteins (protozoa).
Many of the TLRs, especially those that bind to bacterial and fungal cell wall components, stimulate the transcription and translation of inflammatory cytokines such as interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF-alpha), and interleukin-12 (IL-12), as well as chemokines such as interleukin-8 (IL-8), MCP-1, and RANTES. These cytokines trigger innate immune defenses such as inflammation, fever, and phagocytosis in order to provide an immediate response against the invading microorganism (see Figure \(9\)). Because cytokines such as IL-I, TNF-alpha, and IL-12 that trigger an inflammatory response, they are often referred to as inflammatory cytokines. Chemokines are a group of cytokines that enable the migration of leukocytes from the blood to the tissues at the site of inflammation. To counter inflammation, anti-inflammatory cytokines such as IL-1 receptor antagonist, IL-4, and IL-10 are produced.
Another cell surface PRR is CD14. CD14 is found on monocytes, macrophages, and neutrophils and promotes the ability of TLR-4 to respond to LPS. LPS typically binds to LPS-binding protein in the plasma and tissue fluid. The LPS-binding protein promotes the binding of LPS to the CD14 receptors. At that point the LPS-binding protein comes off and the LPS-CD14 bind to TLR-4. Interaction of LPS and CD14 with TLR-4 leads to an elevated synthesis and secretion of inflammatory cytokines such as IL-1, IL-6, IL-8, TNF-alpha, and platelet-activating factor (PAF). These cytokines then bind to cytokine receptors on target cells and initiate inflammation and activate both the complement pathways and the coagulation pathway (see Figure \(9\)).
The signaling process for the CD14 and TLR-4 response to LPS is shown in Figure \(15\).
TLRs also participate in adaptive immunity by triggering various secondary signals needed for humoral immunity (the production of antibodies ) and cell-mediated immunity (the production of cytotoxic T-lymphocytes, activated macrophages, and additional cytokines ). Without innate immune responses there could be no adaptive immunity.
a. T-independent (TI) antigens allow B-lymphocytes to mount an antibody response without the requirement of interaction with effector T4-lymphocytes. The resulting antibody molecules are generally of the IgM isotype and do not give rise to a memory response. There are two basic types of T-independent antigens: TI-1 and TI-2. TI-1 antigens are pathogen-associated molecular patterns (PAMPs) such as lipopolysaccharide (LPS) from the outer membrane of the gram-negative cell wall and lipoteichoic acids from the gram-positive cell wall. These antigens activate B-lymphocytes by binding to their specific toll-like receptors rather than to B-cell receptors (see Figure \(11\)). Antibody molecules generated against TI-1 antigens are often called "natural antibodies" because they are always being made against bacteria present in the body.
b. The activation of naive T-lymphocytes requires co-stimulatory signals involving the interaction of accessory molecules on antigen-presenting cells or APCs with their corresponding ligands on T-lymphocytes. These co-stimulatory molecules are only synthesized when toll-like receptors on APCs bind to pathogen-associated molecular patterns of microbes (see Figure \(12\)).
2. Signaling PRRs found in the membranes of the endosomes (phagolysosomes ) used to degrade pathogens (see Figure \(5\)):
a. TLR-3 - binds double-stranded viral RNA;
b. TLR-7 - binds single-stranded viral RNA, such as in HIV, rich in guanine/uracil nucleotide pairs;
c. TLR-8 - binds single-stranded viral RNA;
d. TLR-9 - binds unmethylated cytosine-guanine dinucleotide sequences (CpG DNA) found in bacterial and viral genomes but uncommon or masked in human DNA and RNA.
Most of the TLRs that bind to viral components trigger the synthesis of cytokines called interferons that block viral replication within infected host cells as well as inflammatory cytokines.
3. Signaling PRRs and DRRsfound in the cytoplasm (see Figure \(5\))
Pattern-recognition receptors or PRRs found in the cytoplasm include:
a. NODs (nucleotide-binding oligomerization domain)
NOD proteins, including NOD-1 and NOD-2, are cytostolic proteins that allow intracellular recognition of peptidoglycan components.
1. NOD-1 recognizes peptidoglycan containing the muramyl dipeptide NAG-NAM-gamma-D-glutamyl-meso diaminopimelic acid, part of the peptidoglycan monomer in common gram-negative bacteria and just a few gram-positive bacteria.
2. NOD-2 recognizes peptidoglycan containing the muramyl dipeptide NAG-NAM-L-alanyl-isoglutamine found in practically all bacteria (see Figure \(5\)).
As macrophages phagocytose either whole bacteria or peptidoglycan fragments released during bacterial growth, the peptidoglycan is broken down into muramyl dipeptides. Binding of the muramyl dipetides to NOD-1 or NOD-2 leads to the activation of genes coding for inflammatory cytokines such as IL-1, TNF-alpha, IL-8, and IL-12 in a manner similar to the cell surface TLRs. Activation of NOD-2 also induces the production of antimicrobial peptides such as defensins as well as microbicidal reactive oxygen species (ROS).
b. CARD-containing proteins
CARD (caspase activating and recruitment domain)-containing proteins, such as RIG-1 (retinoic acid-inducible gene-1) and MDA-5 (melanoma differentiation-associated gene-5), are cytoplasmic sensors of viral RNA molecules that trigger the synthesis of type-1 interferons, antiviral cytokines that block viral replication within infected host cells in a manner similar to the endosomal TLRs. RIG-1 recognizes 5'-PPPs on viral RNAs. The 5'-PPPs on host cell RNAs are either capped or removed and are not recognized by RIG-1. Rig-1 and MDA-5 can also, through another regulatory pathway, stimulate the production of inflammatory cytokines.
Detection of PAMPs by PRRs in the cytosol trigger the formation of multi-protein complexes called inflammasomes which, in turn, leads to the activation of caspase-1. Caspase-1 triggers the formation of inflammatory cytokines and can also result in an inflammatory response-induced cell suicide called pyroptosis. Pyroptosis, unlike apoptosis, leads to the release of PAMPS as well as inflammatory cytokines from the lysed cell.
Pyroptosis is initiated by PAMPs binding to pattern-recognition receptors (PRRs) on various defense cells which then triggers the production of inflammatory cytokines and type-1 interferons. Other PRRs, called nod-like receptors (NLRs) located in the cytosol of these defense cells recognize PAMPs and DAMPs that have entered the host cell’s cytosol. Some NLRs trigger the production of inflammatory cytokines while others activate caspase 1-dependent pyroptosis of the cell causing the release of its intracellular inflammatory cytokines (see Figure \(1\)). The binding of PAMPs or DAMPs to their respective NLRs triggers the assembly of multiprotein complexes called inflammasomes in the cytosol of the host cell. It is these inflammasomes that activate caspase 1 and induce inflammation and pyroptosis. Pyroptosis results in production of proinflammatory cytokines, rupture of the cell’s plasma membrane, and subsequent release of proinflammatory intracellular contents. It plays an essential role in innate immunity by promoting inflammation to control microbial infections. At highly elevated levels, however, it can cause considerable harm to the body and even death.
c. Danger recognition receptors or DRRs
Danger recognition receptors or DRRs found in the cytoplasm recognize danger-associated molecular patterns (DAMPS) in the cytosol such as altered membrane phospholipids, and materials released from damaged phagosomes and damaged lysosomes, including antibodies bound to microbes from opsonization. DAMPs are also produced as a result of tissue injury during cancer, heart attack, and stroke. Detection of DAMPs by DRRs in the cytosol also triggers the activation of inflammasomes, release of inflammatory cytokines, and pyroptosis.
4. Secreted signaling PRRs found in plasma and tissue fluid
In addition to the PRRs found on or within cells, there are also secreted pattern-recognition receptors. These PRRs bind to microbial cell walls and enable them to activate the complement pathways, as well as by phagocytes. For example, mannan-binding lectin -also known as mannan-binding protein - is synthesized by the liver and released into the bloodstream as part of the acute phase response discussed later in Unit 4. Here it can bind to the carbohydrates on bacteria, yeast, some viruses, and some parasites (see Figure \(6\)). This, in turn, activates the lectin complement pathway (discussed later in Unit 4) and results in the production of a variety of activated complement proteins that are able to trigger inflammation, chemotactically attract phagocytes to the infection site, promote the attachment of antigens to phagocytes via enhanced attachment or opsonization, and cause lysis of gram-negative bacteria and infected or transformed human cells.
Other secreted PRRs include C-reactive protein (CRP), surfactant protein A (SP-A), surfactant protein D (SP-D), collectin liver 1 (CL-L1), and ficolins.
Compare and contrast the functions of endocytic pattern-recognition receptors and signaling pattern-recognition receptors. Compare and contrast signaling pattern-recognition receptors found on cell surfaces with those found in the membranes of endosomes (phagolysosomes).
Concept Map for PRRs and DRRs
Summary
1. Early induced innate immunity begins 4 - 96 hours after exposure to an infectious agent and involves the recruitment of defense cells as a result of pathogen-associated molecular patterns or PAMPS binding to pattern-recognition receptors or PRRs and danger-associated molecular patterns or DAMPs binding to danger-recognition receptors or DRRs.
2. Endocytic pattern-recognition receptors are found on the surface of phagocytes and promote the attachment of microorganisms to phagocytes leading to their subsequent engulfment and destruction. They include mannose receptors, scavenger receptors, and opsonin receptors.
3. Binding of microbial PAMPs to signaling PRRs promotes the production of inflammatory cytokines, antiviral cytokines called type-1 interferons (IFN), chemotactic factors, and antimicrobial peptides. They include toll-like receptors (TLRs) and NODs.
4. PRRs found on the surface of the body’s cells typically bind to surface PAMPs on microbes and stimulate the production of inflammatory cytokines.
5. PRRs found within cellular phagolysosomes (endosomes) typically detect nucleic acid PAMPs released during the phagocytic destruction of viruses and stimulate the production of antiviral cytokines called type-1 interferons.
6. PRRs and DRRs found within the cytoplasm of host cells typically trigger the formation of multi-protein complexes called inflammasomes which, in turn, triggers the formation of inflammatory cytokines and can also leads to an inflammatory response-induced cell suicide called pyroptosis.
7. PRRs circulating in the blood and tissue fluid activate the complement pathways and may function as opsonins. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.4%3A_Early_Induced_Innate_Immunity/11.3B%3A_Pattern-Recognition_Receptors_%28PRRs%29.txt |
Learning Objectives
1. Describe the following:
1. cytokines
2. chemokines
3. interferons
2. State what is meant by the phrase "Cytokines are pleiotropic, redundant, and multifunctional."
3. Name the two cytokines that are most important in stimulating acute inflammation.
4. Describe specifically how type I interferons are able to block viral replication within an infected host cell.
Cytokines are low molecular weight, soluble proteins that are produced in response to an antigen and function as chemical messengers for regulating the innate and adaptive immune systems. They are produced by virtually all cells involved in innate and adaptive immunity, but especially by T- helper (Th) lymphocytes. The activation of cytokine-producing cells triggers them to synthesize and secrete their cytokines. The cytokines, in turn, are then able to bind to specific cytokine receptors on other cells of the immune system and influence their activity in some manner.
Cytokines are pleiotropic, redundant, and multifunctional.
• Pleiotropic means that a particular cytokine can act on a number of different types of cells rather than a single cell type.
• Redundant refers to to the ability of a number of different cytokines to carry out the same function.
• Multifunctional means the same cytokine is able to regulate a number of different functions.
Some cytokines are antagonistic in that one cytokine stimulates a particular defense function while another cytokine inhibits that function. Other cytokines are synergistic wherein two different cytokines have a greater effect in combination than either of the two would by themselves. There are three functional categories of cytokines:
1. cytokines that regulate innate immune responses,
2. cytokines that regulate adaptive Immune responses, and
3. cytokines that stimulate hematopoiesis.
Cytokines that regulate innate immunity are produced primarily by mononuclear phagocytes such as macrophages and dendritic cells, although they can also be produced by T-lymphocytes, NK cells, endothelial cells, and mucosal epithelial cells. They are produced primarily in response to pathogen-associated molecular patterns (PAMPs) such as LPS, peptidoglycan monomers, teichoic acids, unmethylated cytosine-guanine dinucleotide or CpG sequences in bacterial and viral genomes, and double-stranded viral RNA. Cytokines produced in response to PRRs on cell surfaces, such as the inflammatory cytokines IL-1, IL-6, IL-8, and TNF-alpha, mainly act on leukocytes and the endothelial cells that form blood vessels in order to promote and control early inflammatory responses (Figure \(1\)). Cytokines produced in response to PRRs that recognize viral nucleic acids, such as type I interferons, primarily block viral replication within infected host cells (see Figure \(2\)A and Figure \(2\)B).
Figure \(1\): Integrins on the surface of the leukocyte bind to adhesion molecules on the inner surface of the vascular endothelial cells. The leukocytes flatten out and squeeze between the endothelial cells to leave the blood vessels and enter the tissue. The increased capillary permeability also allows plasma to enter the tissue.
Examples include:
a. Tumor necrosis factor-alpha (TNF-a)
TNF-a is the principle cytokine that mediates acute inflammation. In excessive amounts it also is the principal cause of systemic complications such as the shock cascade. Functions include acting on endothelial cells to stimulate inflammation and the coagulation pathway; stimulating endothelial cells to produce selectins and ligands for leukocyte integrins during diapedesis ; stimulating endothelial cells and macrophages to produce chemokines that contribute to diapedesis, chemotaxis, and the recruitment of leukocytes; stimulating macrophages to secrete interleukin-1 (IL-1) for redundancy; activating neutrophils and promoting extracellular killing by neutrophils; stimulating the liver to produce acute phase proteins, and acting on muscles and fat to stimulate catabolism for energy conversion. TNF-a stimulates the endothelial cells that form capillaries to express proteins that activate blood clot formation within the capillaries. This occludes local blood flow to help prevent microbes from entering the bloodstream. In addition, TNF is cytotoxic for some tumor cells; interacts with the hypothalamus to induce fever and sleep; stimulates the synthesis of collagen and collagenase for scar tissue formation; and activates macrophages. TNF is produced by monocytes,macrophages, dendritic cells, TH1 cells, and other cells.
b. Interleukin-1 (IL-1)
IL-1 function similarly to TNF in that it mediates acute inflammatory responses. It also works synergistically with TNF to enhance inflammation. Functions of IL-1 include promoting inflammation ; activating the coagulation pathway, stimulating the liver to produce acute phase proteins, catabolism of fat for energy conversion, inducing fever and sleep; stimulates the synthesis of collagen and collagenase for scar tissue formation; stimulates the synthesis of adhesion factors on endothelial cells and leukocytes (see Figure \(1\)) for diapedesis ; and activates macrophages. IL-1 is produced primarily by monocytes, macrophages, dendritic cells, endothelial cells, and some epithelial cell.
c. Chemokines
Chemokines are a group of cytokines that enable the migration of leukocytes from the blood to the tissues at the site of inflammation. They increase the affinity of integrins on leukocytes for ligands on the vascular wall (see Figure \(1\) during diapedesis, regulate the polymerization and depolymerization of actin in leukocytes for movement and migration, and function as chemoattractants for leukocytes. In addition, they trigger some WBCs to release their killing agents for extracellular killing and induce some WBCs to ingest the remains of damaged tissue. Certain chemokines promote angiogenesis. Chemokines also regulate the movement of B-lymphocytes, T-lymphocytes, and dendritic cells through the lymph nodes and the spleen. When produced in excess amounts, chemokines can lead to damage of healthy tissue as seen in such disorders as rheumatoid arthritis, pneumonia, asthma, adult respiratory distress syndrome (ARDS), and septic shock. Examples of chemokines include IL-8, MIP-1a, MIP-1b, MCP-1, MCP-2, MCP-3, GRO-a, GRO-b, GRO-g, RANTES, and eotaxin. Chemokines are produced by many cells including leukocytes, endothelial cells, epithelial cells, and fibroblasts.
d. Interleukin-12 (IL-12)
IL-12 is a primary mediator of early innate immune responses to intracellular microbes. It is also an inducer of cell-mediated immunity. It functions to stimulate the synthesis of interferon-gamma by T-lymphocytes and NK cells ; increases the killing activity of cytotoxic T-lymphocytes and NK cells; and stimulates the differentiation of naive T4-lymphocytes into interferon-gamma producing TH1 cells. It is produced mainly by macrophages and dendritic cells.
e. Type I Interferons
Interferons modulate the activity of virtually every component of the immune system. Type I interferons include 13 subtypes of interferon-alpha, interferon-beta, interferon omega, interferon-kappa, and interferon tau. (There is only one type II interferon, interferon-gamma, which is involved in the inflammatory response.)
The most powerful stimulus for type I interferons is the binding of viral DNA or RNA to toll-like receptors TLR-3, TLR-7, and TLR-9 in endosomal membranes.
a. TLR-3 - binds double-stranded viral RNA;
b. TLR-7 - binds single-stranded viral RNA, such as in HIV, rich in guanine/uracil nucleotide pairs;
c. TLR-9 - binds unmethylated cytosine-guanine dinucleotide sequences (CpG DNA) found in bacterial and viral genomes but uncommon or masked in human DNA and RNA.
Signaling pattern recognition receptors located in the cytoplasm of cells such as RIG-1 and MDA-5 also signal synthesis and secretion of type-I interferons.
Type I interferons, produced abundantly by plasmacytoid dendritic cells, by virtually any virus-infected cell, and by other defense cells provide an early innate immune response against viruses. Interferons induce uninfected cells to produce an enzyme capable of degrading viral mRNA, as well as one that blocks translation in eukaryotic cells. These enzymes remain inactive until the uninfected cell becomes infected with a virus. At this point, the enzymes are activated and begin to degrade viral mRNA and block translation in the host cell. This not only blocks viral protein synthesis, it also eventually kills the infected cell (see Figure \(2\)A and Figure \(2\)B). In addition, type I interferons also cause infected cells to produce enzymes that interfere with transcription of viral RNA or DNA. They also promote body defenses by enhancing the activities of CTLs, macrophages, dendritic cells, NK cells, and antibody-producing cells, as well as induce chemokine production to attract leukocytes to the area.
Type I interferons also induce MHC-I antigen expression needed for recognition of antigens by cytotoxic T-lymphocytes ; augment macrophages, NK cells, cytotoxic T-lymphocytes, and B-lymphocytes activity; and induce fever. Interferon-alpha is produced by T-lymphocytes, B-lymphocytes, NK cells, monocytes/macrophages; interferon-beta by virus-infected cells, fibroblasts, macrophages, epithelial cells, and endothelial cells.
f. Interleukin-6 (IL-6)
IL-6 functions to stimulate the liver to produce acute phase proteins ; stimulates the proliferation of B-lymphocytes ; and increases neutrophil production. IL-6 is produced by many cells including T-lymphocytes, macrophages, monocytes, endothelial cells, and fibroblasts.
g. Interleukin-10 (IL-10)
IL-10 is an inhibitor of activated macrophages and dendritic cells and as such, regulates innate immunity and cell-mediated immunity. IL-10 inhibits their production of IL-12, co-stimulator molecules, and MHC-II molecules, all of which are needed for cell-mediated immunity. IL-10 is produced mainly by macrophages, and TH2 cells.
h. Interleukin 15 (IL-15)
IL-15 stimulates NK cell proliferation and proliferation of memory T8-lymphocytes. IL-15 is produced by various cells including macrophages.
i. Interleukin-18 (IL-18)
IL-18 stimulates the production of interferon-gamma by NK cells and T-lymphocytes and thus induces cell-mediated immunity. It is produced mainly by macrophages.
A number of human cytokines produced by recombinant DNA technologies are now being used to treat various infections or immune disorders. These include:
1. recombinant interferon alfa-2a (Roferon-A): a cytokine used to treat Kaposi's sarcoma, chronic myelogenous leukemia, and hairy cell leukemia.
2. peginterferon alfa-2a (Pegasys) : used to treat hepatitis C (HCV).
3. recombinant interferon-alpha 2b (Intron A): a cytokine produced by recombinant DNA technology and used to treat Hepatitis B; malignant melanoma, Kaposi's sarcoma, follicular lymphoma, hairy cell leukemia, warts, and Hepatitis C.
4. peginterferon alfa-2b (PEG-Intron; PEG-Intron Redipen):
used to treat hepatitis C (HCV).
5. recombinant Interferon alfa-2b plus the antiviral drug ribavirin (Rebetron): used to treat hepatitis C (HCV).
6. recombinant interferon-alpha n3 (Alferon N): used to treat warts.
7. recombinant iInterferon alfacon-1 (Infergen) :
used to treat hepatitis C (HCV).
8. G-CSF (granulocyte colony stimulating factor): for reduction of infection in people after myelotoxic anticancer therapy for solid tumors.
9. GM-CSF (granulocyte-macrophage colony stimulating factor): for hematopoietic reconstruction after bone marrow transplant in people with lymphoid cancers.
Summary
1. Cytokines are low molecular weight, soluble proteins that are produced in response to an antigen and function as chemical messengers for regulating the innate and adaptive immune systems.
2. Cytokines are pleiotropic, meaning meaning that a particular cytokine can act on a number of different types of cells rather than a single cell type.
3. Cytokines are redundant, meaning that a number of different cytokines are able to carry out the same function.
4. Cytokines are multifunctional, meaning that the same cytokine is able to regulate a number of different functions.
5. Tumor necrosis factor-alpha (TNF-a) and interleukin-1 (IL-1) are the principle cytokines that mediates acute inflammation.
6. Chemokines are a group of cytokines that enable the migration of leukocytes from the blood to the tissues at the site of inflammation.
7. Type I interferons, produced abundantly by plasmacytoid dendritic cells, by virtually any virus-infected cell, and by other defense cells provide an early innate immune response against viruses by inducing uninfected cells to produce enzymes capable of degrading viral mRNA and blocking translation in eukaryotic cells. They also enhancing the activities of CTLs, macrophages, dendritic cells, NK cells, and antibody-producing cells and induce chemokine production to attract leukocytes to the area.
8. Type II interferon is involved in stimulating an inflammatory response. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.4%3A_Early_Induced_Innate_Immunity/11.3C%3A_Cytokines_Important_in_Innate_Immunity.txt |
The Ability of Pathogen-Associated Molecular Patterns or PAMPsto Trigger the Synthesis and Secretion of Excessive Levels of Inflammatory Cytokines and Chemokines
As learned in Unit 3 under sepsis and systemic inflammatory response syndrome (SIRS), during severe systemic infections with large numbers of bacteria present, high levels of cell wall PAMPs are released resulting in excessive cytokine production by the defense cells and this can harm the body (see Figure \(10\)). In addition, neutrophils start releasing their proteases and toxic oxygen radicals that kill not only the bacteria, but the surrounding tissue as well. Harmful effects include high fever, hypotension, tissue destruction, wasting, acute respiratory distress syndrome (ARDS), disseminated intravascular coagulation (DIC), and damage to the vascular endothelium. This can result in shock, multiple system organ failure (MOSF), and death.
Harmful Effects Associated with either an Overactive or an Underactive Innate Immune Response
There are a number of harmful effects that are known to occur as a result of either an overactive or an underactive innate immune response. This occurs as a result of people possessing different polymorphisms in the various genes participating in PRR signaling.
• People born with underactive PRRs or deficient PRR immune signaling pathways are at increased risk of infection by specific pathogens due to a decrease innate immune response.
• People born with overactive PRRs or deficient PRR immune signaling pathways are at increased risk of inflammatory damage by lower numbers of specific pathogens.
Examples include:
1. People with an underactive form of TLR-4, the toll-like receptor for bacterial LPS, have been found to be five times as likely to contract a severe bacterial infection over a five year period than those with normal TLR-4. People with overactive TLR-4 receptors may be more prone to developing SIRS from gram-negative bacteria.
2. Most people that die as a result of Legionnaire's disease have been found to have a mutation in the gene coding for TLR-5 that enables the body to recognize the flagella of Legionella pneumophila.
3. B-lymphocytes, the cells responsible for recognizing foreign antigens and producing antibodies against those antigens, normally don't make antibodies against the body's own DNA and RNA. The reason is that any B-lymphocytes that bind the body's own antigens normally undergo apoptosis, a programmed cell suicide. People with the autoimmune disease systemic lupus erythematosus have a mutation in a gene that signals the cell to undergo apoptosis. As a result, these B-cells are able to bind and engulf the body's own DNA and RNA and place them in an endosome or phagolysosome where the the DNA can be recognized by TLR-9 and the RNA by TLR-7. This, in turn, triggers those B-lymphocytes to make antibody molecules against the body's own DNA and RNA. Another gene error enables these B- cells to increase the expression of TLR-7.
4. TLR-4, MyD88, TLR-1 and TLR-2 have been implicated in the production of atherosclerosis in mice and some humans.
5. Mutations resulting in loss-of-function in the gene coding for NOD-2 that prevents the NOD-2 from recognizing muramyl dipeptide make a person more susceptible to Crohn's disease, an inflammatory disease of the large intestines. Mutations resulting in over-activation in the gene coding for NOD-2 can lead to an inflammatory disorder called Blau syndrome.
6. People with chronic sinusitis that does not respond well to treatment have decreased activity of TLR-9 and produce reduced levels of human beta-defensin 2, as well as mannan-binding lectin needed to initiate the lectin complement pathway.
7. Pathogenic strains of Staphylococcus aureus producing leukocidin and protein A, including MRSA, cause an increased inflammatory response. Protein A, a protein that blocks opsonization and functions as an adhesin, binds to cytokine receptors for TNF-alpha. It mimics the cytokine and induces a strong inflammatory response. As the inflammatory response attracts neutrophils to the infected area, the leukocidin causes lysis of the neutrophils. As a result, tissue is damaged and the bacteria are not phagocytosed.
8. People with chronic mucocutaneous candidiasis disease have a mutation either in the gene coding for IL-17F or the gene encoding IL-17F receptor. TH17 cells secrete cytokines such as IL-17 that are important for innate immunity against organisms that infect mucous membranes.
9. A polymorphism in the gene for TLR-2 makes individuals less responsive to Treponema pallidum and Borrelia burgdorferi and possibly more susceptible to tuberculosis and staphylococcal infections.
10. Polymorphisms in a gene locus called A20, a gene that helps to control inflammation, are considered as risk alleles for rheumatoid arthritis, systemic lupus erythematosus, psoriasis, type I diabetes, and Chron’s disease.
11. The innate immune response to Mycobacterium tuberculosis and the severity of tuberculosis depends on the response of TLRs 1/2, TLR 6, and TLR 9 to the bacterium. Polymorphisms in Toll-interacting protein (TOLLIP), a negative regulator of TLR signaling, influence the response of the patient to M. tuberculosis.
Therapeutic Possibilities
Researchers are now looking at various ways to either artificially activate TLRs in order to enhance immune responses or inactivate TLRs to lessen inflammatory disorders. Examples of agents being evaluated in clinical studies or animal studies include:
1. TLR activators to activate immune responses
a. Both TLR-4 and TLR-9 activators are being tried in early clinical trials as vaccine adjuvants to improve the immune response to vaccines. TLR-9 activators are being tried as an adjuvant for the hepatitis B and anthrax vaccines and a TLR-4 activator is being tried as an adjuvant for the vaccine against the human papillomaviruses that cause most cervical cancer.
b. Both TLR-7 and TLR-9 activators are being tried in early clinical trials as an antiviral against hepatitis C. Activation of these TLRs triggers the synthesis and secretion of type I interferons that block viral replication within infected host cells.
c. TLR-9 activators are being tried in early clinical trials as an adjuvant for chemotherapy in the treatment of lung cancer.
d. TLR-9 activators are being tried in early clinical trials to help in the treatment and prevention of allergies and asthma. Activation of TLR-9 in macrophages and other cells stimulates these cells to kill TH2 cells, the subclass of T-helper lymphocytes responsible for most allergies and asthma.
2. TLR inhibitors to suppress immune responses
a. General TLR inhibitors might one day be used to treat autoimmune disorders.
b. A TLR-4 inhibitor, a mimic of the endotoxin from the gram-negative cell wall, is being tried in early clinical trials to block or reduce the death rate from Gram-negative sepsis and SIRS.
c. TLR-4, TLR-2, and MyD88 inhibitors might possibly one day lessen atherosclerotic plaques and the risk of heart disease.
Of course using TLR activators or TLR inhibitors to turn up or turn down immune responses also carries risks. Trying to suppress harmful inflammatory responses may also result in increased susceptibility to infections; trying to activate immune responses could lead to SIRS or autoimmune disease.
Summary
1. In severe bacterial infections, pathogen-associated molecular patterns or PAMPs can trigger the synthesis and secretion of excessive levels of inflammatory cytokines and chemokines leading to systemic inflammatory response syndrome or SIRS.
2. People born with underactive PRRs or deficient PRR immune signaling pathways are at increased risk of infection by specific pathogens due to a decrease innate immune response.
3. People born with overactive PRRs or deficient PRR immune signaling pathways are at increased risk of inflammatory damage by lower numbers of specific pathogens.
4. Researchers are now looking at various ways to either artificially activate underactive PRRs in order to enhance immune responses, or inactivate overactive PRRs to lessen inflammatory disorders. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.4%3A_Early_Induced_Innate_Immunity/11.3D%3A_Harmful_Effects_Associated_with_Abnormal_Pattern-Recognition_Receptor_Responses_Variations_in_Innate_Immune_Signali.txt |
Briefly describe the role of the following as they relate to phagocytosis: inflammation lymph nodules lymph nodes spleen Describe the following steps in phagocytosis: activation chemotaxis attachment (both unenhanced and enhanced) ingestion destruction State what happens when either phagocytes are overwhelmed with microbes or they adhere to cells to large to be phagocytosed. Describe what causes most of the tissue destruction seen during microbial infections. Compare the oxygen-dependent and oxygen-independent killing systems of neutrophils and macrophages. Briefly describe the role of autophagy in removing intracellular microbes.
In addition, Langerhans' cells (immature dendritic cells) are located throughout the epithelium of the skin, the respiratory tract, and the gastrointestinal tract where in their immature form they are attached by long cytoplasmic processes. Upon capturing antigens through pinocytosis and phagocytosis and becoming activated by proinflammatory cytokines, the dendritic cells detach from the epithelium, enter lymph vessels, and are carried to regional lymph nodes. By the time they enter the lymph nodes, they have matured and are now able to present antigen to the ever changing populations of naive T-lymphocytes located in the cortex of the lymph nodes.
The spleen contains many reticular fibers that support fixed macrophages and dendritic cells, as well as ever changing populations of circulating B-lymphocytes and T-lymphocytes. Blood carries microorganisms to the spleen where they are filtered out and phagocytosed by the fixed macrophages and dendritic cells and presented to the circulating B-lymphocytes and T-lymphocytes to initiate adaptive immune responses. There are also specialized macrophages and dendritic cells located in the brain (microglia), lungs (alveolar macrophages), liver (Kupffer cells), kidneys (mesangial cells), bones (osteoclasts), and the gastrointestinal tract (peritoneal macrophages).
The Steps Involved in Phagocytosis
There are a number of distinct steps involved in phagocytosis:
Step 1: Activation of the Phagocyte
Resting phagocytes are activated by inflammatory mediators such as bacterial products (bacterial proteins, capsules, LPS, peptidoglycan, teichoic acids, etc.), complement proteins, inflammatory cytokines, and prostaglandins. As a result, the circulating phagocytes produce surface glycoprotein receptors that increase their ability to adhere to the inner surface of capillary walls, enabling them to squeeze out of the capillary and be attracted to the site of infection.
In addition, they produce endocytic pattern-recognition receptors that recognize and bind to pathogen-associated molecular patterns or PAMPs - components of common microbial molecules such as peptidoglycan, teichoic acids, lipopolysaccharide, and mannose-rich glycans that are not found in human cells - to attach the microbe to the phagocyte for what is called unenhanced attachment (discussed below). They also exhibit increased metabolic and microbicidal activity by increasing their production of ATPs, lysosomal enzymes, lethal oxidants, etc.
Step 2: Chemotaxis of Phagocytes (for wandering macrophages, neutrophils, and eosinophils)
Chemotaxis is the movement of phagocytes toward an increasing concentration of some attractant such as bacterial factors (bacterial proteins, capsules, LPS, peptidoglycan, teichoic acids, etc.), complement proteins (C5a), chemokines (chemotactic cytokines such as interleukin-8 secreted by various cells), fibrin split products, kinins, and phospholipids released by injured host cells.
Movie showing chemotaxis by neutrophil. Chemotaxis of Neutrophils. © From Intimate Strangers: Unseen Life on Earth. Created by Mondo Media. Peter Baker, Executive Producer. Licensed for use, ASM MicrobeLibrary.
You Tube animation summarizing phagocytosis by a macrophage.
You Tube movie illustrating chemotaxis.
Some microbes, such as the influenza A viruses, Mycobacterium tuberculosis, blood invasive strains of Neisseria gonorrhoeae, and Bordetella pertussis have been shown to block chemotaxis.
Step 3: Attachment of the Phagocyte to the Microbe or Cell
Attachment of microorganisms is necessary for ingestion. Attachment may be unenhanced or enhanced.
c. Extracellular trapping with NETs: In response to certain pathogen associated molecular patterns such as LPS, and certain cytokines such as IL-8, neutrophils release DNA and antimicrobial granular proteins. These neutrophil extracellular traps (NETs) bind to bacteria, prevent them from spreading, and kill them with antimicrobial proteins (see Figure \(15\) and Figure \(16\)).
Neutrophil NETS Trapping and Killing Bacteria. In response to certain pathogen associated molecular patterns such as LPS, and certain cytokines such as IL-8, neutrophils release DNA and antimicrobial granular proteins. These neutrophil extracellular traps (NETs) bind to bacteria, prevent them from spreading, and kill them with antimicrobial proteins such as histones and elastins. One hypothesis, shown in this animation, proposes that the NETs are produced by living neutrophils in response to bacteria. Alternately, NETs may be released as a result of necrotic cell death of neutrophils.
Some microorganisms are more resistant to phagocytic attachment.
a. Capsules can resist unenhanced attachment by preventing the endocytic pattern recognition receptors on phagocytes from recognizing the bacterial cell wall components and mannose-containing carbohydrates (see Figure \(14\)). Streptococcus. pneumonia activates the classical complement pathway, but resists C3b opsonization, and complement causes further inflammation in the lungs.
Movie of an encapsulated bacterium resisting engulfment by a neutrophil. Phagocytosis. © James Sullivan, author. Licensed for use, ASM MicrobeLibrary.
b. Some capsules prevent the formation of C3 convertase, an early enzyme in the complement pathways. Without this enzyme, the opsonins C3b and C4b, as well as the other beneficial proteins are not produced.
c. Other capsules, rich in sialic acid, a common component of host cell glycoprotein, have an affinity for serum protein H, a complement regulatory protein that leads to the degradation of the opsonin C3b by factor I and the formation of C3 convertase. (Serum protein H is what normally leads to the degradation of any C3b that binds to host glycoproteins so that we don't stick our own phagocytes to our own cells with C3b.)
d. Some capsules simply cover the C3b that does bind to the bacterial surface and prevent the C3b receptor on phagocytes from making contact with the C3b (see Figure \(3\)). This is seen with the capsule of Streptococcus pneumoniae.
e. Neisseria meningitidis has a capsule composed of sialic acid while Streptococcus pyogenes (group A beta streptococci) has a capsule made of hyaluronic acid. Both of these polysaccharides closely resemble carbohydrates found in human tissue polysaccharides and because they are not recognized as foreign by the lymphocytes that carry out the immune responses, antibodies are not made against these capsules. Likewise, some bacteria are able to coat themselves with host proteins such as fibronectin, lactoferrin, or transferrin and in this way avoid antibodies.
f. An outer membrane molecule of Neisseria gonorrhoeae called Protein II and the M-protein of Streptococcus pyogenes allow these bacteria to be more resistant to phagocytic engulfment. The M-protein of S. pyogenes, for example, binds factor H of the complement pathway and this results in the degradation of the opsonin C3b by factor I and the formation of C3 convertase. S. pyogenes also produces a protease that cleaves the complement protein C5a.
g. Staphylococcus aureus produces protein A while Streptococcus pyogenes produces protein G. Both of these proteins bind to the Fc portion of antibodies (see Figure \(4\)) and in this way the bacteria become coated with antibodies in a way that does not result in opsonization (see Figure \(5\)).
Step 4: Ingestion of the Microbe or Cell by the Phagocyte
Following attachment, polymerization and then depolymerization of actin filaments send pseudopods out to engulf the microbe (see Figure \(6\)) and place it in an endocytic vesicle called a phagosome (see Figure \(7\)).
During this process, an electron pump brings protons (H+) into the phagosome. This lowers the pH within the phagosome to 3.5 - 4.0 so that when a lysosome fuses with the phagosome, the pH is correct for the acid hydrolases to effectively break down cellular proteins. The acidification also releases defensins, cathelicidin, and bacterial permeability inducing protein (BPI), peptides and enzymes that can kill microbes, from a matrix and enabling their activation.
Scanning electron micrographs of a macrophage with pseudopods and a macrophage phagocytozing E. coli on a blood vessel; courtesy of Dennis Kunkel's Microscopy.
Intracellular microbes, such as viruses and bacteria that invade host cells, can also be engulfed once they enter the cytosol of the cell by a process called autophagy. A membrane-bound compartment called an autophagosome grows around the microbe and the surrounding cytosol and subsequently delivers it to lysosomes for destruction (see Figure \(17\)). (This process is also used by eukaryotic cells to engulf and degrade unnecessary or dysfunctional cellular components such as damaged organelles.)
Some microorganisms are more resistant to phagocytic ingestion
a. Pathogenic Yersinia, such as the one that causes plague, contact phagocytes and, by means of a type III secretion system, deliver proteins which depolymerize the actin microfilaments needed for phagocytic engulfment into the phagocytes (see Figure \(8\)). Another Yersinia protein degrades C3b and C5a.
b. Some bacteria, like Mycobacterium tuberculosis, Salmonella, and Listeria monocytogenes can block autophagy.
Blocking Phagosome Formation by Depolymerizing Actin. Molecules of some bacteria, through a type III secretion system, deliver proteins which depolymerize the phagocyte's actin microfilaments used for phagocytic engulfment.
Step 5: Destruction of the Microbe or Cell
Phagocytes contain membranous sacs called lysosomes produced by the Golgi apparatus that contain various digestive enzymes, microbicidal chemicals, and toxic oxygen radicals. The lysosomes travel along microtubules within the phagocyte and fuse with the phagosomes containing the ingested microbes and the microbes are destroyed (see Figure \(9\)).
To view an electron micrograph of a phagolysosome, see the Web page for the University of Illinois College of Medicine.
You Tube animation summarizing phagocytosis by a macrophage.
Some bacteria are more resistant to phagocytic destruction once engulfed.
a. Some bacteria, such as Legionella pneumophilia and Mycobacterium species, cause the phagocytic cell to place them into an endocytic vacuole via a pathway that decreases their exposure to toxic oxygen compounds.
b. Some bacteria, such as Salmonella, are more resistant to toxic forms of oxygen and to defensins (toxic peptides that kill bacteria).
c. Some bacteria, such as Shigella flexneri and the spotted fever Rickettsia, escape from the phagosome into the cytoplasm prior to the phagosome fusing with a lysosome (see Figure \(10\)).
d. Neisseria gonorrhoeae produces Por protein (protein I) that prevents phagosomes from fusing with lysosomes enabling the bacteria to survive inside phagocytes.
e. Some bacteria, such as species of Salmonella, Mycobacterium, Legionella, and Chlamydia, block the vesicular transport machinery that enables the phagosome to fuse with the lysosome.
f. Some bacteria, such as pathogenic Mycobacterium and Legionella pneumophilia, prevent the acidification of the phagosome which is needed for effective killing of microbes by lysosomal enzymes. (Normally after the phagosome forms, the contents become acidified because the lysosomal enzymes used for killing function much more effectively at an acidic pH.)
g. The carotenoid pigments that give Staphylococcus aureus its golden color and group B streptococci (GBS) its orange tint shield the bacteria from the toxic oxidants that neutrophils use to kill bacteria.
h. Cell wall lipids of Mycobacterium tuberculosis, such as lipoarabinomannan, arrest the maturation of phagosomes preventing delivery of the bacteria to lysosomes.
i. Some bacteria are able to kill phagocytes. Bacteria such as Staphylococcus aureus and Streptococcus pyogenes produce the exotoxin leukocidin which damages the cytoplasmic membrane of the phagocyte. On the other hand, bacteria, such as Shigella and Salmonella, induce macrophage apoptosis, a programmed cell death.
If the the infection site contains very large numbers of microorganisms and high levels of inflammatory cytokines and chemokines are being produced in response to PAMPs, the phagocyte will empty the contents of its lysosomes by a process called degranulation in order to kill the microorganisms or cell extracellularly.
These released lysosomal contents, however, also kill surrounding host cells and tissue. Most tissue destruction associated with infections is a result of this process (see Figure \(11\)).
The phagocyte will also empty the contents of its lysosomes for extracellular killing if the cell to which the phagocyte adheres is too large to be engulfed (see Figure \(12\) and Figure \(13\)).
There are 2 killing systems in neutrophils and macrophages: the oxygen-dependent system and the oxygen-independent system.
1. The oxygen-dependent system: production of reactive oxygen species (ROS)
The cytoplasmic membrane of phagocytes contains the enzyme oxidase which converts oxygen into superoxide anion (O2-). This can combine with water by way of the enzyme dismutase to form hydrogen peroxide (H2O2) and hydroxyl (OH) radicals.
In the case of neutrophils, but not macrophages, the hydrogen peroxide can then combine with chloride (Cl2-) ions by the action of the enzyme myeloperoxidase (MPO) to form hypochlorous acid (HOCL), and singlet oxygen.
In macrophages, nitric oxide (NO) can combine with hydrogen peroxide to form peroxynitrite radicals. (In addition to ROS and NO, macrophages secrete inflammatory cytokines such as TNF-alpha, IL-1, IL-8, and IL-12 to promote an inflammatory response.)
These compounds are very microbicidal because they are powerful oxidizing agents which oxidize most of the chemical groups found in proteins, enzymes, carbohydrates, DNA, and lipids. Lipid oxidation can break down cytoplasmic membranes. Collectively, these oxidizing free radicals are called reactive oxygen species (ROS).
Oxidase also acts as an electron pump that brings protons (H+) into the phagosome. This lowers the pH within the phagosome so that when lysosomes fuse with the phagosome, the pH is correct for the acid hydrolases, like elastase, to effectively break down cellular proteins.
In addition to phagocytes using this oxygen-dependant system to kill microbes intracellularly, neutrophils also routinely release these oxidizing agents, as well as acid hydrolases, for the purpose of killing microbes extracellularly. These agents, however, also wind up killing the neutrophils themselves as well as some surrounding body cells and tissues as mentioned above.
2. The oxygen-independent system
Some lysosomes contain defensins ), cationic peptides that alter cytoplasmic membranes; lysozyme, an enzyme that breaks down peptidoglycan, lactoferrin, a protein that deprives bacteria of needed iron; cathepsin G, a protease that causes damage to microbial membranes; elastase, a protease that kills many types of bacteria; cathelicidins, proteins that upon cleavage are directly toxic to a variety of microorganisms; bactericidal permeability inducing protein (BPI ), proteins used by neutrophils to kill certain bacteria by damaging their membranes; collagenase ; and various other digestive enzymes that exhibit antimicrobial activity by breaking down proteins, RNA, phosphate compounds, lipids, and carbohydrates.
Summary
Phagocytosis is the primary method used by the body to remove free microorganisms in the blood and tissue fluids. An inflammatory response to injury and/or infection allows phagocytes to leave the bloodstream, enter the tissue, and go to the site of infection or injury. Microorganisms entering lymph nodules found in the respiratory, gastrointestinal, and genitourinary tract can be phagocytosed by fixed macrophages and dendritic cells and presented to B-lymphocytes and T-lymphocytes to initiate adaptive immune responses.Tissue fluid picks up microbes in the tissue, enters the lymph vessels as lymph, and carries the microbes to regional lymph nodes where they are filtered out and phagocytosed by fixed macrophages and dendritic cells and presented to the circulating B-lymphocytes and T-lymphocytes to initiate adaptive immune responses.
Dendritic cells located throughout the epithelium of the skin, the respiratory tract, and the gastrointestinal tract phagocytize microbes, enter lymph vessels, and carry the microbes to regional lymph nodes where the dendritic cells present antigens associated with the microbes to the ever changing populations of naive T-lymphocytes.Blood carries microorganisms to the spleen where they are filtered out and phagocytosed by fixed macrophages and dendritic cells and presented to the circulating B-lymphocytes and T-lymphocytes to initiate adaptive immune responses. There are also specialized macrophages and dendritic cells located in the brain (microglia), lungs (alveolar macrophages), liver (Kupffer cells), kidneys (mesangial cells), bones (osteoclasts), and the gastrointestinal tract (peritoneal macrophages.
1. Resting phagocytes are activated by inflammatory mediators and produce surface receptors that increase their ability to adhere to the inner surface of capillary walls enabling them to squeeze out of the capillary and enter the tissue, a process called diapedesis.
2. Activation also enables phagocytes to produce endocytic pattern-recognition receptors that recognize and bind to microbial PAMPs in order to attach the microbe to the phagocyte, as well as to exhibit increased metabolic and microbicidal activity.
3. Phagocytes then use chemotaxis to move towards an increasing concentration of some attractant such as bacterial factors or defense molecules.
4. Attachment of phagocytes to the microbes or cells can be through unenhanced attachment or enhanced attachment.
5. Unenhanced attachment is the recognition of pathogen-associated molecular patterns or PAMPs by endocytic pattern-recognition receptors on the surface of the phagocytes.
6. Enhanced attachment, or opsonization, is the attachment of microbes to phagocytes by way of an antibody molecule called IgG, the complement proteins C3b and C4b, and acute phase proteins such as mannose-binding lectin (MBL) and C-reactive protein (CRP).
7. Following attachment, polymerization and then depolymerization of actin filaments send pseudopods out to engulf the microbe and place it in an endocytic vesicle called a phagosome.
8. During this process, an electron pump brings protons (H+) into the phagosome to lowers the pH within the phagosome to a pH that is correct for the acid hydrolases to effectively break down cellular proteins.
9. Phagocytes contain membranous sacs called lysosomes that contain various digestive enzymes, microbicidal chemicals, and toxic oxygen radicals. The lysosomes fuse with the phagosomes containing the ingested microbes and the microbes are destroyed.
10. If the infection site contains very large numbers of microorganisms and high levels of inflammatory cytokines and chemokines are being produced in response to PAMPs, the phagocyte will empty the contents of its lysosomes in order to kill the microorganisms or cell extracellularly.
11. Lysosomal contents released during extracellular killing also kill surrounding host cells and tissue. Most tissue destruction associated with infections is a result of extracellular killing by phagocytes. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.4%3A_Early_Induced_Innate_Immunity/11.3E%3A_Phagocytosis.txt |
Describe how NK cells are able to recognize and kill infected cells and cancer cells lacking MHC-I molecules. State two factors that can result in a nucleated human cell not producing MHC-I molecules. State how iNKT cells recognize glycolipids in order to become activated. Describe the overall function of iNKT cells in terms how they promote both innate and adaptive immunity and may also help to regulate the immune responses.
We will now take a closer look at natural killer (NK) cells and invariant natural killer T-lymphocytes (iNKT cells).
Natural Killer Cells (NK Cells)
NK cells are important in innate immunity because they are able to recognize infected cells, cancer cells, and stressed cells and kill them. In addition, they produce a variety of cytokines, including proinflammatory cytokines, chemokines, colony-stimulating factors, and other cytokines that function as regulators of body defenses. For example, through cytokine production NK cells also suppress and/or activate macrophages , suppress and/or activate the antigen-presenting capabilities of dendritic cells, and suppress and/or activate T-lymphocyte responses.
NK cells use a dual receptor system in determining whether to kill or not kill human cells. When cells are either under stress, are turning into tumors, or are infected, various stress-induced molecules such as MHC class I polypeptide-related sequence A (MICA) and MHC class I polypeptide-related sequence B (MICB) are produced and are put on the surface of that cell.
The first receptor, called the killer-activating receptor, can bind to these stress-induced molecules, and this sends a positive signal that enables the NK cell to kill the cell to which it has bound unless the second receptor cancels that signal.
This second receptor, called the killer-ihibitory receptor, recognizes MHC-I molecules that are usually present on all nucleated human cells. MHC-I molecules, produced by all nucleated cells in the body, possess a deep groove that can bind peptides from proteins found within the cytosol of human cells, transport them to the surface of that cell, and display the MHC-!/peptide complex to receptors on cytotoxic T-lymphocytes or CTLs. If the MHC-I molecules have peptides from the body's own proteins bound to them, CTLs do not recognize those cells as foreign and the cell is not killed. If, on the other hand, the MHC-I molecules have peptides from viral, bacterial, or mutant proteins bound to them, CTLs recognize that cell as foreign and kill that cell. (CTLs will be discussed in greater detail in Unit 6.)
If MHC-I molecules/self peptide complexes are expressed on the cell, the killer-inhibitory receptors on the NK cell recognize this MHC-I/peptide complex and sends a negative signal that overrides the original kill signal and prevents the NK cell from killing the cell to which it has bound (see Figure \(3\)).
Viruses, stress, and malignant transformation, however, can often interfere with the ability of the infected cell or tumor cell to express MHC-I molecules. Without the signal from the killer-inhibitory receptor, the kill signal from the killer-activating signal is not overridden and the NK cell kills the cell to which it has bound (see Figure \(4\)).
The NK cell then releases pore-forming proteins called perforins, proteolytic enzymes called granzymes, and chemokines. Granzymes pass through the pores and activate the enzymes that lead to apoptosis of the infected cell by means of destruction of its structural cytoskeleton proteins and by chromosomal degradation. As a result, the cell breaks into fragments that are subsequently removed by phagocytes (see Figure \(5\)). Perforins can also sometimes result in cell lysis.
Cytokines such as interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) produced by TH1 lymphocytes activate NK cells.
NK cells also play a role in adaptive immune responses. As will be seen in Unit 6, NK cells are also capable of antibody-dependent cellular cytotoxicity or ADCC where they kill cells to which antibody molecules have bound.
Invariant Natural Killer T-Lymphocytes (iNKT Cells)
iNKT cells are a subset of lymphocytes that bridge the gap between innate and adaptive immunity. They have T-cell receptors (TCRs) on their surface for glycolipid antigen recognition. They also have natural killer (NK) cell receptors.
Through the cytokines they produce once activated, iNKT cells are essential in both innate and adaptive immune protection against pathogens and tumors. They also play a regulatory role in the development of autoimmune diseases, asthma, and transplantation tolerance. It has been shown that iNKT cell deficiency or disfunction can lead to the development of autoimmune diseases, human asthma, and cancers.
Pathogens may not directly activate iNKT cells. The TCR of iNKT cells recognize exogenous glycolipid antigens , as well as endogenous self glycolipid antigens presented by MHC-I-like CD1d molecules on antigen presenting dendritic cells. iNKT cells can also be activated by the cytokine interleukin-12 (IL-12) produced by dendritic cells that have themselves become activated by pathogen-associated molecular patterns (PAMPs) of microbes binding to the pattern-recognition receptors (PRRs) of the dendritic cell.
Once activated, the iNKT cells rapidly produce large quantities of cytokines, including interferon-gamma (IFN-?), interleukin-4 (IL-4), interleukin-2 (IL-2), interleukin-10 (IL-10), tumor necrosis factor-alpha (TNF-a), interleukin-13 (IL-13), and chemokines. Through the rapid productions of such cytokines, iNKT cells are able to promote and suppress different innate and adaptive immune responses. For example, large amounts of IFN-? are produced by activated iNKT cells. IFN-? activates NK cells and macrophages as a part of innate immunity.
It has been proposed that if the iNKT cell is repeatedly stimulated by the body's own glycolipids in the absence of microbes that this might stimulate the iNKT cell /dendritic cell interaction to produce tolerizing signals that inhibit the TH1 cell response and possibly stimulate the production of regulatory T-lymphocytes (Treg cells). In this way it might suppress autoimmune responses and prevent tissue damage.
There is also growing evidence that early childhood exposure to microbes is associated with protection against allergic diseases, asthma, and inflammatory diseases such as ulcerative colitis. It has been found that germ-free mice have large accumulations of mucosal iNKT cells in the lungs and intestines and increased morbidity from allergic asthma and inflammatory bowel disease. However, colonization of neonatal germ-free mice with normal microbiota resulted in mucosal iNKT cell tolerance to these diseases. It has been proposed that microbes the human body has been traditionally exposed to from early childhood throughout most of human history might play a role in developing normal iNKT cell numbers and iNKT cell responses.
iNKT cells will be discussed in further detail in Unit 6.
Summary
1. Natural Killer (NK) cells are able to recognize infected cells, cancer cells, and stressed cells and kill them. In addition, they produce a variety of cytokines, including proinflammatory cytokines, chemokines, colony-stimulating factors, and other cytokines that function as regulators of body defenses.
2. When body cells are either under stress, are turning into tumors, or are infected, various stress-induced molecules are produced and are put on the surface of that cell.
3. NK cells use a dual receptor system in determining whether to kill or not kill human cells.
4. The first receptor, called the killer-activating receptor, can bind to these stress-induced molecules, and this sends a positive signal that enables the NK cell to kill the cell to which it has bound unless the second receptor cancels that signal.
5. The second receptor, called the killer-ihibitory receptor, recognizes MHC-I molecules that are usually present on all nucleated human cells. If MHC-I molecules/self peptide complexes are expressed on the cell, the killer-inhibitory receptors on the NK cell recognize this MHC-I/peptide complex and sends a negative signal that overrides the original kill signal and prevents the NK cell from killing the cell to which it has bound.
6. Viruses, stress, and malignant transformation can often interfere with the ability of the infected cell or tumor cell to express MHC-I molecules. Without the signal from the killer-inhibitory receptor, the kill signal from the killer-activating signal is not overridden and the NK cell kills the cell to which it has bound.
7. NK cells kill their target cells by inducing apoptosis, a programmed cell suicide.
8. NK cells also play a role in adaptive immune responses by way of antibody-dependent cellular cytotoxicity or ADCC where they bind to and kill cells to which antibody molecules have bound.
9. Invariant natural killer T-lymphocytes (iNKT cells) are a subset of lymphocytes that have T-cell receptors on their surface for glycolipid antigen recognition. They also have natural killer (NK) cell receptors.
10. Through the cytokines they produce, iNKT cells are able to promote and suppress different innate and adaptive immune responses. They also play a regulatory role in the development of autoimmune diseases, asthma, and transplantation tolerance. iNKT cell deficiency or disfunction can lead to the development of autoimmune diseases, human asthma, and cancers. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.4%3A_Early_Induced_Innate_Immunity/11.3F%3A_Natural_Killer_Cells_%28NK_Cells%29_and_Invariant_Natural_Killer_T-Lymphocytes_%28iNKT_Cells%29.txt |
Describe the 4 processes that make up the inflammatory mechanism. Briefly describe the various beneficial effects of inflammation that are associated with plasma leakage and with diapedesis. Briefly describe the process of diapedesis, indicating the role of P-selectins, integrins, and adhesion molecules. Briefly describe the healing stage of inflammation. Briefly describe the problems that arise from chronic inflammation.
The inflammatory response is an attempt by the body to restore and maintain homeostasis after injury and is an integral part of body defense. Most of the body defense elements are located in the blood and inflammation is the means by which body defense cells and defense chemicals leave the blood and enter the tissue around the injured or infected site. Inflammation is essentially beneficial, however, excess or prolonged inflammation can cause harm.
The Mechanism of Inflammation
Essentially, four processes make up the inflammatory mechanism:
a. Smooth muscles around larger blood vessels contract to slow the flow of blood through the capillary beds at the infected or injured site. This gives more opportunity for leukocytes to adhere to the walls of the capillary and squeeze out into the surrounding tissue.
b. The endothelial cells that make up the wall of the smaller blood vessels contract. This increases the space between the endothelial cells resulting in increased capillary permeability. Since these blood vessels get larger in diameter as a result of this, the process is called vasodilation (see Figure \(1\)).
Scanning electron micrographs of a cross section of a capillary showing an endothelial cell and a capillary with a red blood cell; courtesy of Dennis Kunkel's Microscopy).
c. Molecules called selectins are produced on the membrane of the leukocyte and are able to reversibly bind to corresponding selectin glycoprotein receptors on the inner wall of the venule. This reversible binding enables the leukocyte to roll along the inner wall of the venule. This reversible binding enables the leukocyte to roll along the inner wall of the venule. Adhesion molecules are activated on the surface of the endothelial cells on the inner wall of the capillaries. Corresponding molecules on the surface of leukocytes called integrins attach to these adhesion molecules allowing the leukocytes to flatten and squeeze through the space between the endothelial cells. This process is called diapedesis or extravasation.
d. Activation of the coagulation pathway causes fibrin clots to physically trap the infectious microbes and prevent their entry into the bloodstream. This also triggers blood clotting within the surrounding small blood vessels to both stop bleeding and further prevent the microorganisms from entering the bloodstream.
You Tube movie and animation of leukocyte extravasation (diapedesis)
from ImmuneDocumentary
These four events are triggered and enhanced by a variety of chemical inflammatory mediators. We will now divide the inflammatory response into two stages: early inflammation and late inflammation.
Early Inflammation and Diapedesis
Most leukocyte diapedesis (extravasation) occurs in post-capillary venules because hemodynamic shear forces are lower in these venules. This makes it easier for leukocytes to attach to the inner wall of the vessel and squeeze out between the endothelial cells. We will look at this process in more detail below.
1. During the very early stages of inflammation, stimuli such as injury or infection trigger the release of a variety of mediators of inflammation such as leukotrienes, prostaglandins, and histamine. The binding of these mediators to their receptors on endothelial cells leads to vasodilation, contraction of endothelial cells, and increased blood vessel permeability. In addition, the basement membrane surrounding the capillaries becoming rearranged so as to promote the migration of leukocytes and the movement of plasma macromolecules from the capillaries into the surrounding tissue. Mast cells in the connective tissue as well as basophils, neutrophils and platelets leaving the blood from injured capillaries, release or stimulate the synthesis of vasodilators such as histamine, leukotrienes, kinins, and prostaglandins. Certain products of the complement pathways (C5a and C3a) can bind to mast cells and trigger their release their vasoactive agents. In addition, tissue damage activates the coagulation cascade and production of inflammatory mediators like bradykinins.
2. The binding of histamine to histamine receptors on endothelial cells triggers an upregulation of P-selectin molecules and platelet-activating factor or PAF on the endothelial cells that line the venules.
3. The P-selectins then are able to reversibly bind to corresponding P-selectin glycoprotein ligands (PSGL-1) on leukocytes. This reversible binding enables the leukocyte to now roll along the inner wall of the venule.
4. The binding of PAF to its corresponding receptor PAF-R on the leukocyte upregulates the surface expression of an integrin called leukocyte function-associated molecule-1 (LFA-1) on the surface of the leukocyte.
5. The LFA-1 molecules on the rolling leukocytes can now bind firmly to an an adhesion molecule called intercellular adhesion molecule-1 (ICAM-1) found on the surface of the endothelial cells forming the inner wall of the blood vessel (see Figure \(4\)).
6. The leukocytes flatten out, squeeze between the constricted endothelial cells, and use enzymes to breakdown the matrix that forms the basement membrane surrounding the blood vessel. The leukocytes then migrate towards chemotactic agents such as the complement protein C5a and leukotriene B4 generated by cells at the site of infection or injury (see Figure \(5\)).
Late Inflammation and Diapedesis
1) Usually within two to four hours of the early stages of inflammation, activated macrophages and vascular endothelial cells release inflammatory cytokines such as TNF and IL-1 when their toll-like receptors bind pathogen-associated molecular patterns - molecular components associated with microorganisms but not found as a part of eukaryotic cells. This enables vascular endothelial cells of nearby venules to increase their expression of adhesion molecules such as P-selectins, E-selectins, intercellular adhesion molecules (ICAMs), and chemokines.
2) The binding of TNF and IL-1 to receptors on endothelial cells triggers an maintains the inflammatory response by upregulation the production of the adhesion molecule E-selectin and maintaining P-selectin expression on the endothelial cells that line the venules.
3). The E-selectins on the inner surface of the endothelial cells can now bind firmly to its corresponding integrin E-selectin ligand-1 (ESL-1) on leukocytes (see Figure \(4\)).
4) The leukocytes flatten out, squeeze between the constricted endothelial cells, and move across the basement membrane as they are are attracted towards chemokines such as interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1) generated by cells at the site of infection or injury (see Figure \(5\)). Leakage of fibrinogen and plasma fibronectin then forms a molecular scaffold that enhances the migration and retention of leukocytes at the infected site.
Benefits of Inflammation
As a result of this increased permeability:
a. Plasma flows out of the blood into the tissue.
Beneficial molecules in the plasma (see Figure \(2\)) include:
1. Clotting factors. Tissue damage activates the coagulation cascade causing fibrin clots to form to localize the infection, stop the bleeding, and chemotactically attract phagocytes.
2. Antibodies. These help remove or block the action of microbes through a variety of methods that will be explained in Unit 6.
3. Proteins of the complement pathways. These, in turn: 1) stimulate more inflammation (C5a, C3a, and C4a), 2) stick microorganisms to phagocytes (C3b and C4b), 3) chemotactically attract phagocytes ( C5a), and 4) lyse membrane-bound cells displaying foreign antigens (membrane attack complex or MAC).
4. Nutrients. These feed the cells of the inflamed tissue.
5. Lysozyme, cathelicidins, phospholipase A2,and human defensins. Lysozyme degrades peptidoglycan. Cathelicidins are cleaved into two peptides that are directly toxic to microbes and can neutralize LPS from the gram-negative bacterial cell wall. Phospholipase A2 hydrolyzes the phospholipids in the bacterial cytoplasmic membrane. Human defensins put pores in the cytoplasmic membranes of many bacteria. Defensins also activate cells involved in the inflammatory response.
6. Transferrin.Transferrin deprives microbes of needed iron.
b. Leukocytes enter the tissue through a process called diapedesis or extravasation, discussed above under early inflammation and late inflammation.
Benefits of diapedesis include (see Figure \(2\)):
1. Increased phagocytosis. Neutrophils, monocytes that differentiate into macrophages when they enter the tissue, and eosinophils are phagocytic leukocytes.
2. More vasodilation. Basophils, eosinophils, neutrophils, and platelets enter the tissue and release or stimulate the production of vasoactive agents that promote inflammation.
3. Cytotoxic T-lymphocytes (CTLs), effector T4-cells, and NK cells enter the tissue to kill cells such as infected cells and cancer cells that are displaying foreign antigens on their surface (discussed in Unit 6).
Cytokines called chemokines are especially important in this part of the inflammatory response. They play key roles in diapedesis -enabling white blood cells to adhere to the inner surface of blood vessels, migrate out of the blood vessels into the tissue, and be chemotactically attracted to the injured or infected site. They also trigger extracellular killing by neutrophils.
Finally, within 1 to 3 days, macrophages release the cytokines interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-a). These cytokines stimulate NK cells and T-lymphocytes to produce the cytokine interferon-gamma. (IF-?). The IF-? then binds to receptors on macrophages causing them to produce fibroblast growth factor and angiogenic factors for tissue remodeling. With the proliferation of endothelial cells and fibroblasts, endothelial cells form a fine network of new capillaries into the injured area to supply blood, oxygen, and nutrients to the inflamed tissue. The fibroblasts deposit the protein collagen in the injured area and form a bridge of connective scar tissue to close the open, exposed area. This is called fibrosis or scarring, and represents the final healing stage.
Inflammation is normally carefully regulated by cytokines. Inflammatory cytokines such as interferon-gamma and interleukin-12 enhance the inflammatory response whereas the cytokine interleukin-10 inhibits inflammation by decreasing the expression of inflammatory cytokines.
So as can be seen, acute inflammation is essential to body defense. Chronic inflammation, however, can result in considerable tissue damage and scarring. With prolonged increased capillary permeability, neutrophils continually leave the blood and accumulate in the tissue at the infected or injured site. As they discharge their lysosomal contents and reactive oxygen species or ROS, surrounding tissue is destroyed and eventually replaced with scar tissue. Anti-inflammatory agents such as antihistamines or corticosteroids may have to be given to relieve symptoms or reduce tissue damage.
For example, as learned in Unit 3, during severe systemic infections with large numbers of microorganisms present, high levels of pathogen-associated molecular patterns (PAMPs) are released resulting in excessive cytokine production by macrophages and this can harm the body. In addition, neutrophils start releasing their proteases and reactive oxygen species that kill not only the bacteria, but the surrounding tissue as well. Harmful effects include high fever, hypotension, tissue destruction, wasting, acute respiratory distress syndrome or ARDS, disseminated intravascular coagulation or DIC, damage to the vascular endothelium, hypovolemia, and reduced perfusion of blood through tissues and organs resulting to shock, multiple system organ failure (MOSF), and often death. This excessive inflammatory response is referred to as Systemic Inflammatory Response Syndrome or SIRS or the Shock Cascade.
Briefly describe the mechanisms that enable to slow the flow of blood at an infection site and get phagocytes, complement proteins and antibodies to the infection site. Why is it important to deliver plasma to an infection site? Why is it important for diapedesis to occur during inflammation?
Chronic inflammation also contributes to heart disease, Alzheimer's disease, diabetes, and cancer.
• In the case of cancer,it is proposed that when macrophages produce inflammatory cytokines, such as TNF-alpha, these cytokines activate a gene switch in the cancer cell that turns on the synthesis of proteins that promote cell replication and inflammation while blocking apoptosis of the cancer cell.
• In heart disease, it is thought that macrophages digest low density lipoprotein or LDL, the bad cholesterol, and are then encased in a fibrous cap that forms arterial plaque.
• With diabetes, it is thought that the metabolic stress of obesity triggers innate immune cells and fat cells to produce cytokines such as TNF-alpha that can interfere with the normal function of insulin.
• In the case of Alzheimer's disease, microglial cells, macrophage-like cells in the brain, interact with the beta-amyloid proteins that build up in neurons of those with Alzheimer's and subsequently produce inflammatory cytokines and free radicals that destroy the neurons.
Summary
1. Most of the body defense elements are located in the blood and inflammation is the means by which body defense cells and defense chemicals leave the blood and enter the tissue around the injured or infected site.
2. As part of the mechanism for inflammation, smooth muscles around larger blood vessels contract to slow the flow of blood through the capillary beds at the infected or injured site. This gives more opportunity for leukocytes to adhere to the walls of the capillary and squeeze out into the surrounding tissue.
3. As part of the mechanism for inflammation, the endothelial cells that make up the wall of the smaller blood vessels contract. This increases the space between the endothelial cells resulting in increased capillary permeability.
4. As part of the mechanism for inflammation, adhesion molecules are activated on the surface of the endothelial cells on the inner wall of the capillaries and corresponding molecules on the surface of leukocytes called integrins attach to these adhesion molecules allowing the leukocytes to flatten and squeeze through the space between the endothelial cells. This process is called diapedesis or extravasation.
5. As part of the mechanism for inflammation, activation of the coagulation pathway causes fibrin clots to physically trap the infectious microbes and prevent their entry into the bloodstream.
6. Acute inflammation is essential to body defense.
7. As a result of this increased permeability, plasma flows out of the blood into the tissue delivering clotting factors, antibody molecules, complement pathway proteins, nutrients, antibacterial enzymes and peptides, and transferrin for innate body defense.
8. As a result of this increased permeability, leukocytes enter the tissue delivering phagocytic cells, inflammation-inducing cells, cytotoxic T-lymphocytes, effector T4-lymphocytes, and NK cells.
9. Inflammatory cytokines also, enable endothelial cells form a fine network of new capillaries into the injured area to supply blood, oxygen, and nutrients to the inflamed tissue, and enable fibroblasts to deposit the protein collagen in the injured area and form a bridge of connective scar tissue to close the open, exposed area.
10. Chronic inflammation can result in considerable tissue damage and scarring, primarily to extracellular killing by phagocytes and hypoperfusion.
11. Chronic inflammation is thought to also contribute to heart disease, Alzheimer's disease, diabetes, and cancer. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.4%3A_Early_Induced_Innate_Immunity/11.3G%3A_Inflammation.txt |
Learning Objectives
• Describe at least four ways the body deprives microorganisms of iron.
We will now take a closer look at nutritional immunity. Iron is needed as a cofactor for certain enzymes in both bacteria and humans. Both bacteria and human cells produce iron chelators that trap free iron from their environment and transport it into the cell. During infection, the body makes considerable metabolic adjustment in order to make iron unavailable to microorganisms. Much of this is due to production of a defense chemical called leukocyte-endogenous mediator (LEM). As a result of infection, there is:
1. decreased intestinal absorption of iron from the diet;
2. a decrease of iron in the plasma and an increase in iron in storage as ferritin;
3. increased synthesis of the human iron-binding proteins (iron chelators) such as lactoferrin, transferrin, ferritin, and hemin that trap iron for use by human cells while making it unavailable to most microbes;
4. coupled with the febrile response, decreased ability of bacteria to synthesize their own iron chelators called siderophores;
5. prior stationing of lactoferrin at common sites of microbial invasion such as in the mucous of mucous membranes, and the entry of transferrin into the tissue during inflammation.
This lack of iron, which is needed as a cofactor for certain enzyme reactions, can inhibit the growth of many bacteria.
As seen in Unit 3, some bacteria produce in addition to their own siderophore, receptors for siderophores of other bacteria in this way take iron from other bacteria. Furthermore, a number of pathogenic bacteria are able to bind human transferrin, lactoferrin, ferritin, and hemin and use that as their iron source. For example, Neisseria gonorrhoeae, Neisseria meningitidis, and Haemophilus influenzaeare able to use iron bound to human transferrin and lactoferrin for their iron needs, while pathogenic Yersinia species are able to use transferrin and hemin as iron sources. Borrelia burgdorferi doesn't even use iron as a cofactor, but instead uses manganese. Furthermore, a number of bacteria are able to produce exotoxins that kill host cells only when iron concentrations are low. Perhaps in this way the bacteria can gain access to the iron that was in those cells.
Summary
1. Iron is needed as a cofactor for certain enzymes in both bacteria and humans.
2. Both bacteria and human cells produce iron chelators that trap free iron from their environment and transport it into the cell.
3. During infection, the body makes considerable metabolic adjustment in order to make iron unavailable to microorganisms.
4. The lack of iron can inhibit the growth of many bacteria.
5. Some bacteria in addition to their own siderophores, produce receptors for iron chelators of other bacteria and/or human cells and in this way take iron being trapped for use by other organisms.
6. A number of bacteria are able to produce toxins that kill host cells only when iron concentrations are low and in this way gain access to the iron that was in those cells.
11.3I: Fever
Describe the mechanism behind fever induction and indicate its possible benefits. Define hyperpyrexia.
Activated macrophages and other leukocytes release inflammatory cytokines such as TNF-alpha, IL-1, and IL-6 when their pattern-recognition receptors (PRRs) bind pathogen associated molecular patterns or PAMPs - molecular components associated with microorganisms but not found as a part of eukaryotic cells. These include bacterial molecules such as peptidoglycan, teichoic acids, lipopolysaccharide, mannans, flagellin, and bacterial DNA. There are also pattern-recognition molecules for viral double-stranded RNA (dsRNA) and fungal cell walls components such as lipoteichoic acids, glycolipids, mannans, and zymosan.
These cytokines stimulate the anterior hypothalamus of the brain, the part of the brain that regulates body temperature, to produce prostaglandin E2, which leads to an increase bodily heat production and increased vasoconstriction. This, in turn, decreases the loss of heat from the skin and increases body temperature. Up to a certain point, fever is beneficial:
1. Fever increases the environmental temperature above the optimum growth temperature for many microorganisms. If the microorganisms are growing more slowly, the body's defenses have a better chance of removing them all.
2. Fever leads to the production of heat shock proteins that are recognized by some intraepithelial T-lymphocytes called delta gamma T-cells, resulting in the production of inflammation-promoting cytokines.
3. Fever elevates the temperature of the body increasing the rate of enzyme reactions, and speeding up metabolism within the body. An elevation in the rate of metabolism can increase the production and activity of phagocytes, speed up the multiplication of lymphocytes, increase the rate of antibody and cytokine production, increase the rate at which leukocytes are released from the bone marrow into the bloodstream, and speed up tissue repair.
Too high of a body temperature, however, may cause damage by denaturing the body's enzymes. Hyperpyrexia is a fever with an extreme elevation of body temperature greater than or equal to 41.5 °C (106.7 °F). Body temperature this elevated often indicates a serious underlying condition and may lead to potentially hazardous side effects. As a result, hyperpyrexia is considered as a medical emergency.
Summary
1. Activated macrophages and other leukocytes release inflammatory cytokines such as TNF-alpha, IL-1, and IL-6 when their pattern-recognition receptors (PRRs) bind pathogen associated molecular patterns or PAMPs.
2. These cytokines stimulate the anterior hypothalamus of the brain, the part of the brain that regulates body temperature, to produce prostaglandin E2, which leads to an increase bodily heat production and increased vasoconstriction.
3. Vasoconstriction decreases the loss of heat from the skin and increases body temperature.
4. Fever increases the environmental temperature above the optimum growth temperature for many microorganisms.
5. Fever leads to the production of heat shock proteins that are recognized by some intraepithelial T-lymphocytes resulting in the production of inflammation-promoting cytokines.
6. Fever elevates the temperature of the body increasing the rate of enzyme reactions, and speeding up metabolism within the body including that involved in innate and adaptive immunity as well as tissue repair. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.4%3A_Early_Induced_Innate_Immunity/11.3H%3A_Nutritional_Immunity.txt |
Briefly describe the mechanism behind the acute phase response. State the functions of the following acute phase proteins: C-reactive protein mannose-binding lectin
We will now take a closer look at the acute phase response. The acute phase response is an innate body defense seen during acute illnesses and involves the increased production of certain blood proteins termed acute phase proteins.
Activated macrophages and other leukocytes release inflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), and interleukin-6 (IL-6) when their pattern-recognition receptors (PRRs) bind pathogen associated molecular patterns or PAMPs - molecular components associated with microorganisms but not found as a part of eukaryotic cells. These include bacterial molecules such as peptidoglycan, teichoic acids, lipopolysaccharide, mannans, flagellin, pilin, and bacterial DNA. There are also pattern-recognition molecules for viral double-stranded RNA (dsRNA) and fungal cell walls components such as lipoteichoic acids, glycolipids, mannans, and zymosan.
These cytokines travel through the blood and stimulate hepatocytes in the liver to synthesize and secrete acute phase proteins. This response provides an early defense and enables the body to recognize foreign substances early on in the infection process prior to the full activation and implementation of the immune responses. Two important acute phase proteins are C-reactive protein and mannose-binding protein. They function as soluble pattern-recognition receptors.
1. C-reactive protein (CRP) binds to the phosphorylcholine portion of teichoic acids and lipopolysaccharides of bacterial and fungal cell walls. It also binds to the phosphocholine found on the surface of damaged or dead human cells. It functions as an opsonin, sticking the microorganism to phagocytes, and activates the classical complement pathway by binding C1q, the first component in the pathway.
2. Mannan-bindinglectin (MBL) - also known as mannan-binding protein or MBP -binds to mannose-richglycans (short carbohydrate chains with the sugar mannose or fructose as the terminal sugar). These are common in microbial glycoproteins and glycolipids but rare in those of humans. It functions as an opsonin, sticking the microorganism to phagocytes, and activates the lectin pathway.
Products of the complement pathways, in turn, promote inflammation, attach microbes to phagocytes, cause to MAC cytolysis, and chemotactically attract phagocytes to the infected area.
Summary
1. The acute phase response is an innate body defense seen during acute illnesses and involves the increased production of certain blood proteins termed acute phase proteins.
2. Inflammatory cytokines produced during innate immunity travel through the blood and stimulate hepatocytes in the liver to synthesize and secrete acute phase proteins.
3. Two important acute phase proteins are C-reactive protein and mannose-binding protein, both functioning as soluble pattern-recognition receptors.
4. C-reactive protein (CRP) binds to certain PAMPs bacterial and fungal cell walls as well as to phosphocholine found on the surface of damaged or dead human cells.
5. CRP functions as an opsonin, sticking the microorganism to phagocytes, and activates the classical complement pathway by binding C1q, the first component in the pathway.
6. Mannan-binding lectin (MBL) - also known as mannan-binding protein or MBP - binds to mannose-rich glycans on microbial cell walls.
7. MBL functions as an opsonin, sticking the microorganism to phagocytes, and activates the lectin pathway. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.4%3A_Early_Induced_Innate_Immunity/11.3J%3A_The_Acute_Phase_Response.txt |
Briefly describe how intraepithelial T-lymphocytes (gamma:delta T-lymphocytes) play a role in innate immunity. Briefly describe how B-1 cells play a role in innate immunity.
We will now take a closer look at Intraepithelial T-lymphocytes (e.g., T4 and T8) and B-1 cells. Most of the T-lymphocytes and B-lymphocytes in the body are involved in the adaptive immune responses that will be discussed in Unit 6. In adaptive immunity, specific receptors on T-lymphocytes (T-cell receptors or TCRs) and B-lymphocytes (B-cell receptors or BCRs) recognize specific antigens of specific microbes.
Intraepithelial T-lymphocytes and B-1 cells, on the other hand, are subpopulations of T-lymphocytes and B-lymphocytes that possess a more limited diversity of receptors and are designed to directly recognize the more common microbes that enter the epidermis or the mucosal epithelia. As such, they function more as effector cells for innate immunity rather than adaptive immunity.
1. Intraepithelial T-lymphocytes (IELs) are found in the epidermis of the skin and the mucosal epithelia. These T-lymphocytes, known as gamma:delta T-lymphocytes, differ from the T-lymphocytes (alpha:beta T-lymphocytes) associated with adaptive immunity. The alpha:beta T-lymphocytes are designed to recognize peptide antigens bound to MHC-I molecules of infected cells and tumor cells.
Although their exact function is unknown, it has been proposed that they recognize molecules associated with epithelial cells but expressed only when those cells are infected, such as MHC-I molecules and heat shock proteins. They then trigger apoptosis of these stressed or infected cells using perforins and granzymes similar to cytotoxic T-lymphocytes (CTLs) of adaptive immunity. Rather than recognizing antigens specific to an infectious microorganism, they recognize molecules associated with the epithelium as a consequence of infection. Their T-cell receptors may also function as PRRs for recognizing certain PAMPs. As such, they function more as effector cells for innate immunity rather than adaptive immunity. They probably help defend the body by producing cytokines that play a variety of roles in body defense. IELs are also thought to aid in repair of mucous membranes following inflammatory damage. Excessive or inappropriate activation of IELs can also lead to damage of the intestines as in the case of celiac disease.
2. B-1 lymphocytes, or B-1 cells are found mostly in the peritoneal and pleural cavities . B-1 cells have a limited diversity of antigen receptors that initially produce a class of antibody molecule called IgM against common polysaccharide and lipid antigens of microbes and against PAMPs. As such they function more as effector cells for innate immunity rather than adaptive immunity. Antibodies produced by B-1 cells are often called natural antibodies that help to protect against bacteria in body cavities. Similar B-lymphocytes called marginal zone B cells are found in the marginal zone of the white pulp of the spleen. These are thought to make IgM to protect against bacteria that enter the bloodstream.
Summary
1. Most of the T-lymphocytes and B-lymphocytes in the body are involved in the adaptive immune responses wherein specific receptors on T-lymphocytes (T-cell receptors or TCRs) and B-lymphocytes (B-cell receptors or BCRs) recognize specific antigens of specific microbes.
2. Intraepithelial T-lymphocytes and B-1 cells, however, are subpopulations of T-lymphocytes and B-lymphocytes that possess a more limited diversity of receptors and are designed to directly recognize the more common microbes that enter the epidermis or the mucosal epithelia and function more as effector cells for innate immunity rather than adaptive immunity.
3. Intraepithelial T-lymphocytes (IELs) are found in the epidermis of the skin and the mucosal epithelia.
4. It has been proposed that they recognize molecules such as MHC-I molecules and heat shock proteins associated with epithelial cells but expressed only when those cells are infected and trigger apoptosis of these stressed or infected cells. They may also aid in repair of mucous membranes following inflammatory damage.
5. B-1 lymphocytes, or B-1 cells, are found mostly in the peritoneal and pleural cavities.
6. B-1 cells have a limited diversity of antigen receptors that initially produce a class of antibody molecule called IgM against common polysaccharide and lipid antigens of microbes and against PAMPs.
7. Similar B-lymphocytes called marginal zone B cells are found in the spleen.\ and are thought to make IgM to protect against bacteria that enter the bloodstream. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.4%3A_Early_Induced_Innate_Immunity/11.3K%3A_Intraepithelial_T-lymphocytes_and_B-1_cells.txt |
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied.
11.1: The Innate Immune System: An Overview
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe what is meant by the following:
1. innate immunity (ans)
2. adaptive (acquired) immunity (ans)
2. Define the following:
1. antigen (ans)
2. pathogen-associated molecular patterns or PAMPs (ans)
3. epitope (ans)
3. Multiple Choice (ans)
11.2: Defense Cells in the Blood: The Leukocytes
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. What is the difference between a CBC and a leukocyte differential count? (ans)
2. A person has an elevated white blood cell count with anelevated number of band-form neutrophils. What is the significance of this? (ans)
3. Match the following descriptions and functions with the type of leukocytes:
_____ Important phagocytes; 54%-75% of the leukocytes; granules stain poorly; produce enzymes for the synthesis of bradykinins and prostaglandins that promote inflammation. (ans)
_____ Capable of phagocytosis but primarily kill microorganisms and parasitic worms extracellularly; 1%-4% of the leukocytes; large granules stain red; secrete leukotriens and prostaglandins to promote inflammation. (ans)
_____ Not important in phagocytosis; large granules stain a purplish blue; 0%-1% of the leukocytes; release histamine, leukotriens, and prostaglandins to promote inflammation. (ans)
_____ Important in phagocytosis and aid in the adaptive immune responses; produce cytokines; 4%-8% of the leukocytes; differentiate into macrophages and dendritic cells when they leave the blood and enter the tissue. (ans)
_____ Mediate humoral immunity (antibody production); have B-cell receptors (BCR) on their surface for antigen recognition; differentiate into antibody-secreting plasma cells. (ans)
_____ Regulate the adaptive immune responses through cytokine production; have CD4 molecules and TCRs on their surface for antigen recognition. (ans)
_____ Carry out cell-mediated immunity; have CD8 molecules and TCRs on their surface for antigen recognition; differentiate into cytotoxic T-lymphocytes (CTLs). (ans)
_____ Lymphocytes that lack B-cell receptors and T-cell receptors; kill cells to which the antibody IgG has attached as well as human cells lacking MHC-I molecules on their surface. (ans)
1. B-lymphocytes
2. T4-lymphocytes
3. T8-lymphocytes
4. NK cells
5. basophils
6. neutrophils
7. eosinophils
8. monocytes
4. State what type of cell monocytes differentiate into when they enter tissue. (ans)
5. Multiple Choice (ans)
11.3: Defense Cells in the Tissue: Dendritic Cells, Macrophages, and Mast Cells
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. State 3 different functions of macrophages in body defense.
1. (ans)
2. (ans)
3. (ans)
2. Name the cells in the tissue whose primary function is to present antigen to naive T-lymphocytes. (ans)
3. Name the cells in the tissue whose primary function is to present antigen to effector T-lymphocytes. (ans)
4. State the primary function of mast cells in body defense. (ans)
5. Multiple Choice (ans)
11.3: Immediate Innate Immunity
11.3A: Antimicrobial Enzymes and Antimicrobial Peptides
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Matching
____ Found in in tears, mucous, saliva, plasma, tissue fluid, etc.; breaks down peptidoglycan. (ans)
____ A protein produced by skin and mucosal epithelial cells. The two peptides produced upon cleavage of this protein are directly toxic to a variety of microorganisms. (ans)
____ An enzyme that penetrates the bacterial cell wall and hydrolizes the phospholipids in the bacterial cytoplasmic membrane. (ans)
____ Short cationic peptides that are directly toxic by disrupting the cytoplasmic membrane of a variety of microorganisms causing leakage of cellular needs. They also activate cells for an inflammatory response. (ans)
1. lysozyme
2. phospholipase A2
3. defensins
4. cathelicidins
5. lactotransferrin and transferrin
11.3B: The Complement System
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe how the classical complement pathway is activated. (ans)
2. Match the following:
_____ Complement proteins that trigger inflammation (ans)
_____ Complement proteins that chemotactically attracting phagocytes to the infection site. (ans)
_____ Complement proteins that promote the attachment of antigens to phagocytes (enhanced attachment or opsonization. (ans)
_____ Complement proteins that cause lysis of Gram-negative bacteria and human cells displaying foreign epitopes. (ans)
1. the membrane attack complex (MAC)
2. C5a. and to a lesser extent, C3a and C4a.
3. C3b, and to a lesser extent, C4b.
4. C5a
3. Briefly describe how the lectin complement pathway is activated. (ans)
4. Briefly describe how the alternative complement pathway is activated. (ans)
5. Multiple Choice (ans)
11.3C: Anatomical Barriers to Infection, Mechanical Removal of Microbes, and Bacterial Antagonism by Normal Body Microbiota
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe what is meant by anatomical barriers to infection. (ans)
2. List 4 ways in which the body can physically remove microorganisms or their products. (ans)
3. Describe how bacterial antagonism by normal microbiota acts as a nonspecific body defense mechanism. (ans)
4. Multiple Choice (ans)
11.4: Early Induced Innate Immunity
11.3A: Pathogen-Associated Molecular Patterns (PAMPs) and Danger-Associated Molecular Patterns (DAMPs)
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. State the function of pathogen-associated molecular patterns as they relate to innate immunity. (ans)
2. Name at least 5 PAMPS associated with bacteria. (ans)
3. Name at least 2 PAMPS associated with viruses. (ans)
4. Define DAMP. (ans)
5. Multiple Choice PAMPs and DAMPs (ans)
11.3B: Pattern-Recognition Receptors (PRRs)
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. State the function of the following as they relate to innate immunity.
1. pathogen-associated molecular patterns (ans)
2. pattern recognition receptors (ans)
3. endocytic pattern recognition receptors (ans)
4. signaling pattern recognition receptors (ans)
5. danger-associated molecular patterns
6. danger recognition receptors (ans)
7. inflammasome (ans)
2. Briefly describe the major difference between the effect of the cytokines produced in response to PAMPs that bind to cell surface signaling PRRs and endosomal PRRs. (ans)
3. Multiple Choice (PRRs) (ans)
11.3C: Cytokines Important in Innate Immunity
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following:
_____ Cytokines that promote inflammation by enabling white blood cells to adhere to the inner surface of blood vessels, migrate out of the blood vessels into the tissue, and be chemotactically attracted to the injured or infected site. (ans)
_____ Cytokines that prevent viral replication, activate a variety of cells important in body defense, and exhibit some anti-tumor activity. (ans)
_____ A wide variety of intercellular regulatory proteins produced by many different cells in the body that ultimately control every aspect of body defense. Cytokines activate and deactivate phagocytes and immune defense cells, increase or decrease the functions of the different immune defense cells, and promote or inhibit a variety of nonspecific body defenses. (ans)
1. lysozyme
2. chemokines
3. cytokines
4. interferons
5. human beta-defensins
2. Describe specifically how type-I interferons are able to block viral replication within an infected host cell. (ans)
3. Multiple Choice (ans)
11.3D: Harmful Effects Associated with Abnormal Pattern-Recognition Receptor Responses, Variations in Innate Immune Signaling Pathways, and/or Levels of Cytokine Production
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe two specific examples of how an improper functioning PRR can lead to an increased risk of a specific infection or disease.
1. (ans)
2. (ans)
Questions I
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the role of the following as they relate to phagocytosis:
1. inflammation (ans)
2. lymph nodules (ans)
3. lymph nodes (ans)
4. spleen (ans)
2. Multiple Choice (ans)
Questions II
1. Describe the following steps in phagocytosis:
1. activation (ans)
2. chemotaxis (ans)
3. attachment (both unenhanced and enhanced) (ans)
4. ingestion (ans)
5. destruction (ans)
2. State what happens when either phagocytes are overwhelmed with microbes or they adhere to cells to large to be phagocytosed. (ans)
3. Most of the tissue destruction seen during microbial infections is do to ______________________. (ans)
4. Multiple Choice (ans)
11.3F: Natural Killer Cells (NK Cells) and Invariant Natural Killer T-Lymphocytes (iNKT Cells)
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Matching
_____ Recognize stress induced molecules such as MICA and MICB on the surface of tumor cells or infected cells. (ans)
_____ Recognize MHC-I molecules usually present on all nucleated cells of the body. (ans)
_____ Mechanism by which NK cells kill tumor cells and infected cells. (ans)
1. Apoptosis, a programmed cell suicide
2. Killer-activating receptors
3. Killer-inhibitory receptors
2. Epitopes of glycolipid antigens are recognized by iNKT lymphocytes by way of their _______. (ans)
3. iNKT cells promote both innate and adaptive immunity and may also regulate immune responses by way of the ____________ they produce once activated. (ans)
4. Multiple Choice (ans)
11.3G: Inflammation
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe the following in termsof inflammation:
1. mechanism for inflammation (ans)
2. benefits of plasma leakage (ans)
3. benefits of diapedesis (ans)
4. healing (ans)
2. Briefly describe the process of diapedesis, indicating the role of the following:
1. P-selectins (ans)
2. integrins (ans)
3. adhesion molecules (ans)
3. Briefly describe the problems that arise from chronic inflammation. (ans)
4. Multiple Choice (ans)
11.3H: Nutritional Immunity
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. State three different ways the body deprives microorganisms of iron.
1. (ans)
2. (ans)
3. (ans)
11.3I: Fever
1. Describe the mechanism behind fever. (ans)
2. State 2 benefits of fever.
1. (ans)
2. (ans)
11.3J: The Acute Phase Response
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the mechanism behind the acute phase response. (ans)
2. An acute phase protein that binds to phospholipids in microbial membranes, sticks the micobe to phagocytes, and activates the classical complement pathway is ___________________. (ans)
3. An acute phase protein that binds to mannose in microbial walls, sticks the micobe to phagocytes, and activates the lectin pathway is ___________________. (ans)
4. Multiple Choice (ans)
11.3K: Intraepithelial T-lymphocytes and B-1 cells
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
_____ These cells have a limited diversity of antigen receptors that initially produce a class of antibody molecule called IgM against common polysaccharide and lipid antigens of microbes and against PAMPs of bacteria that invade body cavities. (ans)
_____ These cells have a limited diversity of antigen receptors that recognize molecules associated with epithelial cells but expressed only when those cells arestressed or infected. They kill those cells by inducing apoptosis, a programmed cell suicide. (ans)
1. gamma:delta T-lymphocytes
2. alpha:beta T-lymphocytes
3. B-1 cells
4. marginal zone B cells | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_5%3A_Innate_Immunity/11.E%3A_Innate_Immunity_%28Exercises%29.txt |
Thumbnail: Anterior view of chest showing location and size of adult thymus. (CC-BY 3.0; LearnAnatomy)
12: Introduction to Adaptive Immunity
Compare adaptive (acquired) immunity with innate immunity. Define the following: antigen immunogen epitope humoral immunity cell-mediated immunity
As mentioned in Unit 5, the body has two immune systems: innate immunity and adaptive immunity. Unit 5 dealt with innate immunity. In Unit 6 we will cover adaptive immunity. Let's first again briefly compare acquired and innate immunity.
Innate Immunity
Innate immunity is an antigen-nonspecific defense mechanisms that a host uses immediately or within several hours after exposure to almost any microbe. This is the immunity one is born with and is the initial response by the body to eliminate microbes and prevent infection. Innate immunity can be divided into immediate innate immunity and early induced innate immunity.
Immediate innate immunity begins 0 - 4 hours after exposure to an infectious agent and involves the action of soluble preformed antimicrobial molecules that circulate in the blood, our found in extracellular tissue fluids, and are secreted by epithelial cells. These include: antimicrobial enzymes and peptides; complement system proteins; and anatomical barriers to infection, mechanical removal of microbes, and bacterial antagonism by normal flora bacteria. These preformed innate defense molecules will be discussed in greater detail later in this unit.
Early induced innate immunity begins 4 - 96 hours after exposure to an infectious agent and involves the recruitment of defense cells as a result of pathogen-associated molecular patterns or PAMPS binding to pattern-recognition receptors or PRRs. These recruited defense cells include: phagocytic cells: leukocytes such as neutrophils, eosinophils, and monocytes; tissue phagocytic cells in the tissue such as macrophages; cells that release inflammatory mediators: inflammatory cells in the tissue such as macrophages and mast cells; leukocytes such as basophils and eosinophils; and natural killer cells (NK cells).
Unlike adaptive immunity, innate immunity does not recognize every possible antigen. Instead, it is designed to recognize molecules shared by groups of related microbes that are essential for the survival of those organisms and are not found associated with mammalian cells. These unique microbial molecules are called pathogen-associated molecular patterns or PAMPS and include LPS from the gram-negative cell wall, peptidoglycan and lipotechoic acids from the gram-positive cell wall, the sugar mannose (a terminal sugar common in microbial glycolipids and glycoproteins but rare in those of humans), bacterial and viral unmethylated CpG DNA, bacterial flagellin, the amino acid N-formylmethionine found in bacterial proteins, double-stranded and single-stranded RNA from viruses, and glucans from fungal cell walls. In addition, unique molecules displayed on stressed, injured, infected, or transformed human cells also act as PAMPS. (Because all microbes, not just pathogenic microbes, possess PAMPs, pathogen-associated molecular patterns are sometimes referred to as microbe-associated molecular patterns or MAMPs.)
Most body defense cells have pattern-recognition receptors or PRRs for these common PAMPS (Figure \(1\)) and so there is an immediate response against the invading microorganism. Pathogen-associated molecular patterns can also be recognized by a series of soluble pattern-recognition receptors in the blood that function as opsonins and initiate the complement pathways. In all, the innate immune system is thought to recognize approximately 103 of these microbial molecular patterns.
For More Information: Leukocytes from Unit 5
Examples of innate immunity include anatomical barriers, mechanical removal, bacterial antagonism, antigen-nonspecific defense chemicals, the complement pathways, phagocytosis, inflammation, fever, and the acute-phase response. In this current unit we will look at each of these in greater detail.
Adaptive Immunity
Adaptive (acquired) immunity refers to antigen-specific defense mechanisms that take several days to become protective and are designed to react with and remove a specific antigen. This is the immunity one develops throughout life. There are two major branches of the adaptive immune responses: humoral immunity and cell-mediated immunity.
1. humoral immunity: humoral immunity involves the production of antibody molecules in response to an antigen and is mediated by B-lymphocytes.
2. cell-mediated immunity: Cell-mediated immunity involves the production of cytotoxic T-lymphocytes, activated macrophages, activated NK cells, and cytokines in response to an antigen and is mediated by T-lymphocytes.
During adaptive immunity, antigens are transported to lymphoid organs where they are recognized by naive B-lymphocytes and T-lymphocytes. These activated B- and T-lymphocytes subsequently proliferate and differentiate into effector cells. An antigen is defined as a substance that reacts with antibody molecules and antigen receptors on lymphocytes. An immunogen is an antigen that is recognized by the body as nonself and stimulates an adaptive immune response. For simplicity we will use the term antigen when referring to both antigens and immunogens. The actual portions or fragments of an antigen that react with antibodies and lymphocyte receptors are called epitopes. The size of an epitope is generally thought to be equivalent to 5-15 amino acids in the case of protein antigens (Figure \(2\)); 3-4 sugar residues in the case of polysaccharide antigens (Figure \(3\)).
For More Information: Antigens and Immunogens
For More Information: Antibodies from Unit 6
The body recognizes an antigen as foreign when epitopes of that antigen bind to epitope-specific receptor molecules on the surface of B-lymphocytes and/or T-lymphocytes. The epitope receptor on the surface of a B-lymphocyte is called a B-cell receptor (BCR) and is actually an antibody molecule. The receptor on a T-lymphocyte is called a T-cell receptor (TCR).
• B-cell receptors (BCRs) can bind directly to epitopes on peptide, protein, polysaccharide, nucleic acid, and lipid antigens (Figure \(4\)).
• T-cell receptors ( TCRs) of most T4-lymphocytes and T8-lymphocytes can only recognize peptide epitopes from protein antigens presented by the body's own cells by way of special molecules called MHC molecules (Figure \(4\)).
It is estimated that the human body has the ability to recognize 107 or more different epitopes and make up to 109 different antibodies, each with a unique specificity. In order to recognize this immense number of different epitopes, the body produces 107 or more distinct clones of both B-lymphocytes and T-lymphocytes, each with a unique B-cell receptor or T-cell receptor. Among this large variety of B-cell receptors and T-cell receptors there is bound to be at least one that has an epitope-binding site able to fit, at least to some degree, any antigen the immune system eventually encounters. With the adaptive immune responses, the body is able to recognize any conceivable antigen it may eventually encounter.
The downside to the specificity of adaptive immunity is that only a few B-cells and T-cells in the body recognize any one epitope. These few cells then must rapidly proliferate in order to produce enough cells to mount an effective immune response against that particular epitope, and that typically takes several days. During this time the pathogen could be causing considerable harm, and that is why innate immunity is also essential.
Adaptive immunity usually improves upon repeated exposure to a given infection and involves the following:
• antigen-presenting cells (APCs) such as macrophages and dendritic cells;
• the activation and proliferation of antigen-specific B-lymphocytes;
• the activation and proliferation of antigen-specific T-lymphocytes; and
• the production of antibody molecules, cytotoxic T-lymphocytes (CTLs), activated macrophages, and cytokines.
Acquired immunity includes both humoral immunity and cell-mediated immunity and will be the topic of Unit 5.
We will now take a closer look at adaptive immunity.
Summary
1. The body has two immune systems: the innate immune system and the adaptive immune system.
2. Innate immunity is an antigen-nonspecific defense mechanisms that a host uses immediately or within several hours after exposure to almost any microbe.
3. Innate immunity is the immunity one is born with and is the initial response by the body to eliminate microbes and prevent infection.
4. Immediate innate immunity begins 0 - 4 hours after exposure to an infectious agent and involves the action of soluble preformed antimicrobial molecules that circulate in the blood and in extracellular tissue fluids.
5. Early induced innate immunity begins 4 - 96 hours after exposure to an infectious agent and involves the recruitment of defense cells as a result of pathogen-associated molecular patterns or PAMPS binding to pattern-recognition receptors or PRRs.
6. Adaptive (acquired) immunity refers to antigen-specific defense mechanisms that take several days to become protective and are designed to react with and remove a specific antigen.
7. Adaptive immunity is the immunity one develops throughout life.
8. An antigen is defined as a substance that reacts with antibody molecules and antigen receptors on lymphocytes.
9. The actual portions or fragments of an antigen that react with antibodies and lymphocyte receptors are called epitopes. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/12%3A_Introduction_to_Adaptive_Immunity/12.1%3A_An_Overview_of_Innate_and_Adaptive_Immunity.txt |
Define antigen and immunogen. State what antigens are composed of chemically. List 3 characteristics an antigen must have to be immunogenic. Define epitope. Briefly describe how the body recognizes an antigen as foreign. Compare B-cell receptors and T-cell receptors in terms of how they recognize epitopes. In terms of infectious diseases, list 2 categories of microbial materials that may act as an antigen. List 3 groups of noninfectious materials that may act as an antigen. Define the following: endogenous antigen exogenous antigen autoantigen hapten
An antigen is defined as a substance that reacts with antibody molecules and antigen receptors on lymphocytes. An immunogen is an antigen that is recognized by the body as non-self and stimulates an adaptive immune response. For simplicity, both antigens and immunogens are usually referred to as antigens.
To be immunogenic, an antigen must possess three characteristics:
• be of high molecular weight,
• exhibit chemical complexity, and
• exhibit foreignness (recognized as non-self by the body).
Chemically, antigens are large molecular weight proteins (including conjugated proteins such as glycoproteins, lipoproteins, and nucleoproteins) and polysaccharides (including lipopolysaccharides). These protein and polysaccharide antigens are found on the surfaces of viruses and cells, including microbial cells (bacteria, fungi, protozoans) and human cells.
Epitopes of an antigen
The actual portions or fragments of an antigen that react with receptors on B-lymphocytes and T-lymphocytes, as well as with free antibody molecules, are called epitopes or antigenic determinants. The size of an epitope is generally thought to be equivalent to 5-15 amino acids or 3-4 sugar residues.
Some antigens, such as polysaccharides, usually have many epitopes, but all of the same specificity. This is because polysaccharides may be composed of hundreds of sugars with branching sugar side chains, but usually contain only one or two different sugars. As a result, most "shapes" along the polysaccharide are the same (see Figure \(1\)).
Other antigens such as proteins usually have many epitopes of different specificities. This is because proteins are usually hundreds of amino acids long and are composed of 20 different amino acids. Certain amino acids are able to interact with other amino acids in the protein chain and this causes the protein to fold over upon itself and assume a complex three-dimensional shape. As a result, there are many different "shapes" on the protein (see Figure \(2\)). That is why proteins are more immunogenic than polysaccharides; they are chemically more complex.
A microbe, such as a single bacterium, has many different proteins (and polysaccharides) on its surface that collectively form its various structures, and each different protein may have many different epitopes. Therefore, immune responses are directed against many different epitopes of many different antigens of the same microbe. (For example, a bacterial cell wall alone may contain over 100 different epitopes.) Even simple viruses possess many different epitopes. (see Figure \(3\)).
Recognizing an antigen as foreign
As we saw earlier in Unit 5, the B-lymphocytes and T-lymphocytes are the cells that carry out the immune responses. The body recognizes an antigen as foreign when epitopes of that antigen bind to B-lymphocytes and T-lymphocytes by means of epitope-specific receptor molecules having a shape complementary to that of the epitope (similar to interlocking pieces of a puzzle).
a. B-cell receptors
The antigen receptors on the cytoplasmic membrane of B-lymphocytes are called B-cell receptors and are actually antibody molecules made by that cell and anchored to the outer surface of its cytoplasmic membrane. As will be seen in a later section, antibodies are "Y"-shaped macromolecules composed of four glycoprotein chains connected to one another by disulfide (S-S) bonds and noncovalent bonds (see Figure \(4\)). Additional S-S bonds fold the individual glycoprotein chains into a number of distinct globular domains (see Figure \(5\)).
The two tips of the "Y" are referred to as the Fab portions of the antibody (see Figure \(4\) and Figure \(5\)). The first 110 amino acids or first domain of both the heavy and light chain of the Fab region of the antibody provide specificity for binding an epitope on an antigen. Because they recognize molecular shapes that occur as a result of the 3-dimensional folding of an antigen, B-cell receptors can bind directly to epitopes on peptide, protein, polysaccharide, nucleic acid, and lipid antigens.
The bottom part of the "Y", the C terminal region of each glycoprotein chain, is called the Fc portion. The Fc portion has a constant amino acid sequence that defines the class and subclass of each antibody. The terminal portion of the Fc region of the B-cell receptor is the part that becomes anchored to the cytoplasmic membrane of B-lymphocyte (see Figure \(6\)).
b. T-cell receptors
The receptors on the membrane of T-lymphocytes are called T-cell receptors or TCRs. They are analogous to the B-cell receptor, but are composed of just two glycoprotein chains, each having a variable domain and a constant domain (see Figure \(7\)).
Unlike B-cell receptors that can directly bind to epitopes on antigens, the T-cell receptor or TCR of most T4-lymphocytes and T8-lymphocytes can only recognize peptide epitopes from protein antigens presented by the body's own cells by way of special molecules called MHC molecules as seen in Figure \(6\). The terminal portion of the variable domains provides specificity for binding peptides of protein antigens after the protein has been unfolded, broken into peptides, and bound to a MHC molecule, while the terminus of the constant region becomes anchored to the cytoplasmic membrane of the T-lymphocyte.
The TCR of CD4-CD8- T-lymphocytes and non-MHC restricted CD4+ and CD8+ lymphocytes can recognize epitopes of lipid or glycolipid antigens after they have been attached to CD1 molecules on antigen-presenting cells or in some cases, epitopes directly on antigens.
It is estimated that the human body has the ability to recognize 107or more different epitopes and make up to 109 different antibodies, each with a unique specificity. In order to recognize this immense number of different epitopes, the body produces 107 or more distinct clones of both B-lymphocytes and T-lymphocytes, each with a unique B-cell receptor or T-cell receptor. Among this large variety of B-cell receptors and T-cell receptors there is bound to be at least one that has an epitope-binding site able to fit, at least to some degree, any antigen the immune system eventually encounters. With the adaptive immune responses, the body is able to recognize any conceivable antigen it may eventually encounter.
Substances that act as antigens
In terms of infectious diseases, the following may act as antigens:
a. microbial structures, such as bacterial and fungal cell walls, protozoan cell membranes, bacterial and fungal capsules, microbial flagella, bacterial pili, viral capsids, viral envelope-associated glycoproteins, etc.; and
b. microbial toxins
Certain non-infectious materials may also act as antigens if they are recognized as "nonself" by the body. These include:
a. allergens, including dust, pollen, hair, foods, dander, bee venom, drugs, and other agents causing allergic reactions;
b. foreign tissues and cells from transplants and transfusions; and
c. the body's own cells that the body fails to recognize as "normal self," such as cancer cells, infected cells, cells involved in autoimmune diseases.
There are three broad categories of antigens: endogenous antigens, exogenous antigens, and autoantigens.
1. Endogenous antigens are proteins found within the cytosol of human cells. Examples of endogenous antigens include:
1. viral proteins produced during viral replication;
2. proteins produced by intracellular bacteria such as Rickettsias and Chlamydias during their replication;
3. proteins that have escaped into the cytosol from the phagosome of phagocytes such as antigen-presenting cells;
4. tumor antigens produced by cancer cells; and
5. self-peptides from host cellular proteins.
2. Exogenous antigens are antigens that enter from outside the body, such as bacteria, fungi, protozoa, and free viruses. These exogenous antigens enter macrophages, dendritic cells, and B-lymphocytes through phagocytosis or pinocytosis.
3. Autoantigens are any of an organism’s own antigens (self-antigens) that stimulate an autoimmune reaction, that is humoral immunity or cell-mediated against self.
A hapten is a small molecule that by itself is not immunogenic but can act as an antigen when it binds to a larger protein molecule. The hapten then acts as an epitope on the protein. For example with penicillin and poison ivy allergies, the penicillin molecules and the oil urushiol from the poison ivy plant function as haptens, binding to tissue proteins to form an antigen and stimulating an allergic immune response.
Summary
1. An antigen is defined as a substance that reacts with antibody molecules and antigen receptors on lymphocytes.
2. An immunogen is an antigen that is recognized by the body as non-self and stimulates an adaptive immune response.
3. Chemically, antigens are large molecular weight proteins and polysaccharides.
4. The actual portions or fragments of an antigen that react with receptors on B-lymphocytes and T-lymphocytes, as well as with free antibody molecules, are called epitopes.
5. The size of an epitope is generally thought to be equivalent to 5-15 amino acids or 3-4 sugar residues.
6. Polysaccharides antigens usually have many epitopes but all of the same specificity.
7. Proteins antigens usually have many epitopes of different specificities.
8. Immune responses are directed against many different epitopes of many different antigens of the same microbe.
9. The body recognizes an antigen as foreign when epitopes of that antigen bind to B-lymphocytes and T-lymphocytes by means of epitope-specific receptor molecules having a shape complementary to that of the epitope.
10. The antigen receptors on the cytoplasmic membrane of B-lymphocytes are called B-cell receptors and are actually antibody molecules made by that cell and anchored to the outer surface of its cytoplasmic membrane and is composed of composed of four interconnected glycoprotein chains.
11. The receptors on the membrane of T-lymphocytes are called T-cell receptors or TCRs and are composed of just two glycoprotein chains.
12. During its development, each different B-lymphocyte and T-lymphocyte becomes genetically programmed to produce a B-cell receptor or T-cell receptor with a unique three-dimensional shape.
13. The body produces 107 or more distinct clones of both B-lymphocytes and T-lymphocytes, each with a unique B-cell receptor or T-cell receptor and with this large variety of B-cell receptors and T-cell receptors there is bound to be at least one that has an epitope-binding site able to fit, at least to some degree, any antigen the immune system eventually encounters.
14. In terms of infectious diseases, microbial structures and microbial toxins act as antigens.
15. Certain noninfectious materials also act as antigens, including allergens, foreign tissues and cells from transplants and transfusions, and the body's own cells that the body fails to recognize as "normal self," such as cancer cells, infected cells, and cells involved in autoimmune diseases.
16. Endogenous antigens are antigens found within the cytosol of human cells such as viral proteins, proteins from intracellular bacteria, and tumor antigens.
17. Exogenous antigens are antigens that enter from outside the body, such as bacteria, fungi, protozoa, and free viruses.
18. Autoantigens are any of an organism's own antigens (self-antigens) that stimulate an autoimmune reaction. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/12%3A_Introduction_to_Adaptive_Immunity/12.2%3A_Antigens_and_Epitopes.txt |
We will now take a look at major cells and key cell-surface molecules involved in adaptive immune responses.
12.3: Major Cells and Key Cell Surface Molecules Involved in Adaptive Immune Responses
Learning Objectives
• State which body cells display MHC-I surface molecules and which cells normally display MHC-II surface molecules.
• Define endogenous antigen and exogenous antigen and state which class of MHC molecule primarily binds each.
• State which type of T-lymphocyte recognizes epitopes from protein antigens on MHC-I molecules and which type recognizes epitopes from protein antigens on MHC-II molecules.
• State the role of proteasomes in binding of peptides from endogenous antigens by MHC-I molecules.
• State the role of lysosomes in binding of peptides from exogenous antigens by MHC-II molecules.
The Roles of MHC Molecules In Adaptive Immune Responses
MHC molecules enable T-lymphocytes to recognize epitopes of antigens and discriminate self from non-self. Unlike B-cell receptors on B-lymphocytes that are able to directly bind epitopes on antigens, the T-cell receptors (TCRs) of T-lymphocytes can only recognize epitopes - typically short chains of amino acids called peptides - after they are bound to MHC molecules (Figure \(1\)).
The MHC genes are the most polymorphic genes in the human genome, possessing many alleles for each gene. The MHC genes are co-dominantly expressed so that an individual expresses the alleles inherited from each parent. In this way, the number of MHC molecules that bind peptide for presentation to T-lymphocytes is maximized. In addition, each MHC molecule is able to bind a wide variety of different peptides, both self-peptides and foreign peptides. There are two classes of MHC molecules: MHC-I and MHC-II.
• MHC-I molecules present epitopes to T8-lymphocytes.
• MHC-II molecules presents epitopes to T4-lymphocytes .
The expression of MHC molecules is increased by cytokines produced during both innate immune responses and adaptive immune responses. Cytokines such as interferon-alpha, interferon-beta, interferon-gamma, tumor necrosis factor increase the expression of MHC-I molecules, while interferon-gamma is the main cytokine to increase the expression of MHC-II molecules.
MHC-I molecules
MHC-I molecules are designed to enable the body to recognize infected cells and tumor cells and destroy them with cytotoxic T-lymphocytes or CTLs. CTLs are effector defense cells derived from naive T8-lymphocytes. MHC-I molecules are:
• Made by all nucleated cells in the body.
• Possess a deep groove that can bind peptide epitopes, typically 8-11 amino acids long, typically from endogenous antigens .
• Present MHC-I/peptide complexes to naive T8-lymphocytes and cytotoxic T-lymphocytes possessing a complementary-shaped T-cell receptor or TCR.
• Through the process of cross-presentation, some antigen-presenting dendritic cells can cross-present epitopes of exogenous antigens to MHC-I molecules for eventual presentation to naive T8-lymphocytes.
Endogenous antigens are proteins found within the cytosol of human cells. Examples of endogenous antigens include:
a. Viral proteins produced during viral replication;
b. Proteins produced by intracellular bacteria such as Rickettsias and Chlamydias during their replication;
c. Proteins that have escaped into the cytosol from the phagosome of phagocytes such as antigen-presenting cells;
d. Tumor antigens produced by cancer cells; and
e. Self-peptides from host cellular proteins.
During the replication of viruses and intracellular bacteria within their host cell, as well as during the replication of tumor cells, viral, bacterial, or tumor proteins are degraded into a variety of peptide epitopes by cylindrical organelles called proteasomes. The body's own cytosolic proteins are also degraded into peptides by proteasomes.
These peptide epitopes are then attached to a groove of MHC-I molecules that are then transported to the surface of that cell where they can be recognized by a complementary-shaped T-cell receptor (TCR) and a CD8 molecule, a co-receptor, on the surface of either a naive T8-lymphocyte or a cytotoxic T-lymphocyte (CTL). The TCRs recognize both the foreign peptide antigen and the MHC molecule (Figure \(2\)). TCRs, however, will not recognize self-peptides bound to MHC-I. As a result, normal cells are not attacked and killed.
Dendritic cells bind epitopes from endogenous antigens to MHC-I molecules and present them to naive T8-lymphocytes in order to activate these naive T8-lymphocytes.
1. Antigens are engulfed by dendritic cells and placed in a phagosome. Some of the proteins escape from the phagosome into the cytosol of the dendritic cell where they become endogenous antigens.
2. These endogenous antigens pass through proteasomes where they are degraded into a series of peptides.
3. The peptides are transported into the rough endoplasmic reticulum (ER) by a transporter protein called TAP.
4. The peptides then bind to the grooves of newly synthesized MHC-I molecules.
5. The endoplasmic reticulum transports the MHC-I molecules with bound peptides to the Golgi complex.
6. The Golgi complex, in turn, transports the MHC-I/peptide complexes by way of an exocytic vesicle to the cytoplasmic membrane where they become anchored. Here, the peptide and MHC-I/peptide complexes can be recognized by naive T8-lymphocytes by way of TCRs and CD8 molecules having a complementary shape.
Through the process of cross-presentation, some antigen-presenting dendritic cells can cross-present epitopes of exogenous antigens to MHC-I molecules for eventual presentation to naive T8-lymphocytes.
MHC-I molecule with bound peptide on the surface of antigen-presenting dendritic cells (Figure \(3\)) can be recognized by a complementary-shaped TCR/CD8 on the surface of a naive T8-lymphocyte to initiate cell-mediated immunity (Figure \(4\)). (Certain dendritic cells, as discussed later, can also cross-present exogenous antigens to MHC-I molecules).
MHC-I molecule with bound peptide on the surface of infected cells and tumor cells (Figure \(5\)) can be recognized by a complementary-shaped TCR/CD8 on the surface of a cytotoxic T-lymphocyte or CTL to initiate destruction of the cell containing the endogenous antigen (Figure \(6\)). (CTLs are effector cells derived from naive T8-lymphocytes.)
Cytotoxic T-lymphocytes (CTLs) are then able to recognize peptide/MHC-I complexes by means of their T-cell receptors (TCRs) and CD8 molecules and kill the cells to which they bind.
1. During viral replication within the host cell, endogenous antigens, such as viral proteins, pass through proteasomes where they are degraded into a series of peptides.
2. The peptides are transported into the rough endoplasmic reticulum (ER) by a transporter protein called TAP.
3. The peptides then bind to the grooves of newly synthesized MHC-I molecules.
4. The endoplasmic reticulum transports the MHC-I molecules with bound peptides to the Golgi complex.
5. The Golgi complex, in turn, transports the MHC-I/peptide complexes by way of an exocytic vesicle to the cytoplasmic membrane where they become anchored. Here, the peptide and MHC-I/peptide complexes can be recognized by CTLs by way of TCRs and CD8 molecules having a complementary shape.
MHC-I molecules are coded for by three MHC-I genes, HLA-A, HLA-B, and HLA-C. As mentioned above, however, there are many different alleles for each gene that a person inherits. In this way, the number of MHC-I molecules that bind peptides for presentation to T-8 lymphocytes is maximized. The expression of MHC-I molecules on all cell types is increased by the cytokines interferon-alpha (IFN-a) and interferon-beta (IFN-ß).
Exercise: Think-Pair-Share Questions
All nucleated cells produce MHC-I molecules. MHC-I molecules bind peptide epitopes of antigens found within our cells. Peptide epitopes bound to MHC-I molecules are recognized by TCRs and CD8 molecules on the surfaces of naive T8-lymphocytes and on cytotoxic T-lymphocytes (CTLs).
Why is it important that all nucleated cells in our body are able to produce MHC-I molecules?
MHC-II molecules
MHC-II molecules are designed to enable T4-lymphocytes to recognize epitopes of exogenous antigens and discriminate self from non-self. MHC-II molecules are:
• Made by antigen-presenting cells or APCs, such as dendritic cells, macrophages, and B-lymphocytes .
• Possess a deep groove that can bind peptide epitopes, often 10-30 amino acids long but with an optimum length of 12-16 amino acids, typically from exogenous antigens. The peptides interact along their entire length with the groove.
• Present MHC-II/peptide complexes to naive T4-lymphocytes or effector T4-lymphocytes that have a complementary shaped T-cell receptor or TCR.
• Through the process of cross-presentation, some antigen-presenting dendritic cells can cross-present epitopes of endogenous antigens to MHC-II molecules for eventual presentation to naive T4-lymphocytes.
Exogenous antigens are antigens that enter from outside the body, such as bacteria, fungi, protozoa, and free viruses. These exogenous antigens enter macrophages, dendritic cells, and B-lymphocytes through phagocytosis. The microbes are engulfed and placed in a phagosome which then fuses with lysosomes. Following this fusion, the phagolysosome becomes acidified. Acidification, in turn, activates the proteases within the phagolysosome enabling protein antigens from the microbe to be degraded into a series of short peptides. These peptide epitopes are then attached to MHC-II molecules and are then transported to the surface of the antigen-presenting cell (APC) (Figure \(7\)). (Certain dendritic cells, as discussed later, can also cross-present endogenous antigens to MHC-II molecules.)
Some pathogens, such as Mycobacterium tuberculosis, Mycobacterium leprae, and Leishmania, are able to grow in the endocytic vesicles of macrophages without being killed by lysosomes. These macrophages can, however, become activated by T4-effector lymphocytes called TH1 cells and subsequently use intravesicular proteases to degrade the proteins from these pathogens into peptides for presentation to MHC-II molecules that pass through on their way to the cell surface.
Here the MHC-II molecules with bound peptides can be recognized by a complementary-shaped T-cell receptor and a CD4 molecule, a co-receptor, on the surface of a T4-lymphocyte (Figure \(8\)). T4-lymphocytes are the cells the body uses to regulate both humoral immunity and cell-mediated immunity.
MHC-II molecules are coded for by three MHC-II genes, HLA-DR, HLA-DP, and HLA-DQ. Interferon-gamma (IFN- ?) increases the expression of both MHC-I and MHC-II molecules.
Think-Pair-Share Questions
Only antigen-presenting cells such as dendritic cells, macrophages, and B-lymphocytes produce MHC-II molecules. Peptide epitopes bound to MHC-II molecules are recognized by TCRs and CD4 molecules on the surfaces of naive T4-lymphocytes and on effector T4-lymphocytes.
Why don't all nucleated cells in our body produce MHC-II molecules as well as MHC-I molecules?
Why is it important for dendritic cells to produce both MHC-I and MHC-II molecules?
Summary
1. MHC molecules enable T-lymphocytes to recognize epitopes and discriminate self from non-self.
2. T-cell receptors (TCRs) of T-lymphocytes can only recognize epitopes - typically short chains of amino acids called peptides - after they are bound to MHC molecules.
3. MHC-I presents epitopes to T8-lymphocytes; MHC-II presents epitopes to T4-lymphocytes.
4. MHC-I molecules are designed to enable the body to recognize infected cells and tumor cells and destroy them with cytotoxic T-lymphocytes or CTLs. (CTLs are effector defense cells derived from naïve T8-lymphocytes.)
5. MHC-I molecules are made by all nucleated cells in the body; bind peptide epitopes typically from endogenous antigens; present MHC-I/peptide complexes to naive T8-lymphocytes and cytotoxic T-lymphocytes possessing a complementary-shaped T-cell receptor or TCR.
6. Through the process of cross-presentation, some antigen-presenting dendritic cells can cross-present epitopes of exogenous antigens to MHC-I molecules for eventual presentation to naive T8-lymphocytes.
7. Endogenous antigens are proteins found within the cytosol of human cells and include viral proteins produced during viral replication, proteins produced by intracellular bacteria, proteins that have escaped into the cytosol from the phagosome of phagocytes such as antigen-presenting cells, and tumor antigens produced by cancer cells.
8. During the replication of viruses and intracellular bacteria within their host cell, as well as during the replication of tumor cells, viral, bacterial, or tumor proteins are degraded into a variety of peptide epitopes by cylindrical organelles called proteasomes. The resulting peptide epitopes are then attached to MHC-I molecules that are then transported to the surface of that cell.
9. Exogenous antigens are antigens that enter from outside the body such as bacteria, fungi, protozoa, and free viruses.
10. MHC-II molecules are made by antigen-presenting cells or APCs, such as dendritic cells, macrophages, and B-lymphocytes; bind peptide epitopes typically from exogenous antigens; and present MHC-II/peptide complexes to naive T4-lymphocytes or effector T4-lymphocytes that have a complementary shaped T-cell receptor or TCR.
11. Through the process of cross-presentation, some antigen-presenting dendritic cells can cross-present epitopes of endogenous antigens to MHC-II molecules for eventual presentation to naive T4-lymphocytes.
12. Exogenous antigens enter antigen-presenting macrophages, dendritic cells, and B-lymphocytes through phagocytosis, and are engulfed and placed in a phagosome where protein antigens from the microbe are degraded by proteases into a series of peptides. These peptides are then attached to MHC-II molecules that are then put on the surface of the APC. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/12%3A_Introduction_to_Adaptive_Immunity/12.3%3A_Major_Cells_and_Key_Cell_Surface_Molecules_Involved_in_Adaptive_Immune_Responses/12.3A%3A__Major_Histocompatibil.txt |
Describe the overall function of antigen-presenting cells (APCs) such as dendritic cells, macrophages, and B-lymphocytes in terms of the following: how they "process" exogenous antigens how they "process" endogenous antigens the types of MHC molecule to which they attach peptides the role of proteasomes in the binding of peptides from endogenous antigens by MHC-I molecules. the role of lysosomes in the binding of peptides from exogenous antigens by MHC-II molecules. the types of cells to which they present peptides Name the primary type of cell that functions as an antigen-presenting cell to naive T4-lymphocytes and naive T8-lymphocytes. State the role of T4-effector cells in activating macrophages. State the role of T4-effector cells in the proliferation and differentiation of activated B-lymphocytes.
The primary function of dendritic cells, then, is to capture and present protein antigens to naive T-lymphocytes. (Naive lymphocytes are those that have not yet encountered an antigen.) Since dendritic cells are able to express both MHC-I and MHC-II molecules, they are able to present antigens to both naive T8-lymphocytes and naive T4-lymphocytes.
You Tube movie of a dendritic cell engulfing melanoma cells (red).
These interactions enable the naiveT4-lymphocyte or T8-lymphocyte to become activated, proliferate, and differentiate into effector lymphocytes. (Effector lymphocytes are lymphocytes that have encountered an antigen, proliferated, and matured into a form capable of actively carrying out immune defenses.)
1. MHC-II presentation of protein antigens to naive T4-lymphocytes
a. MHC-II presentation of exogenous antigens to naive T4-lymphocytes
Immature dendritic cells take in protein antigens for attachment to MHC-II molecules and subsequent presentation to naive T4-lymphocytes by:
1. Receptor-mediated phagocytosis, e.g., PAMPs binding to endocytic PRRs, IgG or C3b attachment of microbes to phagocytes during opsonization (see Figure \(2\)).
2. Macropinocytosis, a process where large volumes of surrounding fluid containing microbes are engulfed. This also enables dendritic cells to take in some encapsulated bacteria that might resist classical phagocytosis (see Figure \(3\)).
The binding of microbial PAMPs to the PRRs of the immature dendritic cell activates that dendritic cell and promotes production of the chemokine receptor CCR7 that directs the dendritic cell into local lymphoid tissue. Following maturation, the dendritic cell can now present protein epitopes bound to MHC molecules to all the various naive T-lymphocytes passing through the lymphoid system (See Figure \(4\) and Figure \(5\)).
The MHC-II molecules bind peptide epitopes from exogenous antigens and place them on the surface of the dendritic cell (see Figure \(6\)). Here the MHC-II/peptide complexes can be recognized by complementary shaped TCRs and CD4 molecules on naive T4-lymphocytes (see Figure \(7\)).
b. MHC-II cross-presentation of endogenous antigens to naive T4-lymphocytes
While most dendritic cells present exogenous antigens to naive T4-lymphocytes, certain dendritic cells are capable of cross-presentation of endogenous antigens to naive T4-lymphocytes. In this way, T4-lymphocytes can play a role in defending against both exogenous and endogenous antigens. This is done via autophagy, the cellular process whereby the cell's own cytoplasm is taken into specialized vesicles called autophagosomes (See Figure \(8\)). The autophagosomes subsequently fuse with lysosomes containing proteases that will degrade the proteins in the autophagosome into peptides. From here, the peptides are transported into the vesicles containing MHC-II molecules where they can bind to the MHC-II groove, be transported to the surface of the denritic cell, and interact with the TCRs and CD4 molecules of naive T4-lymphocytes (See Figure \(8\)).
2. MHC-I presentation of protein antigens to naive T8-lymphocytes
Immature dendritic cells take in protein antigens for attachment to MHC-I molecules and subsequent presentation to naive T8-lymphocytes.
a. MHC-I presentation of endogenous antigens to naive T8-lymphocytes
During the replication of viruses and intracellular bacteria within their host cell, as well as during the replication of tumor cells, viral, bacterial, or tumor proteins are degraded into a variety of peptide epitopes by cylindrical organelles called proteasomes. The body's own cytosolic proteins are also degraded into peptides by proteasomes.
These peptide epitopes are then attached to a groove of MHC-I molecules that are then transported to the surface of that cell where they can be recognized by a complementary-shaped T-cell receptor (TCR) and a CD8 molecule, a co-receptor, on the surface of either a naive T8-lymphocyte or a cytotoxic T-lymphocyte (CTL). The TCRs recognize both the foreign peptide antigen and the MHC molecule. TCRs, however, will not recognize self-peptides bound to MHC-I. As a result, normal cells are not attacked and killed.
MHC-I molecule with bound peptide on the surface of antigen-presenting dendritic cells ; see Figure \(9\) can be recognized by a complementary-shaped TCR/CD8 on the surface of a naive T8-lymphocyte to initiate cell-mediated immunity (see Figure \(10\)).
b. MHC-I cross-presentation of exogenous antigens to naive T8-lymphocytes
While most dendritic cells present endogenous antigens to naive T8-lymphocytes, certain dendritic cells are capable of cross-presentation of exogenous antigens to naive T8-lymphocytes. In this way, T8-lymphocytes can play a role in defending against both exogenous and endogenous antigens. There are two proposed mechanisms for cross-presentation of exogenous antigens to T8-lymphocytes:
1. The dendritic cell engulfs the exogenous antigen and places it in a phagosome which then fuses with a lysosome to form a phagolysosome. The antigen is partially degraded in the phagolysosome where proteins are translocated into the cytoplasm where they are processed into peptides by proteasomes, enter the endoplasmic reticulum, and are bound to MHC-I molecules (see Figure \(11\)).
2. The dendritic cell engulfs the exogenous antigen and places it in a phagosome which then fuses with a lysosome to form a phagolysosome. The protein antigens are degraded into peptides within the phagolysosome which then directly fuses with vesicles containing MHC-I molecules to which the peptides subsequently bind (see Figure \(12\)).
In addition, dendritic cells are very susceptible to infection by many different viruses. Once inside the cell, the viruses become endogenous antigens in the cytosol. Once in the cytosol, the viral proteins from the replicating viruses are degraded into peptides by proteasomes where they subsequently bind to MHC-I molecules.
The binding of microbial PAMPs to the PRRs of the immature dendritic cell activates that dendritic cell and promotes production of the chemokine receptor CCR7 that directs the dendritic cell into local lymphoid tissue. Following maturation, the dendritic cell can now present protein epitopes bound to MHC molecules to all the various naive T-lymphocytes passing through the lymphoid system.
To view an electron micrograph of a dendritic cell presenting antigen to T-lymphocytes, #1 see the Web page for the University of Illinois College of Medicine.
To view an electron micrograph of a dendritic cell presenting antigen to T-lymphocytes, #2 see the Web page for the University of Illinois College of Medicine.
Why is this essential for effective adaptive immunity ?
For a Summary of Key Surface Molecules and Cellular Interactions of Antigen-Presenting Dendritic Cells, see Figure \(13\).
Macrophages
As we learned in Unit 5, when monocytes leave the blood and enter the tissue, they become activated and differentiate into macrophages. Those that have recently left the blood during inflammation and move to the site of infection are sometimes referred to as wandering macrophages. In addition, the body has macrophages already stationed throughout the tissues and organs of the body. These are sometimes referred to as fixed macrophages. Many fixed macrophages are part of the mononuclear phagocytic (reticuloendothelial) system. They, along with B-lymphocytes and T-lymphocytes, are found supported by reticular fibers in lymph nodules, lymph nodes, and the spleen where they filter out and phagocytose foreign matter such as microbes. Similar cells derived from stem cells, monocytes, or macrophages are also found in the liver (Kupffer cells), the kidneys (mesangial cells), the brain (microglia), the bones (osteoclasts), the lungs (alveolar macrophages), and the gastrointestinal tract (peritoneal macrophages).
The primary function of macrophages, then, is to capture and present protein antigens to effector T-lymphocytes. (Effector lymphocytes are lymphocytes that have encountered an antigen, proliferated, and matured into a form capable of actively carrying out immune defenses.)
The MHC-II molecules bind peptide epitopes from exogenous antigens and place them on the surface of the macrophages. Here the MHC-II/peptide complexes can be recognized by complementary shaped T-cell receptors (TCRs) and CD4 molecules on an effector T4-lymphocytes ; see Fig.14.
Effector T4-lymphocytes called TH1 cells coordinate immunity against intracellular bacteria and promote opsonization by macrophages.
1. They produce cytokines such as interferon-gamma (IFN-?) that promote cell-mediated immunity against intracellular pathogens, especially by activating macrophages that have either ingested pathogens or have become infected with intracellular microbes such as Mycobacterium tuberculosis, Mycobacterium leprae, Leishmania donovani, and Pneumocystis jiroveci that are able to grow in the endocytic vesicles of macrophages. Activation of the macrophage by TH1 cells greatly enhances their antimicrobial effectiveness (see Figure \(14\)).
2. They produce cytokines that promote the production of opsonizing and complement activating IgG that enhances phagocytosis (see Figure \(15\)).
3. They produce receptors that bind to and kill chronically infected cells, releasing the bacteria that were growing within the cell so they can be engulfed and killed by macrophages.
4. They produce cytokines such as tumor necrosis factor-alpha (TNF-a) that promote diapedesis of macrophages.
5. They produce the chemokine CXCL2 to attract macrophages to the infection site.
There is growing evidence that monocytes and macrophages can be “trained” by an earlier infection to do better in future infections, that is, develop memory. It is thought that microbial pathogen-associated molecular patterns (PAMPs) binding to pattern-recognition (PRRs) on monocytes and macrophages triggers the cell’s epigenome to reprogram or train that cell to react better against new infections.
For a Summary of Key Surface Molecules and Cellular Interactions of Antigen-Presenting Macrophages, see Figure \(16\).
B-lymphocytes
Like all lymphocytes, B-lymphocytes circulate back and forth between the blood and the lymphoid system of the body. B-lymphocytes are able to capture and present peptide epitopes from exogenous antigens to effector T4-lymphocytes. The MHC-II molecules bind peptide epitopes from exogenous antigens and place them on the surface of the B-lymphocytes. Here the MHC-II/peptide complexes can be recognized by complementary shaped T-cell receptors (TCRs) and CD4 molecules on an effector T4-lymphocytes (see Figure \(17\)). This interaction eventually triggers the effector T4-lymphocyte to produce and secrete various cytokines that enable that B-lymphocyte to proliferate and differentiate into antibody-secreting plasma cells (see Figure \(18\)).
For a Summary of Key Surface Molecules and Cellular Interactions of Antigen-Presenting B-Lymphocytes, see Figure \(19\).
Summary
1. Antigen-presenting cells (APCs) include dendritic cells, macrophages, and B-lymphocytes.
2. APCs express both MHC-I and MHC-II molecules and serve two major functions during adaptive immunity: they capture and process antigens for presentation to T-lymphocytes, and they produce signals required for the proliferation and differentiation of lymphocytes.
3. Most dendritic cells are derived from monocytes and are referred to as myeloid dendritic cells and are located under the surface epithelium of the skin, the mucous membranes of the respiratory tract, genitourinary tract, and the gastrointestinal tract, and throughout the body's lymphoid tissues and in most solid organs.
4. The primary function of dendritic cells is to capture and present protein antigens to naive T-lymphocytes which enables the naïve T4-lymphocytes or T8-lymphocytes to become activated, proliferate, and differentiate into effector cells.
5. Naïve lymphocytes are B-lymphocytes and T-lymphocytes that have not yet reacted with an epitope of an antigen.
6. Dendritic cells use MHC-II molecules to present protein antigens to naïve T4-lymphocytes and MHC-I molecules to present protein antigens to naïve T8-lymphocytes.
7. When monocytes leave the blood and enter the tissue, they become activated and differentiate into macrophages.
8. When functioning as APCs, macrophages capture and present peptide epitopes from exogenous antigens to effector T4-lymphocytes.
9. Effector lymphocytes are lymphocytes that have encountered an antigen, proliferated, and matured into a form capable of actively carrying out immune defenses.
10. B-lymphocytes mediate antibody production.
11. When functioning as APCs, B-lymphocytes are able to capture and present peptide epitopes from exogenous antigens to effector T4-lymphocytes.
12. To activate naïve T4-lymphocytes, dendritic cells engulf exogenous antigens, place them in a phagosome, degrade protein antigens into peptides via lysosomes, bind those peptides to MHC-II molecules and transport them to the surface of the dendritic cell where they can be recognized by the T-cell receptors and CD4 molecules of naïve T4-lymphocytes.
13. To activate naïve T8-lymphocytes, dendritic cells degrade endogenous protein antigens into peptides via their proteasomes, bind those peptides to MHC-I molecules and transport them to the surface of the dendritic cell where they can be recognized by the T-cell receptors and CD8 molecules of naïve T8-lymphocytes. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/12%3A_Introduction_to_Adaptive_Immunity/12.3%3A_Major_Cells_and_Key_Cell_Surface_Molecules_Involved_in_Adaptive_Immune_Responses/12.3B%3A_Antigen-Presenting_Cel.txt |
Describe the overall function of T4-lymphocytes and their activation in terms of the following: the role of their TCRs and CD4 molecules what they recognize on antigen-presenting cells (APCs) such as dendritic cells, macrophages, and B-lymphocytes. the role of antigen-presenting dendritic cells in the activation of naive T4-lymphocytes. Compare TH1, TH2, TH17, Treg, and TFH lymphocytes in terms of their primary function(s) in immunity.
• To view an electron micrograph of a dendritic cell presenting antigen to T-lymphocytes, #1 see the Web page for the University of Illinois College of Medicine.
• To view an electron micrograph of a dendritic cell presenting antigen to T-lymphocytes, #2 see the Web page for the University of Illinois College of Medicine.
Those naive T4-lymphocytes not activated by epitopes of antigens on the dendritic cells exit the lymph node (or other lymphoid tissue) and eventually re-enter the bloodstream. However, if a TCR and CD4 molecule of the naive T4-lymphocyte detects a corresponding MHC-II/peptide complex on a mature dendritic cell, this will send a first signal for the activation of that naive T-lymphocyte. Next, a second signal that promotes survival of that T-lymphocyte is sent when co-stimulatory molecules such as B7.1 and B7.2 on the dendritic cell bind to CD28 molecules on the T4-lymphocyte. Finally, the dendritic cell produces cytokines such as interleukin-6 (IL-6), IL-4, IL-12, and T-cell growth factor-beta (TGF-ß) that contribute to proliferation of the T4-lymphocytes and their differentiation into effector T4-lymphocytes, the cells the body uses to regulate both humoral immunity and cell-mediated immunity through the cytokines they produce. (Activated T4-lymphocytes remain in the lymph node as they proliferate (clonal expansion) and only leave the lymphoid tissues and re-enter the bloodstream after they have differentiated into effector T4-lymphocytes.)
CD28-dependent co-stimulation of the T4-lymphocyte also stimulates it to synthesize the cytokine interleukin-2 (IL-2) as well as a high-affinity IL-2 receptor. The binding of IL-2 to its high affinity receptor allows for cell proliferation and formation of a clone of thousands of identical T4-lymphocytes after several days. IL-2 also contributes to survival of those activated T4-lymphocytes and their differentiation into T4-effector cells. In addition, some of the T4-lymphocytes differentiate into circulating T4-memory cells. Circulating T4-memory cells allow for a more rapid and greater production of effector T4-lymphocytes upon subsequent exposure to the same antigen.
Differentiation of naive T4-lymphocyte into T4-effector lymphocytes
Functionally, there are many different types or subpopulations of effector T4-lymphocytes based on the cytokines they produce. Immune reactions are typically dominated by five primary types: TH1 cells, TH2 cells, TH17 cells, Treg cells, and TFH cells.
CD4 TH1 cells
Coordinate immunity against intracellular bacteria and promote opsonization. They:
1. Produce cytokines such as interferon-gamma (IFN-?) that promote cell-mediated immunity against intracellular pathogens, especially by activating macrophages that have either ingested pathogens or have become infected with intracellular microbes such as Mycobacterium tuberculosis, Mycobacterium leprae, Leishmania donovani, and Pneumocystis jjroveci that are able to grow in the endocytic vesicles of macrophages. Activation of the macrophage by TH1 cells greatly enhances their antimicrobial effectiveness.
2. They produce cytokines that promote the production of opsonizing antibodies that enhance phagocytosis (Figure \(9\)).
3. Produce receptors that bind to and kill chronically infected cells, releasing the bacteria that were growing within the cell so the can be engulfed and killed by macrophages.
4. Produce the cytokine interleukin-2 (IL-2) that induces T-lymphocyte proliferation.
5. Produce cytokines such as tumor necrosis factor-alpha (TNF-a) that promote diapedesis of macrophages.
6. Produces the chemokine CXCL2 to attract macrophages to the infection site.
7. Produce cytokines that block the production of TH2 cells.
CD4 TH2 cells
Coordinate immunity against helminths and microbes that colonize mucous membranes
1. Produce the cytokine interleukin-4 (IL-4) that promotes the production of the antibody isotype IgE in response to helminths and allergens. IgE is able to stick eosinophils to helminths for extracellular killing of the helminth (Figure \(10\)); it also promotes many allergic reactions.
2. Produce cytokines that attract and activate eosinophils and mast cells.
3. Promote the production of antibodies that neutralize microbes (Figure \(11\)) and toxins (Figure \(12\)) preventing their attachment to host cells.
4. Produce cytokines that function as B-lymphocyte growth factors such as IL-4, IL-5, IL-9. and IL-13 (Figure \(13\)).
5. Produce interleukin-22 (IL-22) that promotes the removal of microbes in mucosal tissues.
6. Produce cytokines that block the production of TH1 cells.
CD4 TH17 cells
Promote a local inflammatory response to stimulate a strong neutrophil response and promote the integrity of the skin and mucous membranes
Produce cytokines like interleukin-17 (IL-17) and interleukin-6 (IL-6) that trigger local epithelial cells and fibroblasts to produce chemokines that recruit neutrophils to remove extracellular pathogens.
CD4 Treg cells
Suppress immune responses
1. Produce inhibitory cytokines such as Interleukin-10 (IL-10) and TGF-ß that help to limit immune responses and prevent autoimmunity by suppressing T-lymphocyte activity.
2. Promoting anamnestic response (immunologic memory) to resist repeat infections by the same microbe.
3. Protecting beneficial normal flora in the intestines from being destroyed by the immune system.
4. Aiding in sustaining pregnancy so that the immune system doesn't recognize a fetus as foreign and try to destroy it.
5. Controlling established inflammation in tissues.
TFH cells
Promote humoral immunity by stimulating antibody production and antibody isotype switching by B-lymphocytes
1. T follicular helper cells (TFH cells) are located in lymphoid follicles.
2. TFH cells are now thought to be the primary effector T-lymphocytes that stimulate antibody production and isotype switching by B-lymphocytes. They are able to produce cytokines that are characteristic of both TH2 cells and TH1 cells.
3. TFH cells producing (IFN-?) promote the production of opsonizing antibodies; those producing IL-4 promote the production of IgE.
Regulation of effector T4-lymphocyte activity: A role for commensals and helminths?
It is now recognized that genes associated with the normal flora ( microbiota) of the intestinal tract aid in digestion of many foods (especially plant polysaccharides that would normally be indigestible by humans), may play a role in normal growth and regulating appetite, and also help to regulate immune defenses. There is ever growing evidence that commensal bacteria of the gastrointestinal tract, as well as parasitic gastrointestinal helminths, may have coevolved with the human body over the past 200,000 year in such a way that genes from the human microbiota may play a significant role in regulating the human immune responses by providing a series of checks and balances that prevent the immune system from being too aggressive and causing an autoimmune attack upon the body's own cells, while still remaining aggressive enough to recognize and remove harmful pathogens. As exposure to and colonization with these once common human organisms has drastically changed over time as a result of less exposure to mud, animal and human feces,and helminth ova, coupled with ever increasing antibiotic use, improved sanitation, changes in the human diet, increased rate of cesarean sections, and improved methods of processing and preserving of food, the rate of allergies, allergic asthma, and autoimmune diseases (inflammatory bowel disease, Crone's disease, type-1 diabetes, and multiple sclerosis for example) has dramatically increased in developed countries while remaining relatively low in undeveloped and more agrarian parts of the world.
Numerous experiments in germ-free mice (mice with no intestinal commensals) have shown them to be much more susceptible to allergic asthma and autoimmune diseases such as colitis then normal mice. Feeding commensals or nematode ova to newborn germ-free mice, in turn, reduces the occurrence to these disorders. An imbalance in the relationship between proinflammatory TH17 cells and inflammation-suppressing Treg cells appears to increase the risk of inflammatory autoimmune diseases, while an imbalance between TH1 and TH2 cells seems to contribute to the risk of allergies and asthma. For example, a common commensal colon bacterium Bacteroides fragilis produces a molecule called polysaccharide A that dendritic cells engulf, process and present to naive T4-lymphocytes. This interaction stimulates the differentiation of the naive T4-lymphocytes into anti-inflammatory Treg cells that suppress the activity of proinflammatory TH17 cells. Without colonization with B. fragilis, the proinflammatory TH17 cells are not suppressed and there is an increased risk of inflammatory autoimmune diseases.
Normal intestinal microbiota also appear to regulate the intestinal levels of the invariant natural killer (iNKT) cells discussed under innate immune responses in Unit 4. iNKT cells recognize endogenous and exogenous lipid antigens presented on CD1d molecules by dendritic cells and in response, secrete proinflammatory cytokines. Germ free mice show an accumulation of iNKT cells in the colon and in the lungs and have an increased risk of intestinal bowel disease and allergic asthma. Neonatal germ free mice that were subsequently colonized with normal microbiota were protected from this iNKT cell accumulation and the resulting inflammatory pathology.
Summary
1. T-lymphocytes refer to lymphocytes that are produced in the bone marrow but require interaction with the thymus for their maturation.
2. The primary role of T4-lymphocytes is to regulate the body's immune responses through the production of cytokines.
3. T4-lymphocytes display CD4 molecules and T-cell receptors (TCRs) on their surface.
4. The TCR on T4-lymphocytes, in cooperation with CD4, typically bind peptides from exogenous antigens bound to MHC-II molecules.
5. During its development, each T4-lymphocyte becomes genetically programmed to produce a TCR with a unique specificity that is able to bind an epitope/MHC-II complex on an APC such as a dendritic cell, a macrophage, or a B-lymphocyte possessing a corresponding shape.
6. To become activated, naive T4-lymphocytes migrate through lymph nodes where the TCRs on the T4-lymphocyte are able to sample large numbers of MHC-II/peptide complexes on the antigen-presenting dendritic cells for ones that “fit”, thus enabling activation of that naïve T4-lymphocyte.
7. After activation, the dendritic cell produces cytokines that contribute to proliferation of the T4-lymphocytes and their differentiation into effector T4-lymphocytes, the cells the body uses to regulate both humoral immunity and cell-mediated immunity through the cytokines they produce.
8. Some of the T4-lymphocytes differentiate into circulating T4-memory cells that enable a more 9.rapid and greater production of effector T4-lymphocytes upon subsequent exposure to the same antigen.
9. Functionally, there are many different types or subpopulations of effector T4-lymphocytes based on the cytokines they produce. Immune reactions are typically dominated by five primary types: TH1 cells, TH2 cells, TH17 cells, Treg cells, and TFH cells.
10. CD4 TH1 cells coordinate immunity against intracellular bacteria and promote opsonization.
11. CD4 TH2 cells coordinate immunity against helminths and microbes that colonize mucous membranes.
12. CD4 TH17 cells promote a local inflammatory response to stimulate a strong neutrophil response and promote the integrity of the skin and mucous membranes.
13. CD4 Treg cells suppress immune responses.
14. TFH cells promote humoral immunity by stimulating antibody production and antibody isotype switching by B-lymphocytes.
15. There is ever growing evidence that commensal bacteria of the gastrointestinal tract, as well as parasitic gastrointestinal helminths, may have coevolved with the human body over the past 200,000 year in such a way that genes from the human microbiota may play a significant role in regulating the human immune responses by providing a series of checks and balances that prevent the immune system from being too aggressive and causing an autoimmune attack upon the body's own cells, while still remaining aggressive enough to recognize and remove harmful pathogens. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/12%3A_Introduction_to_Adaptive_Immunity/12.3%3A_Major_Cells_and_Key_Cell_Surface_Molecules_Involved_in_Adaptive_Immune_Responses/12.3C%3A_T4-Lymphocytes_%28T4-C.txt |
Describe the overall function of T8-lymphocytes and their activation in terms of the following: the role of their TCRs and CD8 molecules how they are activated by antigen-presenting dendritic cells the type of effector cells into which activated T8-lymphocytes differentiate what CTLs recognize on infected cells and tumor cells how CTLs kill infected cells and tumor cells State the overall function of T8-lymphocytes in adaptive immunity.
The primary role of T8-lymphocytes (T8-Cells; CD8+ Cells; Cytotoxic T-Lymphocytes) is to kill infected cells and tumor cells by inducing apoptosis of those cells. Once naive T8-lymphocytes are activated by dendritic cells , they proliferate and differentiate into T8-effector lymphocytes called cytotoxic T-lymphocytes (CTLs) that bind to and kill infected cells and tumor cells.
T8-lymphocytes are T-lymphocytes displaying a surface molecule called CD8. T8-lymphocytes also have on their surface, T-cell receptors or TCRs similar to those on T4-lymphocytes. The TCR on T8-lymphocytes, in cooperation with CD8, bind peptides from endogenous antigens bound to MHC-I molecules .
During its development, each T8-lymphocyte becomes genetically programmed, by gene-splicing reactions similar to those in B-lymphocytes and T4-lymphocytes, to produce a TCR with a unique shape capable of binding epitope/MHC-I complex with a corresponding shape. It is estimated that the human body has the ability to recognize 107 or more different epitopes . In order to recognize this immense number of different epitopes, the body produces 107 or more distinct clones of T-lymphocytes, each with a unique T-cell receptor. In this variety of T-cell receptors there is bound to be at least one that has an epitope-binding site able to fit, at least to some degree, peptides of any antigen the immune system eventually encounters.
Activation of a naive T8-lymphocyte by a dendritic cell
One of the body's major defenses against viruses, intracellular bacteria, and cancers is the destruction of infected cells and tumor cells by cytotoxic T-lymphocytes or CTLs . These CTLs are effector cells derived from naive T8-lymphocytes during cell-mediated immunity. However, in order to become CTLs, naive T8-lymphocytes must become activated by dendritic cells as shown in Figure \(1\) and Figure \(2\).
• To view an electron micrograph of a dendritic cell presenting antigen to T-lymphocytes, #1 see the Web page for the University of Illinois College of Medicine.
• To view an electron micrograph of a dendritic cell presenting antigen to T-lymphocytes, #2 see the Web page for the University of Illinois College of Medicine.
Certain dendritic cells are capable of cross-presentation of exogenous antigens to naive T8-lymphocytes . In this way, T8-lymphocytes can play a role in defending against both exogenous and endogenous antigens.
Naive T-lymphocytes circulate in the blood. In response to chemokines produced by lymphoid tissues, they leave the vascular endothelium in regions called high endothelial venules and enter lymph nodes (see Figure \(3\)) or other lymphoid tissues, a process called diapedesis .
As naive T8-lymphocytes migrate through the cortical region of lymph nodes, they use surface cell adhesion molecules such as LFA-1 and CD2 to bind transiently to corresponding receptors such as ICAM-1, ICAM-2 and CD58 on the surface of dendritic cells. This transient binding allows time for the TCRs on the T8-lymphocyte to sample large numbers of MHC-I/peptide complexes on the antigen-presenting dendritic cells (see Figure \(4\)).
Those naive T8-lymphocytes not activated by epitopes of antigens on the dendritic cells exit the lymph node (or other lymphoid tissue) and eventually re-enter the bloodstream. However, if a TCR and CD8 molecule of the naive T8-lymphocyte detects a corresponding MHC-I/peptide complex on a mature dendritic cell, this will send a first signal for the activation of that naive T-lymphocyte. Next, a second signal that promotes survival of that T-lymphocyte is sent when co-stimulatory molecules such as B7.1 and B7.2 on the dendritic cell bind to CD28 molecules on the T8-lymphocyte. Finally, the dendritic cell produces cytokines such as interleukin-6 (IL-6), IL-4, IL-12, and T-cell growth factor-beta (TGF-ß) that contribute to proliferation of the T8-lymphocytes and their differentiation into effector T8-lymphocytes called cytotoxic T-lymphocytes (CTLs) that are able to bind to and kill infected cells and tumor cells displaying the same peptide/MHC-I complex on their surface. (Activated T8-lymphocytes remain in the lymph node as they proliferate (clonal expansion) and only leave the lymphoid tissues and re-enter the bloodstream after they have differentiated into CTLs.)
While activated T8-lymphocytes produce interleukin-2 (IL-2) as well as a high-affinity IL-2 receptor themselves, in most cases it is the IL-2 produced by effector T4-lymphocytes that enables cell proliferation and formation of a clone of thousands of identical T8-lymphocytes after several days. IL-2 also contributes to survival of those activated T8-lymphocytes and their differentiation into T8-effector cells called a cytotoxic T-lymphocytes or CTLs .
CTLs leave the secondary lymphoid organs and enter the bloodstream where they can be delivered anywhere in the body via the circulatory system and the inflammatory response. In addition, some of the T8-lymphocytes differentiate into circulating T8-memory cells . Circulating T8-memory cells allow for a more rapid and greater production of CTLs upon subsequent exposure to the same antigen.
Marking an infected cell or tumor cell for destruction by cytotoxic T-lymphocytes (CTLs)
During the replication of viruses and intracellular bacteria within their host cell, as well as during the replication of tumor cells, viral, bacterial, or tumor proteins in the cytosol of that cell are degraded into a variety of peptide epitopes by cylindrical organelles called proteasomes . Other endogenous antigens such as proteins released into the cytosol from the phagosomes of antigen-presenting cells, such as macrophages and dendritic cells as well, as a variety of the human cell's own proteins (self-proteins) are also degraded by proteasomes. As these various endogenous antigens pass through proteasomes, proteases and peptidases chop the protein up into a series of peptides, typically 8-11 amino acids long (see Figure \(5\)).
A transporter protein called TAP located in the membrane of the cell's endoplasmic reticulum then transports these peptide epitopes into the endoplasmic reticulum where they bind to the grooves of various newly made MHC-I molecules. The MHC-I molecules with bound peptides are then transported to the Golgi complex and placed in exocytic vesicles. The exocytic vesicles carry the MHC-I/peptide complexes to the cytoplasmic membrane of the cell where they become anchored to its surface (see Figure \(6\)). A single cell may have up to 250,000 molecules of MHC-I with bound epitope on its surface.
CTLs binding to infected cells or tumor cells and inducing apoptosis
CTLs are, by way of their TCRs and CD8 molecules, able to recognize infected cells and tumor cells displaying MHC-I molecules with bound peptides on their surface (see Figure \(7\)) and destroy them through apoptosis , a programmed cell suicide.
Apoptosis involves a complex of intracellular granules. This complex of granules in a protected state including:
1. Pore-forming proteins called perforins ;
2. Proteolytic enzymes called granzymes ; and
3. A proteoglycan called granulysin.
When the TCR and CD8 of the CTL binds to the MHC-I/epitope on the surface of the virus-infected cell or tumor cell (see Figure \(7\)), this sends a signal through a CD3 molecule which triggers the release of the perforins/granzymes/granulysin complexes from the CTL.
The exact mechanism of entry of the granzymes into the infected cell or tumor cell is still debated. It is, however, dependent on perforins. Possibilities include:
• The perforins/granzymes/granulysin complex may be taken into the target cell by receptor-mediated endocytosis . The perforin molecules may then act on the endosomal membrane allowing granzymes to enter the cytosol.
• The perforin molecules may put pores in the membrane of the target cell allowing the granzymes to directly enter the cytosol (see Figure \(7\)).
Killing of the infected cell or tumor cell by apoptosis involves a variety of mechanisms:
• Certain granzymes can activate the caspase enzymes that lead to apoptosis of the infected cell. The caspases are proteases that destroy the protein structural scaffolding of the cell - the cytoskeleton - and nucleases that degrade both the target cell's nucleoprotein and any microbial DNA within the cell (see Figure \(8\)).
• Granzymes cleave a variety of other cellular substrates that contribute to cell death.
• The perforin molecules may also polymerize and form pores in the membrane of the infected cell, similar to those produced by MAC. This can increase the permeability of the infected cell and contribute to cell death. If enough perforin pores form, the cell might not be able to exclude ions and water and may undergo cytolysis.
• Granulysin has antimicrobial actions and can also induce apoptosis.
Movie illustrating apoptosis. Found on You Tube.
For a Summary of Key Surface Molecules and Cellular Interactions of Naive T8-Lymphocytes, see Figure \(9\).
Summary
1. T-lymphocytes refer to lymphocytes that are produced in the bone marrow but require interaction with the thymus for their maturation.
2. The primary role of T8-lymphocytes is to kill infected cells and tumor cells by inducing apoptosis of those cells.
3. Once naive T8-lymphocytes are activated by dendritic cells, they proliferate and differentiate into T8-effector lymphocytes called cytotoxic T-lymphocytes (CTLs) that bind to and kill infected cells and tumor cells.
4. T8-lymphocytes display CD8 molecules and T-cell receptors (TCRs) on their surface.
5. The TCR on T8-lymphocytes, in cooperation with CD8, typically bind peptides from endogenous antigens bound to MHC-I molecules.
6. During its development, each T8-lymphocyte becomes genetically programmed, by gene-splicing reactions similar to those in B-lymphocytes and T4-lymphocytes, to produce a TCR with a unique shape capable of binding epitope/MHC-I complex with a corresponding shape.
7. To become activated, naive T8-lymphocytes migrate through lymph nodes where the TCRs on the T8-lymphocyte are able to sample large numbers of MHC-I/peptide complexes on the antigen-presenting dendritic cells for ones that “fit”, thus enabling activation of that naïve T8-lymphocyte.
8. After activation, the dendritic cell produces cytokines that contribute to proliferation of the T8-lymphocytes and their differentiation into effector T4-lymphocytes called cytotoxic T-lymphocytes (CTLs) that are able to bind to and kill infected cells and tumor cells displaying the same peptide/MHC-I complex on their surface.
9. Some of the T8-lymphocytes differentiate into circulating T8-memory cells that enable a more rapid and greater production of CTLs upon subsequent exposure to the same antigen.
10. During the replication of viruses and intracellular bacteria within their host cell, as well as during the replication of tumor cells, viral, bacterial, or tumor proteins in the cytosol of that cell are degraded into a variety of peptide epitopes by cylindrical organelles called proteasomes.
11. As these various endogenous antigens pass through proteasomes, proteases and peptidases chop the protein up into a series of peptides that are transported into the endoplasmic reticulum where they bind to newly made MHC-I molecules.
12. The MHC-I molecules with bound peptides are then transported to the Golgi complex and placed in exocytic vesicles that carry the MHC-I/peptide complexes to the cytoplasmic membrane of the cell where they become anchored to its surface.
13. CTLs are, by way of their TCRs and CD8 molecules, are then able to recognize infected cells and tumor cells displaying MHC-I molecules with bound peptides on their surface. This sends a signal that triggers the release of the perforins/granzymes/granulysin complexes from the CTL to destroy the infected cell or tumor cell through apoptosis.
14. The perforin molecules may put pores in the membrane of the target cell allowing the granzymes to directly enter the cytosol, and certain granzymes activate the caspase enzymes that lead to apoptosis of the infected cell or tumor cell by destroying the cytoskeleton of the cell and degrading both the target cell's nucleoprotein and any microbial DNA within the cell. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/12%3A_Introduction_to_Adaptive_Immunity/12.3%3A_Major_Cells_and_Key_Cell_Surface_Molecules_Involved_in_Adaptive_Immune_Responses/12.3D%3A_T8-Lymphocytes_%28T8-C.txt |
Describe the overall function of iNKT cells and their activation in terms of the following: the role of their TCRs how they are activated by antigen-presenting cells how they promote both innate and adaptive immunity and may also help to regulate the immune responses
Once activated, the iNKT cells rapidly produce large quantities of both TH1 cell and TH2 cell cytokines, including interferon-gamma (IFN-?), interleukin-4 (IL-4), interleukin-2 (IL-2), interleukin-10 (IL-10), tumor necrosis factor-alpha (TNF-a), interleukin-13 (IL-13), and chemokines. Through the rapid productions of such cytokines, iNKT cells are able to promote and suppress different innate and adaptive immune responses. For example, large amounts of IFN-? are produced by activated iNKT cells. IFN-? activates NK cells and macrophages as a part of innate immunity; it also promotes the maturation of dendritic cells so that they induce a TH1 cell response to induce adaptive immunity.
It has been proposed that if the iNKT cell is repeatedly stimulated by the body's own glycolipids in the ab sense of microbes that this might stimulate the iNKT cell /dendritic cell interaction to produce tolerizing signals that inhibit the TH1 cell response and possibly stimulate the production of regulatory T-lymphocytes (Treg cells). In this way it might suppress autoimmune responses and prevent tissue damage.
There is also growing evidence that early childhood exposure to microbes is associated with protection against allergic diseases, asthma, and inflammatory diseases such as ulcerative colitis. It has been found that germ-free mice have large accumulations of mucosal iNKT cells in the lungs and intestines and increased morbidity from allergic asthma and inflammatory bowel disease. However, colonization of neonatal germ-free mice with normal microbiota resulted in mucosal iNKT cell tolerance to these diseases. It has been proposed that microbes the human body has been traditionally exposed to from early childhood throughout most of human history might play a role in developing normal iNKT cell numbers and iNKT cell responses.
12.3F: B-Lymphocytes (B-Cel
Learning Objectives
Describe the overall function of B-lymphocytes and their activation by T-dependent antigens in terms of the following:
1. the antigen receptor on their surface
2. how they "process" exogenous antigens
3. the type of MHC molecule to which they attach peptides
4. the role of lysosomes in binding of peptides from exogenous antigens by MHC-II molecules.
5. the type of cell to which they present peptides
6. the types of cells into which activated B-lymphocytes differentiate
B-lymphocytes (B-cells) are responsible for the production of antibody molecules during adaptive immunity. Antibodies are critical in removing extracellular microorganisms and toxins. B-lymphocytes refer to lymphocytes that are produced in the bone marrow and require bone marrow stromal cells and their cytokines for maturation. During its development, each B-lymphocyte becomes genetically programmed through a series of gene-splicing reactions to produce an antibody molecule with a unique specificity - a specific 3-dimensional shape capable of binding a specific epitope of an antigen (Figure \(1\)).
It is estimated that the human body has the ability to recognize 107 or more different epitopes and make up to 109 different antibodies, each with a unique specificity. In order to recognize this immense number of different epitopes, the body produces 107 or more distinct clones of B-lymphocytes, each with a unique B-cell receptor or BCR. In this variety of B-cell receptors there is bound to be at least one that has an epitope-binding site able to fit, at least to some degree, any antigen the immune system eventually encounters.
Typically, over 100,000 identical molecules of that unique antibody are placed on the surface of the B-lymphocyte where they can function as B-cell receptors capable of binding specific epitopes of a corresponding shape (Figure \(2\)). Naive B-lymphocytes can be activated by both T-dependent antigens and T-independent antigens.
Activation of naive B-lymphocytes by T-dependent antigens
In order for naive B-lymphocytes to proliferate, differentiate, and mount an antibody response against T-dependent antigens, such as most proteins, these B-lymphocytes must interact with effector T4-lymphocytes called TFH cells. All classes of antibody molecules can be made against T-dependent antigens and there is usually a memory response against such antigens.
B-Lymphocytes and T4-lymphocytes encounter antigens in secondary lymphoid organs such as the lymph nodes and the spleen. Using a lymph node as an example (Figure \(3\)A), soluble antigens, such as microbial polysaccharides and proteins and toxins, as well as microbes such as bacteria and viruses, enter the lymph node through afferent lymphatic vessels. By this time, complement pathway activation has coated these soluble antigens or microbes with opsonins such as C3b, which in turn can be degraded to C3d.
Located within the lymphoid tissues are specialized macrophages and specialized dendritic cells called follicular dendritic cells (FDCs). These macrophages have poor endocytic ability and produce few lysosomes. The FDCs are nonphagocytic. Both cell types, however, have complement receptors called CR1 and CR2 that bind to the C3b and C3d, enabling the antigens and microbes to stick to the surface of the macrophages and FDCs. However,because of the poor endocytic ability of the macrophages and the lack of endocytosis by the FDCs, the antigens and microbes are not engulfed but rather remain on the surface of the cells. In addition, the macrophages can transfer their bound antigens or microbes to FDCs (Figure \(3\)B).
Here the antigens and microbes in the lymph node can bind to complementary-shaped BCRs on naive B-lymphocytes directly, by way of macrophages, or via the FDCs (Figure \(3\)B).
Circulating naive B-lymphocytes, as a result of chemotaxis, enter lymph nodes through high endothelial venules. Any naive B-lymphocyte that bind antigens become activated and remain in the lymphoid nodes to proliferate and differentiate. Any B-lymphocytes not activated leave the lymphoid node through efferent lymphatic vessels and are returned to the bloodstream.
The first signal for the activation of a naive B-lymphocyte occurs when BCRs on the surface of the B-lymphocyte bind epitopes of antigens having a corresponding shape. A second signal is also needed for the activation of the naive B-lymphocyte. This is provided when the complement protein C3d on the microbial surface or soluble antigen binds to a complement receptor called CR2 on the surface of the naive B-lymphocyte.
Once bound, the antigen is engulfed, placed in a phagosome , and degraded with lysosomes. During this process, protein antigens are broken down into a series of peptide epitopes.These peptides eventually bind to grooves in MHC-II molecules that are then transported to the surface of the B-lymphocyte (Figure \(4\)).
Meanwhile, naïve T4-lymphocytes are being activated by epitopes of antigens bound to MHC-II molecules on antigen-presenting dendritic cells in the T-cell area of the lymph node and subsequently proliferate and differentiate into T4-effector lymphocytes such as TFH cells which remain in the lymph node. The T-cell receptors and CD4 molecules on TFH cells bind to the MHC-II molecules with bound peptide epitope on the B-lymphocyte. The binding of co-receptor molecules such as CD40L and CD28 on the surface of the effector T4-lymphocyte to the corresponding molecules CD40 and B7 on the surface of the B-lymphocyte further contribute to the interaction between these two cells (Figure \(5\)). This enables the TFH cells to produce cytokines such as interleukin-2 (IL-2) , interleukin-4 (IL-4), interleukin-5 (IL-5), and interleukin-6 (IL-6) (Figure \(5\)).
Collectively these cytokines:
1. Enable activated B-lymphocytes to proliferate.
2. Stimulate activated B-lymphocytes to synthesize and secrete antibodies.
3. Promote the differentiation of B-lymphocytes into antibody-secreting plasma cells. See Figure \(6\).
4. Enable antibody producing cells to switch the class or isotype of antibodies being produced.
YouTube animation illustrating production of antibodies by B-lymphocytes.
YouTube animation illustrating production of antibodies by B-lymphocytes against Streptococcus pyogenes.
Effector T4-lymphocytes also enable B-lymphocytes to undergo affinity maturation through a high rate of somatic mutation. This allows the B-lymphocytes to eventually "fine-tune" the shape of the antibody for better fit with the original epitope. After mutation, some antibodies fit better, some worse. To select for B-lymphocytes displaying antibodies with a better fit, the variant B-lymphocytes interact with cells called follicular dendritic cells (FDCs) in the germinal centers of the secondary lymphoid organs. The FDCs display the same antigens that activated the original B-lymphocyte. If the B-lymphocytes have high affinity antibodies for the antigen on the FDC, they are selected to survive. Those B-lymphocytes with low affinity antibodies undergo apoptosis.
With the exception of TFH cells which remain in the germinal centers of the lymph nodes and spleen, progeny of the activated B-lymphocytes and T4 effector lymphocytes leave the secondary lymphoid organs and migrate to tissues where they continue to respond to the invading antigen as long as it is present.
In the case of systemic infections or vaccinations where the antigens enter the bloodstream, plasma cells migrate to the bone marrow where antibodies can be produced for decades. After the antibodies are secreted by the plasma cells, they are found dissolved in the blood plasma and lymph. From here they can be delivered anywhere in the body via the circulatory system and the inflammatory response. In the case of infections of the mucous membranes, however, plasma cells only enter the mucous membranes where antibodies are only produced for a few months to a year or so.
During the proliferation and differentiation that follows lymphocyte activation, some of the B-lymphocytes stop replicating and become circulating, long-lived memory cells. Memory cells are capable of what is called anamnestic response or "memory", that is, they "remember" the original antigen. If that same antigen again enters the body while the B-memory cells (and T4-memory cells) are still present, these memory cells will initiate a rapid, heightened secondary response against that antigen (Figure \(7\)). This is why the body sometimes develops a permanent immunity after an infectious disease and is also the principle behind immunization.
Activation of B-lymphocytes by T-independent antigens
T-independent (TI) antigens are usually large carbohydrate and lipid molecules with multiple, repeating subunits. B-lymphocytes mount an antibody response to T-independent antigens without the requirement of interaction with effector T4-lymphocytes. Bacterial LPS from the Gram-negative cell wall and capsular polysaccharides are examples of TI antigens. The resulting antibody molecules are generally of the IgM isotype and do not give rise to a memory response. There are two basic types of T-independent antigens: TI-1 and TI-2.
a. TI-1 antigens arepathogen-associated molecular patterns or PAMPS such as lipopolysaccharide (LPS) from the outer membrane of the gram-negative cell wall and bacterial nucleic acid. These antigens activate B-lymphocytes by binding to their specific pattern-recognition receptors , in this case toll-like receptors, rather than to B-cell receptors (Figure \(8\)). Antibody molecules generated against TI-1 antigens are often called "natural antibodies" because they are always being made against bacteria present in the body.
b. TI-2 antigens, such as capsular polysaccharides, are molecules with multiple, repeating subunits. These repeating subunits activate B-lymphocytes by simultaneously cross-linking a number of B-cell receptors (Figure \(9\)).
For a Summary of Key Surface Molecules and Cellular Interactions of Naive B-Lymphocytes, see Figure \(10\).
Summary
1. B-lymphocytes are responsible for the production of antibody molecules during adaptive immunity.
2. Antibodies are critical in removing extracellular microorganisms and toxins.
3. B-lymphocytes refer to lymphocytes that are produced in the bone marrow and require bone marrow stromal cells and their cytokines for maturation.
4. During its development, each B-lymphocyte becomes genetically programmed to produce an antibody molecule with a unique 3-dimensional shape capable of binding a specific epitope of an antigen, and puts molecules of that antibody on its surface that function as B-cell receptors or BCRs.
5. Naive B-lymphocytes can be activated by both T-dependent antigens and T-independent antigens.
6. In order for naive B-lymphocytes to proliferate, differentiate, and mount an antibody response against T-dependent antigens, such as most proteins, these B-lymphocytes must interact with effector T4-lymphocytes called TFH cells.
7. The first signal for the activation of a naive B-lymphocyte occurs when BCRs on the surface of the B-lymphocyte bind epitopes of antigens having a corresponding shape.
8. Once bound to the BCR, the antigen is engulfed, placed in a phagosome, and degraded with lysosomes. During this process, protein antigens are broken down into a series of peptide epitopes, bind to MHC-II molecules, and are transported to the surface of the B-lymphocyte.
9. The T-cell receptors and CD4 molecules on TFH cells bind to the MHC-II molecules with bound peptide epitope on the B-lymphocyte which enables the TFH cells to produce cytokines that collectively enable the B-lymphocytes to proliferate, synthesize and secrete antibodies, differentiate into antibody-secreting plasma cells, and switch the class of antibodies being produced.
10. By way of a mutation process called affinity maturation, activated B-lymphocytes are able over time to “fine-tune" the shape of the antibody for better fit with the original epitope.
11. During the proliferation and differentiation that follows lymphocyte activation, some of the B-lymphocytes stop replicating and become circulating, long-lived memory cells that will initiate a rapid, heightened secondary response against that antigen if it again enters the body.
12. T-independent (TI) antigens are usually large carbohydrate and lipid molecules with multiple, repeating subunits. B-lymphocytes mount an antibody response to T-independent antigens without the requirement of interaction with effector T4-lymphocytes, but the resulting antibody molecules are generally of the IgM isotype only and do not give rise to a memory response. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/12%3A_Introduction_to_Adaptive_Immunity/12.3%3A_Major_Cells_and_Key_Cell_Surface_Molecules_Involved_in_Adaptive_Immune_Responses/12.3E%3A_Invarient_Natural_Kill.txt |
Briefly describe how NK cells bind to and kill infected cells and tumor cells through ADCC. Briefly describe how NK cells recognize and kill infected cells and tumor cells that suppress MHC-I production.
NK cells are another group of cytolytic lymphocytes that are distinct from B-lymphocytes and T-lymphocytes, and participate in both innate immunity and adaptive immunity. NK cells are lymphocytes that lack B-cell receptors and T-cell receptors. They are designed to kill certain mutant cells and virus-infected cells in one of two ways:
Antibody-dependent Cellular Cytotoxicity
NK cells kill cells to which antibody molecules have attached through a process called antibody-dependent cellular cytotoxicity (ADCC) as shown in Figure \(1\), Figure \(2\), and Figure \(3\). The Fab portion of the antibody binds to epitopes on the "foreign" cell. The NK cell then binds to the Fc portion of the antibody. The NK cell is then able to contact the cell and by inducing a programmed cell suicide called apoptosis.
As a result, the infected cell breaks into membrane-bound fragments that are subsequently removed by phagocytes. If very large numbers of perforins are inserted into the plasma membrane of the infected cell, this can result in a weakening of the membrane and lead to cell lysis rather than apoptosis. An advantage to killing infected cells by apoptosis is that the cell's contents, including viable virus particles and mediators of inflammation, are not released as they are during cell lysis.
Innate Immunity
In addition, NK cells produce a variety of cytokines, including proinflammatory cytokines, chemokines, colony-stimulating factors, and other cytokines that function as regulators of body defenses. For example, through cytokine production NK cells also suppress and/or activate macrophages, suppress and/or activate the antigen-presenting capabilities of dendritic cells, and suppress and/or activate T-lymphocyte responses.
Summary
1. Natural Killer (NK) cells are able to recognize infected cells, cancer cells, and stressed cells and kill them. In addition, they produce a variety of cytokines, including proinflammatory cytokines, chemokines, colony-stimulating factors, and other cytokines that function as regulators of body defenses.
2. NK cells play a role in adaptive immune responses by way of antibody-dependent cellular cytotoxicity or ADCC where they bind to and kill cells to which antibody molecules have bound.
3. During ADCC, the Fab portion of the antibody binds to epitopes on the "foreign" cell. The NK cell then binds to the Fc portion of the antibody and the NK cell is then able to contact and kill the cell by inducing a programmed cell suicide called apoptosis.
4. During innate immunity, NK cells use a dual receptor system in determining whether to kill or not kill human cells.
5. When body cells are either under stress, are turning into tumors, or are infected, various stress-induced molecules are produced and are put on the surface of that cell.
6. The first receptor, called the killer-activating receptor, can bind to these stress-induced molecules, and this sends a positive signal that enables the NK cell to kill the cell to which it has bound unless the second receptor cancels that signal.
7. The second receptor, called the killer-ihibitory receptor, recognizes MHC-I molecules that are usually present on all nucleated human cells. If MHC-I molecules/self peptide complexes are expressed on the cell, the killer-inhibitory receptors on the NK cell recognize this MHC-I/peptide complex and sends a negative signal that overrides the original kill signal and prevents the NK cell from killing the cell to which it has bound.
8. NK cells kill their target cells by inducing apoptosis, a programmed cell suicide. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/12%3A_Introduction_to_Adaptive_Immunity/12.3%3A_Major_Cells_and_Key_Cell_Surface_Molecules_Involved_in_Adaptive_Immune_Responses/12.3G%3A_Natural_Killer_Cells_%.txt |
Learning Objectives
1. Compare and give examples of the following:
1. primary lymphoid organs
2. secondary lymphoid organs
2. Define the following:
1. plasma
2. tissue fluid
3. lymph
4. lymph vessels
5. MALT
3. Briefly describe the importance of the lymphoid system in adaptive immune responses and how microbes and other antigens encounter naive B-lymphocytes and T-lymphocytes.
The body uses the lymphoid system to enable lymphocytes to encounter antigens and it is here that adaptive immune responses are initiated. The lymphoid system consists of primary lymphoid organs, secondary lymphoid organs, and lymphatic vessels.
The bone marrow and the thymus constitute the primary lymphoid organs. Both B-lymphocytes and T-lymphocytes are produced from stem cells in the bone marrow. B-lymphocytes mature in the bone marrow while T-lymphocytes migrate to the thymus and mature there. After maturation, both naive B-lymphocytes and naive T-lymphocytes circulate between the blood and the secondary lymphoid organs.
Lymphatic vessels are responsible for flow of lymph within the lymphoid system and are a part of the body's fluid recirculation system. The liquid portion of the blood, called plasma, constantly leaks out of capillaries to deliver oxygen and nutrients to cells of the surrounding tissue. Once in the tissue, the plasma is now called tissue fluid. While most of this tissue fluid re-enters capillaries and is returned directly to the bloodstream, some fluid enters lymph vessels as lymph. The lymph flows through regional lymph nodes and eventually enters the circulatory system at the heart to maintain the fluid volume of the circulation.
Secondary lymphoid organs
Adaptive immune responses require antigen-presenting cells, such as macrophages and dendritic cells, and ever changing populations of B-lymphocytes and T- lymphocytes. These cells gather to detect and present antigens in secondary lymphoid organs. The secondary lymphoid organs include highly organized lymphoid organs such as lymph nodes and the spleen, as well as less organized accumulations of lymphoid organs scattered strategically throughout the body.
Lymph nodes (Figure \(1\)) contain many reticular fibers that support fixed macrophages and dendritic cells as well as ever changing populations of circulating B-lymphocytes and T-lymphocytes. When microorganisms and other antigens enter tissues, they are transported by tissue fluid into the lymph vessels. Lymph vessels, in turn, carry these antigens, now in the lymph, to regional lymph nodes. In addition, immature dendritic cells located under the surface epithelium of the skin and the surface epithelium of the mucous membranes of the respiratory tract, genitourinary tract, and the gastrointestinal tract capture antigens through pinocytosis and phagocytosis. The dendritic cells detach from their initial site, enter lymph vessels, and are carried to regional lymph nodes. Here the microbes and other antigens in the lymph encounter changing populations of B-lymphocytes, are filtered out and phagocytosed by the fixed macrophages and dendritic cells, and are presented to changing populations of T-lymphocytes (Figure \(2\)). Approximately 25 billion different lymphocytes migrate through each lymph node every day.
Like the lymph nodes, the spleen contains many reticular fibers that support fixed macrophages and dendritic cells as well as ever changing populations of circulating B-lymphocytes and T-lymphocytes. When microorganisms and other antigens enter the blood, they are transported by the blood vessels to the spleen. Most of the spleen is referred to as red pulp. This area is involved in the disposal of old red blood cells. Scattered throughout the spleen are isolated areas called the white pulp (Figure \(3\)). Here antigens in the blood encounter macrophages, dendritic cells, and ever-changing populations of B-lymphocytes and T-lymphocytes.
Mucosal surfaces within the body, the most common sites of microbial invasion, are protected by the mucosal immune system consisting of the mucosa-associated lymphoid tissue or MALT, an extensive diffuse system of small concentrations of lymphoid tissue found in various sites of the body such as the gastrointestinal tract, thyroid, breast, lung, salivary glands, eye, and skin. MALT is populated by loose clusters of T-lymphocytes, B-lymphocytes, plasma cells, activated TH cells, and macrophages. MALT can be subdivided into:
• GALT (gut-associated lymphoid tissue, such as the Peyer's patches (Figure \(4\)) in the lining of the small intestines, as well as the adenoids, tonsils, and appendix)
• BALT (bronchial-associated lymphoid tissue in the bronchi)
• SALT (skin-associated lymphoid tissue beneath the epidermis)
• NALT (nose-associated lymphoid tissue)
• LALT (larynx-associated lymphoid tissue)
• CALT (conjunctiva-associated lymphoid tissue in the eye)
As can be seen, no matter how microbes and other antigens enter the body, they will eventually encounter the lymphoid system to initiate adaptive immune responses.
Summary
1. The body uses the lymphoid system to enable lymphocytes to encounter antigens and it is here that adaptive immune responses are initiated.
2. The lymphoid system consists of primary lymphoid organs, secondary lymphoid organs, and lymphatic vessels.
3. The bone marrow and the thymus constitute the primary lymphoid organs.
4. While both B-lymphocytes and T-lymphocytes are produced from stem cells in the bone marrow, B-lymphocytes mature in the bone marrow and T-lymphocytes migrate to the thymus to mature.
5. After maturation, both naive B-lymphocytes and naive T-lymphocytes circulate between the blood and the secondary lymphoid organs.
6. Adaptive immune responses require antigen-presenting cells, such as macrophages and dendritic cells, and ever changing populations of B-lymphocytes and T- lymphocytes. These cells gather to detect and present antigens in secondary lymphoid organs.
7. The secondary lymphoid organs include highly organized lymphoid organs such as lymph nodes and the spleen, as well as less organized accumulations of lymphoid organs scattered strategically throughout the body.
8. Lymphatic vessels are responsible for flow of lymph within the lymphoid system and are a part of the body's fluid recirculation system. The lymph flows through regional lymph nodes and eventually enters the circulatory system at the heart to maintain the fluid volume of the circulation. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/12%3A_Introduction_to_Adaptive_Immunity/12.4%3A_The_Lymphoid_System.txt |
Learning Objectives
1. List the 5 general steps involved in the immune responses in their correct order.
2. State where antigens may encounter APCs, B-lymphocytes, and T-lymphocytes if they enter the following:
1. the blood
2. tissues
3. the respiratory tract
4. the gastrointestinal tract
5. the genitourinary tract
3. Briefly describe how the receptor molecules on the surface of naive B-lymphocytes, T4-helper lymphocytes, and T8-lymphocytes eventually recognize or bind epitope, indicating the roles of BCR, TCR, CD4, CD8, MHC-I, and MHC-II molecules in lymphocyte activation.
4. State the overall function of T4-effector lymphocytes and the importance behind rapid proliferation of activated lymphocytes.
5. State what types of effector cells the proliferating B-lymphocytes and T8-lymphocytes differentiate into in order to destroy or neutralize the antigen.
6. Define cytokine.
7. State the function of memory cells.
8. State what is meant by immunologic tolerance.
Whether considering humoral immunity or cell-mediated immunity, there are several general steps involved in the immune responses.
Step 1. The antigen must encounter the B-lymphocytes, T-lymphocytes, and antigen-presenting cells (APCs) capable of carrying out an adaptive immune response.
Fundamental Statement for this Step:
1. Antigens encounter the APCs, B-lymphocytes, and T-lymphocytes in the secondary lymphoid organs of the lymphoid system.
Antigens encounter the APCs, B-lymphocytes, and T-lymphocytes in the secondary lymphoid organs of the lymphoid system. Tissue fluid carries antigens to lymph nodes, blood carries antigens to the spleen, and immature dendritic cells under the skin and mucosal epithelium carry antigens to regional lymph nodes. Here they encounter ever changing populations of naive B-lymphocytes, T4-lymphocytes, and T8-lymphocytes as they circulate back and forth between the blood and the lymphatics.
1. Antigens that enter through the bloodstream, encounter the APCs, B-lymphocytes, and T-lymphocytes in the spleen ; see Figure \(1\).
2. Antigens that enter through the tissue, are picked up by tissue fluid, enter the lymph vessels, and are carried to the lymph nodes where they encounter APCs, B-lymphocytes, and T-lymphocytes; see Figure \(2\).
3. Antigens that enter the respiratory tract, encounter APCs, B-lymphocytes, and T-lymphocytes in the tonsils and the mucosa-associated lymphoid tissue (MALT), including the bronchial-associated lymphoid tissue (BALT), the nose-associated lymphoid tissue (NALT), and the larynx-associated lymphoid tissue (LALT).
4. Antigens that enter the intestinal tract, encounter APCs, B-lymphocytes, and T-lymphocytes in the Peyer's patches (see Figure \(3\)) and other gut-associated lymphoid tissues (GALT).
5. Antigens that enter the genitourinary tract, encounter APCs, B-lymphocytes, and T-lymphocytes in the mucosa-associated lymphoid tissue (MALT) found there.
6. Finally, antigens that penetrate the skin, encounter APCs, B-lymphocytes, and T-lymphocytes of the skin-associated lymphoid tissue (SALT).
Step 2. Naive B-lymphocytes, T4-lymphocytes, and T8-lymphocytes must recognize epitopes of an antigen by means of antigen-specific receptor molecules on their surface and become activated. This is known as clonal selection.
Fundamental Statements for this Step:
1. Dendritic cells bind peptide epitopes to MHC-II molecules to enable them to be recognized by complementary shaped T-cell receptors (TCR) and CD4 molecules on naive T4-lymphocyte.
2. Dendritic cells bind peptide epitopes to MHC-I molecules to enable them to be recognized by complementary shaped T-cell receptors (TCR) and CD8 molecules on naive T8-lymphocytes.
3. These interactions are required to enable the T4-lymphocyte or T8-lymphocyte to become activated, proliferate, and differentiate into effector cells.
4. Naive T4-lymphocytes have T cell receptors (TCRs ) that, in cooperation with CD4 molecules, bind to MHC-II molecules with attached epitope from an antigen found on the surface of an antigen-presenting dendritic cell.
5. Naive T8-lymphocytes have T cell receptors (TCRs) that, in cooperation with CD8 molecules, bind to MHC-I molecules with attached epitope from an antigen found on the surface of antigen-presenting dendritic cells.
6. Most proteins are T-dependent antigens. In order for naive B-lymphocytes to proliferate, differentiate and mount an antibody response against T-dependent antigens, these B-lymphocytes must interact with effector T4-lymphocytes.
7. Specialized macrophages and specialized dendritic cells called FDCs are located in the lymphoid tissues. Antigens and microbes are are found on the surface of these FDCs and macrophages which present them to complementary-shaped BCRs on naive B-lymphocytes.
8. A few antigens are called T-independent antigens. T-independent (TI) antigens are usually large carbohydrate and lipid molecules with multiple, repeating subunits. B-lymphocytes mount an antibody response to T-independent antigens without the requirement of interaction with effector T4-lymphocytes but the antibody response is much more limited than with T-dependent antigens.
a. The role of antigen-presenting dendritic cells
The primary function of dendritic cells is to capture and present protein antigens to naive T-lymphocytes.
• Dendritic cells bind peptide epitopes to MHC-II molecules (see Figure \(4\)) to enable them to be recognized by complementary shaped T-cell receptors (TCR) and CD4 molecules on naive T4-lymphocyte.
• Dendritic cells bind peptide epitopes to MHC-I molecules (see Figure \(5\)) to enable them to be recognized by complementary shaped T-cell receptors (TCR) and CD8 molecules on naive T8-lymphocytes.
These interactions are required to enable the T4-lymphocyte or T8-lymphocyte to become activated, proliferate, and differentiate into effector cells.
Most dendritic cells are derived from monocytes and are referred to as myeloid dendritic cells. They are located under the surface epithelium of the skin and the surface epithelium of the mucous membranes of the respiratory tract, genitourinary tract, and the gastrointestinal tract. They are also found throughout the body's lymphoid tissues and in most solid organs.
Upon capturing antigens through pinocytosis and phagocytosis and becoming activated by proinflammatory cytokines, the dendritic cells detach from the epithelium, enter lymph vessels, and are carried to regional lymph nodes (see Figure \(6\)). By the time they enter the lymph nodes, they have matured and are now able to present antigen to the ever changing populations of naive T-lymphocytes located in the T-cell area of the lymph nodes (see Figure \(7\)).
• To view an electron micrograph of a dendritic cell presenting antigen to T-lymphocytes, #1 see the Web page for the University of Illinois College of Medicine.
• To view an electron micrograph of a dendritic cell presenting antigen to T-lymphocytes, #2 see the Web page for the University of Illinois College of Medicine.
b. Naive T4-helper lymphocytes recognizing peptide epitopes
Naive T4-lymphocytes circulate in the blood. In response to chemokines produced by lymphoid tissues, they leave the vascular endothelium in regions called high endothelial venules and enter lymph nodes or other secondary lymphoid tissues, a process called diapedesis.
Naive T4-lymphocytes have T-cell receptors (TCRs) that, in cooperation with CD4 molecules, bind to MHC-II molecules with attached epitope from an antigen found on the surface of an antigen-presenting dendritic cells ; (see Figure \(8\)). Each T4-lymphocyte is genetically programmed to make a unique TCR. The TCR recognizes the peptide while the CD4 molecule recognizes the MHC-II molecule.
c. Naive T8-lymphocytes recognizing peptide epitopes
Naive T8-lymphocytes circulate in the blood. In response to chemokines produced by lymphoid tissues, they leave the vascular endothelium in regions called high endothelial venules and enter lymph nodes or other secondary lymphoid tissues, a process called diapedesis.
Naive T8-lymphocytes have T-cell receptors (TCRs) that, in cooperation with CD8 molecules, bind to MHC-I molecules with attached epitope from an antigen found on the surface of antigen-presenting dendritic cells (see Figure \(9\)). Each T8-lymphocyte is genetically programmed to make a unique TCR. The TCR recognizes the peptide while the CD8 molecule recognizes the MHC-I molecule.
d. Naive B-lymphocytes recognizing epitopes of antigens
Most proteins are T-dependent antigens. In order for naive B-lymphocytes to proliferate, differentiate and mount an antibody response against T-dependent antigens, these B-lymphocytes must interact with effector T4-lymphocytes. All classes or isotypes of antibody molecules can be made against T-dependent antigens and there is usually a memory response against such antigens.
Naive B-Lymphocytes encounter antigens in secondary lymphoid organs such as the lymph nodes and the spleen. Using a lymph node as an example, soluble antigens, such as microbial polysaccharides and proteins and toxins, as well as microbes such as bacteria and viruses, enter the lymph node through afferent lymphatic vessels. By this time, complement pathway activation has coated these soluble antigens or microbes with opsonins such as C3b, which in turn can be degraded to C3d.
Located within the lymphoid tissues are specialized macrophages and specialized dendritic cells called follicular dendritic cells (FDCs). These macrophages have poor endocytic ability and produce few lysosomes. The FDCs are nonphagocytic. Both cell types, however, have complement receptors called CR1 and CR2 that bind to the C3b and C3d, enabling the antigens and microbes to stick to the surface of the macrophages and FDCs. However,because of the poor endocytic ability of the macrophages and the lack of endocytosis by the FDCs, the antigens and microbes are not engulfed but rather remain on the surface of the cells. In addition, the macrophages can transfer their bound antigens or microbes to FDCs (see Figure \(10\)). Here the antigens and microbes in the lymph node can bind to complementary-shaped BCRs on naive B-lymphocytes directly, by way of macrophages, or via the FDCs (see Figure \(10\)).
A few antigens are called T-independent antigens. T-independent (TI) antigens are usually large carbohydrate and lipid molecules with multiple, repeating subunits. B-lymphocytes mount an antibody response to T-independent antigens without the requirement of interaction with effector T4-lymphocytes. Bacterial lipopolysaccharide (LPS) from the Gram-negative cell wall and capsular polysaccharides are examples of TI antigens. The resulting antibody molecules are generally of the IgM isotype and do not give rise to a memory response. There are two basic types of T-independent antigens: TI-1 and TI-2.
1. TI-1 antigens are pathogen-associated molecular patterns or PAMPS such as lipopolysaccharide (LPS) from the outer membrane of the Gram-negative cell wall and bacterial nucleic acid. These antigens activate B-lymphocytes by binding to their specific pattern-recognition receptors, in this case toll-like receptors, rather than to B-cell receptors (see Figure \(11\)). Antibody molecules generated against TI-1 antigens are often called "natural antibodies" because they are always being made against bacteria present in the body.
2. TI-2 antigens, such as capsular polysaccharides, are molecules with multiple, repeating subunits. These repeating subunits activate B-lymphocytes by simultaneously cross-linking a number of B-cell receptors (see Figure \(12\)).
Those naive B-lymphocytes not activated by epitopes of antigens exit the lymph node or other lymphoid tissue and eventually re-enter the bloodstream.
3. After the naive B-lymphocytes, T4-lymphocytes, and T8-lymphocytes bind their corresponding epitopes, they must proliferate into large clones of identical cells in order to mount a successful immune response against that antigen. This is known as clonal expansion.
Fundamental Statements for this Step:
1. With the exception of T-independent antigens, naive B-lymphocytes must be stimulated to proliferate by means of cytokines called interleukins produced primarily by effector T4- lymphocytes such as TFH cells.
2. In the case of T4-lymphocytes and T8-lymphocytes, dendritic cells produces cytokines that contribute to proliferation of the activated T-lymphocytes. CD28-dependent co-stimulation of the T4-lymphocyte also stimulates it to synthesize the cytokine interleukin-2 (IL-2) as well as a high-affinity IL-2 receptor. The binding of IL-2 to its high affinity receptor allows for cell proliferation and formation of a clone of thousands of identical T-lymphocytes after several days.
With the exception of T-independent antigens, the naive B-lymphocytes that were activated in step 2 must be stimulated to proliferate by means of cytokines called interleukins (such as IL-2, IL-4, IL-5, Il-6, and IL-10) produced primarily by effector T4- lymphocytes such as TFH cells (see Figure \(13\)).
In the case of T4-lymphocytes and T8-lymphocytes, dendritic cells produces cytokines such as interleukin-6 (IL-6), IL-4, IL-12, and T-cell growth factor-beta (TGF-ß) that contribute to proliferation of the activated T-lymphocytes. CD28-dependent co-stimulation of the T4-lymphocyte also stimulates it to synthesize the cytokine interleukin-2 (IL-2) as well as a high-affinity IL-2 receptor. The binding of IL-2 to its high affinity receptor allows for cell proliferation and formation of a clone of thousands of identical T-lymphocytes after several days.
It is thought that in most immune responses, only around 1/1000 to 1/10,000 lymphocytes will have a receptor capable of binding the initiating antigen. Thus, proliferation allows the production of clones of thousands of identical lymphocytes having specificity for the original antigen. This is essential to give enough cells to mount a successful immune response against that antigen.
4. The large clones of identical B-lymphocytes, T4-lymphocytes, and T8-lymphocytes now differentiate into effector cells capable of directing body defenses against the original antigen resulting in its destruction or neutralization.
Fundamental Statements for this Step:
1. Cytokines produced by dendritic cells and T4-effector lymphocytes enable the clones of B-lymphocytes and T-lymphocytes above to differentiate into effector cells.
2.
In the case of humoral immunity, B-lymphocytes differentiate into effector cells called plasma cells. These cells synthesize and secrete vast quantities of antibodies capable of reacting with and eliminating or neutralizing the original antigen.
3. T4-lymphocytes differentiate into T4-effector lymphocytes.
Functionally, there are many different types or subpopulations of effector T4-lymphocytes based on the cytokines they produce. Examples include TH1 cells, TH2 cells, TH17 cells, Treg cells, and TFH cells.
4. In the case of cell-mediated immunity, the T8-lymphocytes differentiate into cytotoxic T-lymphocytes (CTLs) capable of destroying body cells having the original epitope on their surface, such as viral infected cells, bacterial infected cells, and tumor cells by inducing apoptosis.
5. Antibodies, cytokines, activated macrophages, and cytotoxic T-lymphocytes eventually destroy or remove the antigen.
Cytokines produced by dendritic cells and T4-effector lymphocytes enable the clones of B-lymphocytes and T-lymphocytes from step 3 above to differentiate into effector cells.
a. In the case of humoral immunity, B-lymphocytes differentiate into effector cells calledplasma cells. These cells synthesize and secrete vast quantities of antibodies capable of reacting with and eliminating or neutralizing the original antigen (see Figure \(14\)).
b. T4-lymphocytes differentiate into T4-effector lymphocytes. Functionally, there are many different types or subpopulations of effector T4-lymphocytes based on the cytokines they produce. Immune reactions are typically dominated by five primary types: TH1 cells, TH2 cells, TH17 cells, Treg cells, and TFH cells.
1. CD4 TH1 cells: Coordinate immunity against intracellular bacteria and promote opsonization. They:
• Produce cytokines such as interferon-gamma (IFN-?) that promote cell-mediated immunity against intracellular pathogens, especially by activating macrophages that have either ingested pathogens or have become infected with intracellular microbes such as Mycobacterium tuberculosis, Mycobacterium leprae, Leishmania donovani, and Pneumocystis jiroveci that are able to grow in the endocytic vesicles of macrophages. Activation of the macrophage by TH1 cells greatly enhances their antimicrobial effectiveness.
• They produce cytokines that promote the production of opsonizing antibodies that enhance phagocytosis (see Figure \(15\)).
• Produce receptors that bind to and kill chronically infected cells, releasing the bacteria that were growing within the cell so the can be engulfed and killed by macrophages.
• Produce the cytokine interleukin-2 (IL-2) that induces T-lymphocyte proliferation.
• Produce cytokines such as tumor necrosis factor-alpha (TNF-a) that promote diapedesis of macrophages.
• Produces the chemokine CXCL2 to attract macrophages to the infection site.
• Produce cytokines that block the production of TH2 cells.
2. CD4 TH2 cells: Coordinate immunity against helminths and microbes that colonize mucous membranes
• Produce the cytokine interleukin-4 (IL-4) that promotes the production of the antibody isotype IgE in response to helminths and allergens. IgE is able to stick eosinophils to helminths for extracellular killing of the helminth (see Figure \(16\)); it also promotes many allergic reactions.
• Produce cytokines that attract and activate eosinophils and mast cells.
• Promote the production of antibodies that neutralize microbes (see Figure \(17\)) and toxins (see Figure \(18\)) preventing their attachment to host cells.
• Produce cytokines that function as B-lymphocyte growth factors such as IL-4, IL-5, IL-9. and IL-13 (see Figure \(13\)).
• Produce interleukin-22 (IL-22) that promotes the removal of microbes in mucosal tissues.
• Produce cytokines that block the production of TH1 cells.
3. CD4 TH17 cells: Promote a local inflammatory response to stimulate a strong neutrophil response and promote the integrity of the skin and mucous membranes
• Produce cytokines like interleukin-17 (IL-17) and interleukin-6 (IL-6) that trigger local epithelial cells and fibroblasts to produce chemokines that recruit neutrophils to remove extracellular pathogens.
4. CD4 Treg cells: Suppress immune responses
• Produce inhibitory cytokines such as Interleukin-10 (IL-10) and TGF-ß that help to limit immune responses and prevent autoimmunity by suppressing T-lymphocyte activity.
• Promoting anamnestic response (immunologic memory) to resist repeat infections by the same microbe.
• Protecting beneficial normal flora in the intestines from being destroyed by the immune system.
• Aiding in sustaining pregnancy so that the immune system doesn't recognize a fetus as foreign and try to destroy it.
• Controlling established inflammation in tissues.
5. TFH cells: Promote humoral immunity by stimulating antibody production and antibody isotype switching by B-lymphocytes
• T follicular helper cells (TFH cells) are located in lymphoid follicles.
• TFH cells are now thought to be the primary effector T-lymphocytes that stimulate antibody production and isotype switching by B-lymphocytes. They are able to produce cytokines that are characteristic of both TH2 cells and TH1 cells.
• TFH cells producing (IFN-?) promote the production of opsonizing antibodies; those producing IL-4 promote the production of IgE.
c. In the case of cell-mediated immunity , the T8-lymphocytes differentiate into cytotoxic T-lymphocytes (CTLs) capable of destroying body cells having the original epitope on their surface, such as viral infected cells, bacterial infected cells, and tumor cells. They do this by inducing apoptosis, a programmed cell suicide (see Figure \(19\) and Figure \(20\)). T-lymphocytes also secrete various cytokines that participate in various aspects of adoptive and innate immunity.
For More Information: Cytotoxic T-Lymphocytes from Unit 6
Progeny of the original lymphocytes leave the secondary lymphoid organs and migrate to tissues where they continue to respond to the invading antigen.
Antibodies, cytokines, activated macrophages, and cytotoxic T-lymphocytes eventually destroy or remove the antigen. Antibodies and cytokines amplify defense functions and collaborate with cells of the innate immune system, such as phagocytes and NK cells, as well as with molecules of the innate immune system, such as those of the complement system and the acute phase response. Cytotoxic T-lymphocytes (CTLs) destroy body cells having the original epitope on their surface, e.g., viral infected cells, bacterial infected cells, and tumor cells. Cytokines also amplify innate immune defenses such as inflammation, fever, and the acute phase response.
5. Some of the B-lymphocytes, T4-lymphocytes, and T8-lymphocytes differentiate into long-lived, circulating memory cells.
Fundamental Statements for this Step:
1. During the proliferation and differentiation that follows lymphocyte activation, some of the B-lymphocytes and T-lymphocytes stop replicating and become circulating, long-lived memory cells.
2.
Memory cells are capable of what is called anamnestic response or "memory", that is, they "remember" the original antigen. If that same antigen again enters the body while the memory cells are still present, these memory cells will initiate a rapid, heightened secondary response against that antigen.
During the proliferation and differentiation that follows lymphocyte activation, some of the B-lymphocytes and T-lymphocytes stop replicating and become circulating, long-lived memory cells. Memory cells are capable of what is called anamnestic response or "memory", that is, they "remember" the original antigen. If that same antigen again enters the body while the memory cells are still present, these memory cells will initiate a rapid, heightened secondary response against that antigen (see Figure \(14\) and Figure \(21\)).
This is why the body sometimes develops a permanent immunity after an infectious disease and is also the principle behind immunization.
Immune Regulation
The immune responses are carefully regulated by a variety of mechanisms. They are turned on only in response to an antigen and are turned off once the antigen has been removed.
Fundamental Statements for this Process:
1. The immune responses are carefully regulated by a variety of mechanisms. They are turned on only in response to an antigen and are turned off once the antigen has been removed.
2. The immune responses are also able to discriminate between self and non-self in order to prevent autoimmune tissue damage.
3. During the random gene-splicing reactions mentioned earlier, some lymphocytes are bound to produce receptors that fit the body's own proteins and polysaccharides. The body develops immunologic tolerance to these self antigens by triggering apoptosis in self-reactive lymphocytes.
4. Alternately, immature B-lymphocytes with self-reactive B-cell receptors may be stimulated to undergo a new gene rearrangement to make a new receptor that is no longer self-reactive. This process is called receptor editing.
5. Some autoreactive T-lymphocytes are able to slip through the system but a group of T4-effector lymphocytes called Treg cells are able to suppress their action.
6. If there is a breakdown in this normal elimination or suppression of self-reacting cells, autoimmune diseases may develop.
The immune responses are also able to discriminate between self and non-self in order to prevent autoimmune tissue damage. During the random gene-splicing reactions mentioned earlier, some lymphocytes are bound to produce receptors that fit the body's own proteins and polysaccharides. Through mechanisms that are not fully understood, the body develops immunologic tolerance to these self antigens. In other words, the immune system becomes tolerant of the body's own molecules.
During lymphocyte development, the body eliminates self-reactive lymphocytes. Self-reactive B-lymphocytes undergo negative selection. Since the bone marrow, where the B-lymphocytes are produced and mature, is normally free of foreign substances, any B-lymphocytes that bind substances there must be recognizing "self" and are eliminated by apoptosis, a programmed cell suicide. Apoptosis results in the activation of proteases within the target cell which then degrade the cell's structural proteins and DNA. Alternately, immature B-lymphocytes with self-reactive B-cell receptors may be stimulated to undergo a new gene rearrangement to make a new receptor that is no longer self-reactive. This process is called receptor editing.
This negative selection also occurs in secondary lymphoid organs whenever a T-dependent B-lymphocyte binds to an antigen but is then unable to react with its specific T-4 lymphocyte because the T4-lymphocyte does not recognize that antigen as foreign.
Self-reactive T-lymphocytes undergo both negative selection and positive selection. Positive selection occurs in the thymus and eliminates T-lymphocytes that cannot recognize MHC molecules. Because T4-lymphocytes and T8-lymphocytes can only recognize peptide epitopes bound to MHC molecules, any T-lymphocytes that cannot recognize MHC molecules fail this positive selection, do not develop any further, and are eventually eliminated. Then, each T-lymphocyte that passes positive selection by being able to recognize a MHC molecule must undergo negative selection. Any T-lymphocytes recognizing "self" peptides bound to MHC molecules are eliminated by apoptosis. Like with B-lymphocytes, this negative selection also occurs in secondary lymphoid organs whenever a T-lymphocyte binds to a peptide on a MHC molecule but is then unable to react with its specific T-4 lymphocyte because the T4-lymphocyte does not recognize that peptide as foreign.
Some autoreactive T-lymphocytes are able to slip through the system but a group of T4-effector lymphocytes called Treg cells are able to suppress their action. If there is a breakdown in this normal elimination or suppression of self-reacting cells, autoimmune diseases may develop.
We will now look at the various events discussed above in greater detail as they apply to both humoral immunity and cell-mediated immunity with special emphasis on infectious diseases. Keep in mind that some infectious agents live outside human cells (e.g., most bacteria), a few live inside the phagosomes and lysosomes of human cells through which they enter (e.g., Mycobacterium tuberculosis, Mycobacterium leprae), and others live in the fluid interior of human cells (e.g., viruses, Rickettsias, and Chlamydias). Through a combination of humoral immunity and cell-mediated immunity, all types of infectious agents, as well as many types of tumor cells, may be eliminated from the body. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/12%3A_Introduction_to_Adaptive_Immunity/12.5%3A_An_Overview_of_the_Steps_Involved_in_Adaptive_Immune_Responses.txt |
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied.
12.1: An Overview of Innate and Adaptive Immunity
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe what is meant by the following:
1. innate immunity (ans)
2. adaptive (acquired) immunity (ans)
2. Define the following:
1. antigen (ans)
2. immunogen (ans)
3. epitope (ans)
4. humoral immunity (ans)
5. cell-mediated immunity (ans)
12.2: Antigens and Epitopes
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following:
_____ Asubstance that reacts with antibody molecules and antigen receptors on lymphocytes. (ans)
_____ An antigen that is recognized by the body as non-self and stimulates an adaptive immune response. (ans)
_____ The actual portions or fragments of an antigen that react with receptors on B-lymphocytes and T-lymphocytes as well as with free antibody molecules. (ans)
_____ An antibody molecule composed of 4 glycoprotein chains whose Fc portion is anchored to the membrane of certain lymphocytes; able to recognize epitopes on protein and polysaccharide antigens. (ans)
_____ A molecule composed of 2 glycoprotein chains anchored to the membrane of certain lymphocytes; able to recognize peptide epitopes from protein antigens presented by the body's own cells by way of MHC molecules. (ans)
_____ Antigens are proteins found within the cytosol of human cells such as viral proteins, proteins from intracellular bacteria, and tumor antigens. (ans)
_____ An organism’s own antigens (self-antigens) that stimulate an autoimmune reaction. (ans)
_____ Antigens that enter from outside the body, such as bacteria, fungi, protozoa, and free viruses. (ans)
1. B-cell receptor
2. T-cell receptor
3. immunogen
4. hapten
5. epitope
6. antigen
7. autoantigens
8. endogenous antigens
9. exogenous antigens.
2. Briefly describe how the body recognizes an antigen as foreign. (ans)
3. In terms of infectious diseases, describe 2 categories of microbial materials that may act as an antigen.
1. (ans)
2. (ans)
4. Describe 3 groups of noninfectious materials that may act as an antigen.
1. (ans)
2. (ans)
3. (ans)
5. Multiple Choice (ans)
12.3: Major Cells and Key Cell Surface Molecules Involved in Adaptive Immune Responses
12.3B: Antigen-Presenting Cells (APCs)
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following in terms of antigen-presenting dendritic cells presenting antigens to naive T4-lymphocytes:
_____ Dendritic cells engulf ____________ antigens. (ans)
_____ Once engulfed by dendritic cells, protein antigens are degraded into peptides by organelles called ____________. (ans)
_____ Dendritic cells bind peptides to grooves in _________________. (ans)
_____ The dendritic cell then presents the MHC/peptide complex to the ___________________. (ans)
_____ Dendritic cells produce co-stimulatory signals after pathogen-associated molecular patterns bind to ___________________. (ans)
1. TCR of T4-lymphocytes
2. TCR of T8-lymphocytes
3. MHC-I molecules
4. MHC-II molecules
5. exogenous
6. endogenous
7. toll-like receptors
8. lysosomes
9. proteasomes
10. cytosol
2. Match the following in terms of ntigen-presenting dendritic cells presenting antigens to naive T8-lymphocytes:
_____ Dendritic cells engulf ____________ antigens. (ans)
_____ Once engulfed by dendritic cells, protein antigens are degraded into peptides by organelles called ____________. (ans)
_____ Some proteins escape from phagosomes and phagolysosomes into the ____________. (ans)
_____ Once in the cytosol, protein antigens are degraded into peptides by organelles called ____________. (ans)
_____ Dendritic cells then bind peptides to grooves in _________________. (ans)
_____ The Dendritic cell then presents the MHC/peptide complex to the ___________________. (ans)
1. TCR of T4-lymphocytes
2. TCR of T8-lymphocytes
3. MHC-I molecules
4. MHC-II molecules
5. exogenous
6. endogenous
7. toll-like receptors
8. lysosomes
9. proteasomes
10. cytosol
3. Name the primary type of cell that functions as an antigen-presenting cell to naive T4-lymphocytes and naive T8-lymphocytes. (ans)
4. State the role of T4-effector cells in activating macrophages (ans) .
5. State the role of T4-effector cells in the proliferation and differentiation of activated B-lymphocytes. (ans)
6. Multiple Choice (ans)
12.3C: T4-Lymphocytes (T4-Cells)
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following in terms of activation and function of T4-lymphocytes:
_____ Epitopes of antigens are recognized by T4-lymphocytes by way of their ____________. (ans)
_____ The TCR/CD4 molecules of T4-lymphocytes recognize ________________________ on antigen-presenting cells (APCs) such as dendritic cells, macrophages, and B-lymphocytes. (ans)
1. peptides from exogenous antigens bound to MHC-II molecules
2. peptides from endogenous antigens bound to MHC-I molecules
3. MHC-I molecules
4. toll-like receptors
5. B-cell receptors
6. T-cell receptors
7. plasma cells
8. lysosomes
9. proteasomes
2. Matching
_____ Promote cell-mediated immunity against intracellular pathogens; enhance the killing ability of macrophages, promote diapedesis and chemotaxis of macrophages, and promote the production of opsonizing antibodies. (ans)
_____ Help to limit immune responses and prevent autoimmunity by suppressing T-lymphocyte activies, promote immune memory, help to sustain pregnancy, and control established inflammation. (ans)
_____ Promote a local inflammatory response to stimulate a strong neutrophil response and promote the integrity of the skin and mucous membranes. (ans)
_____ Promote the production of the antibody isotype IgE in response to helminthsand allergens, attract and activate eosinophils and mast cells, promote the production of antibodies that neutralize microbesand toxins, and promote the removal of microbes in mucosal tissues. (ans)
1. CD4 TH2 cells
2. CD4 TH1 cells
3. CD4 Treg cells
4. CD4 TH17 cells
5. CD4 TFH cells
3. Multiple Choice (ans)
12.3D: T8-Lymphocytes (T8-Cells)
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following in terms of activation and function of T8-lymphocytes:
_____ Epitopes of antigens are recognized by T8-lymphocytes by way of their ____________. (ans)
_____ The TCR/CD8 molecules of naive T8-lymphocytes recognize ________________________ on antigen-presenting dendritic cells. (ans)
_____ After activation, T8-lymphocytes proliferate and differentiate into _____________________ (ans)
1. peptides from exogenous antigens bound to MHC-II molecules
2. peptides from endogenous antigens bound to MHC-I molecules
3. MHC-I molecules
4. toll-like receptors
5. B-cell receptors
6. T-cell receptors
7. plasma cells
8. cytotoxic T-lymphocytes (CTLs)
9. natural killer cells (NK cells)
2. State the overall function of activated T8-lymphocytes in adaptive immunity. (ans)
3. Multiple Choice (ans)
12.3E: Invarient Natural Killer T-Lymphocytes (iNKT Cells)
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Epitopes of glycolipid antigens are recognized by iNKT lymphocytes by way of their _______. (ans)
2. The TCR molecules of iNKT lymphocytes recognize ________________________ on antigen-presenting dendritic cells. (ans)
3. iNKT lymphocytes can also be activated by the cytokine __________ (ans) produced by activated dendritic cells.
4. iNKT cells promote both innate and adaptive immunity and may also regulate immune responses by way of the ____________ they produce once activated. (ans)
12.3F: B-Lymphocytes (B-Cells)
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following in terms of activation of B-lymphocytes by T-dependent antigens:
_____ Epitopes of antigens are recognized by B-lymphocytes by way of their ____________. (ans)
_____ Once engulfed by APCs, protein antigens are degraded into peptides by organelles called ____________. (ans)
_____ B-lymphocytes bind peptides to grooves in _________________. (ans)
_____ The B-lymphocyte then presents the MHC/peptide complex to the ___________________. (ans)
_____ B-lymphocytes eventually differentiate into antibody-secreting cells called ___________________. (ans)
1. TCR of T4-lymphocytes
2. TCR of T8-lymphocytes
3. MHC-I molecules
4. MHC-II molecules
5. B-cell receptors
6. CD4 molecules
7. plasma cells
8. lysosomes
9. proteasomes
2. State the overall function of B-lymphocytes in adaptive immunity. (ans)
3. Multiple Choice (ans)
12.3G: Natural Killer Cells (NK Cells)
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe how NK cells bind to and kill infected cells and tumor cells through ADCC. (ans)
2. Briefly describe how NK cells recognize and kill infected cells and tumor cells that suppress MHC-I production. (ans)
12.3A: Major Histocompatibility Complex (MHC) Molecules
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following:
_____ Produced by all nucleated cells in the body. (ans)
_____ Produced primarily by antigen-presenting cells such as macrophages, dendritic cells, and B-lymphocytes. (ans)
_____ Primarily bind peptides from exogenous antigens. (ans)
_____ Primarily bind peptides from endogenous antigens. (ans)
_____ Recognize peptides bound to MHC-II molecules. (ans)
_____ Recognize peptides bound to MHC-I molecules. (ans)
1. TCR of T4-lymphocytes
2. TCR of T8-lymphocytes
3. MHC-I molecules
4. MHC-II molecules
2. State the role of proteasomes in binding of peptides from endogenous antigens by MHC-I molecules. (ans)
3. State the role of lysosomes in binding of peptides from exogenous antigens by MHC-II molecules. (ans)
4. Multiple Choice (ans)
12.4: The Lymphoid System
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following with the BEST answer:
_____ Contain antigen-presenting cells, such as macrophages and dendritic cells, and ever changing populations ofB-lymphocytes and T- lymphocytes. Examples include the tonsils, the appendix, Peyer's patches, MALT, SALT, lymph nodes, and the spleen. (ans)
_____ Produce B-lymphocytes and T-lymphocytes. The bone marrow and the thymus. (ans)
_____ The fluid surrounding cells in the body. (ans)
_____ The liquid portion of the blood. (ans)
_____ A diffuse system of small concentrations of lymphoid tissue found in various sites of the body such as the gastrointestinal tract, respiratory tract, eye, and skin. It is populated by loose clusters of T-lymphocytes, B-lymphocytes, plasma cells, activated TH cells, and macrophages. (ans)
_____ The liquid found in lymph vessels. (ans)
_____ Expose antigens found in the lymph to dendritic cells, B-lymphocytes, and T-lymphocytes. (ans)
_____ Expose antigens found in the blood to dendritic cells, B-lymphocytes, and T-lymphocytes. (ans)
1. plasma
2. lymph
3. tissue fluid
4. primary lymphoid organs
5. secondary lymphoid organs
6. the spleen
7. lymph nodes
8. MALT
2. Briefly describe the importance of the lymphoid system in adaptive immune responses. (ans)
3. Multiple Choice (ans)
12.5: An Overview of the Steps Involved in Adaptive Immune Responses
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. State where antigens may encounter APCs, B-lymphocytes, and T-lymphocytes if they enter the following:
1. the blood (ans)
2. tissues (ans)
3. the respiratory tract (ans)
4. the gastrointestinal tract (ans)
5. the genitourinary tract (ans)
2. Match the following:
_____ Use lysosomes to degrade exogenous antigens into peptides, bind them to MHC-II molecules, and present them to naive T4-lymphocytes. (ans)
_____ Uses BCR to recognize epitopes of antigens; a few antigens are recognized by toll-like receptors. (ans)
_____ Uses TCR and CD4 to recognize peptide epitopes from exogenous antigens bound to MHC-II molecules of antigen-presenting dendritic cells, macrophages, and B-lymphocytes. (ans)
_____ Uses TCR and CD8 to recognize peptide epitopes from endogenous antigens bound to MHC-I molecules of cells. (ans)
_____ Cells that allow for a heightened secondary response upon subsequent exposure to the same antigen. (ans)
_____ Once activated itself, secretes cytokines that enable activated B-lymphocytes and T-lymphocytes to proliferate and differentiate. (ans)
_____ Use proteasomes to degrade endogenous antigens into peptides, bind them to MHC-I molecules, and present them to naive T8-lymphocytes. (ans)
_____ Differentiate into antibody secreting plasma cells. (ans)
_____ Differentiate into cytotoxic T-lymphocytes (CTLs). (ans)
1. T4-lymphocytes
2. T8-lymphocytes
3. dendritic cells
4. B-lymphocytes
5. memory cells
3. State the overall function of T4-effector lymphocytes and the importance behind rapid proliferation of activated lymphocytes. (ans)
4. The ability of the body to initiate and direct adaptive immune responses against antigenic molecules foreign to the body but not against antigenic molecules that are a normal component of the body is called ____________________________. (ans)
5. Multiple Choice (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/12%3A_Introduction_to_Adaptive_Immunity/12.E%3A_Introduction_to_Adaptive_Immunity_%28Exercises%29.txt |
Humoral Immunity refers to the production of antibody molecules in response to an antigen. These antibody molecules circulate in the plasma of the blood and enter tissue and organs via the inflammatory response. Humoral immunity is most effective microbes or their toxins located in the extracellular spaces of the body. Antibodies or immunoglobulins are specific glycoprotein configurations produced by B-lymphocytes and plasma cells in response to a specific antigen and capable of reacting with that antigen.
• 13.1: Antibodies (Immunoglobulins)
Humoral Immunity refers to the production of antibody molecules in response to an antigen. Humoral immunity is most effective microbes or their toxins located in the extracellular spaces of the body. Antibodies or immunoglobulins are specific glycoprotein configurations produced by B-lymphocytes and plasma cells in response to a specific antigen and capable of reacting with that antigen.
• 13.2: Ways That Antibodies Help to Defend the Body
The antibodies produced during humoral immunity ultimately defend the body through a variety of different means. These include: Opsonization, MAC Cytolysis, Antibody-dependent Cellular Cytotoxicity (ADCC) by NK Cells, Neutralization of Exotoxins, Neutralization of Viruse, Preventing Bacterial Adherence to Host Cells, Agglutination of Microorganisms, Immobilization of Bacteria and Protozoan, and Promoting an Inflammatory Response.
• 13.3: Naturally and Artificially Acquired Active and Passive Immunity
During passive immunity, the body receives antibodies made in another person or animal and the immunity is short-lived. During active immunity, antigens enter the body and the body responds by making its own antibodies and B-memory cells. In this case, immunity is longer lived although duration depends on the persistence of the antigen and the memory cells in the body.
• 13.E: Humoral Immunity (Exercises)
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied.
13: Humoral Immunity
Humoral Immunity refers to the production of antibody molecules in response to an antigen. These antibody molecules circulate in the plasma of the blood and enter tissue and organs via the inflammatory response. Humoral immunity is most effective microbes or their toxins located in the extracellular spaces of the body. Antibodies or immunoglobulins are specific glycoprotein configurations produced by B-lymphocytes and plasma cells in response to a specific antigen and capable of reacting with that antigen. In this section we will look at the structure of antibodies, the 5 classes or isotypes of human antibodies, generation of antibody diversity, clonal selection and clonal expansion, and memory (anamnestic) response.
13.1: Antibodies (Immunoglobulins)
Learning Objectives
1. Describe an antibody molecule.
2. Draw the "stick figure" structure of IgG, indicating the Fab portion (variable region) and the Fc portion (constant region).
3. State the functions of the Fab and the Fc portions of an antibody.
4. State what is meant by the biological activity of an antibody.
5. Compare the structure of IgM and secretory IgA with that of IgG.
In this section we will look at the structure of antibodies. There are five classes or isotypes of human antibodies :
1. immunoglobulin G (IgG),
2. immunoglobulin M (IgM),
3. immunoglobulin A (IgA),
4. immunoglobulin D (IgD), and
5. immunoglobulin E (IgE).
The simplest antibodies, such as IgG, IgD, and IgE, are "Y"-shaped macromolecules called monomers. A monomer is composed of four glycoprotein chains: two identical heavy chains and two identical light chains. The two heavy chains have a high molecular weight that varies with the class of antibody. The light chains come in two varieties: kappa or lambda and have a lower molecular weight than the heavy chains. The four glycoprotein chains are connected to one another by disulfide (S-S) bonds and non-covalent bonds (Figure \(1\)).
Additional S-S bonds fold the individual glycoprotein chains into a number of distinct globular domains (Figure \(2\)). The area where the top of the "Y" joins the bottom is called the hinge. This area is flexible to enable the antibody to bind to pairs of epitopes various distances apart on an antigen.
The two tips of the "Y" monomer are referred to as the antigen-binding fragments or Fab portions of the antibody (Figures 1-3). The first 110 amino acids or first domain of both the heavy and light chain of the Fab region of the antibody provide specificity for binding an epitope on an antigen. The amino acid sequence of this first domain of both the light chain and the heavy chain shows tremendous variation from antibody to antibody and constitutes the variable region (V region). This is because each B-lymphocyte, early in its development, becomes genetically programmed through a series of gene-splicing reactions to produce a Fab with a unique 3-dimensional shape capable of fitting some epitope with a corresponding shape.
Figure \(3\): Ribbon Drawing of the Antibody Molecule IgG2a, A ribbon drawing of the first intact antibody molecule ever crystallized (IgG2a). The Fab portion of the antibody has specificity for binding an epitope of an antigen. The Fc portion directs the biological activity of the antibody.
The various genes the cell splices together determine the order of amino acids of the Fab portion of both the light and heavy chain; the amino acid sequence determines the final 3-dimensional shape (Figure \(4\)). Therefore, different antibody molecules produced by different B-lymphocytes will have different orders of amino acids at the tips of the Fab to give them unique shapes for binding epitope. The antigen-binding site is large enough to hold an epitope of about 5-7 amino acids or 3-4 sugar residues. Epitopes bind to the Fab portion of the antibody by reversible, non-covalent bonds.
The bottom part of the "Y", the C terminal region of each glycoprotein chain, is called the Fc portion. The Fc portion, as well as one domain of both the heavy and light chain of the Fab region has a constant amino acid sequence and is referred to as the constant region (C region) of the antibody and defines the class and subclass of each antibody. The Fc portion is responsible for the biological activity of the antibody (Figures 1-3), however, the Fc portion only becomes biologically active after the Fab component has bound to its corresponding antigen. Depending on the class and subclass of antibody, biological activities of the Fc portion of antibodies include the ability to:
• Activate the classical complement pathway (IgG & IgM); see Figure \(5\).
• Activate the lectin complement pathway and the alternative complement pathway (IgA)
• Bind to receptors on phagocytes (IgG); see Figure \(6\).
• Bind to receptors on mast cells, basophils, and eosinophils (IgE); see Fig 7 and Figure \(8\).
• Bind to receptors on NK cells (IgG); see Figure \(9\).
• Determine the tissue distribution of the antibodies, that is, to what tissues types the antibody molecules are able to go.
Individual "Y"-shaped antibody molecules are called monomers and can bind to two identical epitopes. Antibodies of the classes IgG, IgD, and IgE are monomers.
Two classes of antibodies are more complex. IgM (see Figure \(10\)) is a pentamer, consisting of 5 "Y"-like molecules connected at their Fc portions by a "J" or joining chain. Secretory IgA (see Figure \(11\)) is a dimer consisting of 2 "Y"-like molecules connected at their Fc portions by a "J" chain and stabilized to resist enzymatic digestion in body secretions by means of a secretory component.
Summary
1. There are 5 classes or isotypes of human antibodies or immunoglobulins: IgG, IgM, IgA, IgD, and IgE.
2. The simplest antibodies, such as IgG, IgD, and IgE, are "Y"-shaped macromolecules called monomers and are composed of four glycoprotein chains: two identical heavy chains and two identical light chains.
3. The two tips of the "Y" monomer are referred to as the antigen-binding fragments or Fab portions of the antibody and these portions provide specificity for binding an epitope on an antigen.
4. Early in its development, each B-lymphocyte becomes genetically programmed through a series of gene-splicing reactions to produce a Fab with a unique 3-dimensional shape capable of fitting some epitope with a corresponding shape.
5. The Fc portion only becomes biologically active after the Fab component has bound to its corresponding antigen. Biological activities include activating the complement pathways, and binding to receptors on phagocytes and other defense cells to promote adaptive immunity.
6. IgM is a pentamer, consisting of 5 monomers joined at their Fc portions.
7. IgA is a dimer, consisting of 2 monomers joined at their Fc portions.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe an antibody molecule. (ans)
2. Match the following:
_____ The region of the antibody that provide specificity for binding an epitope on an antigen. (ans)
_____ The region of the antibody that is responsible for the biological activity of the antibody. (ans)
_____ Composed of four glycoprotein chains. There are two identical heavy chains having a high molecular weight and two identical light chains. (ans)
_____ A pentamer, consisting of 5 "Y"-like molecules connected at their Fc portions by a "J" or joining chain. (ans)
_____ A dimer consisting of 2 "Y"-like molecules connected at their Fc portions by a "J" chain and stabilized to resist enzymatic digestion. (ans)
1. IgM
2. secretory IgA
3. IgG
4. Fab
5. Fc
3. Multiple Choice (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.1%3A_Antibodies_%28Immunoglobulins%29/13.1B%3A_Antibody_Structure.txt |
State which classes (isotypes) of human antibodies possess the following characteristics: are monomers is a pentamer is a dimer activates the classical complement pathway by its Fc portion binds to macrophages and neutrophils by its Fc portion binds to NK cells by its Fc portion crosses the placenta functions as a B-cell receptor the first antibody produced during an adaptive immune response binds to components of mucous by its Fc portion found mainly in body secretions binds to mast cells and basophils by its Fc portion and promotes inflammation, coughing, sneezing, vomiting, and allergic reactions binds to eosinophils by its Fc portion and promotes the removal of parasitic worms and arthropods Match the antibody isotype with its description.
For More Information: Classical complement pathway from Unit 5
For More Information: Opsonization from Unit 6
For More Information: Antibody-dependent cellular cytotoxicity (ADCC) from Unit 6
IgM (Immunoglobulin M)
The Fc portion of secretory IgA binds to components of mucous and contributes to the ability of mucous to trap microbes. The Fc portion of secretory IgA can bind to macrophages and neutrophils for enhanced attachment (opsonization). IgA can activate the lectin complement pathway and the alternative complement pathway.
IgD: (Immunoglobulin D)
IgD makes up approximately 0.2% of the serum antibodies. IgD is a monomer and has 2 epitope-binding sites and is found on the surface of B-lymphocytes (along with monomeric IgM) as a B-cell receptor where it may control of B-lymphocyte activation and suppression. IgD may play a role in eliminating B-lymphocytes generating self-reactive autoantibodies.
IgE (Immunoglobulin E)
IgE makes up about 0.002% of the serum antibodies with a half-life of 2 days. Most IgE is tightly bound to basophils and mast cells via its Fc region . IgE is a monomer and has 2 epitope-binding sites. IgE is made in response to parasitic worms (helminths) and arthropods. It is also often made in response to allergens(allergens are antigens causing allergic reactions). IgE may protect external mucosal surfaces by promoting inflammation, enabling IgG, complement proteins, and leukocytes to enter the tissues, as well as by triggering coughing, sneezing, and vomiting for mechanical removal of microbes and toxins. .
The Fc portion of IgE can bind to mast cells and basophils where it mediates many allergic reactions. Cross linking of cell-bound IgE by antigen triggers the release of vasodilators for an inflammatory response (Fig 7). The Fc portion of IgE made against parasitic worms and arthropods can bind to eosinophils enabling opsonization (Figure \(8\)). This is a major defense against parasitic worms and arthropods.
For More Information: IgE-mediated hypersensitivity (Type-I) from Unit 6
Each day an average adult produces approximately three grams of antibodies, about two-thirds of this IgA.
Summary
1. IgG makes up approximately 80% of the serum antibodies, is a monomer with 2 Fab sites. The Fc portion can activate the classical complement pathway, bind to macrophages and neutrophils to enable opsonization, bind to NK cells to promote ADCC, and can cross the placenta.
2. IgM makes up approximately 13% of the serum antibodies, is the first antibody produced during an immune response, is found mainly in the blood, and is a pentamer with 10 Fab sites. The Fc portion can activate the classical complement pathway. Monomeric forms of IgM are found on the surface of B-lymphocytes as B-cell receptors.
3. IgA makes up approximately 6% of the serum antibodies, is a dimer with 4 epitope-binding sites and is found mainly in body secretions as secretory IgA (sIgA) where it protects internal body surfaces exposed to the environment by blocking the attachment of bacteria and viruses to mucous membranes.
4. The Fc portion of secretory IgA binds to components of mucous and contributes to the ability of mucous to trap microbes, and can bind to macrophages and neutrophils to enable opsonization, and can activate the lectin complement pathway and the alternative complement pathway.
5. IgD makes up approximately 0.2% of the serum antibodies, is a monomer with 2 Fab sites, is found on the surface of B-lymphocytes as a B-cell receptor, and may play a role in eliminating B-lymphocytes generating self-reactive autoantibodies.
6. IgE makes up about 0.002% of the serum antibodies, is a monomer with 2 Fab sites, and is made in response to parasitic worms (helminths) and arthropods. It is also often made in response to allergens. The Fc portion of IgE can bind to mast cells and basophils (see Figure \(8\)) where it mediates many allergic reactions, and the Fc portion of IgE made against parasitic worms can bind to eosinophils enabling opsonization. IgE may also protect external mucosal surfaces by promoting inflammation.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the class (isotype) of human antibody with its description.
2. Multiple Choice (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.1%3A_Antibodies_%28Immunoglobulins%29/13.1C%3A_The_5_Classes_%28Isotypes%29_of_Human_Antibodies.txt |
Define gene translocation and relate it to each B-lymphocyte being able to produce an antibody with a unique shaped Fab. Define the following: combinatorial diversity junctional diversity affinity maturation
In this section we will look at generation of antibody diversity through gene translocation. As mentioned earlier, the immune system of the body has no idea as to what antigens it may eventually encounter. Therefore, it has evolved a system that possesses the capability of responding to any conceivable antigen. The immune system can do this because both B-lymphocytes and T-lymphocytes have evolved a unique system of gene-splicing called gene translocation, a type of gene-shuffling process where various different genes along a chromosome are cut out of one location and joined with other genes along the chromosome.
To demonstrate this gene translocation process, we will look at how each B-lymphocyte becomes genetically programmed to produce an antibody functioning as a B-cell receptor(BCR) having a unique shaped Fab. As mentioned above, the Fab portion of an antibody is composed of 2 protein chains: a heavy and a light (see Figure \(1\)).
The variable heavy chain portion of the Fab is coded for by a combination of 3 genes, called VH (variable heavy), DH (diversity heavy), and JH (joining heavy). The variable light chain portion of the Fab consists of either a kappa chain or a lambda chain coded for by a combination of 2 genes, VL (variable light) and JL (joining light). In the DNA of each B-lymphocyte there are multiple forms of each one of these variable determinant genes. Although the exact number of each gene isn't known and varies from person, there are approximately 38-46 VH genes; 23 DH genes; 6 JH genes; 34-38 kappa VL genes; 5 kappa JL genes; 29-33 lambda VL genes; and 4-5 lambda JL genes.
While a person inherits alleles for the various V(D)J genes from each parent, an individual B-lymphocyte will only express an inherited allele set from one parent. This increases a greater diversity of antibodies in that individual.
Through random gene translocation, any combination of the multiple forms of each gene can join together (see Figure \(2\)) resulting in thousands of possible gene combinations. This is known as combinatorial diversity.
Gene translocation of the V(D)J genes is initiated when an enzyme called V(D)J recombinase recognizes recombination signal sequences located at the 3' end of V genes, the 5' end of J genes, and both ends of D genes. As a result, the chromosome forms a loop allowing different genes from different regions along the chromosome to align (see Figure \(3\)). In the heavy chain any J-heavy gene and any D-heavy gene align and bind together as the genes are cut from one location and pasted into another. Subsequently, any one of the V-heavy genes is attached to this DJ segment. In the light chain, chromosomal looping enables any V-light gene to attach to any J-light gene.
Flash animation showing gene translocation and combinatorial diversity.
html5 version of animation for iPad showing gene translocation and combinatorial diversity.
During gene translocation, specialized enzymes in the B-lymphocyte cause splicing inaccuracies wherein additional nucleotides are added or deleted at the various gene junctions. This change in the nucleotide base sequence generates even greater diversity in Fab shape. This is called junctional diversity.
Furthermore, as B-lymphocytes proliferate, they undergo affinity maturation, a process that "fine tunes" the shape of the Fab epitope binding site. This is because the immunoglobulin V genes of B-lymphocytes have a mutation rate between 1000 to 10,000 times greater than other human genes in the body. This somatic hypermutation creates a great opportunity for selection of variant B-lymphocytes with even better fitting antigen-binding sites that fit the epitope more precisely. The longer and more tightly the antigen binds to the B-cell receptor, the greater the chance that B-lymphocyte has of surviving and replicating. In other words, the "fit" of the antibody can be improved over time. Affinity maturation occurs in the germinal centers of the lymph nodes.
Most likely humans produce at least 1011 different shaped BCRs. Keep in mind that the 3-dimensional shape of a protein is ultimately determined by the sequence of its amino acids and the sequence of amino acids is determined by the order of nitrogenous bases in the genes coding for that protein. Between combinatorial diversity, junctional diversity, and affinity maturation, there are probably billions of possible gene combinations and rearrangements that can code for the Fab portions of an antibody. Chances are, then, each B-lymphocyte will carry out a unique series of gene translocations and be able to produce an antibody with a unique shaped epitope-binding site.
Because gene translocation is a random process, some immature B-lymphocytes do wind up making B-cell receptors that fit the body's own antigens. Immature B-lymphocytes with self-reactive B-cell receptors may be stimulated to undergo a new gene rearrangement to make a new receptor that is no longer self-reactive. Recognition of self antigen can reactivate genes that allow the B-lymphocyte to carry out new light chain V-J recombinations and enabling that cell to express a new B-cell receptor. This process is called receptor editing.
Alternately, self-reactive B-lymphocytes can also undergo negative selection. Since the bone marrow, where the B-lymphocytes are produced and mature, is normally free of foreign substances, any B-lymphocytes that bind substances there must be recognizing "self" and are eliminated by apoptosis, a programmed cell suicide. Apoptosis results in the activation of proteases within the target cell which then degrade the cell's structural proteins and DNA.
Summary
1. The adaptive immune responses have evolved a system that possesses the capability of responding to any conceivable antigen the body might eventually encounter through a process called gene translocation.
2. Gene translocation is a type of gene-shuffling process where various different genes along a chromosome are cut out of one location and joined with other genes along the chromosome to create a maximum number of different B-cell and T-cell receptors.
3. Each B-lymphocyte becomes genetically programmed to produce an antibody functioning as a B-cell receptor (BCR) having a unique shaped Fab.
4. The variable portion of both the heavy and light chain of the antibody is coded for by multiple genes and there are multiple forms of each one of these variable genes.
5. Through random gene translocations, any combination of the multiple forms of each gene can join together resulting in thousands of possible gene combinations. This is known as combinatorial diversity.
6. During gene translocation, specialized enzymes in the B-lymphocyte cause splicing inaccuracies wherein additional nucleotides are added or deleted at the various gene junctions and this change in the nucleotide base sequence generates even greater diversity in Fab shape. This is called junctional diversity.
7. As B-lymphocytes proliferate, they undergo affinity maturation, a process that "fine tunes" the shape of the Fab epitope binding site through a high rate of somatic hypermutation. This creates a great opportunity for selection of variant B-lymphocytes with even better fitting antigen-binding sites that fit the epitope more precisely.
8. Immature B-lymphocytes with self-reactive B-cell receptors may be stimulated to undergo a new gene rearrangement to make a new receptor that is no longer self-reactive through a process called receptor editing. Alternately, self-reactive B-lymphocytes can also undergo negative selection whereby any B-lymphocytes that bind substances recognized as "self" and are eliminated by apoptosis.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Define gene translocation. (ans)
2. Relate gene translocation to each B-lymphocyte being able to produce an antibody with a unique shaped Fab. (ans)
3. Define the following:
1. combinatorial diversity (ans)
2. affinity maturation (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.1%3A_Antibodies_%28Immunoglobulins%29/13.1D%3A_Generation_of_Antibody_Diversity.txt |
Learning Objectives
• Briefly describe the process of clonal selection and clonal expansion.
As mentioned above, during early differentiation of naive B-lymphocytes in the bone marrow, each B-lymphocyte becomes genetically programmed to make an antibody with a unique antigen-binding site (Fab) through a series of gene translocations, and molecules of that antibody are put on its surface to function as the B-cell receptor (Figure \(1\)).
When an antigen encounters the immune system, its epitopes eventually will react only with B-lymphocytes with B-cell receptors on their surface that more or less fit and this activates those B-lymphocytes. This process is known as clonal selection (Figure \(2\)).
Cytokines produced by effector T4-helper lymphocytes enable those activated B-lymphocytes to rapidly proliferate to produce large clones of thousands of identical B-lymphocytes. In this way, even though only a few B-lymphocytes in the body may have an antibody molecule able to fit a particular epitope, eventually many thousands of cells are produced with the right specificity. This is referred to as clonal expansion (Figure \(3\)).
Furthermore, as the B-lymphocytes proliferate, they undergo affinity maturation as a result of somatic hypermutations. This allows the B-lymphocytes to "fine-tune" the shape of the antibody for better fit with the original epitope. B-lymphocytes having better fitting B-cell receptor on their surface bind epitope longer and more tightly allowing these cells to selectively replicate. Eventually these variant B-lymphocytes differentiate intoplasma cells that synthesize and secrete vast quantities of antibodies that have Fab sites which fit the original epitope very precisely (Figure \(4\)). It generally takes 4-5 days for a naive B- lymphocyte that has been activated to complete clonal expansion and differentiate into effector B-lymphocytes.
A single activated B-lymphocyte can, within seven days, give rise to approximately 4000 antibody-secreting cells. Over 2000 antibody molecules can be produced per plasma cell per second for typically up to four to five days. The B-memory cells that eventually form also have these high affinity antibodies on their surface.
Animation Overview: Clonal Selection and Clonal Expansion
During its development, each B-lymphocyte becomes genetically programmed, through a process called gene translocation, to make a unique antibody molecule that will function as a B-cell receptor. Molecules of that antibody are then placed on the cell's surface where it can react with epitopes of an antigen. A B-lymphocyte with an appropriately fitting B-cell receptor can now react with epitopes of an antigen having a corresponding shape. This activates the B-lymphocyte. Cytokines from an activated T4-lymphocyte now enable the activated B-lymphocyte to proliferate into a large clone of identical B-lymphocytes. During this time, "fine-tuning" of the B-cell receptor occurs as a result of affinity maturation. The B-lymphocytes now differentiate into antibody-secreting B-lymphocytes and plasma cells that secrete large quantities of antibodies "fitting" the original epitope. Some B-lymphocytes differentiate into B-memory cells capable of anamnestic response.
As with naive B-lymphocytes, during its development, each naive T4-lymphocyte becomes genetically programmed by gene-splicing reactions similar to those in B-lymphocytes, to produce a TCR with a unique specificity. Identical molecules of that TCR are placed on its surface where they are able to bind an epitope/MHC-II complex on an antigen-presenting dendritic cell with a corresponding shape (Figure \(5\)). This is clonal selection of the T4-lymphocytes that are required for the body's response to T-dependent antigens.
In response to cytokines, these activated T4-lymphocytes now rapidly proliferate and differentiate into effector T4-lymphocytes. This is clonal expansion of the T4-lymphocytes. Before an antigen enters the body, the number of naive T4-lymphocytes specific for any particular antigen is between 1 in 105 to 106 lymphocytes. After antigen exposure, the number of T4-lymphocytes specific for that antigen may increase to 1 in 100 to 1000 lymphocytes.
Summary
1. Each naïve B-cell becomes genetically programmed to make an antibody with a unique antigen-binding site (Fab) through a series of gene translocations, and molecules of that antibody are put on its surface to function as the B-cell receptor.
2. When an antigen encounters the immune system, its epitopes eventually will react only with B-lymphocytes with B-cell receptors on their surface that more or less fit and this activates those B-lymphocytes. This process is known as clonal selection.
3. Cytokines produced by activated T4-helper lymphocytes enable those activated B-lymphocytes to rapidly proliferate to produce large clones of thousands of identical B-lymphocytes.
4. In this way, even though only a few B-lymphocytes in the body may have an antibody molecule able to fit a particular epitope, eventually eventually many thousands of cells are produced with the right specificity. This is referred to as clonal expansion.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the process of clonal selection and clonal expansion. (ans)
2. Multiple Choice (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.1%3A_Antibodies_%28Immunoglobulins%29/13.1E%3A_Clonal_Selection_and_Clonal_Expansion.txt |
Learning Objectives
1. In terms of humoral immunity, statewhat is meant by anamnestic response and discuss its role in immune defense.
2. Briefly describe why there is a heightened secondary response during anamestic response.
As a result of B-lymphocytes recognizing T-dependent antigens (proteins) during humoral immunity, numerous circulating B-memory cells and T4-memory cells develop (Figure \(1\)), which possess anamnestic response or memory. (During cell-mediated immunity, T8-memory cells also develop.) A subsequent exposure to that same antigen results in:
• A more rapid production of antibodies;
• Produced in greater amounts; and
• Produced for a longer period of time.
The primary response to a new antigen generally peaks at 5-10 days. IgM is made first later to be replaced by IgG. Because of the numerous circulating B-memory cells and T4-memory cells from the primary response, however, the secondary anamnestic response peaks in only 1-3 days (Figure \(2\)). There is an increase in the amount of IgG made and under certain conditions, IgA or IgE may be made.
Because of clonal expansion and affinity maturation , there is now a pool of B-memory cells having the "fine-tuned" B-cell receptors on their surface. The pool of B-memory cells migrate to lymph nodes, to mucosal tissue, and circulate in the blood waiting to encounter the original antigen if it again enters the body. B-memory cells have a long life and also replicate and produce antibodies periodically when they are exposed to persisting epitope remaining on the surface of follicular dendritic cells in the lymphoid organs. In addition to the B-memory cells, a pool of circulating T4-effector memory cells (CD4 TEM cells), as well T4 tissue resident memory cells (CD4 TRM cells) located in the mucosa enable an accelerated helper function.
This memory response applies to T-dependent antigens . In the case of the T-independent antigens, there is usually no anamnestic response.
In the case of systemic infections and most vaccinations, many of the plasma cells migrate to the bone marrow where they may continue to secrete antibodies for months or years after the antigen has been eliminated. Plasma cells produced in the mucous membranes generally remain in the mucous membranes and secrete antibodies for only around a year.
Memory is better in preventing systemic infections than preventing mucosal infections because infections limited to the mucous membranes generally do not provide enough time for the development of effector cells such as plasma cells, effector T4-lymphocytes , and cytotoxic T-lymphocytes from the activated memory cells.
Summary
1. As a result of B-lymphocytes recognizing T-dependent antigens (proteins) during humoral immunity, numerous circulating B-memory cells and T4-memory cells develop which possess anamnestic response or memory.
2. A subsequent exposure to that same antigen results in a more rapid production of antibodies that are produced in greater amounts for a longer period of time.
3. The primary response to a new antigen generally peaks at 5 - 10 days.
4. Because of the numerous circulating B-memory cells and T4-memory cells from the primary response, the secondary anamnestic response peaks in only 1 - 3 days.
5. In the case of systemic infections and most vaccinations, many of the plasma cells migrate to the bone marrow where they may continue to secrete antibodies for months or years after the antigen has been eliminated.
6. Plasma cells produced in the mucous membranes generally remain in the mucous membranes and secrete antibodies for only around a year. Therefore, anamnestic response is better at preventing systemic infections than preventing mucosal infections.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. In terms of humoral immunity, discusswhat is meant by anamnestic response. (ans)
2. Briefly describe why there is a heightened secondary response during anamestic response. (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.1%3A_Antibodies_%28Immunoglobulins%29/13.1F%3A_Anamnestic_%28Memory%29_Response.txt |
The antibodies produced during humoral immunity ultimately defend the body through a variety of different means.
13.2: Ways That Antibodies Help to Defend the Body
Learning Objectives
• Discuss how antibodies defend the body by way of opsonization. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.)
• Briefly describe two different ways bacteria may resist opsonization.
Opsonization, or enhanced attachment, refers to the antibody molecules IgG and IgE, the complement proteins C3b and C4b, and other opsonins attaching antigens to phagocytes. This results in a much more efficient phagocytosis.
Opsonization with IgG, IgA, C3b, and C4b
The process starts with antibodies of the isotype IgG, IgA, or IgM being made against a surface antigen of the organism or cell to be phagocytosed. The Fab portions of the antibody react with epitopes of the antigen. The Fc portion of IgG (but not IgM) can then bind to receptors on neutrophils and macrophages thus sticking the antigen to the phagocyte (Figure \(1\)). The Fc portion of secretory IgA can also bind to macrophages and neutrophils for opsonization.
The Fc portion of secretory IgA can also bind to macrophages and neutrophils for opsonization.
Alternately, IgG, IgA, and IgM can activate the complement pathways (Figure \(2\)) and C3b or C4b can stick the antigen to phagocytes (Figure \(1\)). Like IgG, C3b, and to a lesser extent C4b, can function as opsonins, that is, they can attach antigens to phagocytes.One portion of the C3b binds to proteins and polysaccharides on microbial surfaces; another portion attaches to CR1 receptors on phagocytes, B-lymphocytes, and dendritic cells for enhanced phagocytosis (Figure \(3\)). (Remember that C3b and C4b are also produced during the alternative complement pathway and the lectin pathway as was discussed in Unit 5.) Activation of the complement pathway also promotes inflammation to bring phagocytes and defense chemicals from the bloodstream to the infection site as discussed later under this topic.
Actually, C3b molecule can bind to pretty much any protein or polysaccharide. Human cells, however, produce Factor H that binds to C3b and allows Factor I to inactivate the C3b. On the other hand, substances such as LPS on bacterial cells facilitate the binding of Factor B to C3b and this protects the C3b from inactivation by Factor I. In this way, C3b does not interact with our own cells but is able to interact with microbial cells.
Attachment then promotes destruction of the antigen. Microorganisms are placed in phagosomes (Figure \(4\)) where they are ultimately digested by lysosomes (Figure \(5\)). If the antigen is a cell too large to be ingested - such as virus-infected host cells, transplant cells, and cancer cells - the phagocyte empties the contents of its lysosomes directly on the cell for extracellular killing (Figure \(6\) and Figure \(7\)).
Opsonization is especially important against microorganisms with antiphagocytic structures such as capsules since opsonizing antibodies made against the capsule are able to stick capsules to phagocytes (Figure \(8\)). In vaccines against pneumococccal pneumonia and Haemophilus influenzae type b, it is capsular polysaccharide that is given as the antigen in order to stimulate the body to make opsonizing antibodies against the encapsulated bacterium.
Opsonization with IgE and the promotion of inflammation
The antibody isotype IgE is made against parasitic worms (helminths) and arthropods. The Fab portions of IgE bind to epitopes on the helminth or arthropod while the Fc portion binds to receptors on eosinophils enabling opsonization. In other words, IgE sticks phagocytic eosinophils to helminths and arthropods for the extracellular killing of that organism (Figure \(9\)).
The Fc portion of IgE also binds to receptors on mast cells and basophils to trigger the release of inflammatory mediators (Figure \(10\)). The inflammatory response then enables phagocytes and defense chemicals to leave the bloodstream and go to the infected site as will be discussed later under this topic.
Exercise: Think-Pair-Share Questions
Compare and contrast how IgG, IgM, and IgE promote opsonization..
Because of a particular immunodeficiency disorder, a person is unable to produce C3 convertase. Which of the above antibody isotypes could still participate in opsonization? Briefly explain why.
How Bacteria Resist Attachment to Phagocytes
As we learned previously, some bacteria by means of the activities described below are able to resist phagocytic attachment :
• An outer membrane molecule of Neisseria gonorrhoeae called Protein II and the M-protein of Streptococcus pyogenes allow these bacteria to be more resistant to phagocytic engulfment. The M-protein of S. pyogenes, for example, binds factor H of the complement pathway and this leads to the degradation of the opsonin C3b by factor I and the formation of C3 convertase.
• Some capsules simply cover the C3b that does bind to the bacterial surface and prevent the C3b receptor on phagocytes from making contact with the C3b (Figure \(11\)). This is seen with the capsule of Streptococcus pneumoniae.
• Capsules can also resist unenhanced attachment by preventing the glycoprotein receptors on phagocytes from recognizing the bacterial cell wall components and mannose-containing carbohydrates.
• S. pneumonia activates the classical complement pathway, but resists C3b opsonization, and complement causes further inflammation in the lungs.
• Neisseria meningitidis has a capsule composed of sialic acid while Streptococcus pyogenes (group A beta streptococci) has a capsule made of hyaluronic acid. Both of these polysaccharides closely resemble carbohydrates found in human tissue polysaccharides and because they are not recognized as foreign by the lymphocytes that carry out the immune responses, antibodies are not made against these capsules.
• Some bacteria are able to coat themselves with host proteins such as fibronectin, lactoferrin, or transferrin. This prevents antibody molecules from binding to epitopes on the bacterial surface.
• Staphylococcus aureus produces protein A while Streptococcus pyogenes produces protein G. Both of these proteins bind to the Fc portion of antibodies, the portion that normally binds to receptors on phagocytes (Figure \(12\)). In this way the bacteria become coated with antibodies in a way that does not result in opsonization (Figure \(13\)).
• Streptococcus pyogenes produces Mac proteins that are able to bind to the receptors on phagocytes to which IgG and C3b normally attach (Figure \(14\).and Figure \(15\)). This blocks opsonization.
• Pathogenic Yersinia, such as the one that causes plague, contact phagocytes and, by means of a type III secretion system, deliver proteins which depolymerize the actin microfilaments needed for phagocytic engulfment into the phagocytes. Another Yersinia protein degrades C3b and C5a.
Summary
Opsonization, or enhanced attachment, refers to the antibody molecules IgG and IgE, the complement proteins C3b and C4b, and other opsonins attaching antigens to phagocytes. The Fab portions of the antibody IgG react with epitopes of the antigen. The Fc portion of IgG can then bind to neutrophils and macrophages thus sticking the antigen to the phagocyte. The Fc portion of secretory IgA can also bind to macrophages and neutrophils for opsonization. IgG and IgM can activate the classical complement pathway and C3b or C4b can stick the antigen to phagocytes. IgE is made against parasitic worms (helminths) and arthropods. The Fab portions of IgE bind to epitopes on the helminth or arthropod while the Fc portion binds to receptors on eosinophils enabling opsonization.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Discuss how antibodies defend the body by way of opsonization. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.) (ans)
2. We know Streptococcus pneumoniae is encapsulated and capsules resist phagocytosis. Yet the body is eventually able to phagocytose this organism. Describe how. (ans)
3. Staphylococcus aureus produces an exotoxin called Protein A. Protein A is able to react with the Fc portion of IgG. In terms of humoral immunity, discuss how Protein A may help the Staphylococcus resist phagocytosis. (ans)
4. The M-protein of Streptococcus pyogenes binds factor H of the complement pathway and allows these bacteria to be more resistant to phagocytic engulfment. Explain how. (ans)
5. Multiple Choice (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.2%3A_Ways_That_Antibodies_Help_to_Defend_the_Body/13.2A%3A_Opsonization.txt |
Learning Objectives
1. Discuss how antibodies defend the body by way of MAC cytolysis. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.)
2. State specifically how MAC cytolysis protects against the following:
1. Gram-negative bacteria
2. human cells recognized as nonself
3. enveloped viruses
3. Describe one way Gram-negative bacteria may resist cytolysis.
The process starts with the antibody isotypes IgG or IgM being made against epitopes on membranes. The Fab portion of IgG or IgM reacts with the epitopes on the membrane and the Fc portion of the antibody then activates the classical complement pathway. C5b6789n (the membrane attack complex or MAC) then puts holes in the membrane. (Remember that MAC is also produced during the alternative complement pathway and the lectin pathway as was discussed in Unit 5.)
a. In the case of bacteria, MAC can put holes in the outer membrane and possibly the cytoplasmic membrane of the Gram-negative cell wall (Figure \(1\) left) causing lysis ( Figure \(1\) right).
b. With enveloped viruses, the MAC can damage the viral envelope (Figure \(12\).3.2).
c. In the case of human cells recognized as nonself- virus-infected cells, transplanted cells, transfused cells, cancer cells - the MAC causes direct cell lysis (see Figure \(5\) and Figure \(6\)).
Concept Map for Ways in which Antibodies Protect the Body
However, as learned in Unit 3, some bacteria by means of the activities described below are more resistant to MAC lysis.
• The LPS of the cell wall is the principle target for complement in Gram-negative bacteria by activating the alternative complement pathway and serving as a binding site for C3b as well as the site for formation of MAC. Some Gram-negative bacteria attach sialic acid to the LPS O antigen (see Figure \(7\)) and this prevents the formation of the complement enzyme C3 convertase that is needed for the eventual formation of all the beneficial complement proteins such as C3b, C5a, and MAC. Blood-invasive strains of Neisseria gonorrhoeae , as well as Bordetella pertussis and Hemophilus influenzae are examples of Gram-negative bacteria that are able to alter their LPS in this manner.
• Some Gram-negative bacteria, such as Salmonella , lengthen the LPS O antigen side chain (see Figure \(7\)) and this prevents the formation of MAC. Neisseria meningitidis and Group B Streptococcus , on the other hand, produces capsular polysaccharides composed of sialic acid and as mentioned above, sialic acid prevents MAC lysis.
• An outer membrane molecule of Neisseria gonorrhoeae called Protein II binds factor H of the complement pathway and this leads to the degradation of the opsonin C3b by factor I and the formation of C3 convertase. Without C3 convertase, no MAC is produced.
Summary
The Fab portion of IgG or IgM reacts with the epitopes on the membrane and the Fc portion of the antibody then activates the classical complement pathway. C5b6789n (the membrane attack complex or MAC) then puts holes in the membrane. In the case of bacteria, MAC can put holes in the outer membrane and possibly the cytoplasmic membrane of the Gram-negative cell wall causing lysis. In the case of enveloped viruses, MAC can damage the viral envelope. In the case of human cells recognized as nonself - virus-infected cells, transplanted cells, transfused cells, cancer cells- the MAC causes direct cell lysis.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Discuss how antibodies defend the body by way of MAC cytolysis. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.) (ans)
2. Some Gram-negative bacteria attach sialic acid to the LPS O antigen of their outer membrane. Briefly describe how this may protect that Gram-negative bacterium from MAC cytolysis. (ans)
3. How does MAC affect viruses? (ans)
4. Multiple Choice (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.2%3A_Ways_That_Antibodies_Help_to_Defend_the_Body/13.2B%3A_Cytolysis_by_the_Membrane_Attack_Complex_%28MAC%29.txt |
Learning Objectives
1. Discuss how antibodies defend the body by way of ADCC by Natural Killer cells. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.)
Natural killer (NK) cells are capable of antibody-dependent cellular cytotoxicity or ADCC. NK cells have receptors on their surface for the Fc portion of certain subclasses of IgG. When the antibody IgG is made against epitopes on "foreign" membrane-bound cells, such as virus-infected cells and cancer cells, the Fab portions of the antibodies react with the "foreign" cell. The NK cells then bind to the Fc portion of the antibody (Figure \(1\)).
The NK cell then releases pore-forming proteins called perforins, proteolytic enzymes called granzymes, and chemokines. Granzymes pass through the pores and activate the enzymes that lead to apoptosis of the infected cell by means of destruction of its structural cytoskeleton proteins and by chromosomal degradation (Figure \(13\).5.1; right panel and Figure \(2\)). As a result, the cell breaks into fragments that are subsequently removed by phagocytes. Perforins can also sometimes result in cell lysis. (When NK cells are carrying out ADCC, they are sometimes also referred to as killer cells.)
Summary
NK cells are capable of antibody-dependent cellular cytotoxicity or ADCC. When IgG is made against epitopes on "foreign" membrane-bound cells, such as virus-infected cells and cancer cells, the Fab portions of the antibodies react with epitopes on the "foreign" cell and then NK cells bind to the Fc portion of the antibody. The NK cell then releases pore-forming proteins called perforins and proteolytic enzymes called granzymes. Granzymes pass through the pores and activate the enzymes that lead to apoptosis of the infected cell and the cell breaks into fragments that are subsequently removed by phagocytes.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Discuss how antibodies defend the body by way of ADCC by NK cells. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.)
2. Antibody-dependent cellular cytotoxicity (ADCC) is a result of:
1. Antibodies sticking infected cells and cancer cells to phagocytes.
2. Antibodies sticking infected cells and cancer cells to cytotoxic T-lymphocytes (CTLs).
3. Antibodies sticking infected cells and cancer cells to NK cells.
4. MAC lysing the membranes of infected cells and cancer cells.
3. During ADCC, the Fab portion of the antibody _____________while the Fc portion _______________.
1. binds to epitopes of an antigen; activates the complement pathway.
2. activates the complement pathway; binds to epitopes of an antigen.
3. binds to epitopes of an antigen; binds to cytotoxic T-lymphocytes.
4. binds to epitopes of an antigen; binds to NK cells.
4. NK cells kill the cells they bind to by:
1. Triggering apoptosis.
2. Dumping the contents of their lysosomes on the cell.
3. Producing cytolytic exotoxins that lyse the cell.
4. Inducing extracellular killing by eosinophils. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.2%3A_Ways_That_Antibodies_Help_to_Defend_the_Body/13.2C%3A_Antibody-dependent_Cellular_Cytotoxicity_%28ADCC%29_by_Natural_Killer_Cells.txt |
Learning Objectives
• Discuss how antibodies defend the body by way of neutralizing exotoxins. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.)
• Describe how the ability of bacteria to sense their own population density, communicate with each other by way of secreted factors (cell-to-cell signaling), and behave as a population rather than as individual bacteria most likely plays an important role in pathogenicity for many bacteria.
For an exotoxin to cause harm it must first bind to receptors on a susceptible host cell. Antitoxin antibodies are made against microbial exotoxins. The Fab portion binds to the exotoxin molecules before they can interact with host target cells and thus neutralizes the toxin (Figure \(1\)). IgG neutralizes toxins in tissues while IgA neutralizes toxins at mucosal surfaces within the body.
However, as learned in Unit 2, many Gram-negative and Gram-positive are able to sense their own population density, communicate with each other by way of secreted factors, and behave as a population rather than as individual bacteria. This is referred to as quorum sensing and most likely plays an important role in pathogenicity for many bacteria.
• For example, Pseudomonas aeruginosa causes severe nosocomial infections, chronic infections in people with cystic fibrosis, and potentially fatal infections in those who are immunocompromised. Its virulence depends on the secretion of a variety of harmful exotoxins and enzymes. If there was an isolated production of these virulence toxins and enzymes by a small number of Pseudomonas, the body's immune responses would most likely be able effectively neutralize these harmful agents with antibodies. However, through a coordination of the expression of the genes coding for these toxins and enzymes by the entire population of bacteria, P. aeruginosa appears to only secrete these extracellular virulence factors when the density of bacteria is large enough that they can be produced at high enough levels to overcome body defenses.
Summary
For an exotoxin to cause harm it must first bind to receptors on a susceptible host cell. Antitoxin antibodies are made against microbial exotoxins. The Fab portion binds to the exotoxin molecules before they can interact with host target cells and thus neutralizes the toxin.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Discuss how antibodies defend the body by way of neutralizing exotoxins. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.) (ans)
2. Describe how the ability of Pseudomonas aeruginosa to sense its own population density, communicate with other Pseudomonas by way of secreted factors (cell-to-cell signaling), and behave as a population rather than as individual bacteria most likely plays an important role in pathogenicity of this organism. (ans)
3. Multiple Choice (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.2%3A_Ways_That_Antibodies_Help_to_Defend_the_Body/13.2D%3A_Neutralization_of_Exotoxins.txt |
Discuss how antibodies defend the body by way of neutralizing viruses. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.) Briefly describe 2 different ways viruses may resist neutralizing antibodies.
However, as learned in Unit 4, some viruses by means of the activities described below are able to overcome this antibody defense.
• The influenza viruses undergo what is called antigenic drift and antigenic shift. With antigenic drift, mutations cause a gradual change in the hemagglutinin antigen that adsorbs to receptors on host cells. Antigenic shift is caused by a human influenza virus acquiring a new genome segment from an influenza virus capable of infecting other animals such as a ducks or swine. This new genome segment causes a major change in the hemagglutinin antigen. Antibodies made against the original human influenza virus can no longer bind to the new strain of virus or stick the virus to phagocytes.
• Likewise HIV, because of its high rate of mutation and its intracellular recombination with other strains of HIV, as mentioned earlier in this unit, produces altered gp120 to which antibodies made against the earlier strains of HIV can no longer bind.
• The hepatitis C virus (HCV) frequently, through mutation, produces viral variants ("escape mutants") to resist antibodies.
Summary
In order for viruses to infect a cell and replicate, they must first adsorb to receptors on the host cell's plasma membrane. Antibodies are made against viral capsids or envelope glycoproteins where the Fab portion binds to and covers the viral attachment molecules. This prevents viral adsorption to host cells. Neutralizing antibodies are especially important in preventing viral reinfection.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Discuss how antibodies defend the body by way of neutralizing viruses. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.) (ans)
2. Describe one way a virus can resist virus-neutralizing antibodies and give an example. (ans)
3. Multiple Choice (ans)
13.2F: Preventing Bacterial Adherence
Learning Objectives
1. Discuss how antibodies defend the body by way of preventing bacterial adherence to host cells. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.)
2. Briefly describe 2 different ways bacteria may resist antibodies that block bacterial adherence to host cells.
One of the body's innate defenses is the ability to physically remove bacteria from the body through such means as the constant shedding of surface epithelial cells from the skin and mucous membranes, the removal of bacteria by such means as coughing, sneezing, vomiting, and diarrhea, and bacterial removal by bodily fluids such as saliva, blood, mucous, and urine. Bacteria may resist this physical removal producing pili, cell wall adhesin proteins, and/or biofilm-producing capsules.
Antibodies are made against pili, capsules, and cell wall adhesins. The binding of the Fab portion of the antibody to the adhesive tip of the pili, the cell wall adhesins, or the capsular molecules prevents the bacteria from adhering to and colonizing host cells (see Figure \(1\) and Figure \(2\).) IgG blocks adherence of bacteria in tissues while IgA blocks adherence of bacteria at mucosal surfaces within the body.
The body is able to make antibody molecules against the adhesive tips of Escherichia coli pili and yet E. coli is still the most common cause of urinary tract infections. State what might account for this.
However, as learned in Unit 3, some bacteria by means of the activities described below are able to overcome this antibody defense.
• Some bacteria can produce immunoglobulin proteases which can degrade the protective IgA found in mucus. Examples include bacteria that colonize the mucous membranes such as Streptococcus pneumoniae, Hemophilus influenzae, Neisseria gonorrhoeae, Neisseria meningitidis, Helicobacter pylori, Shigella flexneri and enteropathogenic Escherichia coli.
• Another way certain bacteria can evade antibodies is by changing the adhesive tips of their pili as seen with Neisseria gonorrhoeae (see Figure \(3\)). Bacteria can also vary other surface proteins so that antibodies already made will no longer "fit."
Summary
Bacteria resist physical removal by means of pili, cell wall adhesin proteins, and/or biofilm-producing capsules. The binding of the Fab portion of the antibody to the adhesive tip of the pili, the cell wall adhesins, or the capsular molecules prevents the bacteria from adhering to and colonizing host cells.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Discuss how antibodies defend the body by way of preventing bacterial adherence to host cells. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.) (ans)
2. Describe how immunoglobulin proteases may protect bacteria from antibodies that block bacterial adhence to host cells. (ans)
3. Multiple Choice (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.2%3A_Ways_That_Antibodies_Help_to_Defend_the_Body/13.2E%3A_Neutralization_of_Viruses.txt |
Learning Objectives
1. Discuss the how antibodies defend the body by agglutinating microorganisms. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role - if any - of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.)
Agglutination is mainly a function of antibodies with multiple reactive Fab sites such as IgM and IgA. The Fab portion of the antibodies links microorganisms together (causes them to agglutinate) so they can be phagocytosed more effectively (see Figure \(1\)).
Summary
Agglutination is mainly a function of antibodies with multiple reactive Fab sites such as IgM and IgA. The Fab portion of the antibodies links microorganisms together (causes them to agglutinate) so they can be phagocytosed more effectively.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Discuss how antibodies defend the body by agglutinating microorganisms. (Include what classes or isotypes of immunoglobulins are involved, the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.) (ans)
2. State why IgM and IgA are good at causing agglutination of microorganisms. (ans)
13.2H: Immobilization of Bacteria and Protozoans
Learning Objectives
1. Discuss how antibodies defend the body by immobilizing bacteria and protozoans. (Include the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.)
Flagella and cilia are organelles of locomotion and enable motile microorganisms to move towards or away from environmental molecules through a process called taxis. The mucosal surfaces of the bladder and the intestines constantly flush bacteria away in order to prevent colonization.Motile bacteria that can swim chemotactically toward mucosal surfaces may have a better chance to make contact with the mucous membranes, attach, and colonize.
Antibodies are made against the flagella of motile bacteria or the flagella or cilia of motile protozoans. The Fab portions of the antibodies bind to these locomotor organelles and arrest the organism's movement blocking its ability to spread.
Summary
1. Flagella and cilia are organelles of locomotion and enable motile microorganisms to move towards or away from environmental molecules through a process called taxis.
2. Antibodies are made against the flagella of motile bacteria or the flagella or cilia of motile protozoans.
3. The Fab portions of the antibodies bind to these locomotor organelles and arrest the organism's movement blocking its ability to spread.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Discusshow antibodies defend the body by immobilizing bacteria and protozoans. (Include the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.) (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.2%3A_Ways_That_Antibodies_Help_to_Defend_the_Body/13.2G%3A_Agglutination_of_Microorganisms.txt |
Learning Objectives
1. Describe two different ways antibodies defend the body by promoting an inflammatory response and state the importance of inflammation. (Include the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.)
Antigen-antibody reactions can also promote an inflammatory response:
a. IgG and IgM can activate the classical complement pathway (see Figure \(1\)).
As learned under innate immunity in Unit 5, the classical complement pathway is primarily activated when a complement protein complex called C1 interacts with the Fc portion of the antibody molecules IgG or IgM after they have bound to their specific antigen via their Fab portion . C1 is also able to directly bind to the surfaces of some pathogens.
The C1 complex is composed of three complement proteins called C1q, C1r, and C1s.
• The C1q is the portion of the C1 complex that binds to the antibodies or the microbe.
• The binding of C1q activates the C1r portion of C1 which, in turn, activates C1s. This activation gives C1s enzymatic activity to cleave complement protein C4 into C4a and C4b and C2 into C2a and C2b and begin the classical complement pathway.
The beneficial results of the activated complement proteins include :
1. Triggering inflammation : C5a>C3a>C4a.
2. Chemotactically attracting phagocytes to the infection site: C5a;
3. Promoting the attachment of antigens to phagocytes via enhanced attachment or opsonization : C3b>C4b (discussed earlier under opsonization);
4. Causing the lysis of Gram-negative bacteria, viral envelopes, and human cells displaying foreign epitopes (discussed earlier under MAC cytolysis).
b. IgA can activate the lectin complement pathway and the alternative complement pathway and C5a, C3a, and C4a can trigger inflammation.
c. IgE can bind to mast cells and basophils and trigger the release of inflammatory mediators.
The Fc portion of IgE can bind to receptors on mast cells and basophils . Cross linking of the cell-bound IgE by antigen triggers the release of vasodilators and other inflammatory mediators for an inflammatory response (see Fig 2).
As learned under inflammation in Unit 5, most of the body defense elements are located in the blood and inflammation is the means by which body defense cells and defense chemicals leave the blood and enter the tissue around the injured or infected site.
The inflammatory response produces vasodilators that increase capillary permeability. As a result of this increased permeability:
a. Plasma flows out of the blood into the tissue.
Beneficial molecules in the plasma include:
1. Clotting factors. Tissue damage activates the coagulation cascade causing fibrin clots to form to localize the infection, stop the bleeding, and chemotactically attract phagocytes.
2. Antibodies . These help remove or block the action of microbes through a variety of methods described in this section.
3. Proteins of the complement pathways . These, in turn: 1) stimulate more inflammation (C5a, C3a, and C4a), 2) stick microorganisms to phagocytes (C3b and C4b), 3) chemotactically attract phagocytes ( C5a), and 4) lyse membrane-bound cells displaying foreign antigens (membrane attack complex or MAC).
4. Nutrients. These feed the cells of the inflamed tissue.
5. Lysozyme, cathelicidins, phospholipase A2, and human defensins . Lysozyme degrades peptidoglycan. Cathelicidins are cleaved into two peptides that are directly toxic to microbes and can neutralize LPS from the gram-negative bacterial cell wall. Phospholipase A2 hydrolyzes the phospholipids in the bacterial cytoplasmic membrane. Human defensins put pores in the cytoplasmic membranes of many bacteria. Defensins also activate cells involved in the inflammatory response.
6. Transferrin . Transferrin deprives microbes of needed iron.
b. Leukocytes enter the tissue through a process called diapedesis or extravasation.
Benefits of diapedesis include:
1. Increased phagocytosis. Phagocytes such as neutrophils, monocytes that differentiate into macrophages when they enter the tissue, and eosinophils are phagocytic leukocytes.
2. More vasodilation. Basophils, eosinophils, neutrophils, and platelets enter the tissue and release or stimulate the production of vasoactive agents that promote inflammation.
3. Cytotoxic T-lymphocytes (CTLs) , effector T4-cells , and NK cells enter the tissue to kill cells such as infected cells and cancer cells that are displaying foreign antigens on their surface (discussed in Unit 6).
Summary
1. IgG and IgM can activate the classical complement pathway and C5a, C3a, and C4a can trigger inflammation.
2. IgA can activate the lectin complement pathway and the alternative complement pathway and C5a, C3a, and C4a can trigger inflammation.
3. IgE can bind to mast cells and basophils and trigger the release of inflammatory mediators.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe one way antibodies defend the body by promoting an inflammatory response and state the importance of inflammation. (Include the role of the Fab portion of the antibody, the role, if any, of the Fc portion of the antibody, and the role of any complement proteins, if any, involved.) (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.2%3A_Ways_That_Antibodies_Help_to_Defend_the_Body/13.2I%3A_Promoting_an_Inflammatory_Response.txt |
Immunity may be passive or active. During passive immunity, antibodies made in another person or animal enter the body and the immunity is short-lived and Active Immunity: In the case of active immunity, antigens enter the body and the body responds by making its own antibodies and B-memory cells. In this case, immunity is longer lived although duration depends on the persistence of the antigen and the memory cells in the body. Both passive and active immunity can be either naturally or artificially acquired.
• 13.3A: Naturally Acquired Immunity
Active naturally acquired immunity refers to the natural exposure to an infectious agent or other antigen by the body. The body responds by making its own antibodies. There are two examples of passive naturally acquired immunity: The placental transfer of IgG from mother to fetus during pregnancy that generally lasts 4 to 6 months after birth; and The IgA and IgG found in human colostrum and milk of babies who are nursed.
• 13.3B: Artificially Acquired Immunity
Active artificially acquired immunity refers to any immunization with an antigen. During artificially acquired active immunity, one is immunized with one or more of the following: attenuated microbes, killed organisms, fragmented microorganisms, or antigens produced by recombinant DNA technology, or toxoids. Passive artificially acquired immunity refers to the injection of antibody-containing serum, or immune globulin (IG), from another person or animal.
13.3: Naturally and Artificially Acquired Active and Passive Immunity
Learning Objectives
1. Give an example of naturally acquired active immunity.
2. Give two examples of naturally acquired passive immunity and state why this is important to newborns and infants.
Active Naturally Acquired Immunity
Active naturally acquired immunity refers to the natural exposure to an infectious agent or other antigen by the body. The body responds by making its own antibodies.
Passive Naturally Acquired Immunity
There are two examples of passive naturally acquired immunity: (1) The placental transfer of IgG from mother to fetus during pregnancy. These antibodies generally last 4 to 6 months following birth. The immune responses reach full strength at about age 5. (2) The IgA and IgG found in human colostrum and milk of babies who are nursed. In addition to the IgA and IgG, human milk also contains:
• Oligosaccharides and mucins that adhere to bacteria and viruses to interfere with their attachment to host cells;
• Lactoferrin to bind iron and make it unavailable to most bacteria;
• B12 binding protein to deprive bacteria of needed vitamin B12;
• Bifidus factor that promotes the growth of Lactobacillus bifidus, normal flora in the gastrointestinal tract of infants that crowds out harmful bacteria;
• Fibronectin that increases the antimicrobial activity of macrophages and helps repair tissue damage from infection in the gastrointestinal tract;
• Gamma-interferon, a cytokine that enhances the activity of certain immune cells;
• Hormones and growth factors that stimulate the baby's gastrointestinal tract to mature faster and be less susceptible to infection;
• Lysozyme to break down peptidoglycan in bacterial cell walls.
Benefits of Breast Feeding
According to the Centers for Disease Control and Prevention (CDC), breast-fed infants have a lower incidence of gastrointestinal infections, ear infections, atopic dermatitis, respiratory infections, urinary tract infections, meningitis, type 2 diabetes, and sudden infant death syndrome. Benefits to the mother include a decreased risk of breast cancer, ovarian cancer, and type 2 diabetes, as well stopping post-birth bleeding and temporarily suppressing ovulation. It may also be associated with a reduced risk of pediatric overweight.
Summary
Active naturally acquired immunity refers to the natural exposure to an infectious agent or other antigen by the body. The body responds by making its own antibodies. There are two examples of passive naturally acquired immunity: The placental transfer of IgG from mother to fetus during pregnancy that generally lasts 4 to 6 months after birth; and The IgA and IgG found in human colostrum and milk of babies who are nursed. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.3%3A_Naturally_and_Artificially_Acquired_Active_and_Passive_Immunity/13.3A%3A_Naturally_Acquired_Immunity.txt |
Active Artificially Acquired Immunity
Active artificially acquired immunity refers to any immunization with an antigen. By giving a safe form of the antigen artificially, the body will produce its own antibodies and, more importantly, develop circulating, long-lived B-memory cells with high affinity B-cell receptors on their surface. If at a later date the body is again exposed to that same antigen, the memory cells will cause immediate and rapid production of the appropriate antibodies for protection. With artificially acquired active immunity, one is immunized with one or more of the following:
Attenuated microbes
Attenuated microbes are living, non-virulent strains of a microbe. Viruses are attenuated by growing them in non-human cells until they mutate and adapt to the non-human host. In the process, they lose virulence for humans. Viruses can also be attenuated using recombinant DNA techniques to either mutate or delete virulence genes in the viral genome.
Attenuated viral vaccines tend to be immunologically quite effective since the viruses can multiply slowly in the body, thus increasing the amount and persistence of the antigen for a greater antibody response. In addition, attenuated viruses enter the cytosol of cells and peptides from viral antigens can be presented by MHC-I molecules to activate naive T8-lymphocytes and stimulate the production of cytotoxic T-lymphocytes (CTLs). Living attenuated microbes can, however, sometimes be potentially dangerous to highly immunosuppressed individuals in whom they may cause opportunistic infections.
Examples of vaccines that contain attenuated microbes include:
• The MMR vaccine containing attenuated measles, mumps, and rubella viruses;
• The MMRV vaccine containing attenuated measles, mumps, rubella viruses and varicella zoster (chickenpox) viruses;
• The TOPV or trivalent oral polio vaccine containing attenuated poliomyelitis viruses types 1, 2, and 3;
• The yellow fever vaccine containing attenuated yellow fever viruses;
• The Var or varicella zoster virus vaccine containing attenuated varicella zoster viruses.
The body responds by producing antibodies that block viral adsorption to host cells.
Killed organisms, fragmented microorganisms, or antigens produced by recombinant DNA technology
Examples of vaccines containing killed or inactivated microbes include:
• The IPV or inactivated poliomyelitis vaccine containing inactivated poliomyelitis viruses types 1, 2, and 3;
• The rabies vaccines containing whole, killed rabies viruses;
• The influenza vaccines consist of inactivated influenza viruses, either whole or broken down;
• The hepatitis A vaccine containing inactivated hepatitis A virus;
• RV1, an attenuated strain of a human rotavirus. Rotaviruses are the most common cause of gastroenteritis in children.
Examples of vaccines containing fragments of microorganisms include the immunizations for:
• Meningococcal meningitis; contains capsular polysaccharide from 4 strains of Neisseria meningitidis;
• Pneumococcal pneumonia; PCV13 containing capsular material from the 13 most serious strains of Streptococcus pneumoniae in children conjugated to diphtheria toxoid protein; PCV 23 containing capsular material from the 23 most serious strains of S. pneumoniae in adults conjugated to diphtheria toxoid protein;
• Hemophilus influenzae type b containing capsular polysaccharide from H. influenzae type B conjugated to protein (either diphtheria toxoid or an outer membrane protein from Neisseria meningitidis).
These vaccines contain polysaccharide capsular material from the bacteria, usually conjugated to protein for greater immunogenicity. The body responds by producing opsonizing antibodies against the capsule.
While the B-cell receptors of B-lymphocytes can recognize epitopes on polysaccharides, T4-lymphocytes can only recognize peptide epitopes bound to MHC-II molecules. The protein conjugate added to the polysaccharide in the vaccine is degraded into peptides and bound to MHC-II molecules by APCs. They then present the peptide to the TCRs on T4-lymphocytes for their activation. In this way the cytokines produced by the activated T4-lymphocytes become available for use by the B-lymphocytes sensitized to the polysaccharide component of the vaccine.
c. Examples of vaccines produced by recombinant DNA technology include:
• The hepatitis B vaccine, the first human vaccine produced by recombinant DNA technology, contains hepatitis B virus surface antigen (HBsAG);
• The acellular pertussis part of the diphtheria, tetanus, and acellular pertussis vaccine (DTaP) containing diphtheria toxoid, tetanus toxoid, and antigens from the whooping cough bacterium Bordetella pertussis (Acellular pertussis vaccines contain inactivated pertussis toxin (PT) and may contain one or more other bacterial components (e.g., filamentous hemagglutinin [FHA], an outer-membrane protein; pertactin [Pn], and fimbriae [Fim] types 2 and 3);
• The vaccine against Lyme disease;
• Gardasil, a vaccine against human papilloma virus (HPV) types 6, 11 that cause about 90% of genital warts, and types 16, and 18 responsible for around 70% of cervical cancer in the US; and Cervarix, a vaccine against HPV types 16 and 18. Both contain recombinant L1 capsid protein from the different strains of HPV;
• RV5, an oral vaccine against human rotavirus gastroenteritis. Capsid proteins from human rotaviruses have been expressed on the surface of harmless non-human rotavirus strains.
Toxoid
A toxoid is an exotoxin treated so as to be non-poisonous but still immunogenic. Examples of vaccines containing toxoids include the diphtheria and tetanus components of the DTaP and Td vaccines. The body responds by making antibodies capable of neutralizing the exotoxin. The antigen may be adsorbed to an adjuvant, a substance such as aluminum hydroxide or aluminum phosphate that is not immunogenic but enhances the immunogenicity of antigens.
Routine immunization practices protect more than just the individuals receiving the vaccine. When a critical portion of a community becomes immunized against a particular infectious disease, most members of the community - including those who were not immunized - are protected against that disease because there is little opportunity for an outbreak. This is known as herd immunity or community immunity.
Passive Artificially Acquired Immunity
Passive artificially acquired immunity refers to the injection of antibody-containing serum, or immune globulin (IG), from another person or animal. Since the body is not making its own antibodies and memory cells are not produced, passive artificially acquired immunity is short lived and offers only mediate, short term protection. Also, the injection of serum during passive immunization carries a greater risk of allergic reactions than the injection of antigens during active immunization. These allergic reactions are referred to as serum sickness and will be discussed later under hypersensitivities.
Examples include:
• The use of pooled adult human immune globulin (IG) to prevent hepatitis A and measles and to prevent infections in people with certain immunodeficiency diseases;
• Human HBIG to prevent hepatitis B in those not actively immunized with the HepB vaccine;
• Human TIG to prevent tetanus in those not actively immunized with the DTP, DTaP, or Td vaccines;
• RhoGAM to prevent Rh hemolytic disease of newborns;
• VZIG to prevent varicella;
• CMV-IGIV to prevent cytomegalovirus infections in highly immunosuppressed individuals;
• RIG to prevent rabies, given concurrently with active immunization with the rabies vaccine;
• The antisera used for botulism; and
• IVIG (intravenous immune globulin), now being used to reduce infections in people with certain immunosuppressive diseases such as primary immunodeficiency syndrome and chronic lymphocytic leukemia as well as to treat certain autoimmune diseases such as immune thrombocytopenia purpura (ITP) and Kawasaki disease.
Tetanus provides a nice example of how active immunization (DTaP) and passive immunization (TIG) may be used in preventing a disease (Table \(13\).3B.1:).
Table \(13\).3B.1: Tetanus prophylaxis in Routine Wound Management
History of tetanus
toxoid doses
Clean, minor wound All other wounds (1)
Td (2) TIG (3) Td TIG
Unknown or < 3 Yes No Yes Yes
Three or more No (4) No No (5) No
(1) Such as, but not limited to, wounds contaminated with dirt, feces, soil, saliva, etc.: puncture wounds, avulsions, and wounds resulting from missles, crushing, burns, and frostbite.
(2) Tetanus toxoid, diphtheria toxoid (active immunization).
(3) Tetanus Immune Globulin (passive immunization).
(4) Yes, if more than 10 years since last dose.
(5) Yes, if more than 5 years since last dose. (More frequent boosters are not needed and can accentuate side effects.)
There is also some early evidence that immunization may be of value in the treatment of some infections as well as in their prevention, possibly by supercharging the immune system of those already infected. Vaccine therapies in various stages of testing include those against diseases such as herpes, leprosy, tuberculosis, and hepatitis B.
13.E: Humoral Immunity (Exercises)
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied.
13.1: Antibodies (Immunoglobulins)
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Define antibody. (ans)
2. In terms of infectious disease, state what humoral immunity is most effective against. (ans)
13.2: Ways That Antibodies Help to Defend the Body
• List 9 ways that antibodies help to defend the body.
13.3: Naturally and Artificially Acquired Active and Passive Immunity
1. Define the following:
1. active immunity
2. passive immunity
2. Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1.Matching
_____ Antibodies made in another person or animal enter the body and the immunity is short-lived. (ans)
_____ Antigens enter the body and the body responds by making its own antibodies and B-memory cells. (ans)
1. active immunity
2. passive immunity
13.3A: Naturally Acquired Immunity
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Give an example of naturally acquired active immunity. (ans)
2. Give two examples of naturally acquired passive immunity.
3. State why naturally acquired passive immunity is important to newborns and infants. (ans)
4. Multiple Choice (ans)
13.3B: Artificially Acquired Immunity
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Define and give an example of artifically acquired passive immunity. (ans)
2. Define and give an example of artifically acquired active immunity. (ans)
3. List 3 different forms of antigen that may be used for artificially acquired active immunity and state 2 common examples of each.
4. A patient with a deep puncture wound who has never received a DTP vaccinationis given both Td and TIG. Another patient with an identical wound and who had 4 DTP vaccinationsas a child and a Td booster 3 years ago is given nothing. Discuss the reasoning behind this. (hint: see Fig. 1) (ans)
5. Multiple Choice (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/13%3A_Humoral_Immunity/13.3%3A_Naturally_and_Artificially_Acquired_Active_and_Passive_Immunity/13.3B%3A_Artificially_Acquired_Immunity.txt |
Cell-mediated immunity (CMI) is an immune response that does not involve antibodies but rather involves the activation of macrophages and NK-cells, the production of antigen-specific cytotoxic T-lymphocytes, and the release of various cytokines in response to an antigen. Cellular immunity protects the body by:
• Activating antigen-specific cytotoxic T-lymphocytes (CTLs) that are able to lyse body cells displaying epitopes of foreign antigen on their surface, such as virus-infected cells, cells with intracellular bacteria, and cancer cells displaying tumor antigens;
• Activating macrophages and NK cells, enabling them to destroy intracellular pathogens; and
• Stimulating cells to secrete a variety of cytokines that influence the function of other cells involved in adaptive immune responses and innate immune responses.
Cell-mediated immunity is directed primarily microbes that survive in phagocytes and microbes that infect non-phagocytic cells. It is most effective in destroying virus-infected cells, intracellular bacteria, and cancers. It also plays a major role in delayed transplant rejection.
14: Cell-Mediated Immunity
Learning Objectives
1. Briefly compare humoral immunity with cell-mediated immunity.
2. Define cell-mediated immunity and state what it is most effective against.
3. State three different ways by which cell-mediated immunity protects the body.
4. Define gene translocation and relate it to each T-lymphocyte being able to produce T-cell receptor with a unique shape.
5. Define the following:
1. combinatorial diversity
2. junctional diversity
6. In terms of cell-mediated immunity, state what is meant by anamnestic response and discuss its role in immune defense.
7. Briefly describe why there is a heightened secondary response during anamestic response.
Cell-mediated immunity (CMI) is an immune response that does not involve antibodies but rather involves the activation of macrophages and NK-cells, the production of antigen-specific cytotoxic T-lymphocytes, and the release of various cytokines in response to an antigen . Cellular immunity protects the body by:
1. Activating antigen-specific cytotoxic T-lymphocytes (CTLs) that are able to destroy body cells displaying epitopes of foreign antigen on their surface, such as virus-infected cells, cells with intracellular bacteria, and cancer cells displaying tumor antigens;
2. Activating macrophages and NK cells, enabling them to destroy intracellular pathogens; and
3. Stimulating cells to secrete a variety of cytokines that influence the function of other cells involved in adaptive immune responses and innate immune responses.
Cell-mediated immunity is directed primarily microbes that survive in phagocytes and microbes that infect non-phagocytic cells. It is most effective in destroying virus-infected cells, intracellular bacteria, and cancers. It also plays a major role in delayed transplant rejection.
Generation of T-cell receptor (TCR) diversity through gene translocation
As mentioned earlier, the immune system of the body has no idea as to what antigens it may eventually encounter. Therefore, it has evolved a system that possesses the capability of responding to any conceivable antigen. The immune system can do this because both B-lymphocytes and T-lymphocytes have evolved a unique system of gene-splicing called gene translocation, a type of gene-shuffling process where various different genes along a chromosome move and join with other genes from the chromosome. To demonstrate this gene translocation process, we will look at how each T-lymphocyte becomes genetically programmed to produce a T-cell receptor (TCR) having a unique shape to fit a specific epitope.
In a manner similar to B-lymphocytes, T-lymphocytes are able to cut out and splice together different combinations of genes along their chromosomes. Through random gene translocation, any combination of the multiple forms of each gene can join together. This is known as combinatorial diversity. The T-cell receptors or TCRs (Figure \(1\)) of most T-lymphocytes involved in adaptive immunity consist of an alpha (a) and a beta (ß) chain. There are 70-80 different Va genes and 61 different Ja genes that code for the variable portion of the a chain of the TCR. Likewise, there are 52 Vß genes, 1 Dß1 gene, 1 Dß2 gene, and 6-7 Jß genes that can recombine to form the variable portion of the TCR.
During gene translocation, specialized enzymes in the T-lymphocyte cause splicing inaccuracies wherein additional nucleotides are added or deleted at the various gene junctions. This change in the nucleotide base sequence generates even greater diversity in Fab shape. This is called junctional diversity. Unlike the BCR, somatic hypermutation does not occur during the production of the TCRs. As a result of combinatorial diversity and junctional diversity, each T-lymphocyte is able to produce a unique shaped T-cell receptor (TCR) capable of reacting with complementary-shaped peptide bound to a MHC molecule.
Anamnestic Response (Memory)
As a result of T-lymphocytes recognizing epitopes of protein antigens during cell-mediated immunity, numerous circulating T8-memory cells and T4-memory cells develop which possess anamnestic response or memory. These T-memory cells persist for the remainder of a person’s life. Effector memory T-cells (TEM cells) circulate in the blood whereas tissue resident memory T-cells (TRM cells) are found within the epithelium of the skin and mucous membranes. CD8 TRM cells are typically activated by viral antigens and subsequently produce inflammatory cytokines that trigger an innate immune response for nonspecific antiviral activity. CD4 TRM cells are found in clusters surrounding macrophages in the mucosa. Unlike TEM cells, TRM cells do not circulate in the blood and are not replenished from the blood. They remain in peripheral tissues.
A subsequent exposure to that same antigen results in:
• A more rapid and longer production of cytotoxic T-lymphocytes (CTLs);
• A more rapid and longer production of T4-effector lymphocytes; and
• Triggering of nonspecific innate immune responses.
Clonal Selection and Clonal Expansion
As mentioned above, during early differentiation of naive T-lymphocytes in the thymus marrow, each T4-lymphocyte and each T8-lymphocyte becomes genetically programmed to make a T-cell receptor or TCR with a unique shape through a series of gene translocations, and molecules of that TCR are put on its surface of that T-lymphocyte to function as its epitope receptor. When an antigen encounters the immune system, epitopes from protein antigens bound to MHC-I or MHC-II molecules eventually will react with a naive T4- and T8-lymphocyte with TCRs and CD4 or CD8 molecules on its surface that more or less fit and this activates that T-lymphocyte. This process is known as clonal selection.
Cytokines produced by effector T4-helper lymphocytes enable the now activated T4- and T8-lymphocyte to rapidly proliferate to produce large clones of thousands of identical T4- and T8-lymphocytes. In this way, even though only a few T-lymphocytes in the body may have TCR molecule able to fit a particular epitope, eventually many thousands of cells are produced with the right specificity. This is referred to as clonal expansion. These cells then differentiate into effector T4-lymphocytes and cytotoxic T-lymphocytes or CTLs.
Cellular immunity is also the mechanism behind delayed hypersensitivity (discussed later in this unit). Delayed hypersensitivity is generally used to refer to the harmful effects of cell-mediated immunity (tissue and transplant rejections, contact dermatitis, positive skin tests like the PPD test for tuberculosis, granuloma formation during tuberculosis and deep mycoses, and destruction of virus-infected cells).
Summary
1. Cell-mediated immunity (CMI) is an immune response that does not involve antibodies but rather involves the activation of macrophages and NK-cells, the production of antigen-specific cytotoxic T-lymphocytes, and the release of various cytokines in response to an antigen.
2. Cell-mediated immunity is directed primarily microbes that survive in phagocytes and microbes that infect non-phagocytic cells. It is most effective in destroying virus-infected cells, intracellular bacteria, and cancers.
3. In a manner similar to B-lymphocytes, T-lymphocytes are able to randomly cut out and splice together different combinations of genes along their chromosomes through a process called gene translocation. This is known as combinatorial diversity and results in each T-lymphocyte generating a unique T-cell receptor (TCR).
4. During gene translocation, specialized enzymes in the T-lymphocyte cause splicing inaccuracies wherein additional nucleotides are added or deleted at the various gene junctions. This change in the nucleotide base sequence generates even greater diversity in the shape of the TCR. This is called junctional diversity.
5. As a result of combinatorial diversity and junctional diversity, each T-lymphocyte is able to produce a unique shaped T-cell receptor (TCR) capable of reacting with complementary-shaped peptide bound to a MHC molecule.
6. As a result of T-lymphocytes recognizing epitopes of protein antigens during cell-mediated immunity, numerous circulating T8-memory cells and T4-memory cells) develop which possess anamnestic response or memory.
7. A subsequent exposure to that same antigen results in a more rapid and longer production of cytotoxic T-lymphocytes (CTLs), and a more rapid and longer production of T4-effector lymphocytes.
8. When an antigen encounters the immune system, epitopes from protein antigens bound to MHC-I or MHC-II molecules eventually will react with a naive T4- and T8-lymphocyte with TCRs and CD4 or CD8 molecules on its surface that more or less fit and this activates that T-lymphocyte. This process is known as clonal selection.
9. Cytokines produced by effector T4-helper lymphocytes enable the now activated T4- and T8-lymphocyte to rapidly proliferate to produce large clones of thousands of identical T4- and T8-lymphocytes. This is referred to as clonal expansion. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/14%3A_Cell-Mediated_Immunity/14.1%3A_Cell-Mediated_Immunity_-_An_Overview.txt |
Learning Objectives
1. In terms of the role of cytotoxic T-lymphocytes (CTLs) in body defense:
1. State from what cells cytotoxic T-lymphocytes are derived.
2. Describe how they can react with and destroy virus-infected cells, cells containing intracellular bacteria, and cancer cells without harming normal cells. (Indicate the role of following: TCR, CD4, MHC-I, and peptides from endogenous antigens.)
3. State the mechanism by which cytotoxic T-lymphocytes kill the cells to which they bind. (Indicate the role of the following: perforins, granzymes, caspases, and macrophages in the process.)
2. Briefly describe two ways certain viruses may evade cell-mediated immunity.
Marking an Infected Cell or a Tumor Cell for Destruction by Cytotoxic T-Lymphocytes
One of the body's major defenses against viruses, intracellular bacteria, and cancers is the destruction of infected cells and tumor cells by cytotoxic T-lymphocytes (CTLs). These CTLs are effector cells derived from naive T8-lymphocytes during cell-mediated immunity. Both T8-lymphocytes and CTLs produce T-cell receptors or TCRs and CD8 molecules that are anchored to their surface.
1. The TCRs and CD8 molecules on the surface of naive T8-lymphocytes are designed to recognize peptide epitopes bound to MHC-I molecules on antigen-presenting cells or APCs .
2. The TCRs and CD8 molecules on the surface of cytotoxic T-lymphocytes (CTLs) are designed to recognize peptide epitopes bound to MHC-I molecules on infected cells and tumor cells.
During the replication of viruses and intracellular bacteria within their host cell, as well as during the replication of tumor cells, viral, bacterial, or tumor proteins in the cytosol of that cell are degraded into a variety of peptide epitopes by cylindrical organelles called proteasomes . Other endogenous antigens such as proteins released into the cytosol from the phagosomes of antigen-presenting cells, such as macrophages and dendritic cells as well, as a variety of the human cell's own proteins (self-proteins) are also degraded by proteasomes. As these various endogenous antigens pass through proteasomes, proteases and peptidases chop the protein up into a series of peptides, typically 8-11 amino acids long (Figure \(1\)).
of a proteasome degrading proteins into peptides
During the replication of viruses and intracellular bacteria within their host cell, as well as during the replication of tumor cells, viral, bacterial, or tumor proteins, as well proteins released from phagosomes of phagocytes and various human cell or self-proteins, are degraded into a variety of peptide epitopes by cylindrical organelles called proteasomes. As endogenous antigens pass through proteasomes, proteases and peptidases chop the protein up into a series of peptides, typically 8-11 amino acids long.
A transporter protein called TAP located in the membrane of the cell's endoplasmic reticulum then transports these peptide epitopes into the endoplasmic reticulum where they bind to the grooves of various newly made MHC-I molecules. The MHC-I molecules with bound peptides are then transported to the Golgi complex and placed in exocytic vesicles. The exocytic vesicles carry the MHC-I/peptide complexes to the cytoplasmic membrane of the cell where they become anchored to its surface (Figure \(2\)). A single cell may have up to 250,000 molecules of MHC-I with bound epitope on its surface.
During cell-mediated immunity, MHC-I molecule with bound peptide on the surface of infected cells and tumor cells can be recognized by a complementary-shaped TCR/CD8 on the surface of a cytotoxic T-lymphocyte (CTL) to initiate destruction of the cell containing the endogenous antigen (Figure \(3\)).
Endogenous antigens
Endogenous antigens are those located within the cytosol of cells of the body. Examples include:
1. viral proteins produced during viral replication,
2. proteins produced by intracellular bacteria such as Rickettsias and Chlamydias during their replication,
3. proteins that have escaped into the cytosol from the phagosome of phagocytes such as antigen-presenting cells
4. tumor antigens produced by cancer cells,
5. and self peptides from human cell proteins.
The body marks infected cells and tumor cells for destruction by placing peptide epitopes from these endogenous antigens on their surface by way of MHC-I molecules. Cytotoxic T-lymphocytes (CTLs) are then able to recognize peptide/MHC-I complexes by means of their T-cell receptors (TCRs) and CD8 molecules and kill the cells to which they bind.
1. Endogenous antigens, such as viral proteins, pass through proteasomes where they are degraded into a series of peptides.
2. The peptides are transported into the rough endoplasmic reticulum (ER) by a transporter protein called TAP.
3. The peptides then bind to the grooves of newly synthesized MHC-I molecules.
4. The endoplasmic reticulum transports the MHC-I molecules with bound peptides to the Golgi complex.
5. The Golgi complex, in turn, transports the MHC-I/peptide complexes by way of an exocytic vesicle to the cytoplasmic membrane where they become anchored. Here, the peptide and MHC-I/peptide complexes can be recognized by CTLs by way of TCRs and CD8 molecules having a complementary shape.
Cytotoxic T-Lymphocyte (CTL) Destruction of Body Cells Displaying Epitopes of Foreign Antigen on their Surface
The cytotoxic T-lymphocytes (CTLs) produced during cell-mediated immunity are designed to remove body cells displaying "foreign" epitope, such as virus-infected cells, cells containing intracellular bacteria, and cancer cells with mutant surface proteins. The CTLs are able to kill these cells by inducing a programmed cell death known as apoptosis. Using virus-infected cells as an example, the CTLs circulate throughout the body where they encounter virus-infected cells and induce apoptosis. This involves involves a complex of intracellular cytotoxic granules containing:
1. Pore-forming proteins called perforins
2. Proteolytic enzymes called granzymes and
3. Granulysin
When the TCR and CD8 of the CTL binds to the MHC-I/epitope on the surface of the virus-infected cell or tumor cell (Figure \(4\)), this sends a signal through a CD3 molecule which triggers the release of the cytotoxic perforins/granzymes/granulysin granules from the CTL.
The exact mechanism of entry of the granzymes into the infected cell or tumor cell is still debated. It is, however, dependent on perforins. Possibilities include:
1. The perforins/granzymes/granulysin complex may be taken into the target cell by receptor-mediated endocytosis. The perforin molecules may then act on the endosomal membrane allowing granzymes to enter the cytosol.
2. The perforin molecules may put pores in the membrane of the target cell allowing the granzymes to directly enter the cytosol (Figure \(5\)).
Killing of the infected cell or tumor cell by apoptosis involves a variety of mechanisms:
1. Certain granzymes can activate the caspase enzymes that lead to apoptosis of the infected cell. The caspases are proteases that destroy the protein structural scaffolding of the cell - the cytoskeleton - and nucleases that degrade both the target cell's nucleoprotein and any microbial DNA within the cell (Figure \(5\)).
2. Granzymes cleave a variety of other cellular substrates that contribute to cell death.
3. The perforin molecules may also polymerize and form pores in the membrane of the infected cell, similar to those produced by MAC. This can increase the permeability of the infected cell and contribute to cell death. If enough perforin pores form, the cell might not be able to exclude ions and water and may undergo cytolysis.
4. Granulysin has antimicrobial actions and can also induce apoptosis.
• CTLs can also trigger apoptosis through FasL/Fas interactions. Activated lymphocytes express both death receptors called Fas and Fas ligand or FasL (Figure \(6\)) on their surface. This FasL/Fas interaction triggers an intracellular transduction that activates the caspase enzymes that lead to apoptosis. In this way, CTLs can kill other lymphocytes and terminate lymphocyte proliferation after the immune responses have eradicated an infection.
Death by apoptosis does not result in the release of cellular contents such as inflammatory mediators or viruses as occurs during immune-induced cell lysis. Instead, the cell breaks into membrane-bound apoptoptic fragments that are subsequently removed by macrophages. This reduces inflammation and also prevents the release of viruses that have assembled within the infected cell and their spread into uninfected cells. Since the CTLs are not destroyed in these reactions, they can function over and over again to destroy more virus-infected cells.
Exercise: Think-Pair-Share Questions
1. Some viruses inhibit proteasomal activity in the cells they infect. Explain specifically how this might better enable the virus to resist adaptive immunity.
2. Some viruses suppress the production of MHC-I molecules in the cells they infect. Explain specifically how this might better enable the virus to resist adaptive immunity.
3. Some viruses block the TAP transport of peptides into the endoplasmic reticulum of the cells they infect. Explain specifically how this might better enable the virus to resist adaptive immunity.
4. Some viruses block apoptosis of the cells they infect. Explain specifically how this might better enable the virus to resist adaptive immunity.
As with humoral immunity, certain microbes are able to evade to some degree cell-mediated immunity:
• Epstein-Barr virus (EBV) and cytomegalovirus (CMV) inhibit proteasomal activity so that viral proteins are not degraded into viral peptides. (see Figure \(7\)A)
• Herpes simplex viruses (HSV) can block the TAP transport of peptides into the endoplasmic reticulum (see Figure \(7\)B).
• Numerous viruses, such as the cytomegalovirus (CMV) and adenoviruses can block the formation of MHC-I molecules by the infected cell. As a result, no viral peptide is displayed on the infected cell and the CTLs are no longer able to recognize that the cell is infected and kill it (see Figure \(7\)C).
• Epstein-Barr virus (EBV) down regulates several host proteins involved in attaching viral epitopes to MHC-I molecules and displaying them on the host cell's surface (see Figure \(7\)D).
• Adenoviruses and Epstein-Barr Viruses (EBV) code for proteins that blocks apoptosis, the programmed cell suicide mechanism triggered by various defense mechanisms in order to destroy virus-infected cells.
Summary
1. Cell-mediated immunity (CMI) is an immune response that does not involve antibodies but rather involves the activation of macrophages and NK-cells, the production of antigen-specific cytotoxic T-lymphocytes, and the release of various cytokines in response to an antigen.
2. Cell-mediated immunity is directed primarily microbes that survive in phagocytes and microbes that infect non-phagocytic cells.
3. One of the body's major defenses against viruses, intracellular bacteria, and cancers is the destruction of infected cells and tumor cells by cytotoxic T-lymphocytes or CTLs, effector cells derived from naïve T8-lymphocytes during cell-mediated immunity.
4. The TCRs and CD8 molecules on the surface of naive T8-lymphocytes are designed to recognize peptide epitopes bound to MHC-I molecules on antigen-presenting cells (APCs).
5. During the replication of viruses and intracellular bacteria within their host cell, as well as during the replication of tumor cells, viral, bacterial, or tumor proteins (endogenous antigens) in the cytosol of that cell are degraded into a variety of peptide epitopes by cylindrical organelles called proteasomes.
6. These peptide epitopes bind to MHC-I molecules being synthesized in the endoplasmic reticulum which are eventually transported to the cytoplasmic membrane of that cell.
7. During cell-mediated immunity, MHC-I molecule with bound peptide on the surface of infected cells and tumor cells can be recognized by a complementary-shaped TCR/CD8 on the surface of a cytotoxic T-lymphocyte (CTL) to initiate destruction of the cell containing the endogenous antigens.
8. When the TCR and CD8 of the CTL binds to the MHC-I/epitope on the surface of the virus-infected cell or tumor cell, this triggers the release of cytotoxic perforins/granzymes/ granulysin granules from the CTL that lead to apoptosis, a programmed cell suicide of that cell.
9. Cell death by apoptosis does not result in the release of cellular contents such as inflammatory mediators or viruses as occurs during immune-induced cell lysis.
10. During apoptosis, the cell breaks into membrane-bound apoptotic fragments that are subsequently removed by macrophages. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/14%3A_Cell-Mediated_Immunity/14.2%3A_Activating_Antigen-Specific_Cytotoxic_T-_Lymphocytes.txt |
Learning Objectives
1. Describe how TH1 effector cells are able to interact with and activate macrophages.
2. Describe how NK cells are able to recognize and destroy infected cells and cancer cells lacking MHC-I molecules.
After interacting with APCs, some naive T4-lymphocytes differentiate into a subset of effector cells called TH1 cells. TH1 cells function primarily to promote phagocytosis of microbes and the killing of intracellular microbes.
Activation of Macrophages
Effector T4-lymphocytes called TH1 cells coordinate immunity against intracellular bacteria and promote opsonization by macrophages. They produce cytokines such as interferon-gamma (IFN-?) that promote cell-mediated immunity against intracellular pathogens, especially by activating macrophages that have either ingested pathogens or have become infected with intracellular microbes such as Mycobacterium tuberculosis, Mycobacterium leprae, Leishmania donovani, and Pneumocystis jiroveci that are able to grow in the endocytic vesicles of macrophages. Activation of the macrophage by TH1 cells greatly enhances their antimicrobial effectiveness (Figure \(1\)).
They produce cytokines that promote the production of increases the production of opsonizing and complement activating IgG that enhances phagocytosis (Figure \(1\)).
• They produce receptors that bind to and kill chronically infected cells, releasing the bacteria that were growing within the cell so they can be engulfed and killed by macrophages.
• They produce cytokines such as tumor necrosis factor-alpha (TNF-a) that promote diapedesis of macrophages.
• They produce the chemokine CXCL2 to attract macrophages to the infection site.
Activated natural killer T-lymphocytes (NKT cells) also produce large amounts of IFN-gamma to activate macrophages.
Activation of macrophages Increases their production of toxic oxygen radicals, nitric oxide, and hydrolytic lysosomal enzymes enabling the killing of microbes within their phagolysosomes. It also causes the macrophages to secrete cytokines such as TNF-a, IL-1, and IL-12. TNF-a and IL-1 promote inflammation to recruit phagocytic leukocytes. lL-12 enables naive T4-lymphocytes to differentiate into TH1 cells. Moreover activation increases the production of B7 co-stimulator molecules and MHC-1 molecules by macrophages for increased T-lymphocyte activation.
Activation of NK Cells
Cytokines such as interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) produced by TH1 lymphocytes activate NK cells. NK cells are another group of cytolytic lymphocytes, distinct from B-lymphocytes and T-lymphocytes, that participate in both innate immunity and adaptive immunity. NK cells are lymphocytes that lack B-cell receptors and T-cell receptors. They are designed to kill certain mutant cells and virus-infected cells in one of two ways:
1. NK cells kill cells to which antibody molecules have attached through a process called antibody-dependent cellular cytotoxicity (ADCC) as shown in Figure \(3\) , Figure \(4\), and Figure \(5\) . The Fab portion of the antibody binds to epitopes on the "foreign" cell. The NK cell then binds to the Fc portion of the antibody. The NK cell is then able to contact the cell and by inducing a programmed cell suicide called apoptosis.
2. NK cells to use a duel receptor system in determining whether to kill or not kill human cells. When cells are either under stress, are turning into tumors, or are infected, various molecules such as MICA and MICB are produced and are put on the surface of that cell. The first receptor, called the killer-activating receptor, can bind to various molecules such as MICA and MICB that are produced and are put on the surface of that cell, and this sends a positive signal that enables the NK cell to kill the cell to which it has bound unless the second receptor cancels that signal. This second receptor, called the killer-ihibitory receptor, recognizes MHC-I molecules that are also usually present on all nucleated human cells. If MHC-I molecules are expressed on the cell, the killer-inhibitory receptor sends a negative signal that overrides the kill signal and prevents the NK cell from killing that cell (Figure \(6\)).
Viruses and malignant transformation can sometimes interfere with the ability of the infected cell or tumor cell to express MHC-I molecules. Without the signal from the killer-inhibitory receptor, the kill signal from the killer-activating signal is not overridden and the NK cell kills the cell to which it has bound (Figure \(7\)).
The NK cell releases pore-forming proteins called perforins and proteolytic enzymes called granzymes. Granzymes pass through the pores and activate the enzymes that lead to apoptosis of the infected cell by means of destruction of its structural cytoskeleton proteins and by chromosomal degradation. As a result, the cell breaks into fragments that are subsequently removed by macrophages (Figure \(5\)). Perforins can also sometimes result in cell lysis. The distinction between causing apoptosis versus causing cell lysis is important because lysing a virus-infected cell would only release the virions, whereas apoptosis leads to destruction of the virus inside.
NK cells also produce a variety of cytokines, including proinflammatory cytokines, chemokines, colony-stimulating factors, and other cytokines that function as regulators of body defenses. For example, through cytokine production NK cells also suppress and/or activate macrophages, suppress and/or activate the antigen-presenting capabilities of dendritic cells, and suppress and/or activate T-lymphocyte responses.
As with humoral immunity, certain microbes are able to evade to some degree NK cells:
• The cytomegalovirus (CMV) can also trigger its host cell to produce altered MHC-I molecules that are unable to bind viral epitopes, and, therefore, are not recognized by CTLs. However, NK cells are also unable to kill this infected cell because it is still displaying "MHC-I molecules" on its surface.
• CMV also produces microRNAs (miRNAs), small non-coding RNA molecules that down-regulates the production of stress-induced proteins that the killer-activating receptor of NK cells first recognizes. The miRNAs do this by binding to the host cell's mRNA coding for stress-induced proteins ( Figure \(14\).3.9). Without this binding there is no kill signal by the NK cell.
• Cytomegalovirus (CMV) and herpes simplex type 1 virus (HSV-1) produce microRNAs (miRNAs), small non-coding RNA molecules that block protein involved in apoptosis, a programmed cell suicide. The miRNAs do this by binding to the host cell's mRNA coding for apoptosis-inducing proteins (Figure \(9\)).
Figure \(9\): Antisense RNA (microRNA or miRNA). When an antisense RNA (microRNA or miRNA) that is complementary to a mRNA coding for a particular protein or enzyme binds to the mRNA by complementary base pairing, that mRNA cannot be translated and the protein or enzyme is not made.
Summary
1. Effector T4-lymphocytes called TH1 cells coordinate immunity against intracellular bacteria and promote opsonization by macrophages.
2. Cytokines produced by TH1 cells promote cell-mediated immunity against intracellular pathogens by activating macrophages and enhancing their antimicrobial effectiveness, increasing the production of opsonizing and complement activating IgG that enhances phagocytosis, and promoting diapedesis and chemotaxis of macrophages to the infection site.
3. Activation of natural killer T-lymphocytes (NKT cells) produces large amounts of IFN-gamma to activate macrophages.
4. Cytokines such as interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) produced by TH1 lymphocytes activate NK cells.
5. Activated NK cells kill cells to which antibody molecules have attached through a process called antibody-dependent cellular cytotoxicity (ADCC).
6. Activated NK cells also use a duel receptor system in determining whether to kill or not kill cells such as cancer cells and infected cells that are displaying stress molecules and are not producing MHC-I molecules.
7. NK cells kill infected cells and cancer cells by inducing apoptosis, a programmed cell suicide. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/14%3A_Cell-Mediated_Immunity/14.3%3A_Activating_Macrophages_and_NK_Cells.txt |
Learning Objectives
1. Define cytokine and explain what is meant by "cytokines are pleiotropic, redundant, and multifunctional."
2. Name 3 cytokines that regulate innate immune responses by triggering an inflammatory response.
3. Name the group of cytokines that regulates innate immunity by preventing translation of viral mRNA and by degrading both viral and host cell RNA.
4. Name 4 cytokines that regulate adaptive immune responses.
5. Name 2 cytokines that stimulate hematopoiesis.
Cytokines are low molecular weight, soluble proteins that are produced in response to an antigen and function as chemical messengers for regulating the innate and adaptive immune systems. They are produced by virtually all cells involved in innate and adaptive immunity, but especially by T helper (TH) lymphocytes. The activation of cytokine-producing cells triggers them to synthesize and secrete their cytokines. The cytokines, in turn, are then able to bind to specific cytokine receptors on other cells of the immune system and influence their activity in some manner. Cytokines are pleiotropic, redundant, and multifunctional.
• Pleiotropic means that a particular cytokine can act on a number of different types of cells rather than a single cell type.
• Redundant refers to to the ability of a number of different cytokines to carry out the same function.
• Multifunctional means the same cytokine is able to regulate a number of different functions.
Some cytokines are antagonistic in that one cytokine stimulates a particular defense function while another cytokine inhibits that function. Other cytokines are synergistic wherein two different cytokines have a greater effect in combination than either of the two would by themselves. There are three functional categories of cytokines:
1. Cytokines that regulate innate immune responses,
2. Cytokines that regulate adaptive Immune responses, and
3. Cytokines that stimulate hematopoiesis.
Cytokines that Regulate Innate Immunity
a. Cytokines that regulate innate immunity are produced primarily by mononuclear phagocytes such as macrophages and dendritic cells, although they can also be produced by T-lymphocytes, NK cells, endothelial cells, and mucosal epithelial cells. They are produced primarily in response to pathogen-associated molecular patterns (PAMPs) such as LPS, peptidoglycan monomers, teichoic acids, unmethylated cytosine-guanine dinucleotide or CpG sequences in bacterial and viral genomes, and double-stranded viral RNA. Cytokines produced in response to PRRs on cell surfaces, such as the inflammatory cytokines IL-1, IL-6, IL-8, and TNF-alpha, mainly act on leukocytes and the endothelial cells that form blood vessels in order to promote and control early inflammatory responses (Figure \(1\)).
Cytokines produced in response to PRRs that recognize viral nucleic acids, such as type I interferons, primarily block viral replication within infected host cells (Figure \(2\)).
Examples include:
1. Tumor necrosis factor-alpha (TNF-a): TNF-a is the principle cytokine that mediates acute inflammation. In excessive amounts it also is the principal cause of systemic complications such as the shock cascade. Functions include acting on endothelial cells to stimulate inflammation and the coagulation pathway; stimulating endothelial cells to produce selectins and ligands for leukocyte integrins (Figure \(1\)) during diapedesis; stimulating endothelial cells and macrophages to produce chemokines that contribute to diapedesis, chemotaxis, and the recruitment of leukocytes; stimulating macrophages to secrete interleukin-1 (IL-1) for redundancy; activating neutrophils and promoting extracellular killing by neutrophils; stimulating the liver to produce acute phase proteins, and acting on muscles and fat to stimulate catabolism for energy conversion. In addition, TNF is cytotoxic for some tumor cells; interacts with the hypothalamus to induce fever and sleep; stimulates the synthesis of collagen and collagenase for scar tissue formation; and activates macrophages. TNF is produced by monocytes,macrophages, dendritic cells, TH1 cells, and other cells.
2. Interleukin-1 (IL-1): IL-1 function similarly to TNF in that it mediates acute inflammatory responses. It also works synergistically with TNF to enhance inflammation. Functions of IL-1 include promoting inflammation; activating the coagulation pathway, stimulating the liver to produce acute phase proteins, catabolism of fat for energy conversion, inducing fever and sleep; stimulates the synthesis of collagen and collagenase for scar tissue formation; stimulates the synthesis of adhesion factors on endothelial cells and leukocytes (Figure \(1\)) for diapedesis; and activates macrophages. IL-1 is produced primarily by monocytes, macrophages, dendritic cells, endothelial cells, and some epithelial cell.
3. Chemokines: Chemokines are a group of cytokines that enable the migration of leukocytes from the blood to the tissues at the site of inflammation. They increase the affinity of integrins on leukocytes for ligands on the vascular wall (Figure \(1\)) during diapedesis, regulate the polymerization and depolymerization of actin in leukocytes for movement and migration, and function as chemoattractants for leukocytes. In addition, they trigger some WBCs to release their killing agents for extracellular killing and induce some WBCs to ingest the remains of damaged tissue. Chemokines also regulate the movement of B-lymphocytes, T-lymphocytes, and dendritic cells through the lymph nodes and the spleen. When produced in excess amounts, chemokines can lead to damage of healthy tissue as seen in such disorders as rheumatoid arthritis, pneumonia, asthma, adult respiratory distress syndrome (ARDS), and septic shock. Examples of chemokines include IL-8, MIP-1a, MIP-1b, MCP-1, MCP-2, MCP-3, GRO-a, GRO-b, GRO-g, RANTES, and eotaxin. Chemokines are produced by many cells including leukocytes, endothelial cells, epithelial cells, and fibroblasts.
4. Interleukin-12 (IL-12): IL-12 is a primary mediator of early innate immune responses to intracellular microbes. It is also an inducer of cell-mediated immunity. It functions to stimulate the synthesis of interferon-gamma by T-lymphocytes and NK cells; increases the killing activity of cytotoxic T-lymphocytes and NK cells; and stimulates the differentiation of naive T4-lymphocytes into interferon-gamma producing TH1 cells. It is produced mainly by macrophages and dendritic cells.
5. Type I Interferons: Interferons modulate the activity of virtually every component of the immune system. Type I interferons include 13 subtypes of interferon-alpha, interferon-beta, interferon omega, interferon-kappa, and interferon tau. (There is only one type II interferon, interferon-gamma, which is involved in the inflammatory response.)
The most powerful stimulus for type I interferons is the binding of viral DNA or RNA to toll-like receptors TLR-3, TLR-7, and TLR-9 in endosomal membranes.
1. TLR-3 - binds double-stranded viral RNA;
2. TLR-7 - binds single-stranded viral RNA, such as in HIV, rich in guanine/uracil nucleotide pairs;
3. TLR-9 - binds unmethylated cytosine-guanine dinucleotide sequences (CpG DNA) found in bacterial and viral genomes but uncommon or masked in human DNA and RNA.
Signaling pattern recognition receptors located in the cytoplasm of cells such as RIG-1 and MDA-5 also signal synthesis and secretion of type-I interferons.
Type I interferons, produced by virtually any virus-infected cell, provide an early innate immune response against viruses. Interferons induce uninfected cells to produce enzymes capable of degrading mRNA. These enzymes remain inactive until the uninfected cell becomes infected with a virus. At this point, the enzymes are activated and begin to degrade both viral and cellular mRNA. This not only blocks viral protein synthesis, it also eventually kills the infected cell (Figure \(2\)). In addition, type I interferons also cause infected cells to produce enzymes that interfere with transcription of viral RNA or DNA. They also promote body defenses by enhancing the activities of CTLs, macrophages, dendritic cells, NK cells, and antibody-producing cells.
Antiviral Action of Interferon Interferon induces uninfected cells to produce enzymes capable of degrading mRNA. These enzymes remain inactive until the uninfected cell becomes infected with a virus. At this point, the enzymes are activated and begin to degrade both viral and cellular mRNA. This not only blocks viral protein synthesis, it also eventually kills the infected cell.
Type I interferons also induce MHC-I antigen expression needed for recognition of antigens by cytotoxic T-lymphocytes; augment macrophage, NK cell, cytotoxic T-lymphocytes, and B-lymphocyte activity; and induce fever. Interferon-alpha is produced by T-lymphocytes, B-lymphocytes, NK cells, monocytes/macrophages; interferon-beta by virus-infected cells, fibroblasts, macrophages, epithelial cells, and endothelial cells.
6. Interleukin-6 (IL-6): IL-6 functions to stimulate the liver to produce acute phase proteins; stimulates the proliferation of B-lymphocytes; and increases neutrophil production. IL-6 is produced by many cells including T-lymphocytes, macrophages, monocytes, endothelial cells, and fibroblasts.
7. Interleukin-10 (IL-10): IL-10 is an inhibitor of activated macrophages and dendritic cells and as such, regulates innate immunity and cell-mediated immunity. IL-10 inhibits their production of IL-12, co-stimulator molecules, and MHC-II molecules, all of which are needed for cell-mediated immunity. IL-10 is produced mainly by macrophages, and TH2 cells.
8. Interleukin 15 (IL-15): IL-15 stimulates NK cell proliferation and proliferation of memory T8-lymphocytes. IL-15 is produced by various cells including macrophages.
9. Interleukin-18 (IL-18): IL-18 stimulates the production of interferon-gamma by NK cells and T-lymphocytes and thus induces cell-mediated immunity. It is produced mainly by macrophages.
Cytokines that Regulate Adaptive Immune Responses (Humoral Immunity and Cell-Mediated Immunity)
Cytokines that regulate adaptive immunity are produced primarily by T-lymphocytes that have recognized an antigen specific for that cell. These cytokines function in the proliferation and differentiation of B-lymphocytes and T-lymphocytes after antigen recognition and in the activation of effector cells.
Examples include:
1. Interleukin-2 (IL-2): IL-2 is a growth factor for NK cells and antigen-stimulated T-lymphocytes and B-lymphocytes. IL-2 also increases the killing ability of NK cells; increases the synthesis of other cytokines; increases Fas-mediated apoptosis; and stimulates antibody synthesis by B-lymphocytes. IL-2 is produced mainly by T4-lymphocytes and to a lesser extent T8-lymphocytes.
2. Interleukin-4 (IL-4): IL-4 is a major stimulus for production of the antibody isotype IgE and the development of Th2 cells for defense against helminths and arthropods. It also antagonizes the effects of interferon-gamma and thus inhibits cell-mediated immunity. IL-4 is produced mainly by TH2 cells and mast cells.
3. Interleukin-5 (IL-5): IL-5 is a growth and activating factor for eosinophils as a defense against helminths and arthropods. It also stimulates the proliferation and differentiation of antigen-activated B-lymphocytes and the production of IgA. IL-5 is produced mainly by TH2 cells.
4. Interferon-gamma (IFN-?):Interferons modulate the activity of virtually every component of the immune system. Type I interferons include more than 20 types of interferon-alpha, interferon-beta, interferon omega, and interferon tau. There is only one type II interferon, interferon-gamma. Type II interferon is produced by activated T-lymphocytes as part of an immune response and functions mainly to promote the activity of the components of the cell-mediated immune system such as CTLs, macrophages, and NK cells. IFN-? is the principal cytokine for activating macrophages. It also induces the production of MHC-I molecules, MHC-II molecules, and co-stimulatory molecules by APCs in order to promote cell-mediated immunity and activates and increases the antimicrobial and tumoricidal activity of monocytes, macrophages, neutrophils, and NK cells. IFN-? stimulates the differentiation of T4-lymphocytes into TH1 cells and inhibits the proliferation of TH2 cells; stimulates the production of IgG subclasses that activate the complement pathway and promote opsonization; and augments or inhibits other cytokine activities. IFN-? is produced primarily by TH1 cells, CD8+ cells, and NK cells.
5. Transforming growth factor-beta (TGF-ß): TGF-ß functions to inhibit the proliferation and effector function of T-lymphocytes; inhibit the proliferation of B-lymphocytes; and inhibits macrophage function. It also promotes tissue repair. TGF-ß is produced by T-lymphocytes, macrophages, and other cells.
6. Lymphotoxin (LT): LT plays a role in the recruitment and activation of neutrophils and in lymphoid organogenesis. Being chemically similar to TNF, LT is also a mediator of acute inflammatory responses. LT is made by T-lymphocytes.
7. Interleukin-13 (IL-13): IL-13 increases the production of IgE by B-lymphocytes, inhibits macrophages, and increases mucus production. IL-13 is made primarily by TH2 cells.
Cytokines that Stimulate Hematopoiesis
Produced by bone marrow stromal cells, these cytokines stimulate the growth and differentiation of immature leukocytes.
Examples include:
1. Colony-stimulating factors (CSF): Promote the production of colonies of the different leukocytes in the bone marrow and enhance their activity. Examples include granulocyte macrophage colony stimulating factor (GM-CSF), granulocyte colony stimulating factor (G-CSF), and macrophage colony stimulating factor (M-CSF). In addition to their role in promoting production of leukocyte colonies, the CSFs also appear to promote their function. For example, when GM-CSF binds to receptors on neutrophils, eosinophils, and monocytes, it activates these cells and inhibits their apoptosis. GM-CSF increases adhesion of these cells to capillary walls during diapedesis, enhances their phagocytosis and extracellular killing, and increases both superoxide anion generation and antibody-dependent cytotoxicity. The various CSFs are produced by T-lymphocytes, macrophages, and other cells.
2. Stem cell factor: Stem cell factor makes stem cells in the bone marrow mor responsive to the various CSFs. It is made mainly by bone marrow stromal cells.
3. Interleukin-3 (IL-3): IL-3 supports the growth of multilineage bone marrow stem cells. IL-3 is made primarily by T-lymphocytes.
4. Interleukin-7 (IL-7): IL-7 plays a role in the survival and proliferation of immature B-lymphocyte and T-lymphocyte precursors. Il-7 is produced mainly my fibroblasts and bone marrow stromal cells.
Some viruses cause infected host cells to secrete molecules that bind and tie up cytokines, preventing them from binding to normal cytokine receptors on host cells.
• Poxviruses cause infected host cells to secrete molecules that bind interleukin-1 (IL-1) and interferon-gamma (IFN-gamma).
• Cytomegaloviruses (CMV) cause infected host cells to secrete molecules that bind chemokines.
Summary
1. Cytokines are low molecular weight, soluble proteins that are produced in response to an antigen and function as chemical messengers for regulating the innate and adaptive immune systems.
2. Cytokines are pleiotropic, meaning that a particular cytokine can act on a number of different types of cells rather than a single cell type.
3. Cytokines are redundant, meaning that a number of different cytokines to carry out the same function.
4. Cytokines are multifunctional, meaning the same cytokine is able to regulate a number of different functions.
5. There are three functional categories of cytokines: Cytokines that regulate innate immune responses; cytokines that regulate adaptive Immune responses; and cytokines that stimulate hematopoiesis.
6. Type I interferons provide an early innate immune response against viruses. Interferons induce uninfected cells to produce enzymes capable of degrading mRNA. These enzymes remain inactive until the uninfected cell becomes infected with a virus. At this point, the enzymes are activated and begin to degrade both viral and cellular mRNA. This not only blocks viral protein synthesis, it also eventually kills the infected cell.
14.E: Cell-Mediated Immunity (Exercises)
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied.
14.1: Cell-Mediated Immunity: An Overview
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. State three different ways by which cell-mediated immunity protects the body.
2. Define gene translocation. (ans)
3. Relate gene translocation to each T-lymphocyte being able to produce a T-cell receptor with a unique shape. (ans)
4. Define the following:
1. combinatorial diversity (ans)
5. In terms of humoral immunity, discuss what is meant by anamnestic response. (ans)
6. Briefly describe why there is a heightened secondary response during anamestic response. (ans)
14.2: Activating Antigen-Specific Cytotoxic T- Lymphocytes
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. The role of cytotoxic T-lymphocytes (CTLs) in body defense.
1. State from what cells cytotoxic T-lymphocytes are derived. (ans)
2. Describe how they can react with and destroy virus-infected cells, cells containing intracellular bacteria, and cancer cells without harming normal cells. (Indicate the role of following: TCR, CD4, MHC-I, and peptides from endogenous antigens.) (ans)
3. State the mechanism by which cytotoxic T-lymphocytes kill the cells to which they bind. (Indicate the role of the following: perforins, granzymes, caspases, and macrophages in the process.) (ans)
2. Multiple Choice (ans)
14.3: Activating Macrophages and NK Cells
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Viruses and malignant transformation can sometimes interfere with the ability of the infected cell or tumor cell to express MHC-I molecules. This enables them to resist destruction by cytoyoxic T-lymphocytes. However the body is still able to kill these infected cells and tumor cells. Describe how. (ans)
2. Describe how TH1 effector cells are able to interact with and activate macrophages. (ans)
3. Multiple Choice (ans)
14.4: Stimulating Cells to Secrete Cytokines
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Name 4 cytokines that regulate adaptive immune responses. (ans)
2. Name 3 cytokines that regulate innate immune responses by triggering an inflammatory response. (ans)
3. Name 2 cytokines that stimulate hematopoiesis. (ans)
4. Name the group of cytokines that regulates innate immunity by preventing translation of viral mRNA and by degrading both viral and host cell RNA. (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/14%3A_Cell-Mediated_Immunity/14.4%3A_Stimulating_Cells_to_Secrete_Cytokines.txt |
Thumbnail: Scanning electron micrograph of HIV-1 budding (in green) from cultured lymphocyte. This image has been colored to highlight important features; see PHIL 1197 for original black and white view of this image. Multiple round bumps on cell surface represent sites of assembly and budding of virions. Images used with permission (Public Domain; This media comes from the Centers for Disease Control and Prevention's Public Health Image Library (PHIL), with identification number #10000).
15: Immunodeficiency
Learning Objectives
1. Define primary immunodeficiency.
2. Compare and contrast conventional and novel primary immunodeficiencies.
3. Name four categories of conventional immunodeficiencies and give an example of each.
A primary immunodeficiency is usually an immunodeficiency that one is born with. Until recently, primary immunodeficiencies were defined as a rare recessive genetic defect in the immune responses that involved the development of B-lymphocytes, T-lymphocytes, or both and resulted in multiple, recurrent infections during infancy. Depending on the disorder, the lymphocytes in question were either completely absent, present in very low levels, or present but not functioning normally. These disorders represent the conventional immunodeficiencies.
However, based on our increased understanding of the human genome and immune responses it now appears that there are a multitude of common, less severe primary immunodeficiencies involving just one or more of the huge number of genes involved in the immune responses. These so called novel primary immunodeficiencies involve the decreased ability to combat just a single type of infection or a narrow range of infections. The conventional primary immunodeficiencies were grouped as follows:
Conventional: B-lymphocyte Disorders
In the case of B-lymphocyte disorders, there may be may be greatly decreased humoral immunity but cell-mediated immunity , mediated by T-lymphocytes, remains normal.
1. Agammaglobulinemias: Few if any antibodies are produced and there are reduced B-lymphocyte numbers. The person is very susceptible to recurrent infections by common pyogenic bacteria such as Staphylococcus aureus, Streptococcus pyogenes, Streptococcus pneumoniae, Neisseria meningitidis, and Hemophilus influenzae. These bacteria have antiphagocytic capsules that are normally eliminated by antibodies through opsonization. Examples include X-linked agammaglobulinemia and Autosomal recessive agammaglobulinemia.
2. Hypogammaglobulinemias /Isotype Defects: Decreased general antibody production or decrease production of a single isotype of antibody. Examples include:
• IgG2 subclass deficiency: A person is unable to produce the subclass of IgG called IgG2 but can produce other classes of antibodies. There is increased susceptibility to bacterial infections.
• Selective IgA deficiency: A person is unable to make IgA but can produce other classes of antibodies. There is increased susceptibility to bacterial infections and certain protozoan infections.
• Combined Variable Immunodeficiency (CVID): Hypogammaglobulinemia with normal or decreased numbers of B-lymphocytes.
More severe forms such as agammaglobulinemia are treated with artificially-acquired passive immunization - periodic injections of large amounts of immune globulin (IG or IVIG).
Conventional: T-lymphocyte Disorders
In the case of T-lymphocyte disorders, there is little or no cell-mediated immunity if the disorder involves T8-lymphocytes and/or T4-lymphocytes. There may also be decreased humoral immunity if there is a disorder involves T4-lymphocytes.
1. MHC Expression Defects
• MHC-I deficiency. Decreased levels of MHC-I production and reduced T8-lymphocyte numbers.
• Bare lymphocyte syndrome. Decreased levels of MHC-II, decreased numbers of T4-lymphocytes, and decreased T4-dependent antibody production by B-lymphocytes.
2. T-Lymphocyte Signaling Defects
• Wiskott-Aldrich syndrome. Defective T-lymphocyte activation and defective leukocyte mobility.
• Proximal TCR signaling defects. Defective cell-mediated immunity and defective T4-dependent antibody production by B-lymphocytes.
3. Familial Hemophagocytic Lymphohistiocytosis
• Perforin deficiencies. Defective CTL and NK cell function; uncontrolled activation of macrophages and CTLs.
• Granule fusion defects. Defective CTL and NK cell function; uncontrolled activation of macrophages and CTLs.
• X-linked lymphoproliferative syndrome. Defective CTL and NK cell function; uncontrolled activation of macrophages and CTLs. Uncontrolled Epstein-Barr virus - induced B-lymphocyte proliferation.
Conventional: Combined B- and T-lymphocyte Disorders (Severe Combined Immunodeficiency Disease or SCID)
Severe combined immunodeficiency disease or SCID affects both humoral immunity and cell-mediated immunity . There is a defect in both B-lymphocytes and T-lymphocytes, or just T-lymphocytes in which case the humoral deficiency is due to the lack of T4-helper lymphocytes.
1. Cytokine-Signaling Defects
• Autosomal recessive SCID. Shows a marked decrease in T-lymphocytes but normal to increased levels of B-lymphocytes. There is reduced antibody levels due to the lack of T4-helper lymphocytes.
• X-linked recessive SCID. Shows a marked decrease in T-lymphocytes but normal to increased levels of B-lymphocytes. There is reduced antibody levels due to the lack of T4-helper lymphocytes.
2. Defects in Nucleotide Salvage Pathways
• PNP deficiency. Shows a progressive decrease in both T-lymphocytes, B-lymphocytes, and NK cells, as well as reduced antibody levels.
• ADA deficiency. Shows a progressive decrease in both T-lymphocytes, B-lymphocytes, and NK cells, as well as reduced antibody levels.
3. Defects in V(D)J Recombination (Combinatorial Diversity)
• RAG1 or RAG2 deficiency. Shows an absence or deficiency of both T-lymphocytes and B-lymphocytes, as well as reduced antibody levels.
• ARTEMIS defects. Shows an absence or deficiency of both T-lymphocytes and B-lymphocytes, as well as reduced antibody levels.
4. Defective Thymus Development
The thymus is needed for the development of T-lymphocytes from stem cells.
• DiGeorge syndrome. Shows decreased levels of T-lymphocytes, normal levels of B-lymphocytes, and reduced antibody levels.
• Defective pre-TCR checkpoint. Shows decreased levels of T-lymphocytes, normal or reduced levels of B-lymphocytes, and reduced antibody levels.
Conventional: Innate Immunity Disorders
• Chronic granulomatous disease. No oxygen-dependant killing pathway in phagocytes. Recurrent intracellular bacterial and fungal infections.
• Leukocyte adhesion deficiencies. Defective leukocyte adhesion, diapedesis , and migration. Recurrent bacterial and fungal infections.
• Chediak-Higashi syndrome. Defective vesicle fusion and lysosomal function in neutrophils, dendritic cells, macrophages and other cells. Recurrent infections by pyogenic bacteria.
Novel Immunodeficiencies
While the rare conventional primary immunodeficiencies mentioned above are still very important, based on our increased understanding of the human genome and immune responses it now appears that there are a multitude of common, less severe primary immunodeficiencies. These so called novel primary immunodeficiencies relate to an individual’s own unique genetics and can involve one or more of many immunity genes, ranging from any of the huge number of genes conferring protective immunity in general, to individual genes conferring specific immunity to a single pathogen.
It is now thought that almost every person suffers from one form of primary immunodeficiency or another. Unlike the classical primary immunodeficiencies, however, these primary Examples include:
• Disorders of the interleukin-12/interferon-gamma pathway appear to make individuals more susceptible to Mycobacterium and Salmonella infections.
• Disorders of the TLR-3 pathway makes individuals more susceptible to herpes simplex virus encephalitis.
• Disorders of the toll-interleukin 1 receptor/nuclear factor kappa B pathway makes individuals more susceptible to staphylococcal and pneumococcal infections.
• Disorders of properdin and terminal components of the complement pathways make individuals more susceptible to Neisseria infections.
• People with chronic sinusitis that does not respond well to treatment have decreased activity of TLR-9 and produce reduced levels of human beta-defensin 2, as well as mannan-binding lectin needed to initiate the lectin complement pathway.
Summary
1. Immunodeficiency results in an inability to combat certain diseases.
2. A primary immunodeficiency is usually an immunodeficiency that one is born with.
3. Conventional primary immunodeficiencies are rare recessive genetic defect in the immune responses that involved the development of B-lymphocytes, T-lymphocytes, or both and resulted in multiple, recurrent infections during infancy. Depending on the disorder, the lymphocytes in question were either completely absent, present in very low levels, or present but not functioning normally.
4. Conventional primary immunodeficiencies include B-lymphocyte disorders, T-lymphocyte disorders, Severe combined immunodeficiency disease or SCID,and innate immunity disorders.
5. B-lymphocyte disorders may result in greatly decreased humoral immunity but cell-mediated immunity, mediated by T-lymphocytes, remains normal.
6. T-lymphocyte disorders may result in little or no cell-mediated immunity if the disorder involves T8-lymphocytes and/or T4-helper lymphocytes. There may also be decreased humoral immunity if there is a disorder involves T4-helper lymphocytes.
7. Severe combined immunodeficiency disease deficiencies affect both humoral immunity and cell-mediated immunity may result in a defect in both B-lymphocytes and T-lymphocytes, or just T-lymphocytes in which case the humoral deficiency is due to the lack of T4-helper lymphocytes.
8. Innate immunity disorders are due to defects in genes that play a role in innate immune responses.
9. Novel primary immunodeficiencies include a multitude of common, less severe primary immunodeficiencies involving just one or more of the huge number of genes involved in the immune responses resulting in the decreased ability to combat just a single type of infection or a narrow range of infections. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/15%3A_Immunodeficiency/15.1%3A_Primary_Immunodeficiency.txt |
Learning Objectives
1. State what is meant by secondary immunodeficiency and list four possible contributing factors.
2. Briefly give at least four mechanisms of HIV-induced immunodeficiency.
In the case of secondary immunodeficiency, one is born with normal immune responses but some secondary factor or occurrence causes a decrease in immune responses. Secondary immunodeficiency is induced by factors such as:
• Malnutrition. Inhibits lymphocyte maturation and function.
• Some viruses, e.g., HIV. Depletes T4-lymphocytes.
• Irradiation - exposure to X-rays and gamma rays. Causes a decreased production of lymphocyte precursors in the bone marrow.
• Cytotoxic drugs such as many used in cancer chemotherapy. Causes a decreased production of lymphocyte precursors in the bone marrow.
• Corticosteroids – anti-inflammatory steroids. Damages lymphocytes.
• Leukemias, cancers of the lymphoid system, metastases. Reduces areas for lymphocyte development.
• Aging. Adaptive immunity, especially cell-mediated immunity, tends to diminish with aging.
• Removal of the spleen. Decreased ability to remove microbes that enter the blood.
A secondary immunodeficiency of current notoriety is of course Acquired Immunodeficiency Syndrome or AIDS, a secondary immunodeficiency caused by Human Immunodeficiency Virus (HIV). As we saw in Unit 4, HIV, via its gp120, primarily infects cells with CD4 molecules and chemokine receptors on their surface, namely, T4-lymphocytes, macrophages, and dendritic cells. The median incubation period for AIDS is around 10 years.
During early or acute HIV infection the virus primarily infects and destroys memory T4-lymphocytes which express the chemokine receptor CCR5 and are very abundant in mucosal lymphoid tissues. Here HIV also encounters the dendritic cells located throughout the epithelium of the skin and the mucous membranes where in their immature form called Langerhans cells they are attached by long cytoplasmic processes. The envelope glycoproteins gp41 and gp120 of HIV contain mannose-rich glycans that bind to mannan-binding proteins (pattern recognition receptors; also called lectin receptors) on the dendritic cells.
Upon capturing antigens through pinocytosis and phagocytosis and becoming activated by pro-inflammatory cytokines, the dendritic cells detach from the epithelium, enter lymph vessels, and are carried to regional lymph nodes. By the time they enter the lymph nodes, the dendritic cells have matured and are now able to present antigens of HIV to naive T-lymphocytes located in the the lymph nodes in order to induce adaptive immune responses.
At this point the infection has transitioned from the acute phase to the chronic phase. The chronic phase of HIV infection is characterized by viral dissemination, viremia, and induction of adaptive immune responses. The viremia allows the viruses to spread and infect T4-helper lymphocytes, macrophages, and dendritic cells found in peripheral lymphoid tissues.
During the chronic phase of HIV infection, the lymph nodes and the spleen become sites for continuous viral replication and host cell destruction. During most of this phase, the immune system remains active and competent and there are few clinical symptoms. A steady state-infection generally persists where T4-lymphocyte death and T4-lymphocyte replacement by the body are in equilibrium. In a person infected with HIV, somewhere between one and two billion of these T4-cells die each day as a result of HIV infection and must be replaced by the body's lymphopoietic system in the bone marrow. It is estimated that 10 billion virions are produced and cleared in an infected individual each day. However, the enormous turnover of T4-lymphocytes eventually exhausts the lymphopoietic system and it becomes unable to replace the T4-cells being destroyed. A variety of mechanisms then eventually lead to immunodeficiency.
Mechanisms of HIV-induced immunodeficiency include:
• Direct HIV-induced cytopathic effect on infected T4-lymphocytes. This can occur through:
• Increased cell permeability as a result of gp41 expression in the host cell membrane and viral release by budding;
• Inhibition of host cell protein synthesis as a result of viral replication within the infected cell; and
• Fusion of infected T4-cells with numerous uninfected T4-cells resulting in syncytia formation.
• Killing of HIV-infected T4-cells by cytotoxic T-lymphocytes or CTLs.
• Killing of HIV-infected T4-cells by antibody-dependent cytotoxicity or ADCC.
• Apoptosis of T4-cells as a result of chronic activation by HIV and by cytokines.
• Shedding of gp120 molecules by HIV. This subsequently triggers a series of events that cause the adaptive immune system to become less and less effective, primarily by altering the normal balance of immunoregulatory TH1 and TH2 cells in the body.
• Impaired function of HIV infected macrophages and dendritic cells.
To further complicate problems, during the replication of HIV the reverse transcriptase of HIV exhibits a high error rate as it transcribes the RNA genome into DNA. As a result, HIV readily mutates to become more immunoresistant, more drug resistant, and able to change the preferred cell type it is able to infect, e.g., M-tropic to T-tropic as shown in Figure \(2\).
Progression to AIDS is marked by a viral load that progressively increases in number while the immune system weakens as a result of the destruction of increasing numbers of T4-lymphocytes and the inability of the body to continually replace these destroyed cells. The loss of T4-helper lymphocytes leads to a marked decline in cells called cytotoxic T-lymphocytes (CTLs), the primary cells the body's immune responses use to destroy virus-infected cells. Once a person progresses to full-blown AIDS he or she becomes susceptible to a variety of opportunistic infections by:
• bacteria such as Mycobacterium avium complex (MAC), Salmonella, and Nocardia;
• protozoa such as Cryptosporidium and Toxoplasma;
• viruses such as cytomegalovirus (CMV), herpes simplex viruses types 1 and 2 (HSV-1, HSV-2), and varicella zoster virus (VZV);
• Candida, Cryptococcus, Coccidioides, Histoplasma, and Pneumocystis.
There is also an increased incidence of tumors, such Epstein-Barr virus-associated B-cell lymphomas, other lymphomas, cervical cancer, and Kaposi’s sarcoma. Wasting syndrome and encephalopathy are also common.
Summary
A secondary immunodeficiency is one in which a person is born with normal immune responses but some secondary factor or occurrence causes a decrease in immune responses. Causes of secondary immunodeficiencies include malnutrition, some viruses such as HIV, irradiation, cytotoxic drugs used in cancer chemotherapy, anti-inflammatory steroids, leukemias, aging, and removal of the spleen. HIV infects and destroys T4-lymphocytes and when the body becomes unable to replace the T4-lymphocytes as fast as they are being destroyed, secondary immunodeficiency results.
15.E: Immunodeficiency (Exercises)
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied.
15.1: Primary Immunodeficiency
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following:
_____ Rare but severe primary immunodeficiencies occuring as the result of a rare recessive genetic defect in the immune responses that involves the development of B-lymphocytes, T-lymphocytes, or both and results in multiple, recurrent infections during infancy. (ans)
_____ Common, less severe primary immunodeficiencies involving just one or more of the huge number of genes involved in the immune responses. They involve the decreased ability to combat just a single type of infection or a narrow range of infections and relate to an individual’s own unique genetics. (ans)
_____ There may be greatly decreased humoral immunity but cell-mediated immunity remains normal. X-linked agammaglobulinemia and selective IgA deficiency are examples. May be treated with artificially-acquired passive immunization. (ans)
_____ Primary immunodeficiencies that affect both humoral immunity and cell-mediated immunity. There is a defect in both B-lymphocytes and T-lymphocytes, or just T-lymphocytes in which case the humoral deficiency is due to the lack of T4-helper lymphocytes. (ans)
1. B-lymphocyte disorder
2. combined B-lymphocyte and T-lymphocyte disorder
3. novel primary immunodeficiency
4. conventional primary immunodeficiency
2. Infants born with a nonfunctional thymus develop frequent and severe infections. Explain. (ans)
15.2: Secondary Immunodeficiency
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. State what is meant by secondary immunodeficiency and list 4 possible contributing factors. (ans)
2. Briefly give three mechanisms of HIV-induced immunodeficiency. (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/15%3A_Immunodeficiency/15.2%3A_Secondary_Immunodeficiency.txt |
Hypersensitivities are a set of undesirable reactions produced by the normal immune system, including allergies and autoimmunity. These reactions may be damaging, uncomfortable, or occasionally fatal. Hypersensitivity reactions require a pre-sensitized (immune) state of the host. Immediate hypersensitivities refer to humoral immunity (antigen/antibody reactions) causing harm. Delayed hypersensitivities refer to cell-mediated immunity (cytotoxic T-lymphocytes, macrophages, and cytokines) leading to harm.
16: Hypersensitivities
Describe the mechanism for Type I (IgE-mediated) hypersensitivity and give 3 examples. State how they are treated symptomatically. Describe how desensitization (allergy) shots work to lessen the severity of Type I hypersensitivities. Briefly describe how monoclonal antibodies against the Fc portion of IgE may someday be used to prevent Type I allergies. When a person has hay fever, common symptoms include runny eyes, runny nose, swollen sinuses, and difficulty in breathing. In terms of humoral immunity, discuss the mechanism behind these symptoms. Also state the reason for giving antihistamines.
Figure \(4\): Type-I Hypersensitivity, Step-4 . The next time the allergen enters the body, it cross-links the Fab portions of the IgE bound to the mast cell. This triggers the mast cell to degranulate, that is, release its histamine and other inflammatory mediators. The inflammatory mediators are now able to bind to receptors on target cells which leads to dilation of blood vessels, constriction of bronchioles, excessive mucus secretion, and other symptoms of allergy.
Late phase allergic reactions may begin several hours after exposure to antigen. It is thought that basophils play a major role here. Cell-bound IgE on the surface of basophils of sensitive individuals binds a substance called histamine releasing factor (possibly produced by macrophages and B-lymphocytes) causing further histamine release.
The inflammatory agents released or produced cause the following:
1. Dilation of blood vessels. This causes local redness (erythema) at the site of allergen delivery. If dilation is widespread, this can contribute to decreased vascular resistance, a drop in blood pressure, and shock.
2. Increased capillary permeability. This causes swelling of local tissues (edema). If widespread, it can contribute to decreased blood volume and shock.
3. Constriction of bronchial airways. This leads to wheezing and difficulty in breathing.
4. Stimulation of mucous secretion. This leads to congestion of airways.
5. Stimulation of nerve endings. This leads to itching and pain in the skin.
In a systemic anaphylaxis, the allergin is usually picked up by the blood and the reactions occur throughout the body. Examples include severe allergy to insect stings, drugs, and antisera. With a localized anaphylaxis, the allergin is usually found localized in the mucous membranes or the skin. Examples include allergy to hair, pollen, dust, dander, feathers, and food.
Type I hypersensitivity is treated symptomatically with such agents as:
1. Epinephrine. Epinephrine relaxes smooth muscle, constricts blood vessels, and stimulates the heart. It is used for severe systemic reactions.
2. Histamine H1-receptor antagonists. Antihistamines block the binding of histamine to histamine H1-receptors on target cells, e.g., loratadine, fexofenadine, cetirizine.
3. Beta2- agonists. Increase cyclic AMP levels leading to relaxation of bronchial smooth muscles and inhibit mast cell degranulation, e.g., albuterol, salmeterol, formoterol.
4. Leukotriene receptor antagonists. Block smooth muscle constriction, e.g., pranlukast.
5. Sodium cromoglycate. Sodium cromoglycate prevents mast cells from releasing histamines.
6. Nasally administered steroids. Corticosteroids are potent antiinflammatory agents.
Severity may be reduced by desensitization shots (allergy shots). It is thought that when very dilute allergen is given by injection, it stimulates the production of IgG and IgA. IgG and IgA then act as blocking antibodies to bind and neutralize much of the allergen in secretions before it can bind to the deeper cell-bound IgE on the mast cells in the connective tissue. The shots also appear to suppress production of IgE by inducing tolerance and/or by activating T8-suppressor cells.
A new experimental approach to treating and preventing Type-I hypersensitivity involves giving the person with allergies injections of monoclonal antibodies that have been made against the Fc portion of human IgE. This, in turn, blocks the attachment of the IgE to the Fc receptors on mast cells and basophils and the subsequent release of histamine by those cells upon exposure to allergen. In addition, the anti-IgE binds to IgE-producing B-lymphocytes causing apoptosis. The monoclonal antibody is a humanized hybrid molecule consisting of a mouse binding (Fab) portion attached to a human constant (Fc) portion and is known as rhuMab (recombinant human monoclonal antibody).
Summary
1. Immediate hypersensitivities refer to humoral immunity (antigen/antibody reactions) causing harm.
2. During Type I (IgE mediated or anaphylactic-type) hypersensitivity, IgE is made in response to an allergen.
3. In allergic individuals, the levels of IgE may be thousands of times higher than in those without allergies.
4. The Fc portion of IgE binds to the surface of mast cells and basophils and when the allergen subsequently cross-links the Fab portions of the mast cell-bound IgE, this triggers the release of inflammatory mediators such as histamine release by the mast cell, as well as the synthesis of other inflammatory mediators such as platelet-activating factor, leukotrienes, bradykinins, prostaglandins, and cytokines that contribute to inflammation.
5. The inflammatory agents then lead to dilation of blood vessels (redness or erythema, increased capillary permeability (swelling or edema), constriction of bronchial airways (wheezing and difficulty in breathing), stimulation of mucous secretion (congestion of airways), and stimulation of nerve endings (itching and pain in the skin).
6. In a systemic anaphylaxis, the allergin is usually picked up by the blood and the reactions occur throughout the body and can lead to shock. Examples include severe allergy to insect stings, drugs, and antisera.
7. With a localized anaphylaxis, the allergin is usually found localized in the mucous membranes or the skin. Examples include allergy to hair, pollen, dust, dander, feathers, and food.
8. Type I hypersensitivity is treated symptomatically with anti-inflammatory agents such antihistamines and epinephrine.
9. Desensitization shots (allergy shots) are thought to stimulate the production of IgG and IgA which then act as blocking antibodies to bind and neutralize much of the allergen in secretions before it can bind to the deeper cell-bound IgE on the mast cells in the connective tissue.
10. Monoclonal antibodies that have been made against the Fc portion of human IgE have also been used in treatment. They block the attachment of the IgE to the Fc receptors on mast cells and basophils and the subsequent release of histamine by those cells upon exposure to allergen.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe the mechanism for Type I (IgE-mediated) hypersensitivity and give two examples. State how they are treated symptomatically. (ans)
2. When a person has hay fever, common symptoms include runny eyes, runny nose, swollen sinuses, and difficulty in breathing. In terms of humoral immunity, discuss the mechanism behind these symptoms. Also state the reason for giving antihistamines and describe how allergy shots may lessen the severity of this type of hypersensitivity. (ans)
3. Researchers are hoping that the injection of monoclonal antibodies against IgE may someday be used to prevent virtually any Type I hypersensitivity. Explain. (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/16%3A_Hypersensitivities/16.1%3A_Immediate_Hypersensitivities%3A_Type_I.txt |
Describe the mechanism for Type II (antibody-dependent cytotoxicity) hypersensitivity and give 2 examples.
Mechanism: Either IgG or IgM is made against normal self antigens as a result of a failure in immune tolerance , or a foreign antigen resembling some molecule on the surface of host cells enters the body and IgG or IgM made against that antigen then cross reacts with the host cell surface. The binding of these antibodies to the surface of host cells then leads to:
1. Opsonization of the host cells whereby phagocytes stick to host cells by way of IgG, C3b, or C4b and discharge their lysosomes (see Figure \(1\) and Figure \(2\));
1. Activation of the classical complement pathway causing MAC lysis of the cells (see Figure \(3\) and Figure \(4\)); and
1. ADCC destruction of the host cells whereby NK cells attach to the Fc portion of the antibodies. The NK cell then release pore-forming proteins called perforins and proteolytic enzymes called granzymes. Granzymes pass through the pores and activate the enzymes that lead to apoptosis of the infected cell by means of destruction of its structural cytoskeleton proteins and by chromosomal degradation. (see Figure \(5\) , Figure \(5\)A, and Figure \(6\)).
Examples include:
• AB and Rh blood group reactions;
• autoimmune diseases such as:
• rheumatic fever where antibodies result in joint and heart valve damage;
• idiopathic thrombocytopenia purpura where antibodies result in the destruction of platelets;
• myasthenia gravis where antibodies bind to the acetylcholine receptors on muscle cells causing faulty enervation of muscles;
• Goodpasture's syndrome where antibodies lead to destruction of cells in the kidney;
• multiple sclerosis where antibodies are made against the oligodendroglial cells that make myelin, the protein that forms the myelin sheath that insulates the nerve fiber of neurons in the brain and spinal cord; and
• some drug reactions.
Type II hypersensitivity also participates in early transplant rejections.
Summary
1. During type II (antibody-dependent cytotoxicity) hypersensitivity, either IgG or IgM is made against normal self antigens as a result of a failure in immune tolerance, or a foreign antigen resembling some molecule on the surface of host cells enters the body and IgG or IgM made against that antigen then cross reacts with the host cell surface.
2. The binding of these antibodies to the surface of host cells then leads to opsonization of the host cells, membrane attack complex (MAC) lysis of the cells, and antibody-dependent cellular cytotoxicity (ADCC) destruction of the host cells.
3. Examples include AB and Rh blood group reactions and autoimmune diseases such as rheumatic fever, acute glomerulonephritis, myasthenia gravis, and multiple sclerosis.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe the mechanism for Type II (antibody-dependent cytotoxicity) hypersensitivity and give 2 examples. (ans)
16.3: Immediate Hypersensitivities: Type III
Describe the mechanism for Type III (immune complex-mediated) hypersensitivity and give 2 examples.
Mechanism: This is caused when soluble antigen-antibody (IgG or IgM) complexes, which are normally removed by macrophages in the spleen and liver, form in large amounts and overwhelm the body (see Figure \(1\)). These small complexes lodge in the capillaries, pass between the endothelial cells of blood vessels - especially those in the skin, joints, and kidneys - and become trapped on the surrounding basement membrane beneath these cells (see Figure \(2\)). The antigen/antibody complexes then activate the classical complement pathway (see Figure \(3\)). This may cause:
a. Massive inflammation, due to complement protein C5a triggering mast cells to release inflammatory mediators;
b. Influx of neutrophils, due to complement protein C5a, resulting in neutrophils discharging their lysosomes and causing tissue destruction through extracellular killing and causing further inflammation (see Figure \(4\) and Figure \(5\));
c. MAC lysis of surrounding tissue cells, due to the membrane attack complex, C5b6789n;
d. Aggregation of platelets, resulting in more inflammation and the formation of microthrombi that block capillaries; and
e. Activation of macrophages, resulting in production of inflammatory cytokines and extracellular killing causing tissue destruction.
This can lead to tissue death and hemorrhage.
Examples include:
• serum sickness, a combination type I and type III hypersensitivity;
• autoimmune acute glomerulonephritis;
• rheumatoid arthritis;
• systemic lupus erythematosus;
• some cases of chronic viral hepatitis; and
• the skin lesions of syphilis and leprosy.
Summary
1. Type III (immune complex-mediated) hypersensitivity is caused when soluble antigen-antibody (IgG or IgM) complexes, which are normally removed by macrophages in the spleen and liver, form in large amounts and overwhelm the body.
2. These small complexes lodge in the capillaries, pass between the endothelial cells of blood vessels - especially those in the skin, joints, and kidneys - and become trapped on the surrounding basement membrane beneath these cells.
3. The antigen/antibody complexes then trigger excessive activation of the classical complement pathway leading to a massive inflammatory response, influx of neutrophils with extracellular killing of body tissue, MAC lysis of tissue, and aggregation of platelets and macrophages.
4. Examples include Serum sickness, autoimmune acute glomerulonephritis, rheumatoid arthritis, and systemic lupus erythematosus.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe the mechanism for Type III (immune complex-mediated) hypersensitivity and give 2 examples. (ans)
16.4: Immediate Hypersensitivities - Type V
Describe the mechanism for Type V (Stimulatory) hypersensitivity and give an example.
Type V (Stimulatory Hypersensitivity) invovles making Antibodies are made against a particular hormone receptor on a hormone-producing cell. This leads to the overstimulation of those hormone-producing cells. An example is Graves' disease where antibodies are made against thyroid-stimulating hormone receptors of thyroid cells. The binding of the antibodies to the TSH receptors results in constant stimulation of the thyroid leading to hyperthyroidism.
Summary
1. During type V (stimulatory hypersensitivity) antibodies are made against a particular hormone receptor of a hormone-producing cell leading to the overstimulation of those hormone-producing cells.
2. An example is Graves' disease where antibodies are made against thyroid-stimulating hormone receptors of thyroid cells.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe the mechanism for Type V (Stimulatory) hypersensitivity and give an example. (ans)
16.5: Delayed Hypersensitivities - Type IV
Learning Objectives
1. Describe the mechanism for Type IV (delayed) hypersensitivity and give two examples.
Delayed hypersensitivity is cell-mediated rather than antibody-mediated. The underlying Mechanism of delayed hypersensitivity is the same mechanism as cell-mediated immunity. T8-lymphocytes become sensitized to an antigen and differentiate into cytotoxic T-lymphocytes while effector T4-lymphocytes become sensitized to an antigen and produce cytokines . CTLs, cytokines, eosinophils, and/or macrophages then cause harm rather than benefit (Figure \(1\)).
• CTLs use their TCR/CD8 to bind to peptide epitopes bound to MHC-I on infected cells or normal cells having cross-reacting epitopes and kill them through apoptosis.
• TH1 cells activate macrophages causing the production of inflammatory cytokines and extracellular killing by the macrophages leading to tissue damage.
• TH2 cells produce interleukin-4 (IL-4) and interleukin-5 (IL-5) to promote extracellular killing by eosinophils and causing tissue damage.
Examples include:
• the cell or tissue damage done during diseases like tuberculosis, leprosy, smallpox, measles, herpes infections, candidiasis, and histoplasmosis;
• the skin test reactions seen for tuberculosis and other infections;
• contact dermatitis like poison ivy;
• type-1 insulin-dependent diabetes where CTLs destroy insulin-producing cells;
• multiple sclerosis, where T-lymphocytes and macrophages secrete cytokines that destroy the myelin sheath that insulates the nerve fibers of neurons;
• Crohn’s disease and ulcerative colitis; and
• psoriasis.
Delayed hypersensitivity also plays a major role in chronic transplant rejection as a result of CTL destruction of donor cells (host versus graft rejection) or recipient cells (graft versus host rejection). Immunosuppressive drugs such as cyclosporin A or FK-506 (Tacrolimus) are given in an attempt to prevent rejection. Both of these drugs prevent T-lymphocyte proliferation and differentiation by inhibiting the transcription of IL-2.
Summary
1. During delayed hypersensitivity,T8-lymphocytes become sensitized to an antigen and differentiate into cytotoxic T-lymphocytes (CTLs) while effector T4-lymphocytes become sensitized to an antigen and produce cytokines.
2. CTLs, cytokines, eosinophils, and/or macrophages then cause harm rather than benefit.
3. Examples include the cell or tissue damage done during diseases like tuberculosis, leprosy, smallpox, measles, herpes infections, candidiasis, and histoplasmosis, the skin test reactions seen for tuberculosis and other infections, contact dermatitis like poison ivy, type-1 insulin-dependent diabetes where CTLs destroy insulin-producing cells, multiple sclerosis, where T-lymphocytes and macrophages secrete cytokines that destroy the myelin sheath that insulates the nerve fibers of neurons, and Crohn’s disease and ulcerative colitis.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe the mechanism for Type IV (delayed) hypersensitivity and give 2 examples. (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/16%3A_Hypersensitivities/16.2%3A_Immediate_Hypersensitivities%3A_Type_II.txt |
Learning Objectives
1. Define superantigen.
2. Briefly describe the mechanism by which superantigens cause harm to the body.
3. Name 2 superantigens and give an example of a bacterium that produces each.
As was learned earlier under Bacterial Pathogenicity, superantigens are type I toxins that can trigger a harmful immune response. Exotoxins are toxins, often proteins in nature, secreted from a living bacterium but also released upon bacterial lysis. In addition, some bacteria use a type 3 secretion system or a type 4 secretion system to inject toxins directly into human cells. There are three main types of exotoxins:
1. Superantigens (Type I toxins),
2. Exotoxins that damage host cell membranes (Type II toxins)
3. A-B toxins and other toxin that interfere with host cell function (TypeIII toxins).
We will look at superantigens and their role in hypersensitivity.
Learning Objectives
1. Define superantigen.
2. Briefly describe the mechanism by which superantigens cause harm to the body.
3. Name 2 superantigens and give an example of a bacterium that produces each.
Highlighted Bacterium
1. Read the description of Streptococcus pyogenes and match the bacterium with the description of the organism and the infection it causes.
Superantigens are unusual bacterial toxins that interact with exceedingly large numbers of T4-lymphocytes. They bind to the surface of the target cell but do not enter the cell.
Conventional antigens are engulfed by antigen presenting cells (APCs), degraded into epitopes, bind to the peptide groove of MHC-II molecules, and are put on the surface of the APC (Figure \(1\)). Here they are recognized by specific T4-lymphocytes having a TCR with a corresponding shape (Figure \(2\)).
Superantigens, however, bind directly to the outside of MHC-II molecules and activate large numbers of T4-lymphocytes (Figure \(3\)). This activation of very large numbers of T4-lymphocytes results in the secretion of excessive amounts of a cytokine called interleukin-2 (IL-2) as well as the activation of self-reactive T-lymphocytes. The normal response to a conventional antigen results in the activation of maybe 1 in 10,000 T-lymphocytes; superantigens can activate as many as 1 in 5 T-lymphocytes.
Production of high levels of IL-2 can result in circulation of IL-2 in the blood leading to symptoms such as fever, nausea, vomiting, diarrhea, and malaise. However, excess stimulation of IL-2 secretion can also lead to production of inflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), inflammatory chemokines such as IL-8, and platelet-activating factor (PAF), and can lead to the same endothelial damage, acute respiratory distress syndrome, disseminated intravascular coagulation, shock, and multiple organ system failure seen above with LPS and other bacterial cell wall factors. Activation of self-reactive T-lymphocytes can also lead to autoimmune attack.
The following are examples of superantigens.
1. Toxic shock syndrome toxin-1 (TSST-1), produced by some strains of Staphylococcus aureus. This exotoxin causes toxic shock syndrome (TSS). Excessive cytokine production leads to fever, rash, and shock.
2. Streptococcal pyrogenic exotoxin (Spe), produced by rare invasive strains and scarlet fever strains of Streptococcus pyogenes (the group A beta streptococci). S pyogenes produces a number of SPEs that are cytotoxic, pyrogenic, enhance the lethal effects of endotoxins, and contribute to cytokine-induced inflammatory damage. SPEs are responsible for causing streptococcal toxic shock syndrome (STSS) whereby excessive cytokine production leads to fever, rash, and triggering the shock cascade. The SPEs also appear to be responsible for inducing necrotizing fasciitis, a disease that can destroy the skin, fat, and tissue covering the muscle (the fascia). SPE B is also a precursor for a cysteine protease that can destroy muscles tissue.
Read the description of Streptococcus pyogenes, and be able to match the bacterium with its description on an exam.
1. Staphylococcal enterotoxins (SE), producedby many strains of Staphylococcus aureus. These exotoxins cause staphylococcal food poisoning. Excessive Il-2 production results in fever, nausea, vomiting,and diarrhea. The vomiting may also be due to these toxins stimulating the vagus nerve in the stomach lining that controls vomiting.
2. ETEC enterotoxin, produced by enterotoxogenic E. coli (ETEC), one of the most common causes of traveler's diarrhea.
Exercise: Think-Pair-Share Questions
What is the mechanism by which superantigens ultimately lead to SIRS?
Summary
1. Conventional antigens are only recognized by specific T4-cells having a TCR with a corresponding shape.
2. Superantigens are unusual bacterial toxins that interact with exceedingly large numbers of T4-lymphocytes.
3. Activation of very large numbers of T4-lymphocytes results in the secretion of excessive amounts of a cytokine called interleukin-2 (IL-2).
4. Excess stimulation of IL-2 secretion can also lead to production of inflammatory and can lead to the same endothelial damage, acute respiratory distress syndrome, disseminated intravascular coagulation, shock, and multiple organ system failure seen with PAMP-induced inflammation.
5. Examples of superantigens include toxic shock syndrome toxin-1 (TSST-1), Streptococcal pyrogenic exotoxins (SPE), Staphylococcal enterotoxins (SE), and enterotoxogenic E. coli (ETEC) enterotoxin.
Questions
Study the material in this section and then write out the answers to these questions. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Define superantigen (ans).
2. Briefly describe the mechanism by which superantigens cause harm to the body. (ans)
3. Name 2 superantigens and give an example of a bacterium that produces each.
1. (ans)
2. (ans)
4. Multiple Choice (ans)
16.E: Hypersensitivities (Exercises)
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_6%3A_Adaptive_Immunity/16%3A_Hypersensitivities/16.6%3A_Superantigens.txt |
Thumbnail: Supercoiling of the DNA in E.Coli is credited to Garasone and is licensed via a Creative Commons Attribution 3.0 Czech Republic.
Unit 7: Microbial Genetics and Microbial Metabolism
Thumbnail: Scanning electron microscope image of Vibrio cholerae bacteria, which infect the digestive system. (Public Domain; T.J. Kirn, M.J. Lafferty, C.M.P Sandoe and R.K. Taylor).
17: Bacterial Growth and Energy Production
Bacterial Growth
Bacteria replicate by binary fission, a process by which one bacterium splits into two. Therefore, bacteria increase their numbers by geometric progression whereby their population doubles every generation time.Generation time is the time it takes for a population of bacteria to double in number. For many common bacteria, the generation time is quite short, 20-60 minutes under optimum conditions. For most common pathogens in the body, the generation time is probably closer to 5-10 hours. Because bacteria grow by geometric progression and most have a short generation time, they can astronomically increase their number in a short period of time.
The relationship between the number of bacteria in a population at a given time (Nt), the original number of bacterial cells in the population (No), and the number of divisions those bacteria have undergone during that time (n) can be expressed by the following equation:
$N_t = N_o \times 2^n$
For example, Escherichia coli, under optimum conditions, has a generation time of 20 minutes. If one started with only 10 E. coli (No = 10) and allowed them to grow for 12 hours (n = 36; with a generation time of 20 minutes they would divide 3 times in one hour and 36 times in 12 hours), then plugging the numbers in the formula, the number of bacteria after 12 hours (Nt) would be
$10 \times 2^{36} = N_t = 687,194,767,360\: E.\: coli$
In general it is thought that during DNA replication (discussed in Unit 6), each strand of the replicating bacterial DNA attaches to proteins at what will become the cell division plane. For example, Par proteins function to separate bacterial chromosomes to opposite poles of the cell during cell division. They bind to the origin of replication of the DNA and physically pull or push the chromosomes apart, similar to the mitotic apparatus of eukaryotic cells.
In the center of the bacterium, a group of proteins called Fts (filamentous temperature sensitive) proteins interact to form a ring at the cell division plane. These proteins form the cell division apparatus known as the divisome and are directly involved in bacterial cell division by binary fission (see Figure $1$ and Figure $2$).
• electron micrograph of a divisome: see under Bacterial Cell Division, Jon Beckwith's Lab.
The divisome is responsible for directing the synthesis of new cytoplasmic membrane and new peptidoglycan to form the division septum. The function of a number of divisome proteins have been identified, including:
• MinE: Directs formation of the FtsZ ring and divisome complex at the bacterium's division plane.
• FtsZ: Similar to tubulin in eukaryotic cells, FtsZ forms a constricting ring at the division site. As FtsZ depolymerizes, it directs an inward growth of the cell wall to form the division septum. It is found in both Bacteria and Archaea, as well as in mitochondria and chloroplasts.
• ZipA: A protein that connects the FtsZ ring to the bacterial cytoplasmic membrane.
• FtsA: An ATPase that breaks down ATP to provide energy for cell division and also helps connect the FtsZ ring to the bacterial cytoplasmic membrane.
• FtsK: Helps in separating the replicated bacterial chromosome.
• FtsI: Needed for peptidoglycan synthesis.
- Scanning electron micrograph of dividing Escherichia coli; courtesy of CDC.
- Scanning electron micrograph of dividing Salmonella typhimurium; courtesy of CDC.
- To view an transmission electron micrograph of dividing streptococci, see the Rockefeller University home page.
The Population Growth Curve
Although bacteria are capable of replicating geometrically as a result of binary fission, in reality this only occurs as long as their is space to grow, sufficient nutrients, and a way to dispose of waste products. Because these factors limit the ability to replicate geometrically, over time in a closed growth system a bacterial population usually exhibits a predictable pattern of growth - its growth curve - that follows several stages or phases:
1. The lag phase
During the lag phase growth is relatively flat and the population appears either not to be growing or growing quite slowly (see Figure $3$). During this phase the newly inoculated cells are adapting to their new environment and synthesizing the molecules they will need in order to grow rapidly.
2. The exponential growth phase (also called the logarithmic or log phase)
This is the phase where the population increases geometrically as long as there is sufficient food and space for growth (see Figure $3$).
3. The stationary growth phase
Here the population grows slowly or stops growing (see Figure $3$) because of decreasing food, increasing waste, and lack of space. The rate of replication is balanced out by the rate of inhibition or death.
4. The decline or death phase
Here the population dies exponentially from the accumulation of waste products (see Figure $3$), although the rate of death depends on the degree of toxicity and the resistance of the species and viable cells may remain for weeks to months.
Summary
1. Bacteria replicate by binary fission, a process by which one bacterium splits into two.
2. Generation time is the time it takes for a population of bacteria to double in number. For many bacteria the generation time ranges from minutes to hours.
3. Because of binary fission, bacteria increase their numbers by geometric progression whereby their population doubles every generation time.
4. Par proteins function to separate bacterial chromosomes to opposite poles of the cell during bacterial cell division.
5. The bacterial divisome is responsible for directing the synthesis of new cytoplasmic membrane and new peptidoglycan to form the division septum.
6. Although bacteria are capable of replicating geometrically as a result of binary fission, this only occurs as long as their is space to grow, sufficient nutrients, and a way to dispose of waste products.
7. In a closed growth system, a bacterial population usually exhibits a predictable pattern of growth - its growth curve - that follows several stages or phases.
8. During the lag phase growth is relatively flat and the population appears either not to be growing or growing quite slowly as newly inoculated cells are adapt to their new environment.
9. During the exponential growth phase (log phase) the population increases geometrically as long as there is sufficient food and space for growth.
10. During the stationary phase the population grows slowly or stops growing because of decreasing food, increasing waste, and lack of space.
11. During the death (decline) phase the population dies exponentially from the accumulation of waste products. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/17%3A_Bacterial_Growth_and_Energy_Production/17.1%3A_Bacterial_Growth.txt |
Physical requirements
a. Temperature
Bacteria have a minimum, optimum, and maximum temperature for growth and can be divided into 3 groups based on their optimum growth temperature:
1. Psychrophiles are cold-loving bacteria. Their optimum growth temperature is between -5C and 15C. They are usually found in the Arctic and Antarctic regions and in streams fed by glaciers.
2. Mesophiles are bacteria that grow best at moderate temperatures. Their optimum growth temperature is between 25C and 45C. Most bacteria are mesophilic and include common soil bacteria and bacteria that live in and on the body.
3. Thermophiles are heat-loving bacteria. Their optimum growth temperature is between 45C and 70C and are commonly found in hot springs and in compost heaps.
4. Hyperthermophiles are bacteria that grow at very high temperatures. Their optimum growth temperature is between 70C and 110C. They are usually members of the Archaea and are found growing near hydrothermal vents at great depths in the ocean.
b. Oxygen requirements
Bacteria show a great deal of variation in their requirements for gaseous oxygen. Most can be placed in one of the following groups:
1. Obligate aerobes are organisms that grow only in the presence of oxygen. They obtain their energy through aerobic respiration .
2. Microaerophils are organisms that require a low concentration of oxygen (2% to 10%) for growth, but higher concentrations are inhibitory. They obtain their energy through aerobic respiration .
3. Obligate anaerobes are organisms that grow only in the absence of oxygen and, in fact, are often inhibited or killed by its presence. They obtain their energy through anaerobic respiration or fermentation .
4. Aerotolerant anaerobes , like obligate anaerobes, cannot use oxygen to transform energy but can grow in its presence. They obtain energy only by fermentation and are known as obligate fermenters.
5. Facultative anaerobes are organisms that grow with or without oxygen, but generally better with oxygen. They obtain their energy through aerobic respiration if oxygen is present, but use fermentation or anaerobic respiration if it is absent. Most bacteria are facultative anaerobes.
c. pH
Microorganisms can be placed in one of the following groups based on their optimum pH requirements:
1. Neutrophiles grow best at a pH range of 5 to 8.
2. Acidophiles grow best at a pH below 5.5.
3. Alkaliphiles grow best at a pH above 8.5.
d. Osmosis
Osmosis is the diffusion of water across a membrane from an area of higher water concentration (lower solute concentration) to lower water concentration (higher solute concentration). Osmosis is powered by the potential energy of a concentration gradient and does not require the expenditure of metabolic energy. While water molecules are small enough to pass between the phospholipids in the cytoplasmic membrane, their transport can be enhanced by water transporting transport proteins known as aquaporins . The aquaporins form channels that span the cytoplasmic membrane and transport water in and out of the cytoplasm.
To understand osmosis, one must understand what is meant by a solution . A solution consists of a solute dissolved in a solvent . In terms of osmosis, solute refers to all the molecules or ions dissolved in the water (the solvent). When a solute such as sugar dissolves in water, it forms weak hydrogen bonds with water molecules. While free, unbound water molecules are small enough to pass through membrane pores, water molecules bound to solute are not (see Figure \(4\)C and Figure \(4\)D).Therefore, the higher the solute concentration, the lower the concentration of free water molecules capable of passing through the membrane.
A cell can find itself in one of three environments: isotonic , hypertonic , or hypotonic . (The prefixes iso-, hyper-, and hypo- refer to the solute concentration).
• In an isotonic environment (see Figure \(5\)A), both the water and solute concentration are the same inside and outside the cell and water goes into and out of the cell at an equal rate.
http5 version of animation for iPad showing osmosis in an isotonic environment.
• If the environment is hypertonic (see Figure \(5\)B), the water concentration is greater inside the cell while the solute concentration is higher outside (the interior of the cell is hypotonic to the surrounding hypertonic environment). Water goes out of the cell.
html5 version of animation for iPad showing osmosis in a hypertonic environment.
• In an environment that is hypotonic (see Figure \(5\)C), the water concentration is greater outside the cell and the solute concentration is higher inside (the interior of the cell is hypertonic to the hypotonic surroundings). Water goes into the cell.
html5 version of animation for iPad showing osmosis in a hypotonic environment.
Most bacteria require an isotonic environment or a hypotonic environment for optimum growth. Organisms that can grow at relatively high salt concentration (up to 10%) are said to be osmotolerant . Those that require relatively high salt concentrations for growth, like some of the Archaea that require sodium chloride concentrations of 20 % or higher halophiles .
Nutritional requirements
In addition to a proper physical environment, microorganisms also depend on an available source of chemical nutrients. Microorganisms are often grouped according to their energy source and their source of carbon.
a. Energy source
1. Phototrophs use radiant energy (light) as their primary energy source.
2. Chemotrophs use the oxidation and reduction of chemical compounds as their primary energy source.
b. Carbon source
Carbon is the structural backbone of the organic compounds that make up a living cell. Based on their source of carbon bacteria can be classified as autotrophs or heterotrophs.
1. Autotrophs : require only carbon dioxide as a carbon source. An autotroph can synthesize organic molecules from inorganic nutrients.
2. Heterotrophs : require organic forms of carbon. A Heterotroph cannot synthesize organic molecules from inorganic nutrients.
Combining their nutritional patterns, all organisms in nature can be placed into one of four separate groups: photoautotrophs, photoheterotrophs, chemoautotrophs, and chemoheterotrophs.
1. Photoautotrophs use light as an energy source and carbon dioxide as their main carbon source. They include photosynthetic bacteria (green sulfur bacteria, purple sulfur bacteria, and cyanobacteria), algae, and green plants. Photoautotrophs transform carbon dioxide and water into carbohydrates and oxygen gas through photosynthesis .
Cyanobacteria, as well as algae and green plants, use hydrogen atoms from water to reduce carbon dioxide to form carbohydrates, and during this process oxygen gas is given off (an oxygenic process). Other photosynthetic bacteria (the green sulfur bacteria and purple sulfur bacteria) carry out an anoxygenic process, using sulfur, sulfur compounds or hydrogen gas to reduce carbon dioxide and form organic compounds.
2. Photoheterotrophs use light as an energy source but cannot convert carbon dioxide into energy. Instead they use organic compounds as a carbon source. They include the green nonsulfur bacteria and the purple nonsulfur bacteria.
3. Chemolithoautotrophs use inorganic compounds such as hydrogen sulfide, sulfur, ammonia, nitrites, hydrogen gas, or iron as an energy source and carbon dioxide as their main carbon source.
4. Chemooganoheterotrophs use organic compounds as both an energy source and a carbon source. Saprophytes live on dead organic matter while parasites get their nutrients from a living host. Most bacteria, and all protozoans, fungi, and animals are chemoorganoheterotrophs.
c. Nitrogen source
Nitrogen is needed for the synthesis of such molecules as amino acids, DNA, RNA and ATP . Depending on the organism, nitrogen, nitrates, ammonia, or organic nitrogen compounds may be used as a nitrogen source.
d. Minerals
1. Sulfur
Sulfur is needed to synthesizes sulfur-containing amino acids and certain vitamins. Depending on the organism, sulfates, hydrogen sulfide, or sulfur-containing amino acids may be used as a sulfur source.
2. Phosphorus
Phosphorus is needed to synthesize phospholipids , DNA, RNA, and ATP . Phosphate ions are the primary source of phosphorus.
3. Potassium, magnesium, and calcium
These are required for certain enzymes to function as well as additional functions.
4. Iron
Iron is a part of certain enzymes.
5. Trace elements
Trace elements are elements required in very minute amounts, and like potassium, magnesium, calcium, and iron, they usually function as cofactors in enzyme reactions. They include sodium, zinc, copper,molybdenum, manganese, and cobalt ions. Cofactors usually function as electron donors or electron acceptors during enzyme reactions.
f. Growth factors
Growth factors are organic compounds such as amino acids , purines , pyrimidines , and vitamins that a cell must have for growth but cannot synthesize itself. Organisms having complex nutritional requirements and needing many growth factors are said to be fastidious .
Summary
1. Bacteria have a minimum, optimum, and maximum temperature for growth and can be divided into 3 groups based on their optimum growth temperature: psychrophils, mesophils, thermophils, or hyperthermophils.
2. Bacteria show a great deal of variation in their requirements for gaseous oxygen. Most can be placed in one of the following groups: obligate aerobes, microaerophils, obligate anaerobes, aerotolerant anaerobes, or facultative anaerobes.
3. Microorganisms can be placed in one of the following groups based on their optimum pH requirements: neutrophiles, acidophiles, or alkaliphiles.
4. A bacterium's osmotic environment can affect bacterial growth.
5. Bacteria can be grouped according to their energy source as phototrophs or chemotrophs.
6. Bacteria can be grouped according to their carbon source as autotrophs or heterotrophs.
7. Combining their nutritional patterns, all organisms in nature can be placed into one of four separate groups: photoautotrophs, photoheterotrophs, chemoautotrophs, and chemoheterotrophs.
8. Bacteria also need a nitrogen source, various minerals, and water for growth.
9. Organisms having complex nutritional requirements and needing many growth factors are said to be fastidious. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/17%3A_Bacterial_Growth_and_Energy_Production/17.2%3A_Factors_that_Influence_Bacterial_Growth.txt |
State what the letters ADP and ATP stand for and how the two molecules differ. Briefly describe how energy that is released from energy-containing compounds is trapped and stored as ATP and how energy stored in ATP is released to do cellular work.
Adenosine triphosphate (ATP) links most cellular exergonic and endergonic chemical reactions. To obtain energy to do cellular work, organisms take energy-rich compounds such as glucose into the cell and enzymatically break them down to release their potential energy. Therefore, the organism needs a way to trap some of that released energy and store the energy in a form that can be utilized by the cell to do cellular work. Principally, energy is trapped and stored in the form of adenosine triphosphate or ATP.
A tremendous amount of ATP is needed for normal cellular growth. For example,a human at rest uses about 45 kilograms (about 99 pounds) of ATP each day but at any one time has a surplus of less than one gram. It is estimated that each cell will generate and consume approximately 10,000,000 molecules of ATP per second. As can be seen, ATP production is an ongoing cellular process.
To trap energy released from exergonic catabolic chemical reactions, the cell uses some of that released energy to attach an inorganic phosphate group on to adenosine diphosphate (ADP) to make adenosine triphosphate (ATP). Because the phosphate groups are all negatively charged, they repel each other and stress the bond holding them together, much like a bent diving board. Thus, energy is trapped and stored in these stressed bonds known as high-energy phosphate bonds. To obtain energy to do cellular work during endergonic anabolic chemical reactions, the organism enzymatically removes the third phosphate from ATP thus releasing the stored energy and forming ADP and inorganic phosphate once again (see Figure \(1\)).
Depending on the type of organism, cells transfer energy and generate ATP by photophosphorylation, by substrate-level phosphorylation, and/or by oxidative phosphorylation. (Phosphorylation refers to the attachment of a phosphate group to a molecule.)
Summary
1. Cellular energy is primarily trapped and stored in the form of adenosine triphosphate or ATP.
2. A tremendous amount of ATP is needed for normal cellular growth.
3. To trap energy released from exergonic catabolic chemical reactions, the cell uses some of that released energy to attach an inorganic phosphate group on to adenosine diphosphate (ADP) to make adenosine triphosphate (ATP). The energy is stored in these high-energy phosphate bonds.
4. To obtain energy to do cellular work during endergonic anabolic chemical reactions, the organism enzymatically removes the third phosphate from ATP thus releasing the stored energy and forming ADP and inorganic phosphate once again.
5. Depending on the type of organism, cells transfer energy and generate ATP by photophosphorylation, by substrate-level phosphorylation, and/or by oxidative phosphorylation.
Questions
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. The acronym ATP stands for________________. (ans)
2. Describe how cells trap energy released from exergonic catabolic chemical reactions and store it as ATP. (ans)
3. Describe how cells obtain energy to do cellular work during endergonic anabolic chemical reactions. (ans)
4. The hydrolysis of ATP is:
1. an exergonic reaction (ans)
2. an endergonic reaction (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/17%3A_Bacterial_Growth_and_Energy_Production/17.4%3A_Adenosine_Triphosphate_%28ATP%29.txt |
Substrate-Level Phosphorylation
Substrate-level phosphorylation is the production of ATP from ADP by a direct transfer of a high-energy phosphate group from a phosphorylated intermediate metabolic compound in an exergonic catabolic pathway as shown in Figure \(2\). Such intermediate compounds are sometimes called high-energy transfer compounds (HETCs) and several HETCs are found as intermediates during glycolysis and aerobic respiration .
Oxidative Phosphorylation
Oxidative phosphorylation is the production of ATP using energy derived from the transfer of electrons in an electron transport system and occurs by chemiosmosis.
To understand oxidative phosphorylation, it is important to first review the hydrogen atom and the process of oxidation and reduction. An atom of hydrogen contains only one proton (H+) and one electron (e-). Therefore, the term proton and the term hydrogen ion (H+) are interchangeable. Also remember that electrons have stored energy, or potential energy, ready to do work and when an atom or molecule loses that electron (becomes oxidized) that energy is released and able to do cellular work.
Oxidation-reduction reactions are coupled chemical reactions in which one atom or molecule loses one or more electrons (oxidation ) while another atom or molecule gains those electrons (reduction ). The compound that loses electrons becomes oxidized; the compound that gains those electrons becomes reduced. In covalent compounds, however, it is usually easier to lose a whole hydrogen (H) atom - a proton and an electron - rather than just an electron. An oxidation reaction during which both a proton and an electron are lost is called dehydrogenation . A reduction reaction during which both a proton and an electron are gained is called hydrogenation .
Cells use specific molecules to carry the electrons that are removed during the oxidation of an energy source. These molecules are called electron carriers and they alternately become oxidized and reduced during electron and proton transfer. These include three freely diffusible coenzymes known as NAD+, FAD, and NADP+. The reduced forms of these coenzymes (NADH, FADH2, and NADPH) have reducing power because their bonds contain a form of usable energy.
• NAD+ , or nicotinamide adenine dinucleotide, is a coenzyme that often works in conjunction with an enzyme called a dehydrogenase. The enzyme removes two hydrogen atoms (2H+ and 2e-) from its substrate. Both electrons but only one proton are accepted by the NAD+ to produce its reduced form, NADH, plus H+. NADH is used to generate proton motive force (discussed below) that can drive the synthesis of ATP.
• FAD , or flavin adenine dinucleotide, is a coenzyme that also works in conjunction with an enzyme called a dehydrogenase. The enzyme removes two hydrogen atoms (2H+ and 2e-) from its substrate. Both electrons and both protons are accepted by the FAD to produce its reduced form, FADH2. FADH2 is used to generate proton motive force (discussed below) that can drive the synthesis of ATP.
• NADP+, or nicotinamide adenine dinucleotide phosphate, is a coenzyme that uses dehydrogenase to remove two hydrogen atoms (2H+ and 2e-) from its substrate. Both electrons but only one proton are accepted by the NADP+ to produce its reduced form, NADPH, plus H+. NADPH is not used for ATP synthesis but its electrons provide the energy for certain biosynthesis reactions such as ones involved in photosynthesis.
During the process of aerobic respiration, discussed in the next section, coupled oxidation-reduction reactions and electron carriers are often part of what is called an electron transport chain , a series of electron carriers that eventually transfers electrons from NADH and FADH2 to oxygen. The diffusible electron carriers NADH and FADH2 carry hydrogen atoms (protons and electrons) from substrates in exergonic catabolic pathways such as glycolysis and the citric acid cycle to other electron carriers that are embedded in membranes. These membrane-associated electron carriers include flavoproteins, iron-sulfur proteins, quinones, and cytochromes. The last electron carrier in the electron transport chain transfers the electrons to the terminal electron acceptor, oxygen.
The chemiosmotic theory explains the functioning of electron transport chains. According to this theory, the transfer of electrons down an electron transport system through a series of oxidation-reduction reactions releases energy (Figure \(1\)). This energy allows certain carriers in the chain to transport hydrogen ions (H+ or protons) across a membrane.
Depending on the type of cell, the electron transport chain may be found in the cytoplasmic membrane, the inner membrane of mitochondria, and the inner membrane of chloroplasts.
• In prokaryotic cells, the protons are transported from the cytoplasm of the bacterium across the cytoplasmic membrane to the periplasmic space located between the cytoplasmic membrane and the cell wall.
• In eukaryotic cells, protons are transported from the matrix of the mitochondria across the inner mitochondrial membrane to the intermembrane space located between the inner and outer mitochondrial membranes.
• In plant cells and the cells of algae, protons are transported from the stroma of the chloroplast across the thylakoid membrane into the interior space of the thylakoid.
As the hydrogen ions accumulate on one side of a membrane, the concentration of hydrogen ions creates an electrochemical gradient or potential difference (voltage) across the membrane. (The fluid on the side of the membrane where the protons accumulate acquires a positive charge; the fluid on the opposite side of the membrane is left with a negative charge.) The energized state of the membrane as a result of this charge separation is called proton motive force or PMF.
This proton motive force provides the energy necessary for enzymes called ATP synthases (Figure \(5\)), also located in the membranes mentioned above, to catalyze the synthesis of ATP from ADP and phosphate. This generation of ATP occurs as the protons cross the membrane through the ATP synthase complexes and re-enter either the bacterial cytoplasm (Figure \(5\)), the matrix of the mitochondria, or the stroma of the chloroplasts. As the protons move down the concentration gradient through the ATP synthase, the energy released causes the rotor and rod of the ATP synthase to rotate. The mechanical energy from this rotation is converted into chemical energy as phosphate is added to ADP to form ATP.
Proton motive force is also used to transport substances across membranes during active transport and to rotate bacterial flagella.
At the end of the electron transport chain involved in aerobic respiration, the last electron carrier in the membrane transfers 2 electrons to half an oxygen molecule (an oxygen atom) that simultaneously combines with 2 protons from the surrounding medium to produce water as an end product (Figure \(3\)). The electron transport chains involved in photosynthesis ultimately transfer 2 electrons to NADP+ that simultaneously combines with 2 protons from the surrounding medium to produce NADPH.
Summary
1. Photophosphorylation uses the radiant energy of the sun to drive the synthesis of ATP.
2. This is a process seen only in cells capable of photosynthesis.
3. Substrate-level phosphorylation is the production of ATP from ADP by a direct transfer of a high-energy phosphate group from a phosphorylated intermediate metabolic compound in an exergonic catabolic pathway.
1. Oxidative phosphorylation is the production of ATP using energy derived from the transfer of electrons in an electron transport system and occurs by chemiosmosis.
2. An atom of hydrogen contains only one proton (H+) and one electron.
3. Electrons have stored energy, or potential energy, ready to do work. When an atom or molecule loses that electron (becomes oxidized) that energy is released and able to do cellular work.
4. Oxidation-reduction reactions are coupled chemical reactions in which one atom or molecule loses one or more electrons (oxidation) while another atom or molecule gains those electrons (reduction).
5. An oxidation reaction during which both a proton and an electron are lost is called dehydrogenation.
6. A reduction reaction during which both a proton and an electron are gained is called hydrogenation.
7. Cells use specific molecules such as NAD+, FAD, and NADP+ to carry the electrons that are removed during the oxidation of an energy source. These molecules are called electron carriers and they alternately become oxidized and reduced during electron and proton transfer.
8. Coupled oxidation-reduction reactions and electron carriers are often part of what is called an electron transport chain.
9. The chemiosmotic theory explains the functioning of electron transport chains. According to this theory, the transfer of electrons down an electron transport system through a series of oxidation-reduction reactions releases energy. This energy allows certain carriers in the chain to transport hydrogen ions (H+ or protons) across a membrane.
10. In prokaryotic cells, the protons are transported from the cytoplasm of the bacterium across the cytoplasmic membrane to the periplasmic space located between the cytoplasmic membrane and the cell wall; in eukaryotic cells, protons are transported from the matrix of the mitochondria across the inner mitochondrial membrane to the intermembrane space located between the inner and outer mitochondrial membranes; in plant cells and the cells of algae, protons are transported from the stroma of the chloroplast across the thylakoid membrane into the interior space of the thylakoid.
11. As the hydrogen ions accumulate on one side of a membrane, the concentration of hydrogen ions creates an electrochemical gradient or potential difference (voltage) across the membrane called proton motive force (PMF).
12. This proton motive force provides the energy necessary for enzymes called ATP synthases to catalyze the synthesis of ATP from ADP and phosphate.
Questions
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Define photophosphorylation. (ans)
2. Briefly describe the process of substrate-level phosphorylation. (ans)
3. Briefly describe the process of oxidative phosphorylation. (ans)
4. Another name for a hydrogen ion (H+) is: (ans)
5. An atom or molecule gains an electron. This best describes:
1. oxidation (ans)
2. reduction (ans)
3. dehydrogenation (ans)
4. hydrogenation (ans)
6. When a molecule gains electrons or both protons and electrons, we say it becomes:
1. oxidized (ans)
2. reduced (ans)
7. Cells use specific molecules to carry the electrons that are removed during the oxidation of an energy source. These molecules are called electron carriers and they alternately become oxidized and reduced during electron and proton transfer. Name three freely diffusible coenzymes and give both their oxidized and reduced state. (ans)
8. A coenzyme that often works in conjunction with an enzyme called a dehydrogenase. The enzyme removes two hydrogen atoms (2H+ and 2e-) from its substrate. Both electrons but only one proton are accepted to produce its reduced form that is used to generate proton motive force for driving the synthesis of ATP. This best describes:
1. NAD+(ans)
2. FAD (ans)
3. NADP+ (ans)
9. NADH + H+ is the ________________ form of NAD+. (ans)
10. Describe an electron transport chain. (ans)
11. Based on the chemiosmotic theory, briefly describe proton motive force and how it develops within a cell. (ans)
12. Based on the chemiosmotic theory, briefly describe how proton motive force leads to the generation of ATP. (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/17%3A_Bacterial_Growth_and_Energy_Production/17.5%3A_Phosphorylation_Mechanisms_for_Generating_ATP.txt |
Describe the relationship between photosynthesis and aerobic respiration and relate this to the first law of thermodynamics.
For the vast majority of life on earth, the flow of energy begins with sunlight and involves a cycle involving photoautotrophs and chemoorganoheterotrophs. Photoautotrophs use sunlight as a source of energy and through the process of photosynthesis, reduce carbon dioxide to form carbohydrates such as glucose. The radient energy is converted to the chemical bond energy within glucose.
The overall reaction for photosynthesis (in the presence of light and chlorophyll) is as follows:
$6CO_2 + 6H_2O \rightarrow C_6H_{12}O_6 + 6O_2$
Note that carbon dioxide (CO2) is reduced to produce glucose (C6H12O6 ) and water (H2O) is oxidized to produce oxygen (O2).
Both chemoorganoheterotrophs and photoautotrophs then convert the chemical bond energy of glucose to the chemical bond energy of ATP, the form of energy required to do most cellular work. This is done through the process called aerobic respiration.
The overall reaction for aerobic respiration is:
C6H12O6 + 6O2 yields 6CO2 + 6H2O + energy (as ATP)
Note that glucose (C6H12O6 ) is oxidized to produce carbon dioxide (CO2) and oxygen (O2) is reduced to produce water (H2O).
As can be seen, the end products for aerobic respiration, carbon dioxide and water, are the reactants for photosynthesis while the end products of photosynthesis, glucose and oxygen, are the reactants for aerobic respiration. In other words, the nutrients are continuously recycled between the two processes. Energy, however, is converted from one form to another: from radiant energy to the chemical bond energy of glucose to the chemical bond energy of ATP, the first law of thermodynamics.
Summary
1. The overall reaction for photosynthesis is 6CO2 + 6H2O in the presence of light and chlorophyll yields C6H12O6 + 6O2.
2. The overall reaction for aerobic respiration is C6H12O6 + 6O2 yields 6CO2 + 6H2O + energy (as ATP).
3. As can be seen, the end products for aerobic respiration, carbon dioxide and water, are the reactants for photosynthesis while the end products of photosynthesis, glucose and oxygen, are the reactants for aerobic respiration. In other words, the nutrients are continuously recycled between the two processes.
4. Energy, however, is not recycled but rather is converted from one form to another: from radiant energy to the chemical bond energy of glucose to the chemical bond energy of ATP.
Questions
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. 6CO2 + 6H2O are the reactants for ____________________ and the products of ___________________.
1. aerobic respiration / photosynthesis (ans)
2. photosynthesis / aerobic respiration (ans)
2. C6H12O6 + 6O2 are the reactants for ____________________ and the products of ___________________.
1. aerobic respiration / photosynthesis (ans)
2. photosynthesis / aerobic respiration (ans)
17.E: Bacterial Growth and Energy Production (Exercises)
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied.
17.1: Bacterial Growth
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following:
_____ A population doubles every generation time. (ans)
_____ One cell splits in two. (ans)
_____ The time it takes for a population of organisms to double in number. (ans)
1. binary fission
2. generation time
3. geometric progression
2. If you started with 1000 E. coli with a generation time of 30 minutes, how many bacteria would you have after 3 hours? (ans)
3. Match the following:
_____ Phase where the population grows slowly or stops growing because of decreasing food, increasing waste, and lack of space. The rate of replication is balanced out by the rate of inhibition or death. (ans)
_____ Phase where the population dies exponentially from the accumulation of waste products, although the rate of death depends on the degree of toxicity and the resistance of the species. (ans)
_____ Phase where growth is relatively flat and the population appears either not to be growing or growing quite slowly. During this phase the newly inoculated cells are adapting to their new environment and synthesizing the molecules they will need in order to grow rapidly. (ans)
_____ Phase where the population increases geometrically as long as there is sufficient food and space for growth. (ans)
1. Lag phase
2. Exponential (log) growth phase
3. Stationary phase
4. Death (decline) phase
17.2: Factors that Influence Bacterial Growth
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Matching
_____ Bacteria that grow best at moderate temperatures. Their optimum growth temperature is between 25C and 45C. (ans)
_____ Cold-loving bacteria. Their optimum growth temperature is between -5C and 15C. They are usually found in the Arctic and Antarctic regions and in streams fed by glaciers. (ans)
_____ Organisms that grow with or without oxygen, but generally better with oxygen. (ans)
_____ Organisms that grow onlyin the absense of oxygen and, in fact, are often inhibited or killed by its presense. (ans)
_____ An environment where the water concentration is greater outside the cell and the solute concentration is higher inside. Water goes into the cell. (ans)
_____ Organisms that use the oxidation and reduction of chemical compounds as their primary energy source. (ans)
_____ Organisms that use light as an energy source and carbon dioxideas their main carbon source. (ans)
_____ Organisms that use organic compounds as both an energy source and a carbon source. (ans)
_____ Organisms that use lightas an energy source but cannot convert carbon dioxide into energy. Instead they use organic compounds as a carbon source. (ans)
1. photoautotrophs
2. photoheterotrophs
3. chemolithoautotrophs
4. chemooganoheterotrophs
5. phototroph
6. heterotroph
7. hypertonic
8. hypotonic
9. obligate aerobe
10. facultative anaerobe
11. obligate anaerobe
12. psychrophile
13. mesophile
14. thermophile
17.4: Cellular Respiration
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. An exergonic processes by which energy released by the breakdown of organic compounds such as glucose can be used to synthesize ATP, the form of energy required to do cellular work. This best describes:
1. anabolism (ans)
2. catabolism (ans)
2. Intermediate molecules that link catabolic and anabolic pathways; can be either oxidized to generate ATP or can be used to synthesize macromolecular subunits such as amino acids, lipids, and nucleotides. (ans)
3. Define cellular respiration. (ans)
4. Pathways that do not require oxygen are said to be:
1. aerobic (ans)
2. anaerobic (ans)
5. Name an exergonic pathway that requires molecular oxygen (O2). (ans)
6. Name two anaerobic exergonic forms of cellular respiration. (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/17%3A_Bacterial_Growth_and_Energy_Production/17.6%3A_The_Flow_of_Energy_in_Nature.txt |
Thumbnail: Plant cells (bounded by purple walls) filled with chloroplasts (green), which are the site of photosynthesis. (CC BY-SA 3.0; Kristian Peters).
18: Microbial Metabolism
Define catabolism and anabolism and state which is exergonic and which is endergonic. Define precursor metabolites and state their functions in metabolism. Define the following: cellular respiration aerobic anaerobic Name one aerobic and two anaerobic forms of cellular respiration.
As mentioned previously, to grow, function, and reproduce, cells must synthesize new cellular components such as cell walls, cell membranes, nucleic acids, ribosomes, proteins, flagella, etc., and harvest energy and convert it into a form that is usable to do cellular work.
Catabolism refers to the exergonic process by which energy released by the breakdown of organic compounds such as glucose can be used to synthesize ATP, the form of energy required to do cellular work. Anabolism is the endergonic process that uses the energy stored in ATP to synthesize the building blocks of the macromolecules that make up the cell. As can be seen, these two metabolic processes are closely linked. Another factor that links catabolic and anabolic pathways is the generation of precursor metabolites. Precursor metabolites are intermediate molecules in catabolic and anabolic pathways that can be either oxidized to generate ATP or can be used to synthesize macromolecular subunits such as amino acids, lipids, and nucleotides.
In this section we will concentrate primarily on harvesting energy and converting it to energy stored in ATP through the process of cellular respiration, but we will also look at some of the key precursor metabolites that are produced during this process.
Cellular respiration is the process cells use to convert the energy in the chemical bonds of nutrients to ATP energy. Depending on the organism, cellular respiration can be aerobic, anaerobic, or both. Aerobic respiration is an exergonic pathway that requires molecular oxygen (O2). Anaerobic exergonic pathways do not require oxygen and include anaerobic respiration and fermentation. We will now look at these three pathways.
Summary
1. Catabolism refers to the exergonic process by which energy released by the breakdown of organic compounds such as glucose can be used to synthesize ATP, the form of energy required to do cellular work.
2. Anabolism is the endergonic process that uses the energy stored in ATP to synthesize the building blocks of the macromolecules that make up the cell.
3. Precursor metabolites are intermediate molecules in catabolic and anabolic pathways that can be either oxidized to generate ATP or can be used to synthesize macromolecular subunits such as amino acids, lipids, and nucleotides.
4. Cellular respiration is the process cells use to convert the energy in the chemical bonds of nutrients to ATP energy.
5. Aerobic respiration is an exergonic pathway that requires molecular oxygen (O2).
6. Anaerobic exergonic pathways do not require oxygen and include anaerobic respiration and fermentation.
18.3A: Glycolysis
Define aerobic respiration. Give the overall chemical reaction for aerobic respiration. Name the four stages of aerobic respiration.
Aerobic respiration is the aerobic catabolism of nutrients to carbon dioxide, water, and energy, and involves an electron transport system in which molecular oxygen is the final electron acceptor. Most eukaryotes and prokaryotes use aerobic respiration to obtain energy from glucose. The overall reaction is:
$C_6H_{12}O_6 + 6O_2 \rightarrow 6CO_2 + 6H_2O \label{1}$
Note that glucose ($C_6H_{12}O_6$) is oxidized to produce carbon dioxide ($CO_2$) and oxygen ($O_2$) is reduced to produce water ($H_2O$). This reaction is a strongly driven reactions and "releases" energy as ATP molecules. This type of ATP production is seen in aerobes and facultative anaerobes. Obligate aerobes are organisms that require molecular oxygen because they produce ATP only by aerobic respiration. Facultative anaerobes, on the other hand are capable of aerobic respiration but can switch to fermentation, an anaerobic ATP-producing process, if oxygen is unavailable.
Aerobic respiration involves four stages:
• glycolysis,
• a transition reaction that forms acetyl coenzyme A,
• the citric acid (Krebs) cycle, and an electron transport chain and
• chemiosmosis.
We will now look at each of these stages.
Summary
1. Aerobic respiration is the aerobic catabolism of nutrients to carbon dioxide, water, and energy, and involves an electron transport system in which molecular oxygen is the final electron acceptor.
2. The overall reaction is: C6H12O6 + 6O2 yields 6CO2 + 6H2O + energy (as ATP). Glucose (C6H12O6 ) is oxidized to produce carbon dioxide (CO2) and oxygen (O2) is reduced to produce water (H2O).
3. This type of ATP production is seen in aerobes and facultative anaerobes.
4. Aerobic respiration involves four stages: glycolysis, a transition reaction that forms acetyl coenzyme A, the citric acid (Krebs) cycle, and an electron transport chain and chemiosmosis.
18.3: Aerobic Respiration
Learning Objectives
1. Briefly describethe function of glycolysis during aerobic respiration and indicate the reactants and products.
2. State whether or not glycolysis requires oxygen.
3. Compare where glycolysis occurs in prokaryotic cells and in eukaryotic cells.
4. State whether steps 1 and 3 of glycolysis are exergonic or endergonic and indicate why.
5. State why one molecule of glucose is able to produce two molecules of pyruvate during glycolysis.
6. Define substrate-level phosphorylation.
7. State the total number and the net number of ATP produced by substrate-level phosphorylation during glycolysis.
8. During aerobic respiration, state what happens to the 2 NADH produced during glycolysis.
9. During aerobic respiration, state what happens to the two molecules of pyruvate produced during glycolysis.
Glycolysis is a partial breakdown of a six-carbon glucose molecule into two, three-carbon molecules of pyruvate, 2NADH +2H+, and 2 net ATP as a result of substrate-level phosphorylation, as shown in (Figures 1 and 2).
Steps of Glycolysis
1. A phosphate from the hydrolysis of a molecule of ATP is added to glucose, a 6-carbon sugar, to form glucose 6-phosphate.
2. The glucose 6-phosphate molecule is rearranged into an isomer called fructose 6-phosphate.
3. A second phosphate provided by the hydrolysis of a second molecule of ATP is added to the fructose 6-phosphate to form fructose 1,
4. The 6-carbon fructose 1,6-biphosphate is split into two molecules of glyceraldehyde 3-phosphate, a 3-carbon molecule.
5. Oxidation and phosphorylation of each glyceraldehyde 3-phosphate produces 1,3-biphosphoglycerate with a high-energy phosphate bond (wavy red line) and NADH.
6. Through substrate-level phosphorylation, the high-energy phosphate is removed from each 1,3-biphosphoglycerate and transferred to ADP forming ATP and 3-phosphoglycerate.
7. Each 3-phosphoglycerate is oxidized to form a molecule of phosphoenolpyruvate with a high-energy phosphate bond.
8. Through substrate-level phosphorylation, the high-energy phosphate is removed from each phosphoenolpyruvate and transferred to ADP forming ATP and pyruvate.
In summary, one molecule of glucose produces two net ATPs (two ATPs were used at the beginning; four ATPs were produced through substrate-level phosphorylation), two molecules of NADH + 2H+, and two molecules of pyruvate.
Glycolysis occurs in the cytoplasm of the cell. The overall reaction is:
$glucose (6C) + 2 NAD+ 2 ADP + 2 inorganic phosphates (P_i)$
$\rightarrow 2 pyruvate (3C) + 2 NADH + 2 H^+ + 2 ATP$
Glycolysis also produces a number of key precursor metabolites, as shown in Figure $3$. Glycolysis does not require oxygen and can occur under aerobic and anaerobic conditions. However, during aerobic respiration, the two reduced NADH molecules transfer protons and electrons to the electron transport chain to generate additional ATPs by way of oxidative phosphorylation.
The glycolysis pathway involves 9 distinct steps, each catalyzed by a unique enzyme. You are not responsible for knowing the chemical structures or enzymes involved in the steps below. They are included to help illustrate how the molecules in the pathway are manipulated by the enzymes in order to to achieve the required products.
Step 1
To initiate glycolysis in eukaryotic cells (Figure $4$), a molecule of ATP is hydrolyzed to transfer a phosphate group to the number 6 carbon of glucose to produce glucose 6-phosphate. In prokaryotes, the conversion of phosphoenolpyruvate (PEP) to pyruvate provides the energy to transport glucose across the cytoplasmic membrane and, in the process, adds a phosphate group to glucose producing glucose 6-phosphate.
Step 2
The glucose 6-phosphate is rearranged to an isomeric form called fructose 6-phosphate (Figure $5$).
Step 3
A second molecule of ATP is hydrolyzed to transfer a phosphate group to the number 1 carbon of fructose 6-phosphate to produce fructose 1,6-biphosphate (Figure $6$).
Step 4
The 6-carbon fructose 1,6 biphosphate is split to form two, 3-carbon molecules: glyceraldehyde 3-phosphate and dihydroxyacetone phosphate. The dihydroxyacetone phosphate is then converted into a second molecule of glyceraldehyde 3-phosphate (Figure $7$). Two molecules of glyceraldehyde 3-phosphate will now go through each of the remaining steps in glycolysis producing two molecules of each product.
Step 5
As each of the two molecules of glyceraldehyde 3-phosphate are oxidized, the energy released is used to add an inorganic phosphate group to form two molecules of 1,3-biphosphoglycerate, each containing a high-energy phosphate bond. During these oxidations, two molecules of NAD+ are reduced to form 2NADH + 2H+ (Figure $8$). During aerobic respiration, the 2NADH + 2H+ carry protons and electrons to the electron transport chain to generate additional ATP by oxidative phosphorylation.
Step 6
As each of the two molecules of 1,3-biphosphoglycerate are converted to 3-phosphoglycerate, the high-energy phosphate group is added to ADP producing 2 ATP by substrate-level phosphorylation, a shown in Figure $9$.
Step 7
The two molecules of 3-phosphoglycerate are rearranged to form two molecules of 2-phosphoglycerate (Figure $10$).
Step 8
Water is removed from each of the two molecules of 2-phosphoglycerate converting the phosphate bonds to a high-energy phosphate bonds as two molecules of phosphoenolpyruvate are produced (Figure $11$).
Step 9
As the two molecules of phosphoenolpyruvate are converted to two molecules of pyruvate, the high-energy phosphate groups are added to ADP producing 2 ATP by substrate-level phosphorylation, a shown in Figure $12$.
Through an intermediate step called the transition reaction, the two molecules of pyruvate then enter the citric acid cycle to be further broken down and generate more ATPs by oxidative phosphorylation.
Overview
Glycolysis is a partial breakdown of a six-carbon glucose molecule into two, three-carbon molecules of pyruvate, 2NADH +2H+, and 2 net ATP as a result of substrate-level phosphorylation. Glycolysis occurs in the cytoplasm of the cell.
The overall Glycolysis reaction is:
glucose (6C) + 2 NAD+ 2 ADP +2 inorganic phosphates (Pi)
yields 2 pyruvate (3C) + 2 NADH + 2 H+ + 2 net ATP
Outside Links
• YouTube movie of Glycolysis: Overview Reaction for Cellular Respiration
• YouTube movie: How Glycolysis Works
Summary
1. Aerobic respiration is the aerobic catabolism of nutrients to carbon dioxide, water, and energy, and involves an electron transport system in which molecular oxygen is the final electron acceptor.
2. Aerobic respiration involves four stages: glycolysis, a transition reaction that forms acetyl coenzyme A, the citric acid (Krebs) cycle, and an electron transport chain and chemiosmosis.
3. Glycolysis is a partial breakdown of a six-carbon glucose molecule into two, three-carbon molecules of pyruvate, 2NADH +2H+, and 2 net ATP as a result of substrate-level phosphorylation.
4. The overall reaction for glycolysis is: glucose (6C) + 2 NAD+ 2 ADP +2 inorganic phosphates (Pi) yields 2 pyruvate (3C) + 2 NADH + 2 H+ + 2 net ATP.
5. Glycolysis does not require oxygen and can occur under aerobic and anaerobic conditions. However, during aerobic respiration, the two reduced NADH molecules transfer protons and electrons to the electron transport chain to generate additional ATPs by way of oxidative phosphorylation.
6. Glycolysis also produces a number of key precursor metabolites.
7. Through an intermediate step called the transition reaction, the two molecules of pyruvate then enter the citric acid cycle to be further broken down and generate more ATPs by oxidative phosphorylation. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/18%3A_Microbial_Metabolism/18.2%3A_Overview_of_Cellular_Respiration.txt |
Learning Objectives
1. Briefly describethe function of transition reaction during aerobic respiration and indicate the reactants and products.
2. During aerobic respiration, state what happens to the 2 NADH produced during the transition reaction.
3. Compare where the transition reaction occurs in prokaryotic cells and in eukaryotic cells.
4. During aerobic respiration, state what happens to the two molecules of Acetyl-CoA produced during the transition reaction.
Formation of Acetyl-CoA through the Transition Reaction
The transition reaction connects glycolysis to the citric acid (Krebs) cycle. Through a process called oxidative decarboxylation, the transition reaction converts the two molecules of the 3-carbon pyruvate from glycolysis (and other pathways) into two molecules of the 2-carbon molecule acetyl Coenzyme A (acetyl-CoA) and 2 molecules of carbon dioxide. First, a carboxyl group of each pyruvate is removed as carbon dioxide and then the remaining acetyl group combines with coenzyme A (CoA) to form acetyl-CoA.
As the two pyruvates undergo oxidative decarboxylation, two molecules of NAD+ become reduced to 2NADH + 2H+ (Figures \(1\) and \(2\)). The 2NADH + 2H+ carry protons and electrons to the electron transport chain to generate additional ATP by oxidative phosphorylation.
The two molecules of acetyl-CoA then enter the citric acid cycle. The 2NADH molecules that are produced carry electrons to the electron transport system for further production of ATPs by oxidative phosphorylation.
The overall reaction for the transition reaction is:
2 pyruvate + 2 NAD+ + 2 coenzyme A
yields 2 acetyl-CoA + 2 NADH + 2 H+ + 2 CO2
In prokaryotic cells, the transition step occurs in the cytoplasm; in eukaryotic cells the pyruvates must first enter the mitochondria because the transition reaction and the citric acid cycle take place in the matrix of the mitochondria.
The two molecules of acetyl-CoA can now enter the citric acid cycle. Acetyl-CoA is also a precursor metabolite for fatty acid synthesis, as shown in Figure \(3\).
Summary
1. Aerobic respiration involves four stages: glycolysis, a transition reaction that forms acetyl coenzyme A, the citric acid (Krebs) cycle, and an electron transport chain and chemiosmosis.
2. The transition reaction connects glycolysis to the citric acid (Krebs) cycle.
3. The transition reaction converts the two molecules of the 3-carbon pyruvate from glycolysis (and other pathways) into two molecules of the 2-carbon molecule acetyl Coenzyme A (acetyl-CoA) and 2 molecules of carbon dioxide.
4. As the two pyruvates undergo oxidative decarboxylation, two molecules of NAD+ become reduced to 2NADH + 2H+ which carry protons and electrons to the electron transport chain to generate additional ATP by oxidative phosphorylation.
5. The overall reaction for the transition reaction is: 2 pyruvate + 2 NAD+ + 2 coenzyme A yields 2 acetyl-CoA + 2 NADH + 2 H+ + 2 CO2.
6. The two molecules of acetyl-CoA can now enter the citric acid cycle.
18.3C: Citric Acid (Krebs) Cycle
Learning Objectives
• State two other names for the citric acid cycle.
• Briefly describethe function of the citric acid cycle during aerobic respiration and indicate the reactants and products.
• Compare where the citric acid cycle occurs in prokaryotic cells and in eukaryotic cells.
• State the total number of ATP produced by substrate-level phosphorylation for each acetyl-CoA that enters the citric acid cycle.
• State the total number of NADH and FADH2 produced for each acetyl-CoA that enters the citric acid cycle.
• During aerobic respiration, state what happens to the NADH and the FADH2 produced during the citric acid cycle.
The citric acid cycle, also known as the tricarboxylic acid cycle and the Krebs cycle, completes the oxidation of glucose by taking the pyruvates from glycolysis (and other pathways), by way of the transition reaction mentioned previously, and completely breaking them down into $CO_2$ molecules, $H_2O$ molecules, and generating additional ATP by oxidative phosphorylation. In prokaryotic cells, the citric acid cycle occurs in the cytoplasm; in eukaryotic cells the citric acid cycle takes place in the matrix of the mitochondria.
The overall reaction for the citric acid cycle is:
$\text{2 acetyl groups} + 6 NAD^+ + 2 FAD + 2 ADP + 2 P_i$
$\rightarrow 4 CO_2 + 6 NADH + 6 H^+ + 2 FADH_2 + 2 ATP$
The citric acid cycle (Figure Figure $1$) provides a series of intermediate compounds that donate protons and electrons to the electron transport chain by way of the reduced coenzymes $NADH$ and $FADH_2$. The electron transport chain then generates additional ATPs by oxidative phosphorylation. The citric acid cycle also produces 2 ATP by substrate phosphorylation and plays an important role in the flow of carbon through the cell by supplying precursor metabolites for various biosynthetic pathways.
The citric acid cycle involves 8 distinct steps, each catalyzed by a unique enzyme. You are not responsible for knowing the chemical structures or enzymes involved in the steps below. They are included to help illustrate how the molecules in the pathway are manipulated by the enzymes in order to to achieve the required products.
Step 1: The citric acid cycle begins when Coenzyme A transfers its 2-carbon acetyl group to the 4-carbon compound oxaloacetate to form the 6-carbon molecule citrate (Figure Figure $2$).
Step 2: The citrate is rearranged to form an isomeric form, isocitrate (Figure $3$).
Step 3: The 6-carbon isocitrate is oxidized and a molecule of carbon dioxide is removed producing the 5-carbon molecule alpha-ketoglutarate. During this oxidation, $NAD^+$ is reduced to $NADH$ and $H^+$ (Figure $4$).
Step 4: Alpha-ketoglutarate is oxidized, carbon dioxide is removed, and coenzyme A is added to form the 4-carbon compound succinyl-CoA. During this oxidation, NAD+ is reduced to NADH + H+ (Figure $5$).
Step 5: CoA is removed from succinyl-CoA to produce succinate. The energy released is used to make guanosine triphosphate (GTP) from guanosine diphosphate (GDP) and Pi by substrate-level phosphorylation. GTP can then be used to make ATP (Figure $6$).
Step 6: Succinate is oxidized to fumarate. During this oxidation, $FAD$ is reduced to $FADH_2$ (Figure $7$).
Step 7: Water is added to fumarate to form malate (Figure $8$).
Step 8: Malate is oxidized to produce oxaloacetate, the starting compound of the citric acid cycle. During this oxidation, NAD+ is reduced to NADH + H+ (Figure $9$).
The NADH + H+ and FADH2 carry protons and electrons to the electron transport chain to generate additional ATP by oxidative phosphorylation.
Summary
1. Aerobic respiration involves four stages: glycolysis, a transition reaction that forms acetyl coenzyme A, the citric acid (Krebs) cycle, and an electron transport chain and chemiosmosis.
2. The citric acid cycle, also known as the tricarboxylic acid cycle and the Krebs cycle, completes the oxidation of glucose by taking the pyruvates from glycolysis, by way of the transition reaction, and completely breaking them down into CO2 molecules, H2O molecules, and generating additional ATP by oxidative phosphorylation.
3. The citric acid cycle provides a series of intermediate compounds that donate protons and electrons to the electron transport chain by way of the reduced coenzymes NADH and FADH2. The electron transport chain then generates additional ATPs by oxidative phosphorylation. The citric acid cycle also produces 2 ATP by substrate phosphorylation.
4. The overall reaction for the citric acid cycle is:$2 acetyl groups + 6 NAD^+ + 2 FAD + 2 ADP + 2 P_i yields 4 CO_2 + 6 NADH + 6 H^+ + 2 FADH_2 + 2 ATP.$
5. The citric acid cycle also plays an important role in the flow of carbon through the cell by supplying precursor metabolites for various biosynthetic pathways. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/18%3A_Microbial_Metabolism/18.3%3A_Aerobic_Respiration/18.3B%3A_Transition_Reaction.txt |
Learning Objectives
1. Briefly describethe function of the electron transport chain during aerobic respiration.
2. Briefly describethe chemiosmotic theory of generation of ATP as a result of an electron transport chain.
3. Compare where the electron transport chain occurs in prokaryotic cells and in eukaryotic cells.
4. State what is meant by proton motive force.
5. State the function of ATP synthases in chemiosmosis.
6. State the final electron acceptor and the end product formed at the end of aerobic respiration.
During various steps in glycolysis and the citric acid cycle, the oxidation of certain intermediate precursor molecules causes the reduction of NAD+ to NADH + H+ and FAD to FADH2. NADH and FADH2 then transfer protons and electrons to the electron transport chain to produce additional ATPs by oxidative phosphorylation .
As mentioned in the previous section on energy, during the process of aerobic respiration, coupled oxidation-reduction reactions and electron carriers are often part of what is called an electron transport chain , a series of electron carriers that eventually transfers electrons from NADH and FADH2 to oxygen. The diffusible electron carriers NADH and FADH2 carry hydrogen atoms (protons and electrons) from substrates in exergonic catabolic pathways such as glycolysis and the citric acid cycle to other electron carriers that are embedded in membranes. These membrane-associated electron carriers include flavoproteins, iron-sulfur proteins, quinones, and cytochromes. The last electron carrier in the electron transport chain transfers the electrons to the terminal electron acceptor, oxygen.
The chemiosmotic theory explains the functioning of electron transport chains. According to this theory, the transfer of electrons down an electron transport system through a series of oxidation-reduction reactions releases energy (Figure \(1\)). This energy allows certain carriers in the chain to transport hydrogen ions (H+ or protons) across a membrane.
Depending on the type of cell, the electron transport chain may be found in the cytoplasmic membrane or the inner membrane of mitochondria.
• In prokaryotic cells, the protons are transported from the cytoplasm of the bacterium across the cytoplasmic membrane to the periplasmic space located between the cytoplasmic membrane and the cell wall .
• In eukaryotic cells, protons are transported from the matrix of the mitochondria across the inner mitochondrial membrane to the intermembrane space located between the inner and outer mitochondrial membranes (Figure \(2\)).
As the hydrogen ions accumulate on one side of a membrane, the concentration of hydrogen ions creates an electrochemical gradient or potential difference (voltage) across the membrane. (The fluid on the side of the membrane where the protons accumulate acquires a positive charge; the fluid on the opposite side of the membrane is left with a negative charge.) The energized state of the membrane as a result of this charge separation is called proton motive force or PMF.
This proton motive force provides the energy necessary for enzymes called ATP synthases (see Figure \(3\)), also located in the membranes mentioned above, to catalyze the synthesis of ATP from ADP and phosphate. This generation of ATP occurs as the protons cross the membrane through the ATP synthase complexes and re-enter either the bacterial cytoplasm (Figure \(4\)) or the matrix of the mitochondria. As the protons move down the concentration gradient through the ATP synthase, the energy released causes the rotor and rod of the ATP synthase to rotate. The mechanical energy from this rotation is converted into chemical energy as phosphate is added to ADP to form ATP.
Proton motive force is also used to transport substances across membranes during active transport and to rotate bacterial flagella.
At the end of the electron transport chain involved in aerobic respiration, the last electron carrier in the membrane transfers 2 electrons to half an oxygen molecule (an oxygen atom) that simultaneously combines with 2 protons from the surrounding medium to produce water as an end product (Figure \(5\)).
Movie illustrating the electron transport system in the mitochondria of eukaryotic cells.
Summary
1. Aerobic respiration involves four stages: glycolysis, a transition reaction that forms acetyl coenzyme A, the citric acid (Krebs) cycle, and an electron transport chain and chemiosmosis.
2. During various steps in glycolysis and the citric acid cycle, the oxidation of certain intermediate precursor molecules causes the reduction of NAD+ to NADH + H+ and FAD to FADH2. NADH and FADH2 then transfer protons and electrons to the electron transport chain to produce additional ATPs by oxidative phosphorylation.
3. The electron transport chain consists of a series of electron carriers that eventually transfer electrons from NADH and FADH2 to oxygen.
4. The chemiosmotic theory states that the transfer of electrons down an electron transport system through a series of oxidation-reduction reactions releases energy. This energy allows certain carriers in the chain to transport hydrogen ions (H+ or protons) across a membrane.
5. As the hydrogen ions accumulate on one side of a membrane, the concentration of hydrogen ions creates an electrochemical gradient or potential difference (voltage) across the membrane called proton motive force.
6. This proton motive force provides the energy necessary for enzymes called ATP synthases, also located in the membranes mentioned above, to catalyze the synthesis of ATP from ADP and phosphate.
7. During aerobic respiration, the last electron carrier in the membrane transfers 2 electrons to half an oxygen molecule (an oxygen atom) that simultaneously combines with 2 protons from the surrounding medium to produce water as an end product. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/18%3A_Microbial_Metabolism/18.3%3A_Aerobic_Respiration/18.3D%3A_Electron_Transport_Chain_and_Chemisomosis.txt |
The theoretical maximum yield of ATP for the oxidation of one molecule of glucose during aerobic respiration is 38. In terms of substrate-level phosphorylation, oxidative phosphorylation, and the component pathways involved, briefly explain how this number is obtained.
Determining the exact yield of ATP for aerobic respiration is difficult for a number of reasons. In addition to generating ATP by oxidative phosphorylation in prokaryotic cells, proton motive force is also used for functions such as transporting materials across membranes and rotating flagella. Also, some bacteria use different carriers in their electron transport chain than others and the carriers may vary in the number of protons they transport across the membrane. Furthermore, the number of ATP generated per reduced NADH or FADH2 is not always a whole number. For every pair of electrons transported to the electron transport chain by a molecule of NADH, between 2 and 3 ATP are generated. For each pair of electrons transferred by FADH2, between 1 and 2 ATP are generated. In eukaryotic cells, unlike prokaryotes, NADH generated in the cytoplasm during glycolysis must be transported across the mitochondrial membrane before it can transfer electrons to the electron transport chain and this requires energy. As a result, between 1 and 2 ATP are generated from these NADH.
For simplicity, however, we will look at the theoretical maximum yield of ATP per glucose molecule oxidized by aerobic respiration. We will assume that for each pair of electrons transferred to the electron transport chain by NADH, 3 ATP will be generated; for each electron pair transferred by FADH2, 2 ATP will be generated. Keep in mind, however, that less ATP may actually be generated.
As seen above, one molecule of glucose oxidized by aerobic respiration in prokaryotes yields the following:
Glycolysis
2 net ATP from substrate-level phosphorylation
2 NADH yields 6 ATP (assuming 3 ATP per NADH) by oxidative phosphorylation
Transition Reaction
2 NADH yields 6 ATP (assuming 3 ATP per NADH) by oxidative phosphorylation
Citric Acid Cycle
2 ATP from substrate-level phosphorylation
6 NADH yields 18 ATP (assuming 3 ATP per NADH) by oxidative phosphorylation
2 FADH2 yields 4 ATP (assuming 2 ATP per FADH2) by oxidative phosphorylation
Total Theoretical Maximum Number of ATP Generated per Glucose in Prokaryotes
38 ATP: 4 from substrate-level phosphorylation; 34 from oxidative phosphorylation.
In eukaryotic cells, the theoretical maximum yield of ATP generated per glucose is 36 to 38, depending on how the 2 NADH generated in the cytoplasm during glycolysis enter the mitochondria and whether the resulting yield is 2 or 3 ATP per NADH.
18.4: Anaerobic Respiration
Define anaerobic respiration and state the pathways involved. State in what types or organism anaerobic respiration occurs.
Some prokaryotes are able to carry out anaerobic respiration, respiration in which an inorganic molecule other than oxygen (O2) is the final electron acceptor. For example, some bacteria called sulfate reducers can transfer electrons to sulfate (SO42-) reducing it to H2S. Other bacteria, called nitrate reducers, can transfer electrons to nitrate (NO3-) reducing it to nitrite (NO2-). Other nitrate reducers can reduce nitrate even further to nitrous oxide (NO) or nitrogen gas (N2).
Like aerobic respiration, anaerobic respiration involves glycolysis, a transition reaction, the citric acid cycle, and an electron transport chain. The total energy yield per glucose oxidized is less than with aerobic respiration with a theoretical maximum yield of 36 ATP or less.
Summary
1. Cellular respiration is the process cells use to convert the energy in the chemical bonds of nutrients to ATP energy.
2. Aerobic respiration is an exergonic pathway that requires molecular oxygen (O2).
3. Anaerobic exergonic pathways do not require oxygen and include anaerobic respiration and fermentation.
4. Some prokaryotes are able to carry out anaerobic respiration, respiration in which an inorganic molecule other than oxygen (O2) is the final electron acceptor.
5. Some bacteria called sulfate reducers can transfer electrons to sulfate (SO42-) reducing it to H2S. Other bacteria, called nitrate reducers, can transfer electrons to nitrate (NO3-) reducing it to nitrite (NO2-). Other nitrate reducers can reduce nitrate even further to nitrous oxide (NO) or nitrogen gas (N2).
6. Like aerobic respiration, anaerobic respiration involves glycolysis, a transition reaction, the citric acid cycle, and an electron transport chain. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/18%3A_Microbial_Metabolism/18.3%3A_Aerobic_Respiration/18.3E%3A_Theoretical_ATP_Yield.txt |
Learning Objectives
1. Define fermentation.
2. State the mechanism for ATP generation during fermentation.
3. Briefly describethe function of glycolysis during fermentation and indicate the reactants and products.
4. Compare the maximum yield of ATP from one molecule of glucose for aerobic respiration and for fermentation.
Fermentation is an anaerobic breakdown of carbohydrates in which an organic molecule is the final electron acceptor. It does not involve an electron transport system. Furthermore,:
1. Fermentation is a partial breakdown of glucose producing only 2 net ATP's per glucose by way of substrate-level phosphorylation ;
2. Fermentation involves only glycolysis; and
3. Fermentation is found in bacteria that are obligate anaerobes and facultative anaerobes.
Glycolysis during Fermentation
Function: As during aerobic respiration, glycolysis is a partial breakdown of a six-carbon glucose molecule into two, three-carbon molecules of pyruvate, 2NADH +2H+, and 2 net ATP as a result of substrate-level phosphorylation, as shown in (see Figure \(1\) and Figure \(2\)).
Glycolysis occurs in the cytoplasm of the cell. As mentioned above, the overall reaction is:
glucose (6C) + 2 NAD+ +2 ADP +2 inorganic phosphates (Pi)
yields 2 pyruvate (3C) + 2 NADH + 2 H+ + 2 net ATP
Glycolysis also produces a number of key precursor metabolites, as shown in Figure \(3\).
Since there is no electron transport system, the protons and electrons donated by certain intermediate precursor molecules during glycolysis generate no additional molecules of ATP. Instead, they combine with the coenzyme NAD+, the organic molecule which serves as the final electron and proton acceptor, reducing it to NADH + H+ (see Figure \(1\) and Figure \(2\)).
Glycolysis
Glycolysis is a partial breakdown of a six-carbon glucose molecule into two, three-carbon molecules of pyruvate, 2NADH +2H+, and 2 net ATP as a result of substrate-level phosphorylation. Glycolysis occurs in the cytoplasm of the cell.
The 2 pyruvic acids are then converted into one of many different fermentation end products in several non-energy-producing steps.
Fermentation end products
Some fermentation end products produced by microorganisms are very beneficial to humans and are the basis of a number of industries (brewing industry, dairy industry, etc.). Fermentation is used in the production of many food products including bread, alcohol, yogurt, sour cream, cheeses, vinegar, sauerkraut, pickles, olives, soy sauce, poi, and kimchi. Examples of microbial fermentation end products include:
• Saccharomyces: ethyl alcohol and CO2
• Streptococcus and Lactobacillus: lactic acid
• Propionibacterium: proprionic acid, acetic acid, and CO2
• Escherichia coli: acetic acid, lactic acid, succinic acid, ethyl alcohol, CO2, and H2
• Enterobacter: formic acid, ethyl alcohol, 2,3-butanediol, lactic acid, CO2, and H2
• Clostridium: butyric acid, butyl alcohol, acetone, isopropyl alcohol, CO2, and H2
Summary
1. Fermentation is an anaerobic breakdown of carbohydrates in which an organic molecules the final electron acceptor and does not involve an electron transport system.
2. Fermentation is a partial breakdown of glucose producing only 2 net ATP's per glucose by way of substrate-level phosphorylation, involves only glycolysis, and is found in anaerobic and facultative anaerobic bacteria.
3. The overall reaction is glucose (6C) + 2 NAD+ +2 ADP +2 inorganic phosphates (Pi) yields 2 pyruvate (3C) + 2 NADH + 2 H+ + 2 net ATP.
4. Glycolysis also produces a number of key precursor metabolites.
5. Some fermentation end products produced by microorganisms are very beneficial to humans and are the basis of a number of industries (brewing industry, dairy industry, etc.). | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/18%3A_Microbial_Metabolism/18.5%3A_Fermentation.txt |
Photoautotrophs use sunlight as a source of energy and through the process of photosynthesis, reduce carbon dioxide to form carbohydrates such as glucose. The radiant energy is converted to the chemical bond energy within glucose and other organic molecules. Plants, algae, and cyanobacteria are known as oxygenic photoautotrophs because they synthesize organic molecules from inorganic materials, convert light energy into chemical energy, use water as an electron source, and generate oxygen as an end product of photosynthesis. The overall reaction for photosynthesis is as follows:
$6 CO_2 + 12 H_2O \xrightarrow[\text{light}]{\text{chlorophyll}} C_6H_{12}O_6 + 6 O_2 + 6 H_2O$
Note that photosyntehsis is a redox reaction with carbon dioxide ($CO_2$) reduced to produce glucose ($C_6H_{12}O_6$) and water ($H_2O$) oxidized to produce oxygen ($O_2$). Photosynthesis is composed of two stages: the light-dependent reactions and the light independent reactions.
18.7: Photosynthesis
Define the following: oxygenic photoautotroph anoxygenic photoautotroph photon Name the two stages of photosynthesis. State how all radiations in the electromagnetic spectrum travel. State what constitutes visible light. Define photon and describe what happens when photons of visible light energy strike certain atoms of pigments during photosynthesis and how this can lead to the generation of ATP. Describe the structure of a chloroplast and list the pigments it may contain. Give the overall reaction for photosynthesis. State the reactants and the products for photosynthesis and indicate which are oxidized and which are reduced.
Autotrophs are organisms that are able to synthesize organic molecules from inorganic materials. Photoautotrophs absorb and convert light energy into the stored energy of chemical bonds in organic molecules through a process called photosynthesis.
Plants, algae, and bacteria known as cyanobacteria are known as oxygenic photoautotrophs because they synthesize organic molecules from inorganic materials, convert light energy into chemical energy, use water as an electron source, and generate oxygen as an end product of photosynthesis. Some bacteria, such as the green and purple bacteria, are known as anoxygenic phototrophs . Unlike the oxygenic plants, algae, and cyanobacteria, anoxygenic phototrophs do not use water as an electron source and, therefore, do not evolve oxygen during photosynthesis. The electrons come from compounds such as hydrogen gas, hydrogen sulfide, and reduced organic molecules. In this section on photosynthesis, we be concerned with the oxygenic phototrophs.
There are three major groups of photosynthetic bacteria: cyanobacteria, purple bacteria, and green bacteria.
1. The cyanobacteria carry out oxygenic photosynthesis, that is, they use water as an electron donor and generate oxygen during photosynthesis. The photosynthetic system is located in an extensive thylakoid membrane system that is lined with particles called phycobilisomes.
Cyanobacteria, as well as algae and green plants, use hydrogen atoms from water to reduce carbon dioxide to form carbohydrates, and during this process oxygen gas is given off (an oxygenic process). Cyanobacteria were the first organisms on earth to carry out oxygenic photosynthesis. (left) Anabaena flosaquae and (right) Oscillatoria princeps. Images used with permission (Anabaena is Public domain from the US Environmental Protection Agenca and Oscillatoria is CC BY-SA 3.0; Kristian Peters).
2. The green bacteria carry out anoxygenic photosynthesis. They use reduced molecules such as H2, H2S, S, and organic molecules as an electron source and generate NADH and NADPH. The photosynthetic system is located in ellipsoidal vesicles called chlorosomes that are independent of the cytoplasmic membrane.
3. The purple bacteria carry out anoxygenic photosynthesis. They use reduced molecules such as H2, H2S, S, and organic molecules as an electron source and generate NADH and NADPH. The photosynthetic system is located in spherical or lamellar membrane systems that are continuous with the cytoplasmic membrane.
In this section we will concentrate on oxygenic photosynthesis. Oxygenic photosynthesis is composed of two stages: the light-dependent reactions and the light-independent reactions.
1. The light-dependent reactions convert light energy into chemical energy, producing ATP and NADPH.
2. The light-independent reactions use the ATP and NADPH from the light-dependent reactions to reduce carbon dioxide and convert the energy to the chemical bond energy in carbohydrates such as glucose. Before we get to these photosynthetic reactions however, we need to understand a little about the electromagnetic spectrum and chloroplasts.
The Electromagnetic Spectrum
The thylakoid membranes contain several pigments capable of absorbing visible light. Chlorophyll is the primary pigment of photosynthesis. Chlorophyll absorbs light in the blue and red region of the visible light spectrum and reflects green light. There are two major types of chlorophyll, chlorophyll a that initiates the light-dependent reactions of photosynthesis, and chlorophyll b, an accessory pigment that also participates in photosynthesis. The thylakoid membranes also contain other accessory pigments. Carotenoids are pigments that absorb blue and green light and reflect yellow, orange, or red. Phycocyanins absorb green and yellow light and reflect blue or purple. These accessory pigments absorb light energy and transfer it to chlorophyll.
Photosynthetic prokaryotic cells do not possess chloroplasts. Instead, thylakoid membranes are usually arranged around the periphery of the bacterium as infoldings of the cytoplasmic membrane.
Photosynthesis
As mentioned above, photoautotrophs use sunlight as a source of energy and through the process of photosynthesis, reduce carbon dioxide to form carbohydrates such as glucose. The radiant energy is converted to the chemical bond energy within glucose and other organic molecules.
The overall reaction for photosynthesis is as follows:
6 CO2 + 12 H2O in the presence of light and chlorophyll yields C6H12O6 + 6 O2 + 6 H2O
Note that carbon dioxide (CO2) is reduced to produce glucose (C6H12O6 ) while water (H2O) is oxidized to produce oxygen (O2).
Outside Links
• YouTube movie on the structure and functions of chloroplasts.
Summary
1. Autotrophs are organisms that are able to synthesize organic molecules from inorganic materials.
2. Photoautotrophs absorb and convert light energy into the stored energy of chemical bonds in organic molecules through a process called photosynthesis.
3. Plants, algae, and cyanobacteria are known as oxygenic photoautotrophs because they synthesize organic molecules from inorganic materials, convert light energy into chemical energy, use water as an electron source, and generate oxygen as an end product of photosynthesis.
4. Green and purple bacteria, are known as anoxygenic phototrophs that do not use water as an electron source and, therefore, do not evolve oxygen during photosynthesis. The electrons come from compounds such as hydrogen gas, hydrogen sulfide, and reduced organic molecules.
5. Oxygenic photosynthesis is composed of two stages: the light-dependent reactions and the light-independent reactions.
6. The light-dependent reactions convert light energy into chemical energy, producing ATP and NADPH.
7. The light-independent reactions use the ATP and NADPH from the light-dependent reactions to reduce carbon dioxide and convert the energy to the chemical bond energy in carbohydrates such as glucose.
8. Light and other types of radiation are composed of individual packets of energy called photons. When photons of visible light energy strike certain atoms of pigments during photosynthesis, that energy may push an electron from that atom to a higher energy level where it can be picked up by an electron acceptor in an electron transport chain.
9. In eukaryotic cells, photosynthesis takes place in organelles called chloroplasts.
10. The inner membrane of a chloroplast encloses a fluid-filled region called the stroma that contains enzymes for the light-independent reactions of photosynthesis.
11. Infolding of this inner membrane forms interconnected stacks of disk-like sacs called thylakoids. The thylakoid membrane, which encloses a fluid-filled thylakoid interior space, contains chlorophyll and other photosynthetic pigments as well as electron transport chains. The light-dependent reactions of photosynthesis occur in the thylakoids.
12. The overall reaction for photosynthesis is as follows: 6 CO2 + 12 H2O in the presence of light and chlorophyll yields C6H12O6 + 6 O2 + 6 H2O. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/18%3A_Microbial_Metabolism/18.7%3A_Photosynthesis/18.7A%3A_Introduction_to_Photosynthesis.txt |
Briefly describe the overall function of the light-dependent reactions in photosynthesis and state where in the chloroplast they occur. State the reactants and the products for the light-dependent reactions. Describe an antenna complex and state the function of the reaction center. Briefly describe the overall function of Photosystem II in the light-dependent reactions of photosynthesis. Briefly describe how ATP is generated by chemiosmosis during the light-dependent reactions of photosynthesis. Briefly describe the overall function of Photosystem I in the light-dependent reactions of photosynthesis. Compare noncyclic photophosphorylation and cyclic photophosphorylation in terms of Photosystems involved and products produced.
The most common light-dependent reaction in photosynthesis is called noncyclic photophosphorylation. Noncyclic photophosphorylation involves both Photosystem I and Photosystem II and produces ATP and NADPH. During noncyclic photophosphorylation, the generation of ATP is coupled to a one-way flow of electrons from H2O to NADP+. We will now look at Photosystems I and II and their roles in noncyclic photophosphorylation.
2. Meanwhile, photons are also being absorbed by pigment molecules in the antenna complex of Photosystem I and excited electrons from the reaction center are picked up by the primary electron acceptor of the Photosystem I electron transport chain. The electrons being lost by the P700 chlorophyll a molecules in the reaction centers of Photosystem I are replaced by the electrons traveling down the Photosystem II electron transport chain. The electrons transported down the Photosystem I electron transport chain combine with 2H+ from the surrounding medium and NADP+ to produce NADPH + H+ (Figure \(2\)).
Cyclic photophosphorylation occurs less commonly in plants than noncyclic photophosphorylation, most likely occurring when there is too little NADP+ available. It is also seen in certain photosynthetic bacteria. Cyclic photophosphorylation involves only Photosystem I and generates ATP but not NADPH. As the electrons from the reaction center of Photosystem I are picked up by the electron transport chain, they are transported back to the reaction center chlorophyll. As the electrons are transported down the electron transport chain, some of the energy released is used to pump protons across the thylakoid membrane from the stroma of the chloroplast to the thylakoid interior space producing a proton gradient or proton motive force. As the accumulating protons in the thylakoid interior space pass back across the thylakoid membrane to the stroma through ATP synthetase complexes, this energy is used to generate ATP from ADP and Pi (Figure \(4\)).
Summary
1. Plants, algae, and cyanobacteria are known as oxygenic photoautotrophs because they synthesize organic molecules from inorganic materials, convert light energy into chemical energy, use water as an electron source, and generate oxygen as an end product of photosynthesis.
2. The overall reaction for photosynthesis is as follows: 6 CO2 + 12 H2O in the presence of light and chlorophyll yields C6H12O6 + 6 O2 + 6 H2O.
3. Oxygenic photosynthesis is composed of two stages: the light-dependent reactions and the light-independent reactions.
4. The light-dependent reactions convert light energy into chemical energy, producing ATP and NADPH.
5. The light-dependent reactions can be summarized as follows: 12 H2O + 12 NADP+ + 18 ADP + 18 Pi + light and chlorophyll yields 6 O2 + 12 NADPH + 18 ATP.
6. The most common light-dependent reaction in photosynthesis is called noncyclic photophosphorylation.
7. During noncyclic photophosphorylation light-dependent reactions, photons are absorbed by pigment molecules in the antenna complexes of Photosystem II, and excited electrons from the reaction center are picked up by the primary electron acceptor of the Photosystem II electron transport chain. During this process, Photosystem II splits molecules of H2O into 1/2 O2, 2H+, and 2 electrons.
8. According to the chemiosmosis theory, as the electrons are transported down the electron transport chain, some of the energy released is used to pump protons across the thylakoid membrane from the stroma of the chloroplast to the thylakoid interior space producing a proton gradient or proton motive force. As the accumulating protons in the thylakoid interior space pass back across the thylakoid membrane to the stroma through ATP synthetase complexes, this proton motive force is used to generate ATP from ADP and Pi.
9. Meanwhile, photons are also being absorbed by pigment molecules in the antenna complex of Photosystem I and excited electrons from the reaction center are picked up by the primary electron acceptor of the Photosystem I electron transport chain. The electrons being lost by chlorophyll molecules in the reaction centers of Photosystem I are replaced by the electrons traveling down the Photosystem II electron transport chain. The electrons transported down the Photosystem I electron transport chain combine with 2H+ from the surrounding medium and NADP+ to produce NADPH + H+. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/18%3A_Microbial_Metabolism/18.7%3A_Photosynthesis/18.7B%3A_Oxygenic_Photosynthesis%3A_Light-Dependent_Reactions.txt |
Summary
1. Photoautotrophs absorb and convert light energy into the stored energy of chemical bonds in organic molecules through a process called photosynthesis.
2. Plants, algae, and cyanobacteria are known as oxygenic photoautotrophs because they synthesize organic molecules from inorganic materials, convert light energy into chemical energy, use water as an electron source, and generate oxygen as an end product of photosynthesis.
3. Oxygenic photosynthesis is composed of two stages: the light-dependent reactions and the light-independent reactions.
4. The light-independent reactions use the ATP and NADPH from the light-dependent reactions to reduce carbon dioxide and convert the energy to the chemical bond energy in carbohydrates such as glucose.
5. The light-independent reactions can be summarized as follows: 12 NADPH + 18 ATP + 6 CO2 yields C6H12O6 (glucose) + 12 NADP+ + 18 ADP + 18 Pi + 6 H2O.
6. Most plants use the Calvin cycle to fix CO2. To begin the Calvin cycle, a molecule of CO2 reacts with a five-carbon compound called ribulose bisphosphate (RuBP) producing an unstable six-carbon intermediate which immediately breaks down into two molecules of the three-carbon compound phosphoglycerate (PGA).
7. The energy from ATP and the reducing power of NADPH (both produced during the light-dependent reactions) is now used to convert the molecules of PGA to glyceraldehyde-3-phosphate (G3P), another three-carbon compound.
8. Most of the G3P produced during the Calvin cycle are used to regenerate the RuBP so that the cycle may continue, however, some of the molecules of G3P, however, are used to synthesize glucose and other organic molecules.
18.7D: C4 and CAM Pathways in Plants
Learning Objectives
1. Briefly describe the C4 pathway and how it differs from the C3 pathway.
2. Briefly describe the CAM pathway and how it differs from the C4 pathway.
The entry and exit of gasses in plants is through small pores called stomata located on the underside of leaves. Carbon dioxide, the gas required for the Calvin cycle, is not a very abundant gas in nature. Under hot and dry environmental conditions the stomata close to reduce the loss of water vapor, but this also results in a greatly diminished supply of CO2 for the plant. Plants that normally live in dry, hot climates have adapted different ways of initially fixing CO2 prior to its entering the Calvin cycle. These pathways of carbon fixation, know as the C4 and the CAM pathways, take place in the cytoplasm of the cell.
The C4 pathway
The C4 pathway is designed to efficiently fix CO2 at low concentrations and plants that use this pathway are known as C4 plants. These plants first fix CO2 into a four carbon compound (C4) called oxaloacetate (Figure \(1\)). This occurs in cells called mesophyll cells. First, CO2 is fixed to a three-carbon compound called phosphoenolpyruvate to produce the four-carbon compound oxaloacetate. The enzyme catalyzing this reaction, PEP carboxylase, fixes CO2 very efficiently so the C4 plants don't need to to have their stomata open as much.
The oxaloacetate is then converted to another four-carbon compound called malate in a step requiring the reducing power of NADPH. The malate then exits the mesophyll cells and enters the chloroplasts of specialized cells called bundle sheath cells. Here the four-carbon malate is decarboxylated to produce CO2, a three-carbon compound called pyruvate, and NADPH. The CO2 combines with ribulose bisphosphate and goes through the Calvin cycle while the pyruvate re-enters the mesophyll cells, reacts with ATP, and is converted back to phosphoenolpyruvate, the starting compound of the C4 cycle. The C4 cycle is summarized in Figure \(1\).
The CAM pathway
CAM plants live in very dry condition and, unlike other plants, open their stomata to fix CO2 only at night. Like C4 plants, the use PEP carboxylase to fix CO2, forming oxaloacetate. The oxaloacetate is converted to malate which is stored in cell vacuoles. During the day when the stomata are closed, CO2 is removed from the stored malate and enters the Calvin cycle.
Summary
1. Carbon dioxide, the gas required for the Calvin cycle, is not a very abundant gas in nature. Under hot and dry environmental conditions the stomata close to reduce the loss of water vapor, but this also results in a greatly diminished supply of CO2 for the plant.
2. Plants that normally live in dry, hot climates have adapted different ways of initially fixing CO2 prior to its entering the Calvin cycle. These pathways of carbon fixation, know as the C4 and the CAM pathways, take place in the cytoplasm of the cell.
3. The C4 pathway is designed to efficiently fix CO2 at low concentrations and plants that use this pathway are known as C4 plants.
4. CAM plants live in very dry condition and, unlike other plants, open their stomata to fix CO2 only at night. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/18%3A_Microbial_Metabolism/18.7%3A_Photosynthesis/18.7C%3A_Oxygenic_Photosynthesis%3A_Light-Independent_Reactions.txt |
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied.
18.3: Aerobic Respiration
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe aerobic respiration. (ans)
2. Give the overall chemical reaction for aerobic respiration. (ans)
3. During aerobic respiration, glucose is __________ to carbon dioxide.
1. oxidized (ans)
2. reduced (ans)
4. During aerobic respiration, oxygen is __________ to water.
1. oxidized (ans)
2. reduced (ans)
5. Name the four stages of aerobic respiration. (ans)
18.3A: Glycolysis
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the function of glycolysis during aerobic respiration and indicate the reactants and products. (ans)
2. State the reactants in glycolysis. (ans)
3. State the products in glycolysis. (ans)
4. Does glycolysis require oxygen? (ans)
5. Is the following statement true or false?
In eukaryotic cells, glycolysis takes place in the mitochondria. (ans)
6. Steps 1 and 3 of glycolysis are:
1. exergonic (ans)
2. endergonic (ans)
7. State why one molecule of glucose is able to produce two molecules of pyruvate during glycolysis. (ans)
8. The two net ATP produced in glycolysis are generated by:
1. oxidative phosphorylation (ans)
2. substrate-level phosphorylation (ans)
9. State the total number and the net number of ATP produced by substrate-level phosphorylation during glycolysis. (ans)
10. During aerobic respiration, state what happens to the 2 NADH produced during glycolysis. (ans)
11. During aerobic respiration, state what happens to the two molecules of pyruvate produced during glycolysis. (ans)
18.3B: Transition Reaction
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the function of transition reaction during aerobic respiration. (ans)
2. State the reactants in the transition reaction. (ans)
3. State the products in the transition reaction. (ans)
4. Is the following statement true or false?
In eukaryotic cells, the transition reaction occurs inside the mitochondria. (ans)
5. During aerobic respiration, state what happens to the two molecules of Acetyl-CoA produced during the transition reaction. (ans)
18.3C: Citric Acid (Krebs) Cycle
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the function of the citric acid cycle during aerobic respiration. (ans)
2. State the reactants for the citric acid cycle. (ans)
3. State the products for the citric acid cycle. (ans)
4. Is the following statement true or false?
In eukaryotic cells, the citric acid cycle occurs in the cytoplasm. (ans)
5. State the total number of ATP produced by substrate-level phosphorylation for each acetyl-CoA that enters the citric acid cycle. (ans)
6. State the total number of NADH and FADH2 produced for each acetyl-CoA that enters the citric acid cycle. (ans)
7. During aerobic respiration, state what happens to the NADH and the FADH2 produced during the citric acid cycle. (ans)
18.3D: Electron Transport Chain and Chemisomosis
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the function of the electron transport chain during aerobic respiration. (ans)
2. Describethe chemiosmotic theory of generation of ATP as a result of an electron transport chain. In the process, describe proton motive force and indicate the function of ATP synthase. (ans)
3. State whether the following statement is true or false.
In eukaryotic cells, the electron transport chain is located in the inner membrane of the mitochondria. (ans)
4. State the final electron acceptor and the end product formed at the end of aerobic respiration. (ans)
18.3E: Theoretical ATP Yield
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Fill in the blanks.
One molecule of glucose oxidized by aerobic respiration in prokaryotes yields the following:
Glycolysis:
_____ net ATP (ans) from substrate-level phosphorylation
_____ NADH (ans) yields _____ ATP (assuming 3 ATP per NADH) by oxidative phosphorylation (ans)
Transition Reaction:
_____ NADH (ans) yields _____ ATP (assuming 3 ATP per NADH) by oxidative phosphorylation (ans)
Citric Acid Cycle:
_____ ATP from substrate-level phosphorylation (ans)
_____ NADH (ans) yields _____ ATP (assuming 3 ATP per NADH) by oxidative phosphorylation (ans)
_____ FADH2 (ans) yields _____ ATP (assuming 2 ATP per FADH2) by oxidative phosphorylation (ans)
Total Theoretical Maximum Number of ATP Generated per Glucose in Prokaryotes
_____ ATP (ans): _____ from substrate-level phosphorylation (ans); _____ from oxidative phosphorylation (ans).
In eukaryotic cells, the theoretical maximum yield of ATP generated per glucose is _____ to _____. (ans)
18.4: Anaerobic Respiration
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Define anaerobic respiration. (ans)
2. State the pathways involved in anaerobic respiration. (ans)
3. State whether the following statement is true or false.
All organisms are capable of anaerobic respiration. (ans)
18.5: Fermentation
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Define fermentation. (ans)
2. All the ATP generated by fermentation are produced by:
1. substrate-level phosphorylation (ans)
2. oxidative phosphorylation (ans)
3. State the reactants for fermentation. (ans)
4. State the products for fermentation. (ans)
5. Compare the maximum yield of ATP from one molecule of glucose for aerobic respiration and for fermentation. (ans)
18.6: Precursor Metabolites: Linking Catabolic and Anabolic Pathways
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Define precursor metabolites and indicate their importance in metabolism. (ans)
18.7A: Introduction to Photosynthesis
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Organisms that absorb and convert light energy into the stored energy of chemical bonds in organic molecules through a process called photosynthesis best describes:
1. anoxygenic photoautotrophs (ans)
2. oxygenic photoautotrophs (ans)
2. Name the two stages of photosynthesis. (ans)
3. Define photon. (ans)
4. Describe what happens when photons of visible light energy strike certain atoms of pigments during photosynthesis and how this can lead to the generation of ATP. (ans)
5. Fill in the blank.
The inner membrane of a chloroplast encloses a fluid-filled region called the __________ (ans) that contains enzymes for the light-independent reactions of photosynthesis. Infolding of this inner membrane forms interconnected stacks of disk-like sacs called __________ (ans), often arranged in stacks called __________ (ans).
6. Name three different types of pigments that play a role in photosynthesis by absorbing light energy. (ans)
7. State the reactants and the products for photosynthesis and indicate which are oxidized and which are reduced. (ans)
18.7B: Oxygenic Photosynthesis: Light-Dependent Reactions
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the overall function of the light-dependent reactions in photosynthesis. (ans)
2. Where in the chloroplasts do the light-dependent reactions occur?
1. In the thylakoids. (ans)
2. In the stroma. (ans)
3. The parts of a photosystem that are able to trap light and transfer energy to a complex of chlorophyll molecules and proteins called the reaction center are called _____________. (ans)
4. In Photosystem II, the electrons lost by chlorophyll P680 molecules are replaced by:
1. the electrons traveling down the electron transport system of Photosystem I (ans)
2. the electrons released by the splitting of water (ans)
5. The primary function of Photosystem II is to produce:
1. ATP (ans)
2. NADPH (ans)
6. Briefly describe how ATP is generated by chemiosmosis during the light-dependent reactions of photosynthesis. (ans)
7. In Photosystem I, the electrons lost by chlorophyll P700 molecules are replaced by:
1. the electrons traveling down the electron transport system of Photosystem II (ans)
2. the electrons released by the splitting of water (ans)
8. The primary function of Photosystem I is to produce:
1. ATP (ans)
2. NADPH (ans)
9. Involves only Photosystem I and generates ATP but not NADPH. This best describes:
1. cyclic photophosphorylation (ans)
2. noncyclic photophosphorylation (ans)
18.7C: Oxygenic Photosynthesis: Light-Independent Reactions
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the overall function of the light-independent reactions in photosynthesis. (ans)
2. Where in the chloroplasts do the light-independent reactions occur?
1. In the thylakoids. (ans)
2. In the stroma. (ans)
3. State how the light-dependent and light-independent reactions are linked during photosynthesis. (ans)
4. Briefly describe the following stages of the Calvin cycle:
1. CO2 fixation (ans)
2. production of G3P (ans)
3. regeneration of RuBP (ans)
5. State the significance of glyceraldehyde-3-phosphate (G3P) in the Calvin cycle. (ans)
18.7D: C4 and CAM Pathways in Plants
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Is the following statement true or false?
During the C4 pathway for fixing CO2, CO2 from the air combines with ribulose bisphosphate to begin the Calvin cycle. (ans)
2. Plants that live in very dry condition and, unlike other plants, open their stomata to fix CO2 only at night best describes: (ans)
1. C4 plants
2. C3 plants
3. CAM plants
3. C4 and CAM pathways evolved for plants that live in _____________________ climates. (ans)
1. hot, humid
2. cold, dry
3. hot, dry | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/18%3A_Microbial_Metabolism/18.E%3A_Microbial_Metabolism_%28Exercises%29.txt |
Thumbnail: DNA Double Helix. (Public Domain; Apers0n).
19: Review of Molecular Genetics
Learning Objectives
1. Define or describe the following:
1. amino acid
2. "R" group
3. peptide bond
4. peptide
5. polypeptide
6. primary protein structure
7. secondary protein structure
8. tertiary protein structure
9. quaternary protein structure
10. gene
2. Describe how the primary structure of a protein or polypeptide ultimately detemines its final three-dimensional shape.
3. Describe how the order of nucleotide bases in DNA ultimately determines the final three-dimensional shape of a protein or polypeptide.
Amino acids are the building blocks for proteins. All amino acids contain an amino or NH2 group and a carboxyl (acid) or COOH group. There are 20 different amino acids commonly found in proteins and often 300 or more amino acids per protein molecule. Each amino acid differs in terms of its "R" group. The "R" group of an amino acid is the remainder of the molecule, that is, the portion other than the amino group, the acid group, and the central carbon. Each different amino acid has a unique "R" group and the unique chemical properties of an amino acid depend on that of its "R" group (Figure \(1\)).
To form polypeptides and proteins, amino acids are joined together by peptide bonds, in which the amino or NH2 of one amino acid bonds to the carboxyl (acid) or COOH group of another amino acid as shown in (Figure \(2\) and Figure \(3\)).
A peptide is two or more amino acids joined together by peptide bonds, and a polypeptide is a chain of many amino acids. A protein contains one or more polypeptides. Therefore, proteins are long chains of amino acids held together by peptide bonds.
The actual order of the amino acids in the protein is called its primary structure (Figure \(4\)) and is determined by DNA. As will be seen later in this unit, DNA is divided into functional units called genes. A gene is a sequence of deoxyribonucleotide bases along one strand of DNA that codes for a functional product - a specific molecule of messenger RNA, transfer RNA, or ribosomal RNA. The product is usually messenger RNA (mRNA) and mRNA ultimately results in the synthesis of a polypeptide or a protein. Therefore, it is commonly said that the order of deoxyribonucleotide bases in a gene determines the amino acid sequence of a particular protein. Since certain amino acids can interact with other amino acids in the same protein, this primary structure ultimately determines the final shape and therefore the chemical and physical properties of the protein.
The secondary structure of the protein is due to hydrogen bonds that form between the oxygen atom of one amino acid and the nitrogen atom of another. This gives the protein or polypeptide the two-dimensional form of an alpha-helix or a beta-pleated sheet (Figure \(5\)).
In globular proteins such as enzymes, the long chain of amino acids becomes folded into a three-dimensional functional shape or tertiary structure. This is because certain amino acids with sulfhydryl or SH groups form disulfide (S-S) bonds with other amino acids in the same chain. Other interactions between R groups of amino acids such as hydrogen bonds, ionic bonds, covalent bonds, and hydrophobic interactions also contribute to the tertiary structure (Figure \(6\)). In some proteins, such as antibody molecules and hemoglobin, several polypeptides may bond together to form a quaternary structure (Figure \(7\)).
As will be seen later in this unit, during protein synthesis, the order of nucleotide bases along a gene gets transcribed into a complementary strand of mRNA which is then translated by tRNA into the correct order of amino acids for that polypeptide or protein. Therefore, the order of deoxyribonucleotide bases along the DNA determines the order of amino acids in the proteins. Because certain amino acids can interact with other amino acids, the order of amino acids for each protein determines its final three-dimensional shape, which in turn determines the function of that protein (e.g., what substrate an enzyme will react with, what epitopes the Fab of an antibody will combine with, what receptors a cytokine will bind to).
Summary
1. Amino acids are the building blocks for proteins. There are 20 different amino acids commonly found in proteins and often 300 or more amino acids per protein molecule.
2. All amino acids contain an amino or NH2 group and a carboxyl (acid) or COOH group.
3. To form polypeptides and proteins, amino acids are joined together by peptide bonds, in which the amino or NH2 of one amino acid bonds to the carboxyl (acid) or COOH group of another amino acid.
4. A peptide is two or more amino acids joined together by peptide bonds; a polypeptide is a chain of many amino acids; and a protein contains one or more polypeptides. Therefore, proteins are long chains of amino acids held together by peptide bonds.
5. The actual order of the amino acids in the protein is called its primary structure and is determined by DNA.
6. The order of deoxyribonucleotide bases in a gene determines the amino acid sequence of a particular protein. Since certain amino acids can interact with other amino acids in the same protein, this primary structure ultimately determines the final shape and therefore the chemical and physical properties of the protein.
7. The secondary structure of the protein is due to hydrogen bonds that form between the oxygen atom of one amino acid and the nitrogen atom of another and gives the protein or polypeptide the two-dimensional form of an alpha-helix or a beta-pleated sheet.
8. In globular proteins such as enzymes, the long chain of amino acids becomes folded into a three-dimensional functional shape or tertiary structure. This is because certain amino acids with sulfhydryl or SH groups form disulfide (S-S) bonds with other amino acids in the same chain. Other interactions between R groups of amino acids such as hydrogen bonds, ionic bonds, covalent bonds, and hydrophobic interactions also contribute to the tertiary structure.
9. In some proteins, such as antibody molecules, several polypeptides may bond together to form a quaternary structure. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/19%3A_Review_of_Molecular_Genetics/19.1%3A_Polypeptides_and_Proteins.txt |
Learning Objectives
1. Define or describe the following:
1. metabolism
2. catabolic reaction
3. anabolic reaction
4. enzyme
5. substrate
6. apoenzyme
7. haloenzyme
8. cofactor (coenzyme)
2. State how enzymes are able to speed up the rate of chemical reactions.
3. Briefly describe a generalized enzyme-substrate reaction, state the function of an enzyme's active site, and describe how an enzyme is able to speed up chemical reactions.
4. State four characteristics of enzymes.
5. State how the following affect the rate of an enzyme reaction.
1. enzyme concentration
2. substrate concentration
3. temperature
4. pH
5. salt concentration
6. State how chemicals such as chlorine, iodine, iodophores, mercurials, and ethylene oxide inhibit or kill bacteria.
7. State how high temperature and low temperature exert their effect on bacteria.
To live, grow, and reproduce, microorganisms undergo a variety of chemical changes. They alter nutrients so they can enter the cell and they change them once they enter in order to synthesize cell parts and obtain energy. Metabolism refers to all of the organized chemical reactions in a cell. Reactions in which chemical compounds are broken down are called catabolic reactions while reactions in which chemical compounds are synthesized are termed anabolic reactions. All of these reactions are under the control of enzymes.
Enzymes are substances present in the cell in small amounts that function to speed up or catalyze chemical reactions. On the surface of the enzyme is usually a small crevice that functions as an active site or catalytic site to which one or two specific substrates are able to bind. (Anything that an enzyme normally combines with is called a substrate.) The binding of the substrate to the enzyme causes the flexible enzyme to change its shape slightly through a process called induced fit to form a tempore intermediate called an enzyme-substrate complex (Figure \(1\)).
Enzymes speed up the rate of chemical reactions because they lower the energy of activation, the energy that must be supplied in order for molecules to react with one another (Figure \(2\)). Enzymes lower the energy of activation by forming an enzyme-substrate complex allowing products of the enzyme reaction to be formed and released (Figure \(3\)).
Many enzymes require a nonprotein cofactor to assist them in their reaction. In this case, the protein portion of the enzyme, called an apoenzyme, combines with the cofactor to form the whole enzyme or haloenzyme (Figure \(3\)). Some cofactors are ions such as Ca++, Mg++, and K+; other cofactors are organic molecules called coenzymes which serve as carriers for chemical groups or electrons. NAD+, NADP+, FAD, and coenzyme A (CoA) are examples of coenzymes.
Characteristics of Enzymes
Chemically, enzymes are generally globular proteins. (Some RNA molecules called ribozymes can also be enzymes. These are usually found in the nuclear region of cells and catalyze the splitting of RNA molecules). Enzymes are catalysts that breakdown or synthesize more complex chemical compounds. They allow chemical reactions to occur fast enough to support life. Enzymes speed up the rate of chemical reactions because they lower the energy of activation, the energy that must be supplied in order for molecules to react with one another. Anything that an enzyme normally combines with is called a substrate. Enzymes are very efficient. An enzyme generally can typically catalyze between 1 and 10,000 molecules of substrate per second. Enzymes are only present in small amounts in the cell since they are not altered during their reactions. and they are highly specific for their substrate. Generally there is one specific enzyme for each specific chemical reaction.
Enzyme Activity
Enzyme activity is affected by a number of factors including:
• The concentration of enzyme: Assuming a sufficient concentration of substrate is available, increasing enzyme concentration will increase the enzyme reaction rate.
• The concentration of substrate: At a constant enzyme concentration and at lower concentrations of substrates, the substrate concentration is the limiting factor. As the substrate concentration increases, the enzyme reaction rate increases. However, at very high substrate concentrations, the enzymes become saturated with substrate and a higher concentration of substrate does not increase the reaction rate.
• The temperature: Each enzyme has an optimum temperature at which it works best. A higher temperature generally results in an increase in enzyme activity. As the temperature increases, molecular motion increases resulting in more molecular collisions. If, however, the temperature rises above a certain point, the heat will denature the enzyme, causing it to lose its three-dimensional functional shape by denaturing its hydrogen bonds. Cold temperature, on the other hand, slows down enzyme activity by decreasing molecular motion.
• The pH: Each enzyme has an optimal pH that helps maintain its three-dimensional shape. Changes in pH may denature enzymes by altering the enzyme's charge. This alters the ionic bonds of the enzyme that contribute to its functional shape.
• The salt concentration: Each enzyme has an optimal salt concentration. Changes in the salt concentration may also denature enzymes.
Some relationships between bacterial enzymes and the use of disinfectants and extremes of temperature to control bacteria.
1. Many disinfectants, such as chlorine, iodine, iodophores, mercurials, silver nitrate, formaldehyde, and ethylene oxide, inactivate bacterial enzymes and thus block metabolism.
2. High temperatures, such as autoclaving, boiling, and pasteurization, denature proteins and enzymes.
3. Cold temperatures, such as refrigeration and freezing, slow down or stop enzyme reactions.
Summary
1. Enzymes are substances present in the cell in small amounts that function to speed up or catalyze chemical reactions so they occur fast enough to support life.
2. On the surface of the enzyme is typically a small crevice that functions as an active site or catalytic site to which one or two specific substrates are able to bind.
3. Anything that an enzyme normally combines with is called a substrate.
4. The binding of the substrate to the enzyme causes the flexible enzyme to change its shape slightly through a process called induced fit to form a temporary intermediate called an enzyme-substrate complex.
5. Enzymes speed up the rate of chemical reactions because they lower the energy of activation, the energy that must be supplied in order for molecules to react with one another.
6. Many enzymes require a nonprotein cofactor to assist them in their reaction. In this case, the protein portion of the enzyme, called an apoenzyme, combines with the cofactor to form the whole enzyme or haloenzyme.
7. Some cofactors are ions such as Ca++, Mg++, and K+; other cofactors are organic molecules called coenzymes which serve as carriers for chemical groups or electrons. NAD+, NADP+, FAD, and coenzyme A (CoA) are examples of coenzymes.
8. Chemically, enzymes are generally globular proteins. Some RNA molecules called ribozymes can also be enzymes, usually functioning to cleave RNA molecules.
9. Enzymes are only present in small amounts in the cell since they are not altered during their reactions and are highly specific for their substrate.
10. Enzyme activity is affected by a number of factors including the concentration of the enzyme, the concentration of the substrate, the temperature, the pH, and the salt concentration. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/19%3A_Review_of_Molecular_Genetics/19.2%3A_Enzymes.txt |
Learning Objectives
1. State the three basic parts of a deoxyribonucleotide.
2. State which nitrogenous bases are purines and which are pyrimidines.
3. Define complementary base pairing.
4. State why DNA can only be synthesized in a 5' to 3' direction.
5. Compare the prokaryotic nucleoid with the eukaryotic nucleus in terms of the following:
1. number of chromosomes
2. linear or circular chromosomes
3. presence or absence of a nuclear membrane
4. presence or absence of nucleosomes
5. presence or absence of mitosis
6. presence or absence of meiosis
DNA is a long, double-stranded, helical molecule composed of building blocks called deoxyribonucleotides. Each deoxyribonucleotide is composed of three parts: a molecule of the 5-carbon sugar deoxyribose, a nitrogenous base, and a phosphate group (Figure \(1\)).
• Deoxyribose. Deoxyribose is a ringed 5-carbon sugar (Figure \(2\)). The 5 carbons are numbered sequentially clockwise around the sugar. The first 4 carbons actually form the ring of the sugar with the 5' carbon coming off of the 4' carbon in the ring. The nitrogenous base of the nucleotide is attached to the 1' carbon of the sugar and the phosphate group is bound to the 5' carbon. During DNA synthesis, the phosphate group of a new deoxyribonucleotide is covalently attached by the enzyme DNA polymerase to the 3' carbon of a nucleotide already in the chain.
• A nitrogenous base. There are four nitrogenous bases found in DNA: adenine, guanine, cytosine, or thymine. Adenine and guanine are known as purine bases while cytosine and thymine are known as pyrimidine bases (Figure \(3\)).
• A phosphate group (Figure \(4\)).
To synthesize the two chains of deoxyribonucleotides during DNA replication, the DNA polymerase enzymes involved are only able to join the phosphate group at the 5' carbon of a new nucleotide to the hydroxyl (OH) group of the 3' carbon of a nucleotide (Figure \(2\)) already in the chain. The covalent bond that joins the nucleotides is called a phosphodiester bond. Each DNA strand has what is called a 5' end and a 3' end. This means that one end of each DNA strand, called the 5' end , will always have a phosphate group attached to the 5' carbon of its terminal deoxyribonucleotide (Figure \(5\)). The other end of that strand, called the 3' end, will always have a hydroxyl (OH) on the 3' carbon of its terminal deoxyribonulceotide.
As will be seen in the next section, each parent strand, during DNA replication, acts as a template for the synthesis of the other strand by way of complementary base pairing. Complementary base pairing refers to DNA nucleotides with the base adenine only forming hydrogen bonds with nucleotides having the base thymine (A-T). Likewise, nucleotides with the base guanine can hydrogen bond only with nucleotides having the base cytosine (G-C). (In the case of RNA nucleotides, as will be seen later, adenine nucleotides form hydrogen bonds with nucleotides having the base uracil since thymine is not found in RNA.) As a result of this bonding, the DNA assumes its helical shape. Therefore, the two strands of DNA are said to be complementary. Wherever one strand has an adenine-containing nucleotide, the opposite strand will always have a thymine nucleotide; wherever there is a guanine-containing nucleotide, the opposite strand will always have a cytosine nucleotide (Figure \(1\)).
While the two strands of DNA are complementary, they are oriented in opposite directions to each other. One strand is said to run 5' to 3'; the opposite DNA strand runs antiparallel, or 3' to 5' (Figure \(1\)).
We will now briefly compare the genome of prokaryotic cells with that of eukaryotic cells.
The Prokaryotic (Bacterial) Genome
The area within a bacterium where the chromosome can be seen with an electron microscope is called a nucleoid. The chromosome of most prokaryotes is typically one long, single molecule of double stranded, helical, supercoiled DNA which forms a physical and genetic circle. The chromosome is generally around 1000 µm long and frequently contains around 4000 genes (Figure \(8\)). Escherichia coli, which is 2-3 µm in length has a chromosome approximately 1400 µm long. To enable a macromolecule this large to fit within the bacterium, histone-like proteins bind to the DNA, segregating the DNA molecule into around 50 chromosomal domains and making it more compact. A DNA topoisomerase enzyme called DNA gyrase then supercoils the chromosome into a tight bundle forming a compacted, supercoiled mass of DNA approximately 0.2 µm in diameter.
Bacterial enzymes called DNA topoisomerases are essential in the unwinding, replication, and rewinding of the circular, supercoiled bacterial DNA (Figure \(7\)). They are also essential in transcription of DNA into RNA, in DNA repair, and in genetic recombination in bacteria.
Figure \(7\): Circular, Supercoiled Prokaryotic DNA. To enable the large DNA molecyle to fit within the bacterium, a DNA topoisomerase enzyme called DNA gyrase supercoils the chromosome into a tight bundle forming a compacted, supercoiled mass of DNA approximately 0.2 µm in diameter.
The prokaryotic nucleoid has no nuclear membrane surrounding the DNA and the nuclear body does not divide by mitosis. The cytoplasmic membrane plays a role in DNA separation during bacterial replication. Since bacteria are haploid (have only one chromosome), there is also no meiosis.
The Eukaryotic Genome
Prokaryotic and eukaryotic cells differ a great detail in both the amount and the organization of their molecules of DNA. Eukaryotic cells contain much more DNA than do bacteria, and this DNA is organized as multiple chromosomes located within a nucleus.
The nucleus in eukaryotic cells is surrounded by a nuclear membrane (Figure \(7\)) and contains linear chromosomes composed of negatively charged DNA associated with positively charged basic proteins called histones to form structures known as nucleosomes. The nucleosomes are part of what is called chromatin, the DNA and proteins that make up the chromosomes. The nucleus divides my mitosis and haploid sex cells are produced from diploid cells by meiosis.
The DNA in eukaryotic cells is packaged in a highly organized way. It consists of a basic unit called a nucleosome, a beadlike structure 11 nm in diameter that consists of 146 base pairs of DNA wrapped around eight histone molecules. The nucleosomes are linked to one another by a segment of DNA approximately 60 base pairs long called linker DNA (Figure \(9\)). Another histone associated with the linker DNA then packages adjacent nucleotides together to form a nucleosome thread 30nm in diameter. Finally, these packaged nucleosome threads form large coiled loops that are held together by nonhistone scaffolding proteins. These coiled loops on the scaffolding proteins interact to form the condensed chromatin seen in chromosomes during mitosis (Figure \(10\)).
In recent years its been found that the structural nature of the deoxyribonucleoprotein contributes to whether or not DNA is transcribed into RNA. For example, chemical changes to the chromatin can enable portions of it to condense or relax. When a region is condensed, genes cannot be transcribed. In addition, chemical can attach to or be removed from the histone proteins around which the DNA wraps. The attachment or removal of these chemical groups to the histone determines whether nearby gene expression is amplified or repressed.
The epigenome refers to a variety of chemical compounds that modify the genome typically by adding a methyl (CH3) group to the nucleotide base adenine at specific locations along the DNA molecule. This methylation can, in turn, either repress or activate transcription of specific genes. By basically turning genes on or off, the epigenome enables the genome to interact with and respond to the cell's environment. The epigenome can be inherited just like the genome.
Summary
1. Deoxyribonucleic acid (DNA) is a long, double-stranded, helical molecule composed of building blocks called deoxyribonucleotides.
2. A deoxyribonucleotide is composed of 3 parts: a molecule of the 5-carbon sugar deoxyribose, a nitrogenous base, and a phosphate group.
3. There are four nitrogenous bases found in DNA: adenine, guanine, cytosine, or thymine. Adenine and guanine are known as purine bases while cytosine and thymine are known as pyrimidine bases.
4. Deoxyribose is a ringed 5-carbon sugar. The 5 carbons are numbered sequentially clockwise around the sugar. The first 4 carbons actually form the ring of the sugar with the 5' carbon coming off of the 4' carbon in the ring. The nitrogenous base of the nucleotide is attached to the 1' carbon of the sugar and the phosphate group is bound to the 5' carbon.
5. During DNA synthesis, the enzyme DNA polymerase can only attach the phosphate group of a new deoxyribonucleotide to the 3' carbon of a nucleotide already in the chain.
6. During DNA replication, each parent strand acts as a template for the synthesis of the other strand by way of complementary base pairing.
7. Complementary base pairing refers to DNA nucleotides with the base adenine only forming hydrogen bonds with nucleotides having the base thymine (A-T). Likewise, nucleotides with the base guanine can hydrogen bond only with nucleotides having the base cytosine (G-C).
8. While the two strands of DNA are complementary, they are oriented in opposite directions to each other. One strand is said to run 5' to 3'; the opposite DNA strand runs antiparallel, or 3' to 5'.
9. In prokaryotic cells there is no nuclear membrane surrounding the DNA. Prokaryotic cells lack mitosis and meiosis.
10. To enable a macromolecule this large to fit within the bacterium, histone-like proteins bind to the DNA, segregating the DNA molecule into around 50 chromosomal domains and making it more compact. Then an enzyme called DNA gyrase supercoils each domain around itself forming a compacted, supercoiled mass of DNA. A topoisomerase called DNA gyrase catalyzes the negative supercoiling of the circular DNA found in bacteria. Topoisomerase IV, on the other hand, is involved in the relaxation of the supercoiled circular DNA, enabling the separation of the interlinked daughter chromosomes at the end of bacterial DNA replication.
11. The DNA in eukaryotic cells is packaged in basic units called a nucleosomes, a beadlike structure consisting of DNA wrapped around eight histone molecules. The DNA is organized as multiple chromosomes located within a nucleus surrounded by a nuclear membrane. The nucleus divides by mitosis and gametes are produced by meiosis in eukaryotes reproducing sexually.
12. The structural nature of the deoxyribonucleoprtein contributes to whether or not DNA is transcribed into RNA. For example, chemical changes to the chromatin can enable portions of it to condense or relax. When a region is condensed, genes cannot be transcribed. In addition, chemical can attach to or be removed from the histone proteins around which the DNA wraps. The attachment or removal of these chemical groups to the histone determines whether nearby gene expression is amplified or repressed. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/19%3A_Review_of_Molecular_Genetics/19.3%3A_Deoxyribonucleic_Acid_%28DNA%29.txt |
Learning Objectives
1. Briefly describe the process of DNA replication.
2. State why DNA can only be synthesized in a 5' to 3' direction.
3. State the function of the following enzymes in bacterial DNA replication:
1. DNA polymeraseIII
2. DNA polymerase II
3. DNA helicase
4. primase
5. DNA ligase
In general, DNA is replicated by uncoiling of the helix, strand separation by breaking of the hydrogen bonds between the complementary strands, and synthesis of two new strands by complementary base pairing. Replication begins at a specific site in the DNA called the origin of replication (oriC). DNA replication is bidirectional from the origin of replication. To begin DNA replication, unwinding enzymes called DNA helicases cause short segments of the two parent DNA strands to unwind and separate from one another at the origin of replication to form two "Y"-shaped replication forks. These replication forks are the actual site of DNA copying (Figure \(1\)).
All the proteins involved in DNA replication aggregate at the replication forks to form a replication complex called a replisome (Figure \(2\)).
Single-strand binding proteins bind to the single-stranded regions so the two strands do not rejoin. Unwinding of the double-stranded helix generates positive supercoils ahead of the replication fork. Enzymes called topoisomerases counteract this by producing breaks in the DNA and then rejoin them to form negative supercoils in order to relieve this stress in the helical molecule during replication.
As the strands continue to unwind and separate in both directions around the entire DNA molecule, new complementary strands are produced by the hydrogen bonding of free DNA nucleotides with those on each parent strand. As the new nucleotides line up opposite each parent strand by hydrogen bonding, enzymes called DNA polymerases join the nucleotides by way of phosphodiester bonds. Actually, the nucleotides lining up by complementary base pairing are deoxynucleotide triphosphates, composed of a nitrogenous base, deoxyribose, and three phosphates. As the phosphodiester bond forms between the 5' phosphate group of the new nucleotide and the 3' OH of the last nucleotide in the DNA strand, two of the phosphates are removed providing energy for bonding (Figure \(3\)). In the end, each parent strand serves as a template to synthesize a complementary copy of itself, resulting in the formation of two identical DNA molecules (Figure \(4\)).
In bacteria, Par proteins function to separate bacterial chromosomes to opposite poles of the cell during cell division. They bind to the origin of replication of the DNA and physically pull or push the chromosomes apart, similar to the mitotic apparatus of eukaryotic cells. Fts proteins, such as FtsK in the divisome, also help in separating the replicated bacterial chromosome.
Animation: Replication of DNA by Complementary Base Pairing. As the DNA strands unwind and separate, new complementary strands are produced by the hydrogen bonding of free DNA nucleotides with those on each parent strand. As the new nucleotides line up opposite each parent strand by hydrogen bonding, enzymes called DNA polymerases join the nucleotides by way of phosphodiester bonds. The DNA polymerase responsible for these events is not shown here.
In reality, DNA replication is more complicated than this because of the nature of the DNA polmerases. DNA polymerase enzymes are only able to join the phosphate group at the 5' carbon of a new nucleotide to the hydroxyl (OH) group of the 3' carbon of a nucleotide already in the chain. As a result, DNA can only be synthesized in a 5' to 3' direction while copying a parent strand running in a 3' to 5' direction.
Remember, as mentioned above, each DNA strand has two ends. The 5' end of the DNA is the one with the terminal phosphate group on the 5' carbon of the deoxyribose; the 3' end is the one with a terminal hydroxyl (OH) group on the deoxyribose of the 3' carbon of the deoxyribose. The two strands are antiparallel, that is they run in opposite directions. Therefore, one parent strand - the one running 3' to 5' and called the leading strand - can be copied directly down its entire length (Figure \(5\)).
However, the other parent strand - the one running 5' to 3' and called the lagging strand - must be copied discontinuously in short fragments (Okazaki fragments) of around 100-1000 nucleotides each as the DNA unwinds. This occurs, as mentioned above, at the replisome. The lagging DNA strand loops out from the leading strand and this enables the replisome to move along both strands pulling the DNA through as replication occurs. It is the actual DNA, not the DNA polymerase that moves during bacterial DNA replication (Figure \(2\)).
In addition, DNA polymerase enzymes cannot begin a new DNA chain from scratch. They can only attach new nucleotides onto 3' OH group of a nucleotide in a preexisting strand. Therefore, to start the synthesis of the leading strand and each DNA fragment of the lagging strand, an RNA polymerase complex called a primase is required. The primase, which is capable of joining RNA nucleotides without requiring a preexisting strand of nucleic acid, first adds several complementary RNA nucleotides opposite the DNA nucleotides on the parent strand. This forms what is called an RNA primer (Figure \(6\)).
DNA polymerase III then replaces the primase and is able to add DNA nucleotides to the RNA primer (Figure \(7\)).
Later, DNA polymerase II digests away the RNA primer and replaces the RNA nucleotides of the primer with the proper DNA nucleotides to fill the gap (Figure \(8\)).
Finally, the DNA fragments themselves are hooked together by the enzyme DNA ligase (Figure \(6\)). Yet even with this complicated procedure, a 1000 micrometer-long macromolecule of tightly-packed, supercoiled DNA can make an exact copy of itself in only about 10 minutes time under optimum conditions, inserting nucleotides at a rate of about 1000 nucleotides per second!
Animation: Replication of Leading and Lagging DNA Strands. The leading strand is made continuously in a 5' to 3' direction by DNA polymerase III as the DNA helicase unwinds the parental DNA helix. However, because the parental DNA strands are antiparallel, the lagging strand must be made in short fragments. RNA polymerase (primase) synthesizes a short RNA primer which is extended by DNA polymerase III. DNA polymerase II then digests the RNA primer and replaces it with DNA. Finally, DNA ligase joins the fragments of the lagging strand together.
There is a great deal of genetic information in the bacterial chromosome. For example Escherichia coli, the most studied of all bacteria, has a genome containing 4,639,221 base pairs, which code for at least 4288 proteins.
Summary
1. During DNA replication, each parent strand acts as a template for the synthesis of the other strand by way of complementary base pairing.
2. Complementary base pairing refers to DNA nucleotides with the base adenine only forming hydrogen bonds with nucleotides having the base thymine (A-T). Likewise, nucleotides with the base guanine can hydrogen bond only with nucleotides having the base cytosine (G-C).
3. Each DNA strand has two ends. The 5' end of the DNA is the one with the terminal phosphate group on the 5' carbon of the deoxyribose; the 3' end is the one with a terminal hydroxyl (OH) group on the deoxyribose of the 3' carbon of the deoxyribose.
4. To synthesize the two chains of deoxyribonucleotides during DNA replication, the DNA polymerase enzymes involved are only able to join the phosphate group at the 5' carbon of a new nucleotide to the hydroxyl (OH) group of the 3' carbon of a nucleotide already in the chain.
5. While the two strands of DNA are complementary, they are oriented in opposite directions to each other. One strand is said to run 5' to 3'; the opposite DNA strand runs antiparallel, or 3' to 5'.
6. To begin DNA replication, unwinding enzymes called DNA helicases cause short segments of the two parent DNA strands to unwind and separate from one another at the origin of replication to form two "Y"-shaped replication forks.
7. Single-strand binding proteins bind to the now unpaired single-stranded regions so the two strands do not rejoin.
8. As the strands continue to unwind and separate in both directions around the entire DNA molecule, new complementary strands are produced by the hydrogen bonding of free DNA nucleotides with those on each parent strand.
9. As the new nucleotides line up opposite each parent strand by hydrogen bonding, enzymes called DNA polymerases join the nucleotides by way of phosphodiester bonds.
10. The two strands are antiparallel, that is they run in opposite directions. Therefore, one parent strand - the one running 3' to 5' and called the leading strand- can be copied directly down its entire length. However, the other parent strand - the one running 5' to 3' and called the lagging strand- must be copied discontinuously in short fragments (Okazaki fragments) of around 100-1000 nucleotides each as the DNA unwinds.
11. Furthermore, DNA polymerase enzymes cannot begin a new DNA chain from scratch. They can only attach new nucleotides onto 3' OH group of a nucleotide in a preexisting strand. Therefore, to start the synthesis of the leading strand and each DNA fragment of the lagging strand, an RNA polymerase complex called a primase is required. The primase, which is capable of joining RNA nucleotides without requiring a preexisting strand of nucleic acid, first adds several comlementary RNA nucleotides opposite the DNA nucleotides on the parent strand forming what is called an RNA primer.
12. DNA polymerase III then replaces the primase and is able to add DNA nucleotides to the RNA primer. Later, DNA polymerase II digests away the RNA primer and replaces the RNA nucleotides of the primer with the proper DNA nucleotides to fill the gap.
13. The DNA fragments themselves are hooked together by the enzyme DNA ligase to complete the process. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/19%3A_Review_of_Molecular_Genetics/19.4%3A_DNA_Replication_in_Prokaryotic_Cells.txt |
Learning Objectives
1. Briefly describe the process of DNA replication.
2. Compare prokaryotic and eukaryotic DNA replication in terms of origins of replication.
3. Define telomeres and state whether they are found in prokaryotic or eukaryotic DNA.
4. Name the stages of mitosis and state what happens during each.
As in prokaryotes, the linear chromosomes of eukaryotes replicate by strand separation and complementary base pairing of free deoxyribonucleotides with those on each parent DNA strand. As with prokaryotes, DNA replication in eukaryotic cells is bidirectional. However, unlike the circular DNA in prokaryotic cells that usually has a single origin of replication, the linear DNA of a eukaryotic cell contains multiple origins of replication (Figure \(11\)).
As discussed earlier under prokaryotic DNA replication, DNA can only be synthesized in a 5' to 3' direction and all DNA polymerase requires a primer. To solve this problem, the ends of the linear eukaryotic DNA strands, called telomeres , have short, repetitive, noncoding DNA base sequences. A unique enzyme called telomerase binds to the telomeric DNA at the 3' end. The telomerase contains a small RNA template as a cofactor which is copied by DNA nucleotides to extend the 3' end. Once the extension is long enough, primase can assemble a short RNA primer on the lagging strand and DNA replication can proceed in a manner similar to the lagging strand of prokaryotic DNA.
Animation: Replication of DNA by Complementary Base Pairing. As the DNA strands unwind and separate, new complementary strands are produced by the hydrogen bonding of free DNA nucleotides with those on each parent strand. As the new nucleotides line up opposite each parent strand by hydrogen bonding, enzymes called DNA polymerases join the nucleotides by way of phosphodiester bonds. The DNA polymerase responsible for these events is not shown here.
Once the chromosomes have replicated, the nucleus divides by mitosis (see Figure \(12\) through 16). The eukaryotic cell cycle is divided into two major phases: interphase and cell division.
Interphase
Ninety percent or more of the cell cycle is spent in interphase. During interphase, cellular organelles double in number, the DNA replicates, and protein synthesis occurs. The chromosomes are not visible and the DNA appears as uncoiled chromatin.
Interphase in a plant cell: see Figure \(17\)
Interphase in an animal cell: see Figure \(18\)
Interphase is divided into the following stages: G1, S, and G2.
1. G1 phase: During G1 phase, the period that immediately follows cell division, the cell grows and differentiates. New organelles are made but the chromosomes have not yet replicated in preparation for cell division.
2. S phase: DNA synthesis occurs during S phase. The chromosomes replicate in preparation for cell division.
3. G2 phase: During G2 phase, molecules that will be required for cell replication are synthesized.
Cell Division
Cell division consists of nuclear division and cytoplasmic division. Nuclear division is referred to as mitosis while cytoplasmic division is called cytokenesis.
1. Mitosis (nuclear division)
Mitosis is the nuclear division process in eukaryotic cells and ensures that each daughter cell receives the same number of chromosomes as the original parent cell. Mitosis can be divided into the following phases: prophase, metaphase, anaphase, and telophase.
a. Prophase: During prophase, the chromatin condenses and the chromosomes become visible. Also the nucleolus disappears, the nuclear membrane fragments, and the spindle apparatus forms and attaches to the centromeres of the chromosomes.
Prophase in a plant cell: see Figure \(19\) and Figure \(20\)
Prophase in an animal cell: see Figure \(21\) and Figure \(22\)
b. Metaphase: During metaphase, the nuclear membrane fragmentation is complete and the duplicated chromosomes line up along the cell's equator.
Metaphase in a plant cell: see Figure \(23\)
Metaphase in an animal cell: see Figure \(24\)
c. Anaphase: During anaphase, diploid sets of daughter chromosomes separate and are pushed and pulled toward opposite poles of the cell. This is accomplished by the polymerization and depolymerization of the microtubules that help to form the spindle apparatus.
Anaphase in a plant cell: see Figure \(25\) and Figure \(26\)
d. Telophase: During telophase, the nuclear membrane and nucleoli reform, cytokinesis is nearly complete, and the chromosomes eventually uncoil to chromatin. Usually cytokinesis occurs during telophase.
Telophase in a plant cell: see Figure \(28\) and Figure \(29\)
Telophase in an animal cell: see Figure \(30\)
YouTube movie illustrating mitosis.
2. Cytokinesis (cytoplasmic division)
During cytokinesis, the dividing cell separates into two diploid daughter cells. In animal cells, which lack a cell wall and are surrounded only by a cytoplasmic membrane, microfilaments of actin and myosin attached to the membrane form constricting rings around the central portion of the dividing cell and eventually divide the cytoplasm into two daughter cells. In the case of plant cells , which are surrounded by a cell wall in addition to the cytoplasmic membrane, carbohydrate-filled vesicles accumulate and fuse along the equator of the cell forming a cell plate that separates the cytoplasm into two daughter cells.
Summary
1. During DNA replication, each parent strand acts as a template for the synthesis of the other strand by way of complementary base pairing.
2. Complementary base pairing refers to DNA nucleotides with the base adenine only forming hydrogen bonds with nucleotides having the base thymine (A-T). Likewise, nucleotides with the base guanine can hydrogen bond only with nucleotides having the base cytosine (G-C).
3. Each DNA strand has two ends. The 5' end of the DNA is the one with the terminal phosphate group on the 5' carbon of the deoxyribose; the 3' end is the one with a terminal hydroxyl (OH) group on the deoxyribose of the 3' carbon of the deoxyribose.
4. To synthesize the two chains of deoxyribonucleotides during DNA replication, the DNA polymerase enzymes involved are only able to join the phosphate group at the 5' carbon of a new nucleotide to the hydroxyl (OH) group of the 3' carbon of a nucleotide already in the chain.
5. While the two strands of DNA are complementary, they are oriented in opposite directions to each other. One strand is said to run 5' to 3'; the opposite DNA strand runs antiparallel, or 3' to 5'.
6. Unlike the circular DNA in prokaryotic cells that usually has a single origin of replication, the linear DNA of a eukaryotic cell contains multiple origins of replication.
7. Because DNA can only be synthesized in a 5' to 3' direction and all DNA polymerase requires a primer, the ends of the linear eukaryotic DNA strands, called telomeres, have short, repetitive, noncoding DNA base sequences. A unique enzyme called telomerase binds to the telomeric DNA at the 3' end. The telomerase contains a small RNA template as a cofactor which is copied by DNA nucleotides to extend the 3' end. Once the extension is long enough, primase can assemble a short RNA primer on the lagging strand and DNA replication can proceed in a manner similar to the lagging strand of prokaryotic DNA.
8. Once the chromosomes have replicated, the nucleus divides by mitosis.
9. During interphase, cellular organelles double in number, the DNA replicates, and protein synthesis occurs. The chromosomes are not visible and the DNA appears as uncoiled chromatin.
10. During G1 phase, the period that immediately follows cell division, the cell grows and differentiates and new organelles are made.
11. DNA synthesis (chromosome replication) occurs during S phase.
12. During G2 phase, molecules that will be required for cell replication are synthesized.
13. Nuclear division is referred to as mitosis while cytoplasmic division is called cytokenesis.
14. During prophase, the chromatin condenses and the chromosomes become visible, the nucleolus disappears, the nuclear membrane fragments, and the spindle apparatus forms and attaches to the centromeres of the chromosomes.
15. During metaphase, the nuclear membrane fragmentation is complete and the duplicated chromosomes line up along the cell's equator.
16. During anaphase, diploid sets of daughter chromosomes separate and are pushed and pulled toward opposite poles of the cell.
17. During telophase, the nuclear membrane and nucleoli reform, cytokinesis is nearly complete, and the chromosomes eventually uncoil to chromatin.
18. During cytokinesis, the dividing cell separates into two diploid daughter cells. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/19%3A_Review_of_Molecular_Genetics/19.5%3A_DNA_Replication_in_Eukaryotic_Cells_and_the_Eukaryotic_Cell_Cycle.txt |
Learning Objectives
1. State the 3 basic parts of a ribonucleotide.
2. State 3 ways RNA differs from DNA.
3. State the function of each of the following:
1. tRNA
2. mRNA
3. rRNA
RNA is a single-stranded molecule composed of building blocks called ribonucleotides. A ribonucleotide is composed of three parts: a molecule of the sugar ribose, a nitrogenous base, and a phosphate group (Figure \(1\)).
Ribose is a ringed 5-carbon sugar (Figure \(2\)) similar to deoxyribose except it has a hydroxyl (OH) group) on its 2' carbon. The nitrogenous base is attached to the 1' carbon of the sugar and the phosphate group is bound to the 5' carbon. During RNA synthesis, the phosphate group of a new ribonucleotide is attached by the enzyme RNA polymerase to the 3' carbon of a ribonucleotide.
There are four nitrogenous bases found in RNA: adenine, guanine, cytosine, or uracil. Adenine and guanine are known as purine bases while cytosine and uracil are known as pyrimidine bases (Figure \(3\)).
A phosphate group (Figure \(4\)).
RNA differs from DNA in several ways. First of all, RNA is single-stranded, not double-stranded. Unlike DNA polymerases, RNA polymerases are able to join RNA nucleotides together without requiring a preexisting strand of RNA. In addition, RNA has the base uracil in place of thymine. Uracil, like thymine, can form hydrogen bond with adenine. Also, RNA and has the sugar ribose instead of deoxyribose. Finally, there are three functionally different types of RNA:
• Messenger RNA (mRNA): Messenger RNA copies the genetic information in the DNA by complementary base pairing and carries this "message" to the ribosomes where the proteins are assembled.
• Transfer RNA (tRNA): Transfer RNAs picks up specific amino acids, transfers the amino acids to the ribosomes, and insert the correct amino acids in the proper place according to the mRNA message.
• Ribosomal RNA (rRNA): Ribosomal RNA and ribosomal proteins form the ribosomal subunits.
• Other RNA transcripts: A variety of other RNA molecules transcribed off of DNA have also been found. These RNA molecules are not translated into proteins, but rather perform a wide range of direct genetic regulatory functions. Examples include antisense RNAs, microRNAs, and riboswitch RNAs.
RNA has the base uracil in place of thymine in DNA.
Summary
1. RNA is a single-stranded molecule composed of building blocks called ribonucleotides.
2. A ribonucleotide is composed of 3 parts: a molecule of the sugar ribose, a nitrogenous base, and a phosphate group.
3. RNA differs from DNA in several ways: RNA is single-stranded, not double-stranded; unlike DNA polymerases, RNA polymerases are able to join RNA nucleotides together without requiring a preexisting strand of RNA; RNA has the base uracil in place of thymine, but like thymine, uracil can form hydrogen bond with adenine; and RNA and has the sugar ribose instead of deoxyribose.
4. There are three functionally different types of RNA: messenger RNA (mRNA), transfer RNA (tRNA), and ribosomal RNA (rRNA).
5. Messenger RNA copies the genetic information in the DNA by complementary base pairing and carries this "message" to the ribosomes where the proteins are assembled.
6. Transfer RNAs picks up specific amino acids, transfers the amino acids to the ribosomes, and insert the correct amino acids in the proper place according to the mRNA message.
7. Ribosomal RNA and ribosomal proteins form the ribosomal subunits.
8. A variety of other RNA molecules transcribed off of DNA have also been found, including antisense RNAs, microRNAs, and riboswitch RNAs. These RNA molecules are not translated into proteins but rather perform a wide range of direct genetic regulatory functions | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/19%3A_Review_of_Molecular_Genetics/19.6%3A_Ribonucleic_Acid_%28RNA%29.txt |
DNA is divided into functional units called genes. A gene is a segment of DNA that codes for a functional product (mRNA, tRNA, or rRNA). Since the vast majority of genes are transcribed into mRNA and mRNA is subsequently translated into polypeptides or proteins, most genes code for protein synthesis. The term polypeptide refers to many amino acids connected by peptide bonds. While all proteins are polypeptides, not all polypeptides are proteins. In some cases, smaller polypeptides coded for by two or more genes must be joined together to produce a functional protein. In other cases, as will be mentioned below, mRNA carries a transcript of several genes resulting in the synthesis of a large polypeptide that must subsequently be cleaved by enzymes called proteases into two or more smaller functional proteins. For simplicity, we will use the term protein when referring to the end product of transcription and translation. In this section we will see how the sequence of deoxyribonucleotide bases along one strand of DNA ultimately codes for the amino acid sequence of a particular polypeptide or protein.
During protein synthesis, the order of nucleotide bases along a gene gets transcribed into a complementary strand of mRNA which is then translated by tRNA into the correct order of amino acids for that polypeptide or protein. Therefore, the order of deoxyribonucleotide bases along the DNA determines the order of amino acids in the proteins, that is, its primary structure. Because certain amino acids can interact with other amino acids, the order of amino acids for each protein determines its final three-dimensional shape, which in turn determines the function of that protein. Protein synthesis can be divided into two stages: transcription and translation. In the next two sections we will look at these stages in greater detail.
19.7: Polypeptide and Protein Synthesis
Learning Objectives
1. Define the following:
1. gene
2. transcription
2. Briefly describethe function of the following in terms of transcription:
1. mRNA
2. 3' end
3. 5' end
4. RNA polymerase
5. phosphodiester bond
6. promoter
7. leader sequence
8. coding sequence
9. transcription terminator
10. codon
3. Define the following in terms of transcription in eukaryotic cells:
1. introns
2. exons
3. precurser mRNA
4. cap
5. poly-A tail
6. mature mRNA
Transcription in Prokaryotic Cells
Description: Messenger RNA (mRNA) is synthesized by complementary base pairing of ribonucleotides with deoxyribonucleotides to match a portion of one strand of DNA called a gene. Although genes are present on both strands of DNA, only one strand is transcribed for any given gene. Following transcription, 30S and 50S ribosomal subunits attach to the mRNA and tRNA inserts the correct amino acids which are subsequently joined to form a polypeptide or a protein through a process called translation.
The enzyme RNA polymerase transcribes DNA. This enzyme initiates transcription, joins the RNA nucleotides together, and terminates transcription. To initiate transcription in bacteria, a variety of proteins called sigma factors bind to RNA polymerases. This complex can then bind to a specific sequence of usually about 40 deoxyribonucleotide bases called the promoter located along the DNA prior to the coding region of the gene. The promotor determines what region of the DNA and which strand of DNA will be transcribed into RNA.
Like DNA polymerase, RNA polymerase can only synthesize nucleic acid in a 5' to 3' direction while "reading" a DNA template in the 3' to 5' direction. As mentioned earlier in this unit, the 3' end of a strand of nucleic acid has a hydroxyl (OH) group on the 3' carbon of the deoxyribose or ribose and is not linked to another nucleotide. The 5' end of that strand has a phosphate group attached to the 5' carbon of the sugar and is not linked to another nucleotide (Figure \(1\)).
Once the RNA polymerase/sigma factor complex recognizes the correct promoter, the sigma factor dissociate from the RNA polymerase and the enzyme begins to unwind the helix of the DNA creating a region of nonpaired deoxyribonucleotides that serve as a template for RNA synthesis (Figures 2 and 3).
Unwinding of the DNA Helix by RNA Polymerase
Once the RNA polymerase/sigma factor complex recognizes the correct promoter, the sigma factor dissociate from the RNA polymerase and the enzyme begins to unwind the helix of the DNA creating a region of nonpaired deoxyribonucleotides that serve as a template for RNA synthesis
While the RNA polymerase does not transcribe the promoter itself, it does transcribe a short noncoding leader sequence just prior to the coding sequence of the gene. The leader sequence is the portion of DNA that is transcribed into the ribosome-binding site of the mRNA (below under translation.) The coding sequence contains the actual message for protein synthesis.
Once the actual transcription begins, ribonucleotides containing 3 phosphate groups hydrogen bond through the process of complementary base pairing with the exposed deoxyribonucleotides on the unwound strand that is to be transcribed (Figure \(4\)). The ribonucleotides are then covalently bonded together by phosphodiester bonds, the energy being supplied by the cleavage of two phosphate groups from the ribonucleotide triphosphate (Figure \(5\)). (The phosphodiester bond refers to the phosphate on the 5'C of the newly inserted nucleotide covalently bonding to the 3'C of the last ribonucleotide in the mRNA chain.) The mRNA polymerizes at a rate of about 30 nucleotides per second.
As the RNA polymerase moves down the DNA, the previous stretch of DNA again pairs with its complementary strand. This process continues until the RNA polymerase encounters a "stop" signal or transcription terminator at the end of the gene. This causes the completed mRNA to drop off the gene.
Transcription of mRNA Complementary to DNA
Once the RNA polymerase/sigma factor complex recognizes the correct promoter, the sigma factor dissociate from the RNA polymerase and the enzyme begins to unwind the helix of the DNA creating a region of nonpaired deoxyribonucleotides that serve as a template for RNA synthesis. Transcription is under control of the enzyme RNA polymerase which is not shown here.
Once the RNA polymerase moves beyond the promotor region, a new molecule of RNA polymerase can bind to the promotor and start a new round of transcription. In this way, a single gene can be transcribed multiple times.
YouTube movie illustrating complementary base pairing during transcription.
YouTube movie illustrating transcription and protein assembly.
YouTube movie illustrating transcription in bacteria
There are 22 amino acids that can be encoded by the genetic information carried on mRNA. The mRNA molecule is divided up into codons. A codon is a series of three consecutive mRNA bases coding for one specific amino acid. The various codons and the amino acids for which they code are shown in Table \(16\).8.1. There are 64 codons. One codon, AUG, also serves as a start codon to initiate translation, and three codons, UAG, UAA, and UGA, function as stop or nonsense codons to terminate translation. (Alternative start codons are different from the standard AUG codon and are found occasionally in both prokaryotes and eukaryotes.)
Table \(16\).8.1: The Genetic Code - Codons
U
C
A
G
U
UUU = Phe
UUC = Phe
UUA = Leu
UUG = Leu
UCU = Ser
UCC = Ser
UCA = Ser
UCG = Ser
UAU = Tyr
UAC = Tyr
UAA = Stop
UAG = Stop
UGU = Cys
UGC = Cys
UGA = Stop
UGG = Trp
U
C
A
G
C
CUU = Leu
CUC = Leu
CUA = Leu
CUG = Leu
CCU = Pro
CCC = Pro
CCA = Pro
CCG = Pro
CAU = His
CAC = His
CAA = Gln
CAG = Gln
CGU = Arg
CGC = Arg
CGA = Arg
CGG = Arg
U
C
A
G
A
AUU = Ile
AUC = Ile
AUA = Ile
AUG = Met
ACU = Thr
ACC = Thr
ACA = Thr
ACG = Thr
AAU = Asn
AAC = Asn
AAA = Lys
AAG = Lys
AGU = Ser
AGC = Ser
AGA = Arg
AGG = Arg
U
C
A
G
G
GUU = Val
CUC = Val
GUA = Val
GUG = Val
GCU = Ala
GCC = Ala
GCA = Ala
GCG = Ala
GAU = Asp
GAC = Asp
GAA = Glu
GAG = Glu
GGU = Gly
GGC = Gly
GGA = Gly
GGG = Gly
U
C
A
G
Phe = phenylalanine
Leu = leucine
Ile = isoleucine
Met = methionine
Val = valine
Ser = serine
Pro = proline
Thr = threonine
Ala = alanine
Tyr = tyrosine
His = histidine
Gln = glutamine
Asn = asparagine
Lys = lysine
Asp = aspartic acid
Glu = glutamic acid
Cys = cysteine
Trp = tryptophan
Arg = arginine
Gly = glycine
AUG = start codon, UAA, UAG, and UGA = stop (nonsense) codons
In bacteria, a mRNA can be monocistronic or polycistronic. A monocistronic mRNA is a transcript of a single gene. A polycistronic mRNA carries a transcript of multiple genes, often involved in a single biochemical pathway. Groups of related genes that are transcribed together to form a polycistronic mRNA are known as operons. There are also specific genes along the DNA from which each of the different transfer RNAs (tRNAs) and the ribosomal RNAs (rRNAs) are transcribed.
Most mRNAs in prokaryotes have a half-life on the order of a few minutes. Molecules of rRNA and tRNA, on the other hand, are much more stabile. Because rRNA and tRNA are highly folded molecules, unlike mRNA, they are much more resistant to degradation by ribonucleases. Once transcribed, the mRNA can be translated into protein.
Transcription in Eukaryotic Cells
Transcription is more complex in eukaryotic cells than in those that are prokaryotic. Activator proteins bind to genes known as enhancers which help determine which genes are switched on and speed up transcription. Repressor proteins bind to genes called silencers which interfere with activator proteins and slow down transcription. Coactivators, adapter molecules which coordinate signals from activator and repressor proteins, relay this information to basal factors which then position RNA polymerase at the start of the coding region of the gene to begin transcription.
Once the actual transcription begins, ribonucleotides containing 3 phosphate groups form hydrogen bonds through the process of complementary base pairing with the exposed deoxyribonucleotides on the unwound strand that is to be transcribed. The ribonucleotides are then covalently bonded together by phosphodiester bonds, the energy being supplied by the cleavage of two phosphate groups from the ribonucleotide triphosphate (Figure \(16\).8B.5). (The phosphodiester bond refers to the phosphate on the 5'C of the newly inserted nucleotide covalently bonding to the 3'C of the last ribonucleotide in the mRNA chain.)
Unlike prokaryotes, most genes in higher eukaryotic cells contain large amounts - as much as 98% in the human genome - of regionscalled introns that are not part of the code for the final protein. These are interspersed among the coding regions or exons that actually code for the final protein.
RNA polymerase copies both the exons and the introns to form what is called precursor mRNA or pre-mRNA. Early in transcription, a cap in the form of an unusual nucleotide, 7-methylguanylate, is added to the 5' end of the pre-mRNA. This cap helps ribosomes attach for translation. As transcription is nearly completed, a series of 100-250 adenine ribonucleotides called a poly-A tail is added to the 3' end of the pre-mRNA. This poly-A tail is thought to help transport the mRNA out of the nucleus and may stabilize the mRNA against degradation in the cytoplasm. After transcription of the precursor mRNA, non-protein coding regions (introns) are excised and coding regions (exons) are joined together by complexes of ribonucleoproteins called spliceosomes to produce what is termed mature mRNA as shown in Figure \(9\). This process is called RNA processing.
YouTube movie illustrating complementary base pairing during transcription.
YouTube movie illustrating transcription and protein assembly.
YouTube movie illustrating RNA processing in eukaryotic cells #1.
YouTube movie illustrating RNA processing in eukaryotic cells #2.
The mature mRNA then passes through the pores in the nuclear membrane to be translated into protein by tRNA on eukaryotic 80S ribosomes (composed of 60S and 40S subunits) in a manner similar to prokaryotes.
The mRNA molecule is divided up into codons. A codon is a series of three consecutive mRNA bases coding for one specific amino acid. The various codons and the amino acids for which they code are shown in Figure \(8\). There are 64 codons. One codon, AUG, also serves as a start codon to initiate translation, and three codons, UAG, UAA, and UGA, function as stop or nonsense codons to terminate translation. (Alternative start codons are different from the standard AUG codon and are found occasionally in both prokaryotes and eukaryotes.)
In addition to the genes that are transcribed into mRNA to be translated into polypeptides and proteins, there are also specific genes in the DNA from which each of the different transfer RNAs (tRNAs) and the ribosomal RNAs (rRNAs) are transcribed.
Once transcribed, the mRNA can be translated into protein.
As mentioned above, introns make up the majority of DNA in higher eukaryotic cells and for decades was considered to be "junk DNA" accumulated over millions of years of evolution. Over recent years however, it has been discovered that much of this intergenic DNA, although it does not code for protein synthesis, is transcribed into functional molecules of RNA with names such as antisense RNA microRNA, and riboswitch RNA that play important roles in whether or not a protein is actually made.
Antisense RNA is RNA transcribed off of the strand of DNA complementary to the one being transcribed into mRNA. In other words, it is an RNA molecule complementary to a mRNA and as such may complementary base pair with the mRNA and prevents it from being translated into protein.
MicroRNA, often transcribed from intron DNA, folds over upon itself to resemble double-stranded RNA, a form of RNA produced by many viruses during their life cycle. Viral double-stranded RNA activates a host defense mechanism that degrades that viral RNA. The MicroRNA frequently binds to mRNA and tricks this defense mechanism into degrading that mRNA so it can not be translated into protein.
Riboswitch RNA, often transcribed from introns, exists in an inactive form until a specific target chemical binds. The binding of the target chemical turns the riboswitch RNA to an active form that can be translated into a specific protein.
Summary
1. During protein synthesis, the order of nucleotide bases along a gene gets transcribed into a complementary strand of mRNA which is then translated by tRNA into the correct order of amino acids for that polypeptide or protein.
2. The order of deoxyribonucleotide bases along the DNA determines the order of amino acids in the proteins, that is, its primary structure.
3. Because certain amino acids can interact with other amino acids, the order of amino acids for each protein determines its final three-dimensional shape, which in turn determines the function of that protein.
4. Messenger RNA (mRNA) is synthesized by complementary base pairing of ribonucleotides with deoxyribonucleotides to match a portion of one strand of DNA called a gene.
5. Although genes are present on both strands of DNA, only one strand is transcribed for any given gene.
6. The enzyme RNA polymerase transcribes DNA.
7. To initiate transcription in bacteria, a variety of proteins called sigma factors bind to RNA polymerases. This complex can then bind to a specific DNA sequence called the promoter located along the DNA prior to the coding region of the gene. The promotor determines what region of the DNA and which strand of DNA will be transcribed into RNA.
8. Like DNA polymerase, RNA polymerase can only synthesize nucleic acid in a 5' to 3' direction while "reading" a DNA template in the 3' to 5' direction.
9. Once the RNA polymerase/sigma factor complex recognizes the correct promoter, the sigma factor dissociate from the RNA polymerase and the enzyme begins to unwind the helix of the DNA creating a region of nonpaired deoxyribonucleotides that serve as a template for RNA synthesis.
10. During transcription, ribonucleotides hydrogen bond through the process of complementary base pairing with the exposed deoxyribonucleotides on the unwound strand that is to be transcribed. The ribonucleotides are then covalently bonded together by phosphodiester bonds.
11. This process continues until the RNA polymerase encounters a "stop" signal or transcription terminator at the end of the gene.
12. A single gene can be transcribed multiple times.
13. The mRNA molecule is divided up into codons. A codon is a series of three consecutive mRNA bases coding for one specific amino acid.
14. Three codons, UAG, UAA, and UGA, function as stop or nonsense codons to terminate translation.
15. In bacteria, a mRNA can be monocistronic or polycistronic. A monocistronic mRNA is a transcript of a single gene; a polycistronic mRNA carries a transcript of multiple genes, often involved in a single biochemical pathway. Once transcribed, the mRNA can be translated into protein by tRNA on 70S ribosomes (composed of 50S and 30S subunits).
16. Transcription is more complex in eukaryotic cells than in those that are prokaryotic. Activator proteins bind to genes known as enhancers which help determine which genes are switched on and speed up transcription. Repressor proteins bind to genes called silencers which interfere with activator proteins and slow down transcription. Coactivators, adapter molecules which coordinate signals from activator and repressor proteins, relay this information to basal factors which then position RNA polymerase at the start of the coding region of the gene to begin transcription.
17. Most genes in higher eukaryotic cells contain regions called introns that are not part of the code for the final protein. These are interspersed among the coding regions or exons that actually code for the final protein.
18. After transcription of the precursor mRNA, non-protein coding regions (introns) are excised and coding regions (exons) are joined together by complexes of ribonucleoproteins called spliceosomes to produce what is termed mature mRNA.
19. The mature mRNA then passes through the pores in the nuclear membrane to be translated into protein by tRNA on 80S ribosomes (composed of 60S and 40S subunits) in a manner similar to prokaryotes. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/19%3A_Review_of_Molecular_Genetics/19.7%3A_Polypeptide_and_Protein_Synthesis/19.7A%3A_Transcription.txt |
Define translation. Briefly describethe function of the following in terms of translation: 30S ribosomal subunit ribosome binding site start codon initiation complex 50S ribosomal subunit tRNA aminoacyl-tRNA anticodon P-site of ribosome A-site of ribosome E-site of ribosome peptidyl transferase nonsense (stop) codon release factors
Figure \(3\): Translation of mRNA by tRNA: Formation of the Initiation Complex. To initiate translation, a 30S ribosomal subunitbinds to a short nucleotide sequence on the mRNA called the ribosome binding site. However, translation doesn't usually begin until the 30S ribosomal subunit reaches the first AUG sequence in the mRNA. For this reason, AUG is known as the start codon. At this point, an initiation complex composed of the 30S subunit, a tRNA having the anticodon UAC and carrying an altered form of the amino acid methionine (N-formylmethionine or f-Met), and proteins called initiation factors is formed.
A 50S ribosomal subunit then attaches to the initiation complex and the initiation factors leave. This forms the 70S ribosome. (see Figure \(4\)).
The joining of individual amino acids to form a protein or polypeptide is known as the elongation phase of translation. There are three sites on the 70S ribosome. The A or acceptor or aminoacyl site is where an aminoacyl-tRNA first attaches. The P or peptide site is where a tRNA is temporarily holding the growing amino acid chain as the next codon in the mRNA is being read. The E or exit site is where the uncharged tRNA that has released its amino acid exits the ribosome. During peptide bond formation, the amino acid chain or peptide moves from the tRNA at the P-site and forms a peptide bond with the new amino acid attached to the tRNA at the A-site. The peptide bond is formed by a ribozyme , an enzyme composed of the 23S rRNA itself, called peptidyl transferase. The now uncharged tRNA at the P-site leaves the ribosome through the E-site to eventually pick up a new amino acid and be recycled. Meanwhile, the 70S ribosome moves a distance of one codon down the mRNA through a process called translocation to allow decoding of the next codon in the message (see Figure \(5\)A - 5F). The growing polypeptide chain actually passes through a tunnel in the 50S ribosomal subunit.
This process continues over and over again in the 5' to 3' direction until the ribosome hits a stop codon. A stop codon is a series of three mRNA bases coding for no amino acid and thus terminates the protein chain. UAA, UAG, UGA are the three stop codons in the genetic code. Stop codons do not code for an amino acid because a tRNA cannot recognize them.
Proteins called release factors free the protein from the tRNA and the two ribosomal subunits come apart to be recycled (see Figure \(5\)F). During this elongation process, the protein has assumed its three-dimensional functional shape. Proteins called chaperonins assist in the protein folding.
Once the ribosome is clear of the ribosome binding site and the AUG start codon, another 30S ribosomal subunit attaches to the ribosome binding site of the mRNA to initiate another round of translation. In this way, multiple copies of a protein can be produced from a single molecule of mRNA. A mRNA with multiple ribosomes attached is known as a polyribosome or polysome .
Summary
1. During translation, specific tRNAs pick up specific amino acids, transfer those amino acids to the ribosomes, and insert them in their proper place according to the mRNA genetic "message."
2. This is done by the anticodon portion of the tRNA molecules complementary base pairing with the codons along the mRNA.
3. Transfer RNA (tRNA) is a three-dimensional, inverted cloverleaf-shaped molecule of RNA.
4. At the top, or 3' end, a specific amino acid can be attached to a specific tRNA by means of specific enzymes called aminoacyl-tRNA synthetases.
5. At the bottom loop of the cloverleaf is a series of three unpaired tRNA bases called the anticodon. An anticodon is a series of three tRNA bases complementary to a mRNA codon.
6. The anticodons of some tRNAs are able to recognize more than one codon because the tRNA's recognition of the third nucleotide of the codon is not always precise, however, the right amino acid is still inserted because there are 61 codons that code for the 22 different amino acids.
7. To initiate translation in prokaryotic cells, a 30S ribosomal subunit binds to a short nucleotide sequence on the mRNA called the ribosome binding site.
8. AUG is known as the start codon. At this point, an initiation complex composed of the 30S subunit, a tRNA having the anticodon UAC and carrying an altered form of the amino acid methionine (N-formylmethionine or f-Met), and proteins called initiation factors is formed. A 50S ribosomal subunit then attaches to the initiation complex and the initiation factors leave. This forms the 70S ribosome.
9. The A or acceptor or aminoacyl site of the ribosome is where an aminoacyl-tRNA first attaches.
10. The P or peptide site of the ribosome is where a tRNA is temporarily holding the growing amino acid chain as the next codon in the mRNA is being read.
11. The E or exit site of the ribosome is where the uncharged tRNA that has released its amino acid exits the ribosome.
12. During peptide bond formation, the amino acid chain or peptide moves from the tRNA at the P-site and forms a peptide bond with the new amino acid attached to the tRNA at the A-site.
13. A stop codon is a series of three mRNA bases coding for no amino acid and thus terminates the protein chain. UAA, UAG, UGA are the three stop codons in the genetic code. (Stop codons do not code for an amino acid because a tRNA cannot recognize them.)
14. A mRNA with multiple ribosomes attached is known as a polyribosome or polysome.
19.8: Enzyme Regulation
Briefly compare the genetic control of enzyme activity in bacteria with control of enzyme activity through feedback inhibition. Briefly compare an inducible operon with a repressible operon. Briefly compare competitive inhibition with noncompetitive inhibition.
Figure \(5\): An Inducible Operon in the Absence of an Inducer (The Lactose Operon). Step 1: The regulator gene codes for an active repressor protein.
Step 2: The repressor protein then binds to the operator region of the operon.
The regulator gene codes for an active repressor protein. The repressor protein then binds to the operator region of the operon. With the active repressor protein bound to the operator region, RNA polymerase (the enzyme responsible for the transcription of genes) is unable to bind to the promoter region of the operon. If RNA polymerase does not bind to the promoter region, the three enzyme genes (Z, Y, and A) are not transcribed into mRNA. Without the transcription of the three enzyme genes, the three enzymes needed for the utilization of the sugar lactose by the bacterium are not synthesized.
• The regulator gene codes for an active repressor protein.
• Lactose, the inducer molecule binds to the active repressor protein.
• The binding of the inducer alters the shape of the allosteric repressor causing it to become inactivated.
• The inactivated repressor protein is then unable to bind to the operator region of the operon.
• Since the inactive repressor protein is unable to bind to the operator region, RNA polymerase (the enzyme responsible for the transcription of genes) is now able to bind to the promoter region of the operon.
• RNA polymerase is now able to transcribe the three enzyme genes (Z, Y, and A) into mRNA.
• With the transcription of these genes, the three enzymes needed for the bacterium to utilize the sugar lactose are now synthesized. (The Z gene codes for beta-galactosidase, an enzyme that breaks down lactose into glucose and galactose. The Y gene codes for permease, an enzyme which transports lactose into the bacterium. The A gene codes for transacetylase, an enzyme which is thought to aid in the release of galactosides.) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/19%3A_Review_of_Molecular_Genetics/19.7%3A_Polypeptide_and_Protein_Synthesis/19.7B%3A_Translation.txt |
Define the following: genotype phenotype allele mutation spontaneous mutation induced mutation Describe two different mechanisms of spontaneous mutation and, in terms of protein synthesis, describe the four possible results that may occur as a result of these mutations. Briefly describe three ways chemical mutagens work. Compare ultraviolet radiation and gamma radiation in terms of how they induce mutation.
As we learned earlier, the sequence of deoxyribonucleotide bases in the genes that make up a bacterium's DNA determines the order of amino acids in the proteins and polypeptides made by that organism. This order of DNA bases constitutes the organism's genotype. A particular organism may possess alternate forms of some genes. Such alternate forms of genes are referred to as alleles. The physical characteristics an organism possesses, based on its genotype and the interaction with its environment, make up its phenotype.
Mutation is an error during DNA replication that results in a change in the sequence of deoxyribonucleotide bases in the DNA. Spontaneous mutation occurs naturally (a normal mistake rate) about one in every million to one in every billion divisions and is probably due to low level natural mutagens normally present in the environment. Induced mutation is caused by mutagens, substances that cause a much higher rate of mutation.
Mechanisms of Mutation
There are two general mechanisms of mutation.
1. Substitution of a nucleotide (point mutations ): substitution of one deoxyribonucleotide for another during DNA replication (see Figure \(1\)). This is the most common mechanism of mutation. Substitution of one nucleotide for another is a result of tautomeric shift, a rare process by which the hydrogen atoms of a deoxyribonucleotide base move in a way that changes the properties of its hydrogen bonding. For example, a shift in the hydrogen atom of adenine enables it to form hydrogen bonds with cytosine rather than thymine. Likewise, a shift in the hydrogen atom in thymine allows it to bind with guanine rather than adenine.
2. Deletion or addition of a nucleotide (frameshift mutations ): deletion or addition of a deoxyribonucleotide during DNA replication (see Figure \(2\) and Figure \(3\)).
Results of Mutation
One of four things can happen as a result of these mechanisms of mutation and the resulting change in the deoxyribonucleotide base sequence mentioned above:
• A missense mutation occurs. This is usually seen with a single substitution mutation and results in one wrong codon and one wrong amino acid (Figure \(4\)).
Figure \(4\): Results in one wrong codon and one wrong amino acid.
• A nonsense mutation occurs. If the change in the deoxyribonucleotide base sequence results in transcription of a stop or nonsense codon, the protein would be terminated at that point in the message (Figure \(5\)).
Figure \(5\): Results in a "stop" codon and premature termination of the protein.
• A sense mutation occurs. This is sometimes seen with a single substitution mutation when the change in the DNA base sequence results in a new codon still coding for the same amino acid (Figure \(6\)). (With the exception of methionine, all amino acids are coded for by more than one codon.)
Figure \(6\): Results in a new codon which still codes for the same amino acid.
• A frameshift mutation occurs. This is seen when a number of DNA nucleotides not divisible by three is added or deleted. Remember, the genetic code is a triplet code where three consecutive nucleotides code for a specific amino acid. This causes a reading frame shift and all of the codons and all of the amino acids after that mutation are usually wrong (Figure \(7\)); frequently one of the wrong codons turns out to be a stop or nonsense codon and the protein is terminated at that point.
Figure \(7\): Results in a reading frame shift. All codons and all amino acid after the shift are usually wrong.
YouTube movie illustrating frameshift mutations (www.youtube.com/v/o-otJTJ3N_E)
Induced mutation is caused by mutagens, substances that cause a much higher rate of mutation. Chemical mutagens generally work in one of three ways.
1. Some chemical mutagens, such as nitrous acid and nitrosoguanidine work by causing chemical modifications of purine and pyrimidine bases that alter their hydrogen-bonding properties. For example, nitrous acid converts cytosine to uracil which then forms hydrogen bonds with adenine rather than guanine.
2. Other chemical mutagens function as base analogs. They are compounds that chemically resemble a nucleotide base closely enough that during DNA replication, they can be incorporated into the DNA in place of the natural base. Examples include 2-amino purine, a compound that resembles adenine, and 5-bromouracil, a compound that resembles thymine. The base analogs, however, do not have the hydrogen-bonding properties of the natural base.
3. Still other chemical mutagens function as intercalating agents. Intercalating agents are planar three-ringed molecules that are about the same size as a nucleotide base pair. During DNA replication, these compounds can insert or intercalate between adjacent base pairs thus pushing the nucleotides far enough apart that an extra nucleotide is often added to the growing chain during DNA replication. An example is ethidium bromide.
When under stress from antibiotics or other harmful chemicals, some bacteria switch on genes whose protein products can increase the mutation rate within the bacterium 10,000 times as fast as the mutation rate that occurs during normal binary fission. This causes a sort of hyperevolution where mutation acts as a self defense mechanism for the bacterial population by increasing the chance of forming an antibiotic-resistant mutant that is able to survive at the expense of the majority of the population. (Remember that most mutations are harmful to a cell; see SOS repair below.)
Certain types of radiation can also function as mutagens.
1. Ultraviolet Radiation. The ultraviolet portion of the light spectrum includes all radiations with wavelengths from 100 nm to 400 nm. It has low wave length and low energy. The microbicidal activity of ultraviolet (UV) light depends on the length of exposure: the longer the exposure the greater the cidal activity. It also depends on the wavelength of UV used. The most cidal wavelengths of UV light lie in the 260 nm - 270 nm range where it is absorbed by nucleic acid.
In terms of its mode of action, UV light is absorbed by microbial DNA and causes adjacent thymine bases on the same DNA strand to covalently bond together, forming what are called thymine-thymine dimers (see Figure \(8\)). As the DNA replicates, nucleotides do not complementary base pair with the thymine dimers and this terminates the replication of that DNA strand. However, most of the damage from UV radiation actually comes from the cell trying to repair the damage to the DNA by a process called SOS repair. In very heavily damaged DNA containing large numbers of thymine dimers, a process called SOS repair is activated as kind of a last ditch effort to repair the DNA. In this process, a gene product of the SOS system binds to DNA polymerase allowing it to synthesize new DNA across the damaged DNA. However, this altered DNA polymerase loses its proofreading ability resulting in the synthesis of DNA that itself now contains many misincorporated bases. (Most of the chemical mutagens mentioned above also activate SOS repair.)
Video illustrating frameshift mutations (www.youtube.com/v/azszodOhXqk)
2. Ionizing Radiation. Ionizing radiation, such as X-rays and gamma rays, has much more energy and penetrating power than ultraviolet radiation. It ionizes water and other molecules to form radicals (molecular fragments with unpaired electrons) that can break DNA strands and alter purine and pyrimidine bases.
Summary
1. The sequence of deoxyribonucleotide bases in the genes that make up an organism's DNA determines the order of amino acids in the proteins and polypeptides made by that organism. This order of DNA bases constitutes the bacterium's genotype.
2. A particular organism may possess alternate forms of some genes referred to as alleles.
3. The physical characteristics an organism possesses, based on its genotype and the interaction with its environment, make up an organism's phenotype.
4. Mutation is an error during DNA replication that results in a change in the sequence of deoxyribonucleotide bases in the DNA.
5. Spontaneous mutation occurs naturally (a normal mistake rate) about one in every million to one in every billion divisions and is probably due to low level natural mutagens normally present in the environment; induced mutation is caused by mutagens, substances that cause a much higher rate of mutation.
6. There are two primary mechanisms of mutation: substitution of a deoxyribonucleotide (point mutations) whereby one deoxyribonucleotide is substituted for another during DNA replication; and deletion or addition of a nucleotide (frameshift mutations) where deoxyribonucleotides are either added or deleted during DNA replication. Point mutations are most common.
7. There are four possible results from a mutation: missense, nonsense, sense, or frameshift.
8. A missense mutation usually seen with a single substitution mutation and results in one wrong codon and one wrong amino acid.
9. A nonsense mutation occurs when the change in the deoxyribonucleotide base sequence results in transcription of a stop or nonsense codon. The protein would be terminated at that point in the message.
10. A sense mutation occurs is sometimes seen with a single substitution mutation when the change in the DNA base sequence results in a new codon still coding for the same amino acid.
11. A frameshift mutation occurs when a number of DNA nucleotides not divisible by three is added or deleted. This causes a reading frame shift and all of the codons and all of the amino acids after that mutation are usually wrong; frequently one of the wrong codons turns out to be a stop or nonsense codon and the protein is terminated at that point.
12. When under stress from harmful chemicals, some bacteria switch on genes whose protein products can increase the mutation rate within the bacterium 10,000 times as fast as the mutation rate that occurs during normal binary fission. This causes a hyperevolution where mutation acts as a self defense mechanism for the bacterial population by increasing the chance of forming an antibiotic-resistant mutant that is able to survive at the expense of the majority of the population. | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/19%3A_Review_of_Molecular_Genetics/19.9%3A_Mutation.txt |
These are homework exercises to accompany Kaiser's "Microbiology" TextMap. Microbiology is the study of microorganisms, which are defined as any microscopic organism that comprises either a single cell (unicellular), cell clusters or no cell at all (acellular). This includes eukaryotes, such as fungi and protists, and prokaryotes. Viruses and prions, though not strictly classed as living organisms, are also studied.
19.1: Polypeptides and Proteins
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Describe an amino acid and state what all amino acids have in common. (ans)
2. State what makes one amino acid different from another. (ans)
3. Describe how amino acids are joined by peptide bonds. (ans)
4. Compare the terms peptide, polypeptide, and protein. (ans)
5. Due to hydrogen bonds that form between the oxygen atom of one amino acid and the nitrogen atom of another, this gives the protein or polypeptide the two-dimensional form of an alpha-helix or a beta-pleated sheet. This best describes:
1. the primary structure of a protein (ans)
2. the secondary structure of a protein (ans)
3. the tertiary structure of a protein (ans)
4. the quaternary structure of a protein (ans)
6. In some cases, such as with antibody molecules and hemoglobin, several polypeptides may bond together to form a quaternary structure. This best describes:
1. the primary structure of a protein (ans)
2. the secondary structure of a protein (ans)
3. the tertiary structure of a protein (ans)
4. the quaternary structure of a protein (ans)
7. The actual order of the amino acids in the protein that is determined by DNA. This best describes:
1. the primary structure of a protein (ans)
2. the secondary structure of a protein (ans)
3. the tertiary structure of a protein (ans)
4. the quaternary structure of a protein (ans)
8. In globular proteins such as enzymes, the long chain of amino acids becomes folded into a three-dimensional functional shape. This is because certain amino acids with sulfhydryl or SH groups form disulfide (S-S) bonds with other amino acids in the same chain. Other interactions between R groups of amino acids such as hydrogen bonds, ionic bonds, covalent bonds, and hydrophobic interactions also contribute to this structure. This best describes:
1. the primary structure of a protein (ans)
2. the secondary structure of a protein (ans)
3. the tertiary structure of a protein (ans)
4. the quaternary structure of a protein (ans)
9. Define gene. (ans)
10. Describe how the order of nucleotide bases in DNA ultimately determines the final three-dimensional shape of a protein or polypeptide. (ans)
19.2: Enzymes
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Define enzyme and state how enzymes are able to speed up the rate of chemical reactions. (ans)
2. Fill in the blanks.
Many enzymes require a nonprotein cofactor to assist them in their reaction. In this case, the protein portion of the enzyme, called an _______________ (ans), combines with the cofactor to form the whole enzyme or ____________ (ans). Some cofactors are ions such as Ca++, Mg++, and K+; other cofactors are organic molecules called _____________ (ans) which serve as carriers for chemical groups or electrons. Anything that an enzyme normally combines with is called a _____________ (ans).
3. Briefly describe a generalized enzyme-substrate reaction, state the function of an enzyme's active site, and describe how an enzyme is able to speed up chemical reactions. (ans)
4. State four characteristics of enzymes. (ans)
5. State how the following will affect the rate of an enzyme reaction.
1. increasing temperature (ans)
2. decreasing temperature (ans)
3. pH (ans)
4. salt concentration (ans)
19.3: Deoxyribonucleic Acid (DNA)
Questions
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. State the 3 basic parts of a deoxyribonucleotide. (ans)
2. State which nitrogenous bases are purines.
1. cytosine and thymine (ans)
2. adenine and guanine (ans)
3. In the complement base pairing of nucleotides, adenine can form hydrogen bonds with ____________ (ans) and guanine can form hydrogen bonds with ____________ (ans).
4. State what is meant by the 3' (3-prime) and 5' (5-prime) ends of a DNA strand. (ans)
5. State why DNA can only be synthesized in a 5' to 3' direction. (ans)
6. What is a nucleosome? (ans)
7. State whether the following characteristics are seen in prokaryotic or eukaryotic DNA.
1. linear chromosomes (ans)
2. no nuclear membrane (ans)
3. presence of nucleosomes (ans)
4. no mitosis (ans)
5. produce gametes through meiosis (ans)
19.4: DNA Replication in Prokaryotic Cells
Questions
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the process of DNA replication. (ans)
2. State what enzyme carries out the following functions during DNA replication.
1. Unwinds the helical DNA by breaking the hydrogen bonds between complementary bases. (ans)
2. Synthesizes a short RNA primer at the beginning of each origin of replication. (ans)
3. Adds DNA nucleotides to the RNA primer. (ans)
4. Digests away the RNA primer and replaces the RNA nucleotides of the primer with the proper DNA nucleotides. (ans)
5. Links the DNA fragments of the lagging strand together. (ans)
3. The DNA strand replicated in short fragments called Okazaki fragments is called the:
1. lagging strand (ans)
2. leading strand (ans)
19.5: DNA Replication in Eukaryotic Cells and the Eukaryotic Cell Cycle
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Briefly describe the process of DNA replication. (ans)
2. State which cell type has multiple origins of replication in its genome.
1. prokaryotic (ans)
2. eukaryotic (ans)
3. Identify the following stages of mitosis.
1. During this final stage of mitosis, the nuclear membrane and nucleoli reform, cytokinesis is nearly complete, and the chromosomes eventually uncoil to chromatin. (ans)
2. Refers to all stages of the cell cycle other than mitosis. During this phase, cellular organelles double in number, the DNA replicates, and protein synthesis occurs. The chromosomes are not visible and the DNA appears as uncoiled chromatin. (ans)
3. During this phase of mitosis, the nuclear membrane fragmention is complete and the duplicated chromosomes line up along the cell's equator. (ans)
4. During the first stage of mitosis, the chromatin condenses and the chromosomes become visible. Also the nucleolus disappears, the nuclear membrane fragments, and spindle fibers are assembled. (ans)
5. During this phase of mitosis, diploid sets of daughter chromosomes move toward opposite poles of the cell and cytokinesis (cytoplasmic cleavage) begins. (ans)
19.6: Ribonucleic Acid (RNA)
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. State the 3 basic parts of a ribonucleotide. (ans)
2. State 3 ways RNA differs from DNA. (ans)
3. Copies the genetic information in the DNA by complementary base pairing and carries this "message" to the ribosomes where the proteins are assembled. This best describes:
1. tRNA (ans)
2. mRNA (ans)
3. rRNA (ans)
4. Picks up specific amino acids, transfers the amino acids to the ribosomes, and insert the correct amino acids in the proper place according to the mRNA message. This best describes:
1. tRNA (ans)
2. mRNA (ans)
3. rRNA (ans)
19.7: Polypeptide and Protein Synthesis
Questions: Transcription
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Define transcription. (ans)
2. Match the following with their role in transcription.
_____ The end of a strand of nucleic acid that has a hydroxyl (OH) group on the number 3 carbon of the deoxyribose or ribose and is not linked to another nucleotide. (ans)
_____ The covalent bond that links ribonucleotides together to form RNA. (ans)
_____ The portion of DNA that contains the actual message for protein synthesis. (ans)
_____ A molecule synthesized by complementary base pairing of ribonucleotides with deoxyribonucleotides to match a portion of one strand of DNA coding for a polypeptide or protein. (ans)
_____ A series of three consecutive mRNA bases coding for one specific amino acid. (ans)
_____ A segment of DNA that determines what region of the DNA and which strand of DNA will be transcribed into RNA. (ans)
_____ The enzyme that initiates transcription, joins the RNA nucleotides together, and terminates transcription. (ans)
_____ A "stop" signal at the end of a gene that causes the completed mRNA to drop off the gene. (ans)
1. mRNA
2. 3' end
3. 5' end
4. RNA polymerase
5. phosphodiester bond
6. promoter
7. leader sequence
8. coding sequence
9. transcription terminator
10. codon
3. Match the following with their role in transcription in eukaryotic cells.
_____ The RNA synthesized after RNA polymerase copies both the exons and the interons of a gene. (ans)
_____ The RNA produced after non-protein coding regions (introns) are excised and coding regions (exons) are joined together by complexes of ribonucleoproteins called spliceosomes. (ans)
_____ An unusual nucleotide, 7-methylguanylate, that is added to the 5' end of the pre-mRNA early in transcription. It helps ribosomes attach for translation. (ans)
_____ Non-protein coding regions of DNA that are not part of the code for the final protein that are interspersed among the coding regions of DNA in most genes of higher eukaryotic cells. (ans)
_____ The coding regions of DNA in most genes of higher eukaryotic cells that actually code for the final protein. (ans)
_____ A series of 100-250 adenine ribonucleotides that is added to the 3' end of the pre-mRNA. This series of nucleotides is thought to help transport the mRNA out of the nucleus and may stabilize the mRNA against degradation in the cytoplasm. (ans)
1. introns
2. exons
3. precurser mRNA
4. cap
5. poly-A tail
6. mature mRNA
4. What amino acid sequence would the DNA base sequence 5' ATAGCCACC 3'code for? Hint: see Figure 8. (ans)
Questions: Translation
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Define translation. (ans)
2. Match the following with their role in translation.
_____ A series of three tRNA bases complementary to a mRNA codon. (ans)
_____ The ribozyme that forms peptide bonds between amino acids during translation. (ans)
_____ The ribosomal subunit that binds to mRNA to form the initiation complex. (ans)
_____ The ribosomal site where an aminoacyl-tRNA first attaches during translation. (ans)
_____ The ribosomal site where the growing amino acid chain is temporarily being held by a tRNA as the next codon in the mRNA is being read. (ans)
_____ A complex of an amino acid and a tRNA molecule. (ans)
_____ The sequence of bases on mRNA to which a 30S or 40S ribosomal subunit first attaches. (ans)
_____ A series of three mRNA bases coding for no amino acid and thus terminates the protein chain: UAA, UAG, UGA. (ans)
_____ A complex consisting of a 30S or 40S ribosomal subunit, a tRNA having the anticodon UAC and carrying an altered form of the amino acid methionine (N-formylmethionine or f-Met), and proteins called initiation factors. (ans)
_____ A three-dimensional, inverted cloverleaf-shaped molecule about 70 nucleotides long to which a specific amino acid can be attached; transports amino acids to the ribosome during translation. (ans)
1. 30S or 40S ribosomal subunit
2. ribosome binding site
3. initiation complex
4. 50S or 60S ribosomal subunit
5. tRNA
6. aminoacyl-tRNA
7. anticodon
8. P-site of ribosome
9. A-site of ribosome
10. peptidyl transferase
11. nonsense (stop) codon
12. release factors
13. start cocon
3. What amino acid sequence would the DNA base sequence AAAGAGCCT code for? Hint: see Fig. 2. (ans)
19.8: Enzyme Regulation
Questions
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Matching
_____ Regulatory proteins that block transcription of mRNA by binding to a portion of DNA called the operator that lies downstream of a promoter. (ans)
_____ A molecule that alters the shape of the regulatory protein in a way that blocks its binding to the operator and thus permits transcription. (ans)
_____ Regulatory proteins that promote transcription of mRNA. (ans)
_____ A molecule that alters the shape of the regulatory protein to a form that can bind to the operator and block transcription. (ans)
_____ Producing antisense RNA that is complementary to the mRNA coding for the enzyme. When the antisense RNA binds to the mRNA by complementary base pairing, the mRNA cannot be translated into protein and the enzyme is not made. (ans)
_____ The induction or repression of enzyme synthesis by regulatory proteins that can bind to DNA and either block or enhance the function of RNA polymerase. (ans)
_____ The inhibitor is the end product of a metabolic pathway that is able to bind to a second site (the allosteric site) on an enzyme. Binding of the inhibitor to the allosteric site alters the shape of the enzyme's active site thus preventing binding of the first substrate in the metabolic pathway. (ans)
_____ The inhibitor is the end product of an enzymatic reaction. That end product is also capable of reacting with the enzyme's active site and prevents the enzyme from binding its normal substrate. (ans)
1. activators
2. competitive inhibition
3. corepressors
4. genetic control
5. inducer
6. noncompetitive inhibition
7. repressors
8. translational control
2. Describe how the lac operon in E. coli functions as an inducible operon. (ans)
19.9: Mutation
Study the material in this section and then write out the answers to these question. Do not just click on the answers and write them out. This will not test your understanding of this tutorial.
1. Match the following:
_____The sequence of deoxyribonucleotide bases in the genes that make up a organism's DNA. (ans)
_____ An error during DNA replication that results in a change in the sequence of deoxyribonucleotide bases in the DNA. (ans)
_____ Alternate forms of a gene. (ans)
_____ Mutations caused by mutagens, substances that cause a high rate of mutation. (ans)
_____ The physical characteristics of an organism. (ans)
1. genotype
2. phenotype
3. allele
4. mutation
5. spontaneous mutation
6. induced mutation
2. Describe 2 different mechanisms of spontaneous mutation. (ans)
3. Match the following:
_____ This is usually seen with a single substitution mutation and results in one wrong codon and one wrong amino acid (ans)
_____ If the change in the deoxyribonucleotide base sequence results in transcription of a stop, the protein is terminated at that point in the message. (ans)
_____ This is sometimes seen with a single substitution mutation when the change in the DNA base sequence results in a new codon still coding for the same amino acid. (ans)
_____ This is seen when a number of DNA nucleotides not divisible by three is added or deleted and all of the codons and all of the amino acids after that addition or deletion are usually wrong. (ans)
1. sense mutation
2. nonsense mutation
3. frameshift mutation
4. missense mutation
4. Briefly describe 3 ways chemical mutagens work. (ans)
5. Compare ultraviolet radiation and gamma radiation in terms of how they induce mutation. (ans)
6. As a result of a substitution mutation, a DNA base triplet AGA is changed to AGG. State specifically what effect this would have on the resulting protein (see Figure 9). (ans)
7. A third triplet in a bacterial gene is TTT. A substitution mutation changes it to ATT. State specifically what effect this would have on the resulting protein (see Figure 9). (ans) | textbooks/bio/Microbiology/Microbiology_(Kaiser)/Unit_7%3A_Microbial_Genetics_and_Microbial_Metabolism/19%3A_Review_of_Molecular_Genetics/19.E%3A_Review_of_Molecular_Genetics_%28Exercises%29.txt |
From boiling thermal hot springs to deep beneath the Antarctic ice, microorganisms can be found almost everywhere on earth in great quantities. Microorganisms (or microbes, as they are also called) are small organisms. Most are so small that they cannot be seen without a microscope.
Most microorganisms are harmless to humans and, in fact, many are helpful. They play fundamental roles in ecosystems everywhere on earth, forming the backbone of many food webs. People use them to make biofuels, medicines, and even foods. Without microbes, there would be no bread, cheese, or beer. Our bodies are filled with microbes, and our skin alone is home to trillions of them1. Some of them we can’t live without; others cause diseases that can make us sick or even kill us.
Although much more is known today about microbial life than ever before, the vast majority of this invisible world remains unexplored. Microbiologists continue to identify new ways that microbes benefit and threaten humans.
• 1.1: What Our Ancestors Knew
Microorganisms (or microbes) are living organisms that are generally too small to be seen without a microscope. Throughout history, humans have used microbes to make fermented foods such as beer, bread, cheese, and wine. Long before the invention of the microscope, some people theorized that infection and disease were spread by living things that were too small to be seen. They also correctly intuited certain principles regarding the spread of disease and immunity.
• 1.2: A Systematic Approach
Carolus Linnaeus developed a taxonomic system for categorizing organisms into related groups. Binomial nomenclature assigns organisms Latinized scientific names with a genus and species designation. A phylogenetic tree is a way of showing how different organisms are thought to be related to one another from an evolutionary standpoint. The first phylogenetic tree contained kingdoms for plants and animals; Ernst Haeckel proposed adding a kingdom for protists.
• 1.3: Types of Microorganisms
Microorganisms are very diverse and are found in all three domains of life: Archaea, Bacteria, and Eukarya. Archaea and bacteria are classified as prokaryotes because they lack a cellular nucleus. Archaea differ from bacteria in evolutionary history, genetics, metabolic pathways, and cell wall and membrane composition. Archaea inhabit nearly every environment on earth, but no archaea have been identified as human pathogens.
• 1.E: An Invisible World (Exercises)
Footnotes
1. 1 J. Hulcr et al. “A Jungle in There: Bacteria in Belly Buttons are Highly Diverse, but Predictable.” PLoS ONE 7 no. 11 (2012): e47712. doi:10.1371/journal.pone.0047712.
Thumbnail: A cluster of Escherichia coli bacteria magnified 10,000 times. (Public Domain; Eric Erbe, digital colorization by Christopher Pooley, both of USDA, ARS, EMU).
01: An Invisible World
Learning Objectives
• Describe how our ancestors improved food with the use of invisible microbes
• Describe how the causes of sickness and disease were explained in ancient times, prior to the invention of the microscope
• Describe key historical events associated with the birth of microbiology
Clinical Focus: Part 1
Cora, a 41-year-old lawyer and mother of two, has recently been experiencing severe headaches, a high fever, and a stiff neck. Her husband, who has accompanied Cora to see a doctor, reports that Cora also seems confused at times and unusually drowsy. Based on these symptoms, the doctor suspects that Cora may have meningitis, a potentially life-threatening infection of the tissue that surrounds the brain and spinal cord.
Meningitis has several potential causes. It can be brought on by bacteria, fungi, viruses, or even a reaction to medication or exposure to heavy metals. Although people with viral meningitis usually heal on their own, bacterial and fungal meningitis are quite serious and require treatment.
Cora’s doctor orders a lumbar puncture (spinal tap) to take three samples of cerebrospinal fluid (CSF) from around the spinal cord (Figure \(1\)). The samples will be sent to laboratories in three different departments for testing: clinical chemistry, microbiology, and hematology. The samples will first be visually examined to determine whether the CSF is abnormally colored or cloudy; then the CSF will be examined under a microscope to see if it contains a normal number of red and white blood cells and to check for any abnormal cell types. In the microbiology lab, the specimen will be centrifuged to concentrate any cells in a sediment; this sediment will be smeared on a slide and stained with a Gram stain. Gram staining is a procedure used to differentiate between two different types of bacteria (gram-positive and gram-negative).
About 80% of patients with bacterial meningitis will show bacteria in their CSF with a Gram stain.1 Cora’s Gram stain did not show any bacteria, but her doctor decides to prescribe her antibiotics just in case. Part of the CSF sample will be cultured—put in special dishes to see if bacteria or fungi will grow. It takes some time for most microorganisms to reproduce in sufficient quantities to be detected and analyzed.
Exercise \(1\)
What types of microorganisms would be killed by antibiotic treatment?
Most people today, even those who know very little about microbiology, are familiar with the concept of microbes, or “germs,” and their role in human health. Schoolchildren learn about bacteria, viruses, and other microorganisms, and many even view specimens under a microscope. But a few hundred years ago, before the invention of the microscope, the existence of many types of microbes was impossible to prove. By definition, microorganisms, or microbes, are very small organisms; many types of microbes are too small to see without a microscope, although some parasites and fungi are visible to the naked eye.
Humans have been living with—and using—microorganisms for much longer than they have been able to see them. Historical evidence suggests that humans have had some notion of microbial life since prehistoric times and have used that knowledge to develop foods as well as prevent and treat disease. In this section, we will explore some of the historical applications of microbiology as well as the early beginnings of microbiology as a science.
Fermented Foods and Beverages
People across the world have enjoyed fermented foods and beverages like beer, wine, bread, yogurt, cheese, and pickled vegetables for all of recorded history. Discoveries from several archeological sites suggest that even prehistoric people took advantage of fermentation to preserve and enhance the taste of food. Archaeologists studying pottery jars from a Neolithic village in China found that people were making a fermented beverage from rice, honey, and fruit as early as 7000 BC.2
Production of these foods and beverages requires microbial fermentation, a process that uses bacteria, mold, or yeast to convert sugars (carbohydrates) to alcohol, gases, and organic acids (Figure \(2\)). While it is likely that people first learned about fermentation by accident—perhaps by drinking old milk that had curdled or old grape juice that had fermented—they later learned to harness the power of fermentation to make products like bread, cheese, and wine.
The Iceman Treateth
Prehistoric humans had a very limited understanding of the causes of disease, and various cultures developed different beliefs and explanations. While many believed that illness was punishment for angering the gods or was simply the result of fate, archaeological evidence suggests that prehistoric people attempted to treat illnesses and infections. One example of this is Ötzi the Iceman, a 5300-year-old mummy found frozen in the ice of the Ötzal Alps on the Austrian-Italian border in 1991. Because Ötzi was so well preserved by the ice, researchers discovered that he was infected with the eggs of the parasite Trichuris trichiura, which may have caused him to have abdominal pain and anemia. Researchers also found evidence of Borrelia burgdorferi, a bacterium that causes Lyme disease.3 Some researchers think Ötzi may have been trying to treat his infections with the woody fruit of the Piptoporus betulinus fungus, which was discovered tied to his belongings.4 This fungus has both laxative and antibiotic properties. Ötzi was also covered in tattoos that were made by cutting incisions into his skin, filling them with herbs, and then burning the herbs.5 There is speculation that this may have been another attempt to treat his health ailments.
Early Notions of Disease, Contagion, and Containment
Several ancient civilizations appear to have had some understanding that disease could be transmitted by things they could not see. This is especially evident in historical attempts to contain the spread of disease. For example, the Bible refers to the practice of quarantining people with leprosy and other diseases, suggesting that people understood that diseases could be communicable. Ironically, while leprosy is communicable, it is also a disease that progresses slowly. This means that people were likely quarantined after they had already spread the disease to others.
The ancient Greeks attributed disease to bad air, mal’aria, which they called “miasmatic odors.” They developed hygiene practices that built on this idea. The Romans also believed in the miasma hypothesis and created a complex sanitation infrastructure to deal with sewage. In Rome, they built aqueducts, which brought fresh water into the city, and a giant sewer, the Cloaca Maxima, which carried waste away and into the river Tiber (Figure \(3\)). Some researchers believe that this infrastructure helped protect the Romans from epidemics of waterborne illnesses.
Even before the invention of the microscope, some doctors, philosophers, and scientists made great strides in understanding the invisible forces—what we now know as microbes—that can cause infection, disease, and death.
The Greek physician Hippocrates (460–370 BC) is considered the “father of Western medicine” (Figure \(\PageIndex{4a}\)). Unlike many of his ancestors and contemporaries, he dismissed the idea that disease was caused by supernatural forces. Instead, he posited that diseases had natural causes from within patients or their environments. Hippocrates and his heirs are believed to have written the Hippocratic Corpus, a collection of texts that make up some of the oldest surviving medical books.6 Hippocrates is also often credited as the author of the Hippocratic Oath, taken by new physicians to pledge their dedication to diagnosing and treating patients without causing harm.
While Hippocrates is considered the father of Western medicine, the Greek philosopher and historian Thucydides (460–395 BC) is considered the father of scientific history because he advocated for evidence-based analysis of cause-and-effect reasoning (Figure \(\PageIndex{4b}\)). Among his most important contributions are his observations regarding the Athenian plague that killed one-third of the population of Athens between 430 and 410 BC. Having survived the epidemic himself, Thucydides made the important observation that survivors did not get re-infected with the disease, even when taking care of actively sick people.7 This observation shows an early understanding of the concept of immunity.
Marcus Terentius Varro (116–27 BC) was a prolific Roman writer who was one of the first people to propose the concept that things we cannot see (what we now call microorganisms) can cause disease (Figure \(\PageIndex{4c}\)). In Res Rusticae (On Farming), published in 36 BC, he said that “precautions must also be taken in neighborhood swamps . . . because certain minute creatures [animalia minuta] grow there which cannot be seen by the eye, which float in the air and enter the body through the mouth and nose and there cause serious diseases.”8
Exercise \(2\)
1. Give two examples of foods that have historically been produced by humans with the aid of microbes.
2. Explain how historical understandings of disease contributed to attempts to treat and contain disease.
The Birth of Microbiology
While the ancients may have suspected the existence of invisible “minute creatures,” it wasn’t until the invention of the microscope that their existence was definitively confirmed. While it is unclear who exactly invented the microscope, a Dutch cloth merchant named Antonie van Leeuwenhoek (1632–1723) was the first to develop a lens powerful enough to view microbes. In 1675, using a simple but powerful microscope, Leeuwenhoek was able to observe single-celled organisms, which he described as “animalcules” or “wee little beasties,” swimming in a drop of rain water. From his drawings of these little organisms, we now know he was looking at bacteria and protists. (We will explore Leeuwenhoek’s contributions to microscopy further in Chapter 2: How We See the Invisible World.)
Nearly 200 years after van Leeuwenhoek got his first glimpse of microbes, the “Golden Age of Microbiology” spawned a host of new discoveries between 1857 and 1914. Two famous microbiologists, Louis Pasteur and Robert Koch, were especially active in advancing our understanding of the unseen world of microbes (Figure \(5\)). Pasteur, a French chemist, showed that individual microbial strains had unique properties and demonstrated that fermentation is caused by microorganisms. He also invented pasteurization, a process used to kill microorganisms responsible for spoilage, and developed vaccines for the treatment of diseases, including rabies, in animals and humans. Koch, a German physician, was the first to demonstrate the connection between a single, isolated microbe and a known human disease. For example, he discovered the bacteria that cause anthrax (Bacillus anthracis), cholera (Vibrio cholera), and tuberculosis (Mycobacterium tuberculosis).9 We will discuss these famous microbiologists, and others, in later chapters.
As microbiology has developed, it has allowed the broader discipline of biology to grow and flourish in previously unimagined ways. Much of what we know about human cells comes from our understanding of microbes, and many of the tools we use today to study cells and their genetics derive from work with microbes.
Exercise \(3\)
How did the discovery of microbes change human understanding of disease?
Microbiology Toolbox
Because individual microbes are generally too small to be seen with the naked eye, the science of microbiology is dependent on technology that can artificially enhance the capacity of our natural senses of perception. Early microbiologists like Pasteur and Koch had fewer tools at their disposal than are found in modern laboratories, making their discoveries and innovations that much more impressive. Later chapters of this text will explore many applications of technology in depth, but for now, here is a brief overview of some of the fundamental tools of the microbiology lab.
• Microscopes produce magnified images of microorganisms, human cells and tissues, and many other types of specimens too small to be observed with the naked eye.
• Stains and dyes are used to add color to microbes so they can be better observed under a microscope. Some dyes can be used on living microbes, whereas others require that the specimens be fixed with chemicals or heat before staining. Some stains only work on certain types of microbes because of differences in their cellular chemical composition.
• Growth media are used to grow microorganisms in a lab setting. Some media are liquids; others are more solid or gel-like. A growth medium provides nutrients, including water, various salts, a source of carbon (like glucose), and a source of nitrogen and amino acids (like yeast extract) so microorganisms can grow and reproduce. Ingredients in a growth medium can be modified to grow unique types of microorganisms.
• A Petri dish is a flat-lidded dish that is typically 10–11 centimeters (cm) in diameter and 1–1.5 cm high. Petri dishes made out of either plastic or glass are used to hold growth media (Figure \(6\)).
• Test tubes are cylindrical plastic or glass tubes with rounded bottoms and open tops. They can be used to grow microbes in broth, or semisolid or solid growth media.
• A Bunsen burner is a metal apparatus that creates a flame that can be used to sterilize pieces of equipment. A rubber tube carries gas (fuel) to the burner. In many labs, Bunsen burners are being phased out in favor of infrared microincinerators, which serve a similar purpose without the safety risks of an open flame.
• An inoculation loop is a handheld tool that ends in a small wire loop (Figure \(6\)). The loop can be used to streak microorganisms on agar in a Petri dish or to transfer them from one test tube to another. Before each use, the inoculation loop must be sterilized so cultures do not become contaminated.
Key Concepts and Summary
• Microorganisms (or microbes) are living organisms that are generally too small to be seen without a microscope.
• Throughout history, humans have used microbes to make fermented foods such as beer, bread, cheese, and wine.
• Long before the invention of the microscope, some people theorized that infection and disease were spread by living things that were too small to be seen. They also correctly intuited certain principles regarding the spread of disease and immunity.
• Antonie van Leeuwenhoek, using a microscope, was the first to actually describe observations of bacteria, in 1675.
• During the Golden Age of Microbiology (1857–1914), microbiologists, including Louis Pasteur and Robert Koch, discovered many new connections between the fields of microbiology and medicine.
Footnotes
1. 1 Rebecca Buxton. “Examination of Gram Stains of Spinal Fluid—Bacterial Meningitis.” American Society for Microbiology. 2007. www.microbelibrary.org/librar...ial-meningitis
2. 2 P.E. McGovern et al. “Fermented Beverages of Pre- and Proto-Historic China.” Proceedings of the National Academy of Sciences of the United States of America 1 no. 51 (2004):17593–17598. doi:10.1073/pnas.0407921102.
3. 3 A. Keller et al. “New Insights into the Tyrolean Iceman's Origin and Phenotype as Inferred by Whole-Genome Sequencing.”Nature Communications, 3 (2012): 698. doi:10.1038/ncomms1701.
4. 4 L. Capasso. “5300 Years Ago, the Ice Man Used Natural Laxatives and Antibiotics.” The Lancet, 352 (1998) 9143: 1864. doi: 10.1016/s0140-6736(05)79939-6.
5. 5 L. Capasso, L. “5300 Years Ago, the Ice Man Used Natural Laxatives and Antibiotics.” The Lancet, 352 no. 9143 (1998): 1864. doi: 10.1016/s0140-6736(05)79939-6.
6. 6 G. Pappas et al. “Insights Into Infectious Disease in the Era of Hippocrates.” International Journal of Infectious Diseases 12 (2008) 4:347–350. doi: http://dx.doi.org/10.1016/j.ijid.2007.11.003.
7. 7 Thucydides. The History of the Peloponnesian War. The Second Book. 431 BC. Translated by Richard Crawley. http://classics.mit.edu/Thucydides/p....2.second.html.
8. 8 Plinio Prioreschi. A History of Medicine: Roman Medicine. Lewiston, NY: Edwin Mellen Press, 1998: p. 215.
9. 9 S.M. Blevins and M.S. Bronze. “Robert Koch and the ‘Golden Age’ of Bacteriology.” International Journal of Infectious Diseases. 14 no. 9 (2010): e744-e751. doi:10.1016/j.ijid.2009.12.003.
Glossary
microbe
generally, an organism that is too small to be seen without a microscope; also known as a microorganism
microorganism
generally, an organism that is too small to be seen without a microscope; also known as a microbe | textbooks/bio/Microbiology/Microbiology_(OpenStax)/01%3A_An_Invisible_World/1.01%3A_What_Our_Ancestors_Knew.txt |
Learning Objecctives
• Describe how microorganisms are classified and distinguished as unique species
• Compare historical and current systems of taxonomy used to classify microorganisms
Once microbes became visible to humans with the help of microscopes, scientists began to realize their enormous diversity. Microorganisms vary in all sorts of ways, including their size, their appearance, and their rates of reproduction. To study this incredibly diverse new array of organisms, researchers needed a way to systematically organize them.
The Science of Taxonomy
Taxonomy is the classification, description, identification, and naming of living organisms. Classification is the practice of organizing organisms into different groups based on their shared characteristics. The most famous early taxonomist was a Swedish botanist, zoologist, and physician named Carolus Linnaeus (1701–1778). In 1735, Linnaeus published Systema Naturae, an 11-page booklet in which he proposed the Linnaean taxonomy, a system of categorizing and naming organisms using a standard format so scientists could discuss organisms using consistent terminology. He continued to revise and add to the book, which grew into multiple volumes (Figure \(1\)).
In his taxonomy, Linnaeus divided the natural world into three kingdoms: animal, plant, and mineral (the mineral kingdom was later abandoned). Within the animal and plant kingdoms, he grouped organisms using a hierarchy of increasingly specific levels and sublevels based on their similarities. The names of the levels in Linnaeus’s original taxonomy were kingdom, class, order, family, genus (plural: genera), and species. Species was, and continues to be, the most specific and basic taxonomic unit.
Evolving Trees of Life (Phylogenies)
With advances in technology, other scientists gradually made refinements to the Linnaean system and eventually created new systems for classifying organisms. In the 1800s, there was a growing interest in developing taxonomies that took into account the evolutionary relationships, or phylogenies, of all different species of organisms on earth. One way to depict these relationships is via a diagram called a phylogenetic tree (or tree of life). In these diagrams, groups of organisms are arranged by how closely related they are thought to be. In early phylogenetic trees, the relatedness of organisms was inferred by their visible similarities, such as the presence or absence of hair or the number of limbs. Now, the analysis is more complicated. Today, phylogenic analyses include genetic, biochemical, and embryological comparisons, as will be discussed later in this chapter.
Linnaeus’s tree of life contained just two main branches for all living things: the animal and plant kingdoms. In 1866, Ernst Haeckel, a German biologist, philosopher, and physician, proposed another kingdom, Protista, for unicellular organisms (Figure \(2\)). He later proposed a fourth kingdom, Monera, for unicellular organisms whose cells lack nuclei, like bacteria.
Nearly 100 years later, in 1969, American ecologist Robert Whittaker (1920–1980) proposed adding another kingdom—Fungi—in his tree of life. Whittaker’s tree also contained a level of categorization above the kingdom level—the empire or superkingdom level—to distinguish between organisms that have membrane-bound nuclei in their cells (eukaryotes) and those that do not (prokaryotes). Empire Prokaryota contained just the Kingdom Monera. The Empire Eukaryota contained the other four kingdoms: Fungi, Protista, Plantae, and Animalia. Whittaker’s five-kingdom tree was considered the standard phylogeny for many years.
Figure \(3\) shows how the tree of life has changed over time. Note that viruses are not found in any of these trees. That is because they are not made up of cells and thus it is difficult to determine where they would fit into a tree of life.
Exercise \(1\)
Briefly summarize how our evolving understanding of microorganisms has contributed to changes in the way that organisms are classified.
Clinical Focus: Part 2
Antibiotic drugs are specifically designed to kill or inhibit the growth of bacteria. But after a couple of days on antibiotics, Cora shows no signs of improvement. Also, her CSF cultures came back from the lab negative. Since bacteria or fungi were not isolated from Cora’s CSF sample, her doctor rules out bacterial and fungal meningitis. Viral meningitis is still a possibility.
However, Cora now reports some troubling new symptoms. She is starting to have difficulty walking. Her muscle stiffness has spread from her neck to the rest of her body, and her limbs sometimes jerk involuntarily. In addition, Cora’s cognitive symptoms are worsening. At this point, Cora’s doctor becomes very concerned and orders more tests on the CSF samples.
Exercise \(2\)
What types of microorganisms could be causing Cora’s symptoms?
The Role of Genetics in Modern Taxonomy
Haeckel’s and Whittaker’s trees presented hypotheses about the phylogeny of different organisms based on readily observable characteristics. But the advent of molecular genetics in the late 20th century revealed other ways to organize phylogenetic trees. Genetic methods allow for a standardized way to compare all living organisms without relying on observable characteristics that can often be subjective. Modern taxonomy relies heavily on comparing the nucleic acids (deoxyribonucleic acid [DNA] or ribonucleic acid [RNA]) or proteins from different organisms. The more similar the nucleic acids and proteins are between two organisms, the more closely related they are considered to be.
In the 1970s, American microbiologist Carl Woese discovered what appeared to be a “living record” of the evolution of organisms. He and his collaborator George Fox created a genetics-based tree of life based on similarities and differences they observed in the small subunit ribosomal RNA (rRNA) of different organisms. In the process, they discovered that a certain type of bacteria, called archaebacteria (now known simply as archaea), were significantly different from other bacteria and eukaryotes in terms of the sequence of small subunit rRNA. To accommodate this difference, they created a tree with three Domains above the level of Kingdom: Archaea, Bacteria, and Eukarya (Figure \(4\)). Genetic analysis of the small subunit rRNA suggests archaea, bacteria, and eukaryotes all evolved from a common ancestral cell type. The tree is skewed to show a closer evolutionary relationship between Archaea and Eukarya than they have to Bacteria.
Exercise \(3\)
1. In modern taxonomy, how do scientists determine how closely two organisms are related?
2. Explain why the branches on the “tree of life” all originate from a single “trunk.”
Naming Microbes
In developing his taxonomy, Linnaeus used a system of binomial nomenclature, a two-word naming system for identifying organisms by genus and species. For example, modern humans are in the genus Homo and have the species name sapiens, so their scientific name in binomial nomenclature is Homo sapiens. In binomial nomenclature, the genus part of the name is always capitalized; it is followed by the species name, which is not capitalized. Both names are italicized.
Taxonomic names in the 18th through 20th centuries were typically derived from Latin, since that was the common language used by scientists when taxonomic systems were first created. Today, newly discovered organisms can be given names derived from Latin, Greek, or English. Sometimes these names reflect some distinctive trait of the organism; in other cases, microorganisms are named after the scientists who discovered them. The archaeon Haloquadratum walsbyi is an example of both of these naming schemes. The genus, Haloquadratum, describes the microorganism’s saltwater habitat (halo is derived from the Greek word for “salt”) as well as the arrangement of its square cells, which are arranged in square clusters of four cells (quadratum is Latin for “foursquare”). The species, walsbyi, is named after Anthony Edward Walsby, the microbiologist who discovered Haloquadratum walsbyi in in 1980. While it might seem easier to give an organism a common descriptive name—like a red-headed woodpecker—we can imagine how that could become problematic. What happens when another species of woodpecker with red head coloring is discovered? The systematic nomenclature scientists use eliminates this potential problem by assigning each organism a single, unique two-word name that is recognized by scientists all over the world.
In this text, we will typically abbreviate an organism’s genus and species after its first mention. The abbreviated form is simply the first initial of the genus, followed by a period and the full name of the species. For example, the bacterium Escherichia coli is shortened to E. coli in its abbreviated form. You will encounter this same convention in other scientific texts as well.
Bergey’s Manuals
Whether in a tree or a web, microbes can be difficult to identify and classify. Without easily observable macroscopic features like feathers, feet, or fur, scientists must capture, grow, and devise ways to study their biochemical properties to differentiate and classify microbes. Despite these hurdles, a group of microbiologists created and updated a set of manuals for identifying and classifying microorganisms. First published in 1923 and since updated many times, Bergey’s Manual of Determinative Bacteriology and Bergey’s Manual of Systematic Bacteriology are the standard references for identifying and classifying different prokaryotes. (Appendix D of this textbook is partly based on Bergey’s manuals; it shows how the organisms that appear in this textbook are classified.) Because so many bacteria look identical, methods based on nonvisual characteristics must be used to identify them. For example, biochemical tests can be used to identify chemicals unique to certain species. Likewise, serological tests can be used to identify specific antibodies that will react against the proteins found in certain species. Ultimately, DNA and rRNA sequencing can be used both for identifying a particular bacterial species and for classifying newly discovered species.
Exercise \(4\)
• What is binomial nomenclature and why is it a useful tool for naming organisms?
• Explain why a resource like one of Bergey’s manuals would be helpful in identifying a microorganism in a sample.
Same Name, Different Strain
Within one species of microorganism, there can be several subtypes called strains. While different strains may be nearly identical genetically, they can have very different attributes. The bacteriumEscherichia coli is infamous for causing food poisoning and traveler’s diarrhea. However, there are actually many different strains of E. coli, and they vary in their ability to cause disease.
One pathogenic (disease-causing) E. coli strain that you may have heard of is E. coli O157:H7. In humans, infection from E. coli O157:H7 can cause abdominal cramps and diarrhea. Infection usually originates from contaminated water or food, particularly raw vegetables and undercooked meat. In the 1990s, there were several large outbreaks of E. coli O157:H7 thought to have originated in undercooked hamburgers.
While E. coli O157:H7 and some other strains have given E. coli a bad name, most E. coli strains do not cause disease. In fact, some can be helpful. Different strains of E. coli found naturally in our gut help us digest our food, provide us with some needed chemicals, and fight against pathogenic microbes.
Summary
• Carolus Linnaeus developed a taxonomic system for categorizing organisms into related groups.
• Binomial nomenclature assigns organisms Latinized scientific names with a genus and species designation.
• A phylogenetic tree is a way of showing how different organisms are thought to be related to one another from an evolutionary standpoint.
• The first phylogenetic tree contained kingdoms for plants and animals; Ernst Haeckel proposed adding a kingdom for protists.
• Robert Whittaker’s tree contained five kingdoms: Animalia, Plantae, Protista, Fungi, and Monera.
• Carl Woese used small subunit ribosomal RNA to create a phylogenetic tree that groups organisms into three domains based on their genetic similarity.
• Bergey’s manuals of determinative and systemic bacteriology are the standard references for identifying and classifying bacteria, respectively.
• Bacteria can be identified through biochemical tests, DNA/RNA analysis, and serological testing methods.
Glossary
binomial nomenclature
a universal convention for the scientific naming of organisms using Latinized names for genus and species
eukaryote
an organism made up of one or more cells that contain a membrane-bound nucleus and organelles
phylogeny
the evolutionary history of a group of organisms
prokaryote
an organism whose cell structure does not include a membrane-bound nucleus
taxonomy
the classification, description, identification, and naming of living organisms | textbooks/bio/Microbiology/Microbiology_(OpenStax)/01%3A_An_Invisible_World/1.02%3A_A_Systematic_Approach.txt |
Learning Objectives
• List the various types of microorganisms and describe their defining characteristics
• Give examples of different types of cellular and viral microorganisms and infectious agents
• Describe the similarities and differences between archaea and bacteria
• Provide an overview of the field of microbiology
Most microbes are unicellular and small enough that they require artificial magnification to be seen. However, there are some unicellular microbes that are visible to the naked eye, and some multicellular organisms that are microscopic. An object must measure about 100 micrometers (µm) to be visible without a microscope, but most microorganisms are many times smaller than that. For some perspective, consider that a typical animal cell measures roughly 10 µm across but is still microscopic. Bacterial cells are typically about 1 µm, and viruses can be 10 times smaller than bacteria (Figure \(1\)). See Table \(1\) for units of length used in microbiology.
Table \(1\): Units of Length Commonly Used in Microbiology
Metric Unit Meaning of Prefix Metric Equivalent
meter (m) 1 m = 100 m
decimeter (dm) 1/10 1 dm = 0.1 m = 10−1 m
centimeter (cm) 1/100 1 cm = 0.01 m = 10−2 m
millimeter (mm) 1/1000 1 mm = 0.001 m = 10−3 m
micrometer (μm) 1/1,000,000 1 μm = 0.000001 m = 10−6 m
nanometer (nm) 1/1,000,000,000 1 nm = 0.000000001 m = 10−9 m
Microorganisms differ from each other not only in size, but also in structure, habitat, metabolism, and many other characteristics. While we typically think of microorganisms as being unicellular, there are also many multicellular organisms that are too small to be seen without a microscope. Some microbes, such as viruses, are even acellular (not composed of cells).
Microorganisms are found in each of the three domains of life: Archaea, Bacteria, and Eukarya. Microbes within the domains Bacteria and Archaea are all prokaryotes (their cells lack a nucleus), whereas microbes in the domain Eukarya are eukaryotes (their cells have a nucleus). Some microorganisms, such as viruses, do not fall within any of the three domains of life. In this section, we will briefly introduce each of the broad groups of microbes. Later chapters will go into greater depth about the diverse species within each group.
Prokaryotic Microorganisms
Bacteria are found in nearly every habitat on earth, including within and on humans. Most bacteria are harmless or helpful, but some are pathogens, causing disease in humans and other animals. Bacteria are prokaryotic because their genetic material (DNA) is not housed within a true nucleus. Most bacteria have cell walls that contain peptidoglycan.
Bacteria are often described in terms of their general shape. Common shapes include spherical (coccus), rod-shaped (bacillus), or curved (spirillum, spirochete, or vibrio). Figure \(2\) shows examples of these shapes.
They have a wide range of metabolic capabilities and can grow in a variety of environments, using different combinations of nutrients. Some bacteria are photosynthetic, such as oxygenic cyanobacteria and anoxygenic green sulfur and green nonsulfur bacteria; these bacteria use energy derived from sunlight, and fix carbon dioxide for growth. Other types of bacteria are nonphotosynthetic, obtaining their energy from organic or inorganic compounds in their environment.
Archaea are also unicellular prokaryotic organisms. Archaea and bacteria have different evolutionary histories, as well as significant differences in genetics, metabolic pathways, and the composition of their cell walls and membranes. Unlike most bacteria, archaeal cell walls do not contain peptidoglycan, but their cell walls are often composed of a similar substance called pseudopeptidoglycan. Like bacteria, archaea are found in nearly every habitat on earth, even extreme environments that are very cold, very hot, very basic, or very acidic (Figure \(3\)). Some archaea live in the human body, but none have been shown to be human pathogens.
Exercise \(1\)
1. What are the two main types of prokaryotic organisms?
2. Name some of the defining characteristics of each type.
Eukaryotic Microorganisms
The domain Eukarya contains all eukaryotes, including uni- or multicellular eukaryotes such as protists, fungi, plants, and animals. The major defining characteristic of eukaryotes is that their cells contain a nucleus.
Protists
Protists are unicellular eukaryotes that are not plants, animals, or fungi. Algae and protozoa are examples of protists.
Algae (singular: alga) are plant-like protists that can be either unicellular or multicellular (Figure \(4\)). Their cells are surrounded by cell walls made of cellulose, a type of carbohydrate. Algae are photosynthetic organisms that extract energy from the sun and release oxygen and carbohydrates into their environment. Because other organisms can use their waste products for energy, algae are important parts of many ecosystems. Many consumer products contain ingredients derived from algae, such as carrageenan or alginic acid, which are found in some brands of ice cream, salad dressing, beverages, lipstick, and toothpaste. A derivative of algae also plays a prominent role in the microbiology laboratory. Agar, a gel derived from algae, can be mixed with various nutrients and used to grow microorganisms in a Petri dish. Algae are also being developed as a possible source for biofuels.
Protozoa (singular: protozoan) are protists that make up the backbone of many food webs by providing nutrients for other organisms. Protozoa are very diverse. Some protozoa move with help from hair-like structures called cilia or whip-like structures called flagella. Others extend part of their cell membrane and cytoplasm to propel themselves forward. These cytoplasmic extensions are called pseudopods (“false feet”). Some protozoa are photosynthetic; others feed on organic material. Some are free-living, whereas others are parasitic, only able to survive by extracting nutrients from a host organism. Most protozoa are harmless, but some are pathogens that can cause disease in animals or humans (Figure \(5\)).
Fungi
Fungi (singular: fungus) are also eukaryotes. Some multicellular fungi, such as mushrooms, resemble plants, but they are actually quite different. Fungi are not photosynthetic, and their cell walls are usually made out of chitin rather than cellulose.
Unicellular fungi—yeasts—are included within the study of microbiology. There are more than 1000 known species. Yeasts are found in many different environments, from the deep sea to the human navel. Some yeasts have beneficial uses, such as causing bread to rise and beverages to ferment; but yeasts can also cause food to spoil. Some even cause diseases, such as vaginal yeast infections and oral thrush (Figure \(6\)).
Other fungi of interest to microbiologists are multicellular organisms called molds. Molds are made up of long filaments that form visible colonies (Figure \(6\)). Molds are found in many different environments, from soil to rotting food to dank bathroom corners. Molds play a critical role in the decomposition of dead plants and animals. Some molds can cause allergies, and others produce disease-causing metabolites called mycotoxins. Molds have been used to make pharmaceuticals, including penicillin, which is one of the most commonly prescribed antibiotics, and cyclosporine, used to prevent organ rejection following a transplant.
Exercise \(2\)
1. Name two types of protists and two types of fungi.
2. Name some of the defining characteristics of each type.
Helminths
Multicellular parasitic worms called helminths are not technically microorganisms, as most are large enough to see without a microscope. However, these worms fall within the field of microbiology because diseases caused by helminths involve microscopic eggs and larvae. One example of a helminth is the guinea worm, or Dracunculus medinensis, which causes dizziness, vomiting, diarrhea, and painful ulcers on the legs and feet when the worm works its way out of the skin (Figure \(7\)). Infection typically occurs after a person drinks water containing water fleas infected by guinea-worm larvae. In the mid-1980s, there were an estimated 3.5 million cases of guinea-worm disease, but the disease has been largely eradicated. In 2014, there were only 126 cases reported, thanks to the coordinated efforts of the World Health Organization (WHO) and other groups committed to improvements in drinking water sanitation.12
Viruses
Viruses are acellular microorganisms, which means they are not composed of cells. Essentially, a virus consists of proteins and genetic material—either DNA or RNA, but never both—that are inert outside of a host organism. However, by incorporating themselves into a host cell, viruses are able to co-opt the host’s cellular mechanisms to multiply and infect other hosts. Viruses can infect all types of cells, from human cells to the cells of other microorganisms. In humans, viruses are responsible for numerous diseases, from the common cold to deadly Ebola (Figure \(8\)). However, many viruses do not cause disease.
Exercise \(3\)
1. Are helminths microorganisms? Explain why or why not.
2. How are viruses different from other microorganisms?
Microbiology as a Field of Study
Microbiology is a broad term that encompasses the study of all different types of microorganisms. But in practice, microbiologists tend to specialize in one of several subfields. For example, bacteriology is the study of bacteria; mycology is the study of fungi;protozoology is the study of protozoa; parasitology is the study of helminths and other parasites; and virology is the study of viruses (Figure \(9\)). Immunology, the study of the immune system, is often included in the study of microbiology because host–pathogen interactions are central to our understanding of infectious disease processes. Microbiologists can also specialize in certain areas of microbiology, such as clinical microbiology, environmental microbiology, applied microbiology, or food microbiology.
In this textbook, we are primarily concerned with clinical applications of microbiology, but since the various subfields of microbiology are highly interrelated, we will often discuss applications that are not strictly clinical.
Bioethics in Microbiology
In the 1940s, the U.S. government was looking for a solution to a medical problem: the prevalence of sexually transmitted diseases (STDs) among soldiers. Several now-infamous government-funded studies used human subjects to research common STDs and treatments. In one such study, American researchers intentionally exposed more than 1300 human subjects in Guatemala to syphilis, gonorrhea, and chancroid to determine the ability of penicillin and other antibiotics to combat these diseases. Subjects of the study included Guatemalan soldiers, prisoners, prostitutes, and psychiatric patients—none of whom were informed that they were taking part in the study. Researchers exposed subjects to STDs by various methods, from facilitating intercourse with infected prostitutes to inoculating subjects with the bacteria known to cause the diseases. This latter method involved making a small wound on the subject’s genitals or elsewhere on the body, and then putting bacteria directly into the wound.3 In 2011, a U.S. government commission tasked with investigating the experiment revealed that only some of the subjects were treated with penicillin, and 83 subjects died by 1953, likely as a result of the study.4
Unfortunately, this is one of many horrific examples of microbiology experiments that have violated basic ethical standards. Even if this study had led to a life-saving medical breakthrough (it did not), few would argue that its methods were ethically sound or morally justifiable. But not every case is so clear cut. Professionals working in clinical settings are frequently confronted with ethical dilemmas, such as working with patients who decline a vaccine or life-saving blood transfusion. These are just two examples of life-and-death decisions that may intersect with the religious and philosophical beliefs of both the patient and the health-care professional.
No matter how noble the goal, microbiology studies and clinical practice must be guided by a certain set of ethical principles. Studies must be done with integrity. Patients and research subjects provide informed consent (not only agreeing to be treated or studied but demonstrating an understanding of the purpose of the study and any risks involved). Patients’ rights must be respected. Procedures must be approved by an institutional review board. When working with patients, accurate record-keeping, honest communication, and confidentiality are paramount. Animals used for research must be treated humanely, and all protocols must be approved by an institutional animal care and use committee. These are just a few of the ethical principles explored in the Eye on Ethics boxes throughout this book.
Clinical Focus: Resolution
Cora’s CSF samples show no signs of inflammation or infection, as would be expected with a viral infection. However, there is a high concentration of a particular protein, 14-3-3 protein, in her CSF. An electroencephalogram (EEG) of her brain function is also abnormal. The EEG resembles that of a patient with a neurodegenerative disease like Alzheimer’s or Huntington’s, but Cora’s rapid cognitive decline is not consistent with either of these. Instead, her doctor concludes that Cora hasCreutzfeldt-Jakob disease (CJD), a type of transmissible spongiform encephalopathy (TSE).
CJD is an extremely rare disease, with only about 300 cases in the United States each year. It is not caused by a bacterium, fungus, or virus, but rather by prions—which do not fit neatly into any particular category of microbe. Like viruses, prions are not found on the tree of life because they are acellular. Prions are extremely small, about one-tenth the size of a typical virus. They contain no genetic material and are composed solely of a type of abnormal protein.
CJD can have several different causes. It can be acquired through exposure to the brain or nervous-system tissue of an infected person or animal. Consuming meat from an infected animal is one way such exposure can occur. There have also been rare cases of exposure to CJD through contact with contaminated surgical equipment5 and from cornea and growth-hormone donors who unknowingly had CJD.67 In rare cases, the disease results from a specific genetic mutation that can sometimes be hereditary. However, in approximately 85% of patients with CJD, the cause of the disease is spontaneous (or sporadic) and has no identifiable cause.8 Based on her symptoms and their rapid progression, Cora is diagnosed with sporadic CJD.
Unfortunately for Cora, CJD is a fatal disease for which there is no approved treatment. Approximately 90% of patients die within 1 year of diagnosis.9 Her doctors focus on limiting her pain and cognitive symptoms as her disease progresses. Eight months later, Cora dies. Her CJD diagnosis is confirmed with a brain autopsy.
Summary
• Microorganisms are very diverse and are found in all three domains of life: Archaea, Bacteria, and Eukarya.
• Archaea and bacteria are classified as prokaryotes because they lack a cellular nucleus. Archaea differ from bacteria in evolutionary history, genetics, metabolic pathways, and cell wall and membrane composition.
• Archaea inhabit nearly every environment on earth, but no archaea have been identified as human pathogens.
• Eukaryotes studied in microbiology include algae, protozoa, fungi, and helminths.
• Algae are plant-like organisms that can be either unicellular or multicellular, and derive energy via photosynthesis.
• Protozoa are unicellular organisms with complex cell structures; most are motile.
• Microscopic fungi include molds and yeasts.
• Helminths are multicellular parasitic worms. They are included in the field of microbiology because their eggs and larvae are often microscopic.
• Viruses are acellular microorganisms that require a host to reproduce.
• The field of microbiology is extremely broad. Microbiologists typically specialize in one of many subfields, but all health professionals need a solid foundation in clinical microbiology.
Footnotes
1. 1 C. Greenaway “Dracunculiasis (Guinea Worm Disease).” Canadian Medical Association Journal 170 no. 4 (2004):495–500.
2. 2 World Health Organization. “Dracunculiasis (Guinea-Worm Disease).” WHO. 2015. http://www.who.int/mediacentre/factsheets/fs359/en/. Accessed October 2, 2015.
3. 3 Kara Rogers. “Guatemala Syphilis Experiment: American Medical Research Project”. Encylopaedia Britannica. www.britannica.com/event/Guat...lis-experiment. Accessed June 24, 2015.
4. 4 Susan Donaldson James. “Syphilis Experiments Shock, But So Do Third-World Drug Trials.” ABC World News. August 30, 2011. http://abcnews.go.com/Health/guatema...ry?id=14414902. Accessed June 24, 2015.
5. 5 Greg Botelho. “Case of Creutzfeldt-Jakob Disease Confirmed in New Hampshire.” CNN. 2013. http://www.cnn.com/2013/09/20/health...brain-disease/.
6. 6 P. Rudge et al. “Iatrogenic CJD Due to Pituitary-Derived Growth Hormone With Genetically Determined Incubation Times of Up to 40 Years.” Brain 138 no. 11 (2015): 3386–3399.
7. 7 J.G. Heckmann et al. “Transmission of Creutzfeldt-Jakob Disease via a Corneal Transplant.” Journal of Neurology, Neurosurgery & Psychiatry 63 no. 3 (1997): 388–390.
8. 8 National Institute of Neurological Disorders and Stroke. “Creutzfeldt-Jakob Disease Fact Sheet.” NIH. 2015. http://www.ninds.nih.gov/disorders/c....htm#288133058.
9. 9 National Institute of Neurological Disorders and Stroke. “Creutzfeldt-Jakob Disease Fact Sheet.” NIH. 2015. http://www.ninds.nih.gov/disorders/c....htm#288133058. Accessed June 22, 2015.
Glossary
acellular
not consisting of a cell or cells
algae
(singular: alga) any of various unicellular and multicellular photosynthetic eukaryotic organisms; distinguished from plants by their lack of vascular tissues and organs
archaea
any of various unicellular prokaryotic microorganisms, typically having cell walls containing pseudopeptidoglycan
bacteria
(singular: bacterium) any of various unicellular prokaryotic microorganisms typically (but not always) having cell wells that contain peptidoglycan
bacteriology
the study of bacteria
Eukarya
the domain of life that includes all unicellular and multicellular organisms with cells that contain membrane-bound nuclei and organelles
fungi
(singular: fungus) any of various unicellular or multicellular eukaryotic organisms, typically having cell walls made out of chitin and lacking photosynthetic pigments, vascular tissues, and organs
helminth
a multicellular parasitic worm
immunology
the study of the immune system
microbiology
the study of microorganisms
mold
a multicellular fungus, typically made up of long filaments
mycology
the study of fungi
parasitology
the study of parasites
pathogen
a disease-causing microorganism
protist
a unicellular eukaryotic microorganism, usually a type of algae or protozoa
protozoan
(plural: protozoa) a unicellular eukaryotic organism, usually motile
protozoology
the study of protozoa
virology
the study of viruses
virus
an acellular microorganism, consisting of proteins and genetic material (DNA or RNA), that can replicate itself by infecting a host cell
yeast
any unicellular fungus | textbooks/bio/Microbiology/Microbiology_(OpenStax)/01%3A_An_Invisible_World/1.03%3A_Types_of_Microorganisms.txt |
1.1: What Our Ancestors Knew
Microorganisms (or microbes) are living organisms that are generally too small to be seen without a microscope. Throughout history, humans have used microbes to make fermented foods such as beer, bread, cheese, and wine. Long before the invention of the microscope, some people theorized that infection and disease were spread by living things that were too small to be seen. They also correctly intuited certain principles regarding the spread of disease and immunity.
Multiple Choice
Which of the following foods is NOT made by fermentation?
1. beer
2. bread
3. cheese
4. orange juice
Answer
D
Who is considered the “father of Western medicine”?
1. Marcus Terentius Varro
2. Thucydides
3. Antonie van Leeuwenhoek
4. Hippocrates
Answer
D
Who was the first to observe “animalcules” under the microscope?
1. Antonie van Leeuwenhoek
2. Ötzi the Iceman
3. Marcus Terentius Varro
4. Robert Koch
Answer
A
Who proposed that swamps might harbor tiny, disease-causing animals too small to see?
1. Thucydides
2. Marcus Terentius Varro
3. Hippocrates
4. Louis Pasteur
Answer
B
Fill in the Blank
Thucydides is known as the father of _______________.
Answer
scientific history
Researchers think that Ötzi the Iceman may have been infected with _____ disease.
Answer
Lyme
The process by which microbes turn grape juice into wine is called _______________.
Answer
fermentation
Short Answer
What did Thucydides learn by observing the Athenian plague?
Why was the invention of the microscope important for microbiology?
What are some ways people use microbes?
Critical Thinking
Explain how the discovery of fermented foods likely benefited our ancestors.
What evidence would you use to support this statement: Ancient people thought that disease was transmitted by things they could not see.
1.2: A Systematic Approach
Carolus Linnaeus developed a taxonomic system for categorizing organisms into related groups. Binomial nomenclature assigns organisms Latinized scientific names with a genus and species designation. A phylogenetic tree is a way of showing how different organisms are thought to be related to one another from an evolutionary standpoint. The first phylogenetic tree contained kingdoms for plants and animals; Ernst Haeckel proposed adding a kingdom for protists.
Multiple Choice
Which of the following was NOT a kingdom in Linnaeus’s taxonomy?
1. animal
2. mineral
3. protist
4. plant
Answer
C
Which of the following is a correct usage of binomial nomenclature?
1. Homo Sapiens
2. homo sapiens
3. Homo sapiens
4. Homo Sapiens
Answer
C
Which scientist proposed adding a kingdom for protists?
1. Carolus Linnaeus
2. Carl Woese
3. Robert Whittaker
4. Ernst Haeckel
Answer
D
Which of the following is NOT a domain in Woese and Fox’s phylogenetic tree?
1. Plantae
2. Bacteria
3. Archaea
4. Eukarya
Answer
A
Which of the following is the standard resource for identifying bacteria?
1. Systema Naturae
2. Bergey’s Manual of Determinative Bacteriology
3. Woese and Fox’s phylogenetic tree
4. Haeckel’s General Morphology of Organisms
Answer
B
Short Answer
What is a phylogenetic tree?
Which of the five kingdoms in Whittaker’s phylogenetic tree are prokaryotic, and which are eukaryotic?
What molecule did Woese and Fox use to construct their phylogenetic tree?
Name some techniques that can be used to identify and differentiate species of bacteria.
Critical Thinking
Why is using binomial nomenclature more useful than using common names?
Label the three Domains found on modern phylogenetic trees.
1.3: Types of Microorganisms
Microorganisms are very diverse and are found in all three domains of life: Archaea, Bacteria, and Eukarya. Archaea and bacteria are classified as prokaryotes because they lack a cellular nucleus. Archaea differ from bacteria in evolutionary history, genetics, metabolic pathways, and cell wall and membrane composition. Archaea inhabit nearly every environment on earth, but no archaea have been identified as human pathogens.
Multiple Choice
Which of the following types of microorganisms is photosynthetic?
1. yeast
2. virus
3. helminth
4. alga
Answer
D
Which of the following is a prokaryotic microorganism?
1. helminth
2. protozoan
3. cyanobacterium
4. mold
Answer
C
Which of the following is acellular?
1. virus
2. bacterium
3. fungus
4. protozoan
Answer
A
Which of the following is a type of fungal microorganism?
1. bacterium
2. protozoan
3. alga
4. yeast
Answer
D
Which of the following is not a subfield of microbiology?
1. bacteriology
2. botany
3. clinical microbiology
4. virology
Answer
B
Fill in the Blank
A ________ is a disease-causing microorganism.
Answer
pathogen
Multicellular parasitic worms studied by microbiologists are called ___________.
Answer
helminths
The study of viruses is ___________.
Answer
virology
The cells of prokaryotic organisms lack a _______.
Answer
nucleus
Short Answer
Describe the differences between bacteria and archaea.
Name three structures that various protozoa use for locomotion.
Describe the actual and relative sizes of a virus, a bacterium, and a plant or animal cell.
Critical Thinking
Contrast the behavior of a virus outside versus inside a cell.
Where would a virus, bacterium, animal cell, and a prion belong on this chart? | textbooks/bio/Microbiology/Microbiology_(OpenStax)/01%3A_An_Invisible_World/1.E%3A_An_Invisible_World_%28Exercises%29.txt |
When we look at a rainbow, its colors span the full spectrum of light that the human eye can detect and differentiate. Each hue represents a different frequency of visible light, processed by our eyes and brains and rendered as red, orange, yellow, green, or one of the many other familiar colors that have always been a part of the human experience. But only recently have humans developed an understanding of the properties of light that allow us to see images in color.
Over the past several centuries, we have learned to manipulate light to peer into previously invisible worlds—those too small or too far away to be seen by the naked eye. Through a microscope, we can examine microbial cells and colonies, using various techniques to manipulate color, size, and contrast in ways that help us identify species and diagnose disease.
Figure \(1\) illustrates how we can apply the properties of light to visualize and magnify images; but these stunning micrographs are just two examples of the numerous types of images we are now able to produce with different microscopic technologies. This chapter explores how various types of microscopes manipulate light in order to provide a window into the world of microorganisms. By understanding how various kinds of microscopes work, we can produce highly detailed images of microbes that can be useful for both research and clinical applications.
• 2.1: The Properties of Light
Visible light consists of electromagnetic waves that behave like other waves. Hence, many of the properties of light that are relevant to microscopy can be understood in terms of light’s behavior as a wave. An important property of light waves is the wavelength, or the distance between one peak of a wave and the next peak. The height of each peak (or depth of each trough) is called the amplitude.
• 2.2: Peering into the Invisible World
Italian scholar Girolamo Fracastoro is regarded as the first person to formally postulate that disease was spread by tiny invisible seminaria. He proposed that these seeds could attach themselves to certain objects that supported their transfer from person to person. However, since the technology for seeing such tiny objects did not yet exist, the existence of the seminaria remained hypothetical for a little over a century—an invisible world waiting to be revealed.
• 2.3: Instruments of Microscopy
The 20th century saw the development of microscopes that leveraged nonvisible light, such as fluorescence microscopy, which uses an ultraviolet light source, and electron microscopy, which uses short-wavelength electron beams. These advances led to major improvements in magnification, resolution, and contrast. In this section, we will survey the broad range of modern microscopic technology and common applications for each type of microscope.
• 2.4: Staining Microscopic Specimens
In their natural state, most of the cells and microorganisms that we observe under the microscope lack color and contrast. This makes it difficult, if not impossible, to detect important cellular structures and their distinguishing characteristics without artificially treating specimens. Here, we will focus on the most clinically relevant techniques developed to identify specific microbes, cellular structures, DNA sequences, or indicators of infection in tissue samples, under the microscope.
• 2.E: How We See the Invisible World (Exercises)
Thumbnail: A compound microscope in a Biology lab. (CC -BY-SA 4.0; Acagastya).
02: How We See the Invisible World
Learning Objectives
• Identify and define the characteristics of electromagnetic radiation (EMR) used in microscopy
• Explain how lenses are used in microscopy to manipulate visible and ultraviolet (UV) light
Clinical Focus: Part I
Cindy, a 17-year-old counselor at a summer sports camp, scraped her knee playing basketball 2 weeks ago. At the time, she thought it was only a minor abrasion that would heal, like many others before it. Instead, the wound began to look like an insect bite and has continued to become increasingly painful and swollen.
The camp nurse examines the lesion and observes a large amount of pus oozing from the surface. Concerned that Cindy may have developed a potentially aggressive infection, she swabs the wound to collect a sample from the infection site. Then she cleans out the pus and dresses the wound, instructing Cindy to keep the area clean and to come back the next day. When Cindy leaves, the nurse sends the sample to the closest medical lab to be analyzed under a microscope.
Exercise \(1\)
What are some things we can learn about these bacteria by looking at them under a microscope?
Visible light consists of electromagnetic waves that behave like other waves. Hence, many of the properties of light that are relevant to microscopy can be understood in terms of light’s behavior as a wave. An important property of light waves is the wavelength, or the distance between one peak of a wave and the next peak. The height of each peak (or depth of each trough) is called the amplitude. In contrast, the frequency of the wave is the rate of vibration of the wave, or the number of wavelengths within a specified time period (Figure \(1\)).
Interactions of Light
Light waves interact with materials by being reflected, absorbed, or transmitted. Reflection occurs when a wave bounces off of a material. For example, a red piece of cloth may reflect red light to our eyes while absorbing other colors of light. Absorbanceoccurs when a material captures the energy of a light wave. In the case of glow-in-the-dark plastics, the energy from light can be absorbed and then later re-emitted as another form of phosphorescence. Transmission occurs when a wave travels through a material, like light through glass (the process of transmission is called transmittance). When a material allows a large proportion of light to be transmitted, it may do so because it is thinner, or more transparent (having more transparency and less opacity). Figure \(2\) illustrates the difference between transparency and opacity.
Light waves can also interact with each other by interference, creating complex patterns of motion. Dropping two pebbles into a puddle causes the waves on the puddle’s surface to interact, creating complex interference patterns. Light waves can interact in the same way.
In addition to interfering with each other, light waves can also interact with small objects or openings by bending or scattering. This is called diffraction. Diffraction is larger when the object is smaller relative to the wavelength of the light (the distance between two consecutive peaks of a light wave). Often, when waves diffract in different directions around an obstacle or opening, they will interfere with each other.
Exercise \(2\)
1. If a light wave has a long wavelength, is it likely to have a low or high frequency?
2. If an object is transparent, does it reflect, absorb, or transmit light?
Lenses and Refraction
In the context of microscopy, refraction is perhaps the most important behavior exhibited by light waves. Refraction occurs when light waves change direction as they enter a new medium (Figure \(3\)). Different transparent materials transmit light at different speeds; thus, light can change speed when passing from one material to another. This change in speed usually also causes a change in direction (refraction), with the degree of change dependent on the angle of the incoming light.
The extent to which a material slows transmission speed relative to empty space is called the refractive index of that material. Large differences between the refractive indices of two materials will result in a large amount of refraction when light passes from one material to the other. For example, light moves much more slowly through water than through air, so light entering water from air can change direction greatly. We say that the water has a higher refractive index than air (Figure \(4\)).
When light crosses a boundary into a material with a higher refractive index, its direction turns to be closer to perpendicular to the boundary (i.e., more toward a normal to that boundary; Figure \(5\)). This is the principle behind lenses. We can think of a lens as an object with a curved boundary (or a collection of prisms) that collects all of the light that strikes it and refracts it so that it all meets at a single point called the image point (focus). A convex lens can be used to magnify because it can focus at closer range than the human eye, producing a larger image. Concave lenses and mirrors can also be used in microscopes to redirect the light path. Figure \(5\) shows the focal point (the image point when light entering the lens is parallel) and the focal length (the distance to the focal point) for convex and concave lenses.
The human eye contains a lens that enables us to see images. This lens focuses the light reflecting off of objects in front of the eye onto the surface of the retina, which is like a screen in the back of the eye. Artificial lenses placed in front of the eye (contact lenses, glasses, or microscopic lenses) focus light before it is focused (again) by the lens of the eye, manipulating the image that ends up on the retina (e.g., by making it appear larger).
Images are commonly manipulated by controlling the distances between the object, the lens, and the screen, as well as the curvature of the lens. For example, for a given amount of curvature, when an object is closer to the lens, the focal points are farther from the lens. As a result, it is often necessary to manipulate these distances to create a focused image on a screen. Similarly, more curvature creates image points closer to the lens and a larger image when the image is in focus. This property is often described in terms of the focal distance, or distance to the focal point.
Exercise \(3\)
1. Explain how a lens focuses light at the image point.
2. Name some factors that affect the focal length of a lens.
Electromagnetic Spectrum and Color
Visible light is just one form of electromagnetic radiation (EMR), a type of energy that is all around us. Other forms of EMR include microwaves, X-rays, and radio waves, among others. The different types of EMR fall on the electromagnetic spectrum, which is defined in terms of wavelength and frequency. The spectrum of visible light occupies a relatively small range of frequencies between infrared and ultraviolet light (Figure \(6\)).
Whereas wavelength represents the distance between adjacent peaks of a light wave, frequency, in a simplified definition, represents the rate of oscillation. Waves with higher frequencies have shorter wavelengths and, therefore, have more oscillations per unit time than lower-frequency waves. Higher-frequency waves also contain more energy than lower-frequency waves. This energy is delivered as elementary particles called photons. Higher-frequency waves deliver more energetic photons than lower-frequency waves.
Photons with different energies interact differently with the retina. In the spectrum of visible light, each color corresponds to a particular frequency and wavelength (Figure \(6\)).The lowest frequency of visible light appears as the color red, whereas the highest appears as the color violet. When the retina receives visible light of many different frequencies, we perceive this as white light. However, white light can be separated into its component colors using refraction. If we pass white light through a prism, different colors will be refracted in different directions, creating a rainbow-like spectrum on a screen behind the prism. This separation of colors is called dispersion, and it occurs because, for a given material, the refractive index is different for different frequencies of light.
Certain materials can refract nonvisible forms of EMR and, in effect, transform them into visible light. Certain fluorescent dyes, for instance, absorb ultraviolet or blue light and then use the energy to emit photons of a different color, giving off light rather than simply vibrating. This occurs because the energy absorption causes electrons to jump to higher energy states, after which they then almost immediately fall back down to their ground states, emitting specific amounts of energy as photons. Not all of the energy is emitted in a given photon, so the emitted photons will be of lower energy and, thus, of lower frequency than the absorbed ones. Thus, a dye such as Texas red may be excited by blue light, but emit red light; or a dye such as fluorescein isothiocyanate (FITC) may absorb (invisible) high-energy ultraviolet light and emit green light (Figure \(7\)). In some materials, the photons may be emitted following a delay after absorption; in this case, the process is called phosphorescence. Glow-in-the-dark plastic works by using phosphorescent material.
Exercise \(4\)
1. Which has a higher frequency: red light or green light?
2. Explain why dispersion occurs when white light passes through a prism.
3. Why do fluorescent dyes emit a different color of light than they absorb?
Magnification, Resolution, and Contrast
Microscopes magnify images and use the properties of light to create useful images of small objects. Magnification is defined as the ability of a lens to enlarge the image of an object when compared to the real object. For example, a magnification of 10⨯ means that the image appears 10 times the size of the object as viewed with the naked eye.
Greater magnification typically improves our ability to see details of small objects, but magnification alone is not sufficient to make the most useful images. It is often useful to enhance the resolution of objects: the ability to tell that two separate points or objects are separate. A low-resolution image appears fuzzy, whereas a high-resolution image appears sharp. Two factors affect resolution. The first is wavelength. Shorter wavelengths are able to resolve smaller objects; thus, an electron microscope has a much higher resolution than a light microscope, since it uses an electron beam with a very short wavelength, as opposed to the long-wavelength visible light used by a light microscope. The second factor that affects resolution is numerical aperture, which is a measure of a lens’s ability to gather light. The higher the numerical aperture, the better the resolution.
Even when a microscope has high resolution, it can be difficult to distinguish small structures in many specimens because microorganisms are relatively transparent. It is often necessary to increase contrast to detect different structures in a specimen. Various types of microscopes use different features of light or electrons to increase contrast—visible differences between the parts of a specimen (see Instruments of Microscopy). Additionally, dyes that bind to some structures but not others can be used to improve the contrast between images of relatively transparent objects (see Staining Microscopic Specimens).
Exercise \(5\)
1. Explain the difference between magnification and resolution.
2. Explain the difference between resolution and contrast.
3. Name two factors that affect resolution.
Key Concepts and Summary
• Light waves interacting with materials may be reflected, absorbed, or transmitted, depending on the properties of the material.
• Light waves can interact with each other (interference) or be distorted by interactions with small objects or openings (diffraction).
• Refraction occurs when light waves change speed and direction as they pass from one medium to another. Differences in the refraction indices of two materials determine the magnitude of directional changes when light passes from one to the other.
• A lens is a medium with a curved surface that refracts and focuses light to produce an image.
• Visible light is part of the electromagnetic spectrum; light waves of different frequencies and wavelengths are distinguished as colors by the human eye.
• A prism can separate the colors of white light (dispersion) because different frequencies of light have different refractive indices for a given material.
• Fluorescent dyes and phosphorescent materials can effectively transform nonvisible electromagnetic radiation into visible light.
• The power of a microscope can be described in terms of its magnification and resolution.
• Resolution can be increased by shortening wavelength, increasing the numerical aperture of the lens, or using stains that enhance contrast.
Glossary
absorbance
when a molecule captures energy from a photon and vibrates or stretches, using the energy
amplitude
the height of a wave
contrast
visible differences between parts of a microscopic specimen
diffraction
the changing of direction (bending or spreading) that occurs when a light wave interacts with an opening or barrier
dispersion
the separation of light of different frequencies due to different degrees of refraction
fluorescent
the ability of certain materials to absorb energy and then immediately release that energy in the form of light
focal length
the distance from the lens to the image point when the object is at a definite distance from the lens (this is also the distance to the focal point)
focal point
a property of the lens; the image point when light entering the lens is parallel (i.e., the object is an infinite distance from the lens)
frequency
the rate of vibration for a light wave or other electromagnetic wave
image point (focus)
a property of the lens and the distance of the object to the lens; the point at which an image is in focus (the image point is often called the focus)
interference
distortion of a light wave due to interaction with another wave
magnification
the power of a microscope (or lens) to produce an image that appears larger than the actual specimen, expressed as a factor of the actual size
numerical aperture
a measure of a lens’s ability to gather light
opacity
the property of absorbing or blocking light
phosphorescence
the ability of certain materials to absorb energy and then release that energy as light after a delay
reflection
when light bounces back from a surface
refraction
bending of light waves, which occurs when a light wave passes from one medium to another
refractive index
a measure of the magnitude of slowing of light waves by a particular medium
resolution
the ability to distinguish between two points in an image
transmittance
the amount of light that passes through a medium
transparency
the property of allowing light to pass through
wavelength
the distance between one peak of a wave and the next peak | textbooks/bio/Microbiology/Microbiology_(OpenStax)/02%3A_How_We_See_the_Invisible_World/2.01%3A_The_Properties_of_Light.txt |
Learning Objectives
• Describe historical developments and individual contributions that led to the invention and development of the microscope
• Compare and contrast the features of simple and compound microscopes
Some of the fundamental characteristics and functions of microscopes can be understood in the context of the history of their use. Italian scholar Girolamo Fracastoro is regarded as the first person to formally postulate that disease was spread by tiny invisibleseminaria, or “seeds of the contagion.” In his book De Contagione (1546), he proposed that these seeds could attach themselves to certain objects (which he called fomes [cloth]) that supported their transfer from person to person. However, since the technology for seeing such tiny objects did not yet exist, the existence of the seminaria remained hypothetical for a little over a century—an invisible world waiting to be revealed.
Early Microscopes
Antonie van Leeuwenhoek, sometimes hailed as “the Father of Microbiology,” is typically credited as the first person to have created microscopes powerful enough to view microbes (Figure \(1\)). Born in the city of Delft in the Dutch Republic, van Leeuwenhoek began his career selling fabrics. However, he later became interested in lens making (perhaps to look at threads) and his innovative techniques produced microscopes that allowed him to observe microorganisms as no one had before. In 1674, he described his observations of single-celled organisms, whose existence was previously unknown, in a series of letters to the Royal Society of London. His report was initially met with skepticism, but his claims were soon verified and he became something of a celebrity in the scientific community.
While van Leeuwenhoek is credited with the discovery of microorganisms, others before him had contributed to the development of the microscope. These included eyeglass makers in the Netherlands in the late 1500s, as well as the Italian astronomer Galileo Galilei, who used a compound microscope to examine insect parts (Figure \(1\)). Whereas van Leeuwenhoek used a simple microscope, in which light is passed through just one lens, Galileo’s compound microscope was more sophisticated, passing light through two sets of lenses.
Van Leeuwenhoek’s contemporary, the Englishman Robert Hooke (1635–1703), also made important contributions to microscopy, publishing in his book Micrographia (1665) many observations using compound microscopes. Viewing a thin sample of cork through his microscope, he was the first to observe the structures that we now know as cells (Figure \(2\)). Hooke described these structures as resembling “Honey-comb,” and as “small Boxes or Bladders of Air,” noting that each “Cavern, Bubble, or Cell” is distinct from the others (in Latin, “cell” literally means “small room”). They likely appeared to Hooke to be filled with air because the cork cells were dead, with only the rigid cell walls providing the structure.
Exercise \(1\)
1. Explain the difference between simple and compound microscopes.
2. Compare and contrast the contributions of van Leeuwenhoek, Hooke, and Galileo to early microscopy.
Who Invented the Microscope?
While Antonie van Leeuwenhoek and Robert Hooke generally receive much of the credit for early advances in microscopy, neither can claim to be the inventor of the microscope. Some argue that this designation should belong to Hans and Zaccharias Janssen, Dutch spectacle-makers who may have invented the telescope, the simple microscope, and the compound microscope during the late 1500s or early 1600s (Figure \(3\)). Unfortunately, little is known for sure about the Janssens, not even the exact dates of their births and deaths. The Janssens were secretive about their work and never published. It is also possible that the Janssens did not invent anything at all; their neighbor, Hans Lippershey, also developed microscopes and telescopes during the same time frame, and he is often credited with inventing the telescope. The historical records from the time are as fuzzy and imprecise as the images viewed through those early lenses, and any archived records have been lost over the centuries.
By contrast, van Leeuwenhoek and Hooke can thank ample documentation of their work for their respective legacies. Like Janssen, van Leeuwenhoek began his work in obscurity, leaving behind few records. However, his friend, the prominent physician Reinier de Graaf, wrote a letter to the editor of the Philosophical Transactions of the Royal Society of London calling attention to van Leeuwenhoek’s powerful microscopes. From 1673 onward, van Leeuwenhoek began regularly submitting letters to the Royal Society detailing his observations. In 1674, his report describing single-celled organisms produced controversy in the scientific community, but his observations were soon confirmed when the society sent a delegation to investigate his findings. He subsequently enjoyed considerable celebrity, at one point even entertaining a visit by the czar of Russia.
Similarly, Robert Hooke had his observations using microscopes published by the Royal Society in a book called Micrographia in 1665. The book became a bestseller and greatly increased interest in microscopy throughout much of Europe.
Summary
• Antonie van Leeuwenhoek is credited with the first observation of microbes, including protists and bacteria, with simple microscopes that he made.
• Robert Hooke was the first to describe what we now call cells.
• Simple microscopes have a single lens, while compound microscopes have multiple lenses.
Glossary
compound microscope
a microscope that uses multiple lenses to focus light from the specimen
simple microscope
a type of microscope with only one lens to focus light from the specimen | textbooks/bio/Microbiology/Microbiology_(OpenStax)/02%3A_How_We_See_the_Invisible_World/2.02%3A_Peering_into_the_Invisible_World.txt |
Learning Objectives
• Identify and describe the parts of a brightfield microscope
• Calculate total magnification for a compound microscope
• Describe the distinguishing features and typical uses for various types of light microscopes, electron microscopes, and scanning probe microscopes
The early pioneers of microscopy opened a window into the invisible world of microorganisms. But microscopy continued to advance in the centuries that followed. In 1830, Joseph Jackson Lister created an essentially modern light microscope. The 20th century saw the development of microscopes that leveraged nonvisible light, such as fluorescence microscopy, which uses an ultraviolet light source, and electron microscopy, which uses short-wavelength electron beams. These advances led to major improvements in magnification, resolution, and contrast. By comparison, the relatively rudimentary microscopes of van Leeuwenhoek and his contemporaries were far less powerful than even the most basic microscopes in use today. In this section, we will survey the broad range of modern microscopic technology and common applications for each type of microscope.
Light Microscopy
Many types of microscopes fall under the category of light microscopes, which use light to visualize images. Examples of light microscopes include brightfield microscopes, darkfield microscopes, phase-contrast microscopes, differential interference contrast microscopes, fluorescence microscopes, confocal scanning laser microscopes, and two-photon microscopes. These various types of light microscopes can be used to complement each other in diagnostics and research.
Brightfield Microscopes
The brightfield microscope, perhaps the most commonly used type of microscope, is a compound microscope with two or more lenses that produce a dark image on a bright background. Some brightfield microscopes are monocular (having a single eyepiece), though most newer brightfield microscopes are binocular (having two eyepieces), like the one shown in Figure $1$; in either case, each eyepiece contains a lens called an ocular lens. The ocular lenses typically magnify images 10 times (10⨯). At the other end of the body tube are a set of objective lenses on a rotating nosepiece. The magnification of these objective lenses typically ranges from 4⨯ to 100⨯, with the magnification for each lens designated on the metal casing of the lens. The ocular and objective lenses work together to create a magnified image. The total magnification is the product of the ocular magnification times the objective magnification:
$\text{ocular magnification} \times \text{objective magnification} \nonumber$
For example, if a $40 \times$ objective lens is selected and the ocular lens is $10\times$, the total magnification would be
$(40×)(10×)=400× \nonumber$
Components of a typical brightfield microscope.
The item being viewed is called a specimen. The specimen is placed on a glass slide, which is then clipped into place on the stage(a platform) of the microscope. Once the slide is secured, the specimen on the slide is positioned over the light using the x-y mechanical stage knobs. These knobs move the slide on the surface of the stage, but do not raise or lower the stage. Once the specimen is centered over the light, the stage position can be raised or lowered to focus the image. The coarse focusing knob is used for large-scale movements with 4⨯ and 10⨯ objective lenses; the fine focusing knob is used for small-scale movements, especially with 40⨯ or 100⨯ objective lenses.
When images are magnified, they become dimmer because there is less light per unit area of image. Highly magnified images produced by microscopes, therefore, require intense lighting. In a brightfield microscope, this light is provided by an illuminator, which is typically a high-intensity bulb below the stage. Light from the illuminator passes up through condenser lens (located below the stage), which focuses all of the light rays on the specimen to maximize illumination. The position of the condenser can be optimized using the attached condenser focus knob; once the optimal distance is established, the condenser should not be moved to adjust the brightness. If less-than-maximal light levels are needed, the amount of light striking the specimen can be easily adjusted by opening or closing a diaphragm between the condenser and the specimen. In some cases, brightness can also be adjusted using the rheostat, a dimmer switch that controls the intensity of the illuminator.
A brightfield microscope creates an image by directing light from the illuminator at the specimen; this light is differentially transmitted, absorbed, reflected, or refracted by different structures. Different colors can behave differently as they interact withchromophores (pigments that absorb and reflect particular wavelengths of light) in parts of the specimen. Often, chromophores are artificially added to the specimen using stains, which serve to increase contrast and resolution. In general, structures in the specimen will appear darker, to various extents, than the bright background, creating maximally sharp images at magnifications up to about 1000⨯. Further magnification would create a larger image, but without increased resolution. This allows us to see objects as small as bacteria, which are visible at about 400⨯ or so, but not smaller objects such as viruses.
At very high magnifications, resolution may be compromised when light passes through the small amount of air between the specimen and the lens. This is due to the large difference between the refractive indices of air and glass; the air scatters the light rays before they can be focused by the lens. To solve this problem, a drop of oil can be used to fill the space between the specimen and an oil immersion lens, a special lens designed to be used with immersion oils. Since the oil has a refractive index very similar to that of glass, it increases the maximum angle at which light leaving the specimen can strike the lens. This increases the light collected and, thus, the resolution of the image (Figure $2$). A variety of oils can be used for different types of light.
Microscope Maintenance: Best Practices
Even a very powerful microscope cannot deliver high-resolution images if it is not properly cleaned and maintained. Since lenses are carefully designed and manufactured to refract light with a high degree of precision, even a slightly dirty or scratched lens will refract light in unintended ways, degrading the image of the specimen. In addition, microscopes are rather delicate instruments, and great care must be taken to avoid damaging parts and surfaces. Among other things, proper care of a microscope includes the following:
• cleaning the lenses with lens paper
• not allowing lenses to contact the slide (e.g., by rapidly changing the focus)
• protecting the bulb (if there is one) from breakage
• not pushing an objective into a slide
• not using the coarse focusing knob when using the 40⨯ or greater objective lenses
• only using immersion oil with a specialized oil objective, usually the 100⨯ objective
• cleaning oil from immersion lenses after using the microscope
• cleaning any oil accidentally transferred from other lenses
• covering the microscope or placing it in a cabinet when not in use
Darkfield Microscopy
A darkfield microscope is a brightfield microscope that has a small but significant modification to the condenser. A small, opaque disk (about 1 cm in diameter) is placed between the illuminator and the condenser lens. This opaque light stop, as the disk is called, blocks most of the light from the illuminator as it passes through the condenser on its way to the objective lens, producing a hollow cone of light that is focused on the specimen. The only light that reaches the objective is light that has been refracted or reflected by structures in the specimen. The resulting image typically shows bright objects on a dark background (Figure $3$)
An opaque light stop inserted into a brightfield microscope is used to produce a darkfield image. The light stop blocks light traveling directly from the illuminator to the objective lens, allowing only light reflected or refracted off the specimen to reach the eye.
Darkfield microscopy can often create high-contrast, high-resolution images of specimens without the use of stains, which is particularly useful for viewing live specimens that might be killed or otherwise compromised by the stains. For example, thin spirochetes like Treponema pallidum, the causative agent of syphilis, can be best viewed using a darkfield microscope (Figure $4$).
Use of a darkfield microscope allows us to view living, unstained samples of the spirochete Treponema pallidum. Similar to a photographic negative, the spirochetes appear bright against a dark background. (credit: Centers for Disease Control and Prevention/C.W. Hubbard)
Exercise $1$
Identify the key differences between brightfield and darkfield microscopy.
Clinical Focus: Part 2
Wound infections like Cindy’s can be caused by many different types of bacteria, some of which can spread rapidly with serious complications. Identifying the specific cause is very important to select a medication that can kill or stop the growth of the bacteria.
After calling a local doctor about Cindy’s case, the camp nurse sends the sample from the wound to the closest medical laboratory. Unfortunately, since the camp is in a remote area, the nearest lab is small and poorly equipped. A more modern lab would likely use other methods to culture, grow, and identify the bacteria, but in this case, the technician decides to make a wet mount from the specimen and view it under a brightfield microscope. In a wet mount, a small drop of water is added to the slide, and a cover slip is placed over the specimen to keep it in place before it is positioned under the objective lens.
Under the brightfield microscope, the technician can barely see the bacteria cells because they are nearly transparent against the bright background. To increase contrast, the technician inserts an opaque light stop above the illuminator. The resulting darkfield image clearly shows that the bacteria cells are spherical and grouped in clusters, like grapes.
• Why is it important to identify the shape and growth patterns of cells in a specimen?
• What other types of microscopy could be used effectively to view this specimen?
Phase-Contrast Microscopes
Phase-contrast microscopes use refraction and interference caused by structures in a specimen to create high-contrast, high-resolution images without staining. It is the oldest and simplest type of microscope that creates an image by altering the wavelengths of light rays passing through the specimen. To create altered wavelength paths, an annular stop is used in the condenser. The annular stop produces a hollow cone of light that is focused on the specimen before reaching the objective lens. The objective contains a phase plate containing a phase ring. As a result, light traveling directly from the illuminator passes through the phase ring while light refracted or reflected by the specimen passes through the plate. This causes waves traveling through the ring to be about one-half of a wavelength out of phase with those passing through the plate. Because waves have peaks and troughs, they can add together (if in phase together) or cancel each other out (if out of phase). When the wavelengths are out of phase, wave troughs will cancel out wave peaks, which is called destructive interference. Structures that refract light then appear dark against a bright background of only unrefracted light. More generally, structures that differ in features such as refractive index will differ in levels of darkness (Figure $5$).
This diagram of a phase-contrast microscope illustrates phase differences between light passing through the object and background. These differences are produced by passing the rays through different parts of a phase plate. The light rays are superimposed in the image plane, producing contrast due to their interference.
Because it increases contrast without requiring stains, phase-contrast microscopy is often used to observe live specimens. Certain structures, such as organelles in eukaryotic cells and endospores in prokaryotic cells, are especially well visualized with phase-contrast microscopy (Figure $6$).
This figure compares a brightfield image (left) with a phase-contrast image (right) of the same unstained simple squamous epithelial cells. The cells are in the center and bottom right of each photograph (the irregular item above the cells is acellular debris). Notice that the unstained cells in the brightfield image are almost invisible against the background, whereas the cells in the phase-contrast image appear to glow against the background, revealing far more detail. (credit: “Clearly kefir”/Wikimedia Commons)
Differential Interference Contrast Microscopes
Differential interference contrast (DIC) microscopes (also known as Nomarski optics) are similar to phase-contrast microscopes in that they use interference patterns to enhance contrast between different features of a specimen. In a DIC microscope, two beams of light are created in which the direction of wave movement (polarization) differs. Once the beams pass through either the specimen or specimen-free space, they are recombined and effects of the specimens cause differences in the interference patterns generated by the combining of the beams. This results in high-contrast images of living organisms with a three-dimensional appearance. These microscopes are especially useful in distinguishing structures within live, unstained specimens. (Figure $7$).
A DIC image of Fonsecaea pedrosoi grown on modified Leonian’s agar. This fungus causes chromoblastomycosis, a chronic skin infection common in tropical and subtropical climates.
Exercise $2$
What are some advantages of phase-contrast and DIC microscopy?
Fluorescence Microscopes
A fluorescence microscope uses fluorescent chromophores called fluorochromes, which are capable of absorbing energy from a light source and then emitting this energy as visible light. Fluorochromes include naturally fluorescent substances (such as chlorophylls) as well as fluorescent stains that are added to the specimen to create contrast. Dyes such as Texas red and FITC are examples of fluorochromes. Other examples include the nucleic acid dyes 4’,6’-diamidino-2-phenylindole (DAPI) and acridine orange.
The microscope transmits an excitation light, generally a form of EMR with a short wavelength, such as ultraviolet or blue light, toward the specimen; the chromophores absorb the excitation light and emit visible light with longer wavelengths. The excitation light is then filtered out (in part because ultraviolet light is harmful to the eyes) so that only visible light passes through the ocular lens. This produces an image of the specimen in bright colors against a dark background.
Fluorescence microscopes are especially useful in clinical microbiology. They can be used to identify pathogens, to find particular species within an environment, or to find the locations of particular molecules and structures within a cell. Approaches have also been developed to distinguish living from dead cells using fluorescence microscopy based upon whether they take up particular fluorochromes. Sometimes, multiple fluorochromes are used on the same specimen to show different structures or features.
One of the most important applications of fluorescence microscopy is a technique called immunofluorescence, which is used to identify certain disease-causing microbes by observing whether antibodies bind to them. (Antibodies are protein molecules produced by the immune system that attach to specific pathogens to kill or inhibit them.) There are two approaches to this technique: direct immunofluorescence assay (DFA) and indirect immunofluorescence assay (IFA). In DFA, specific antibodies (e.g., those that the target the rabies virus) are stained with a fluorochrome. If the specimen contains the targeted pathogen, one can observe the antibodies binding to the pathogen under the fluorescent microscope. This is called a primary antibody stain because the stained antibodies attach directly to the pathogen.
In IFA, secondary antibodies are stained with a fluorochrome rather than primary antibodies. Secondary antibodies do not attach directly to the pathogen, but they do bind to primary antibodies. When the unstained primary antibodies bind to the pathogen, the fluorescent secondary antibodies can be observed binding to the primary antibodies. Thus, the secondary antibodies are attached indirectly to the pathogen. Since multiple secondary antibodies can often attach to a primary antibody, IFA increases the number of fluorescent antibodies attached to the specimen, making it easier visualize features in the specimen (Figure $8$).
Exercise $3$
Why must fluorochromes be used to examine a specimen under a fluorescence microscope?
Confocal Microscopes
Whereas other forms of light microscopy create an image that is maximally focused at a single distance from the observer (the depth, or z-plane), a confocal microscope uses a laser to scan multiple z-planes successively. This produces numerous two-dimensional, high-resolution images at various depths, which can be constructed into a three-dimensional image by a computer. As with fluorescence microscopes, fluorescent stains are generally used to increase contrast and resolution. Image clarity is further enhanced by a narrow aperture that eliminates any light that is not from the z-plane. Confocal microscopes are thus very useful for examining thick specimens such as biofilms, which can be examined alive and unfixed (Figure $9$).
Two-Photon Microscopes
While the original fluorescent and confocal microscopes allowed better visualization of unique features in specimens, there were still problems that prevented optimum visualization. The effective sensitivity of fluorescence microscopy when viewing thick specimens was generally limited by out-of-focus flare, which resulted in poor resolution. This limitation was greatly reduced in the confocal microscope through the use of a confocal pinhole to reject out-of-focus background fluorescence with thin (<1 μm), unblurred optical sections. However, even the confocal microscopes lacked the resolution needed for viewing thick tissue samples. These problems were resolved with the development of the two-photon microscope, which uses a scanning technique, fluorochromes, and long-wavelength light (such as infrared) to visualize specimens. The low energy associated with the long-wavelength light means that two photons must strike a location at the same time to excite the fluorochrome. The low energy of the excitation light is less damaging to cells, and the long wavelength of the excitation light more easily penetrates deep into thick specimens. This makes the two-photon microscope useful for examining living cells within intact tissues—brain slices, embryos, whole organs, and even entire animals.
Currently, use of two-photon microscopes is limited to advanced clinical and research laboratories because of the high costs of the instruments. A single two-photon microscope typically costs between $300,000 and$500,000, and the lasers used to excite the dyes used on specimens are also very expensive. However, as technology improves, two-photon microscopes may become more readily available in clinical settings.
Exercise $4$
What types of specimens are best examined using confocal or two-photon microscopy?
Electron Microscopy
The maximum theoretical resolution of images created by light microscopes is ultimately limited by the wavelengths of visible light. Most light microscopes can only magnify 1000⨯, and a few can magnify up to 1500⨯, but this does not begin to approach the magnifying power of an electron microscope (EM), which uses short-wavelength electron beams rather than light to increase magnification and resolution.
Electrons, like electromagnetic radiation, can behave as waves, but with wavelengths of 0.005 nm, they can produce much better resolution than visible light. An EM can produce a sharp image that is magnified up to 100,000⨯. Thus, EMs can resolve subcellular structures as well as some molecular structures (e.g., single strands of DNA); however, electron microscopy cannot be used on living material because of the methods needed to prepare the specimens.
There are two basic types of EM: the transmission electron microscope (TEM) and the scanning electron microscope (SEM)(Figure $10$). The TEM is somewhat analogous to the brightfield light microscope in terms of the way it functions. However, it uses an electron beam from above the specimen that is focused using a magnetic lens (rather than a glass lens) and projected through the specimen onto a detector. Electrons pass through the specimen, and then the detector captures the image (Figure $11$).
For electrons to pass through the specimen in a TEM, the specimen must be extremely thin (20–100 nm thick). The image is produced because of varying opacity in various parts of the specimen. This opacity can be enhanced by staining the specimen with materials such as heavy metals, which are electron dense. TEM requires that the beam and specimen be in a vacuum and that the specimen be very thin and dehydrated. The specific steps needed to prepare a specimen for observation under an EM are discussed in detail in the next section.
SEMs form images of surfaces of specimens, usually from electrons that are knocked off of specimens by a beam of electrons. This can create highly detailed images with a three-dimensional appearance that are displayed on a monitor (Figure $12$). Typically, specimens are dried and prepared with fixatives that reduce artifacts, such as shriveling, that can be produced by drying, before being sputter-coated with a thin layer of metal such as gold. Whereas transmission electron microscopy requires very thin sections and allows one to see internal structures such as organelles and the interior of membranes, scanning electron microscopy can be used to view the surfaces of larger objects (such as a pollen grain) as well as the surfaces of very small samples (Figure $13$). Some EMs can magnify an image up to 2,000,000⨯.1
Exercise $5$
1. What are some advantages and disadvantages of electron microscopy, as opposed to light microscopy, for examining microbiological specimens?
2. What kinds of specimens are best examined using TEM? SEM?
Using Microscopy to Study Biofilms
A biofilm is a complex community of one or more microorganism species, typically forming as a slimy coating attached to a surface because of the production of an extrapolymeric substance (EPS) that attaches to a surface or at the interface between surfaces (e.g., between air and water). In nature, biofilms are abundant and frequently occupy complex niches within ecosystems (Figure $14$). In medicine,biofilms can coat medical devices and exist within the body. Because they possess unique characteristics, such as increased resistance against the immune system and to antimicrobial drugs, biofilms are of particular interest to microbiologists and clinicians alike.
Because biofilms are thick, they cannot be observed very well using light microscopy; slicing a biofilm to create a thinner specimen might kill or disturb the microbial community. Confocal microscopy provides clearer images of biofilms because it can focus on one z-plane at a time and produce a three-dimensional image of a thick specimen. Fluorescent dyes can be helpful in identifying cells within the matrix. Additionally, techniques such as immunofluorescence and fluorescence in situ hybridization (FISH), in which fluorescent probes are used to bind to DNA, can be used.
Electron microscopy can be used to observe biofilms, but only after dehydrating the specimen, which produces undesirable artifacts and distorts the specimen. In addition to these approaches, it is possible to follow water currents through the shapes (such as cones and mushrooms) of biofilms, using video of the movement of fluorescently coated beads (Figure $15$).
Scanning Probe Microscopy
A scanning probe microscope does not use light or electrons, but rather very sharp probes that are passed over the surface of the specimen and interact with it directly. This produces information that can be assembled into images with magnifications up to 100,000,000⨯. Such large magnifications can be used to observe individual atoms on surfaces. To date, these techniques have been used primarily for research rather than for diagnostics.
There are two types of scanning probe microscope: the scanning tunneling microscope (STM) and the atomic force microscope (AFM). An STM uses a probe that is passed just above the specimen as a constant voltage bias creates the potential for an electric current between the probe and the specimen. This current occurs via quantum tunneling of electrons between the probe and the specimen, and the intensity of the current is dependent upon the distance between the probe and the specimen. The probe is moved horizontally above the surface and the intensity of the current is measured. Scanning tunneling microscopy can effectively map the structure of surfaces at a resolution at which individual atoms can be detected.
Similar to an STM, AFMs have a thin probe that is passed just above the specimen. However, rather than measuring variations in the current at a constant height above the specimen, an AFM establishes a constant current and measures variations in the height of the probe tip as it passes over the specimen. As the probe tip is passed over the specimen, forces between the atoms (van der Waals forces, capillary forces, chemical bonding, electrostatic forces, and others) cause it to move up and down. Deflection of the probe tip is determined and measured using Hooke’s law of elasticity, and this information is used to construct images of the surface of the specimen with resolution at the atomic level (Figure $16$).
Exercise $6$
1. Which has higher magnification, a light microscope or a scanning probe microscope?
2. Name one advantage and one limitation of scanning probe microscopy.
Key Concepts and Summary
• Numerous types of microscopes use various technologies to generate micrographs. Most are useful for a particular type of specimen or application.
• Light microscopy uses lenses to focus light on a specimen to produce an image. Commonly used light microscopes include brightfield, darkfield, phase-contrast, differential interference contrast, fluorescence, confocal, and two-photon microscopes.
• Electron microscopy focuses electrons on the specimen using magnets, producing much greater magnification than light microscopy. The transmission electron microscope (TEM) and scanning electron microscope (SEM) are two common forms.
• Scanning probe microscopy produces images of even greater magnification by measuring feedback from sharp probes that interact with the specimen. Probe microscopes include the scanning tunneling microscope (STM) and the atomic force microscope (AFM).
Footnotes
1. 1 “JEM-ARM200F Transmission Electron Microscope,” JEOL USA Inc, www.jeolusa.com/PRODUCTS/Tran...specifications. Accessed 8/28/2015.
Glossary
atomic force microscope
a scanning probe microscope that uses a thin probe that is passed just above the specimen to measure forces between the atoms and the probe
binocular
having two eyepieces
brightfield microscope
a compound light microscope with two lenses; it produces a dark image on a bright background
coarse focusing knob
a knob on a microscope that produces relatively large movements to adjust focus
chromophores
pigments that absorb and reflect particular wavelengths of light (giving them a color)
condenser lens
a lens on a microscope that focuses light from the light source onto the specimen
confocal microscope
a scanning laser microscope that uses fluorescent dyes and excitation lasers to create three-dimensional images
darkfield microscope
a compound light microscope that produces a bright image on a dark background; typically a modified brightfield microscope
diaphragm
a component of a microscope; typically consists of a disk under the stage with holes of various sizes; can be adjusted to allow more or less light from the light source to reach the specimen
differential interference-contrast microscope
a microscope that uses polarized light to increase contrast
electron microscope
a type of microscope that uses short-wavelength electron beams rather than light to increase magnification and resolution
fine focusing knob
a knob on a microscope that produces relatively small movements to adjust focus
fluorescence microscope
a microscope that uses natural fluorochromes or fluorescent stains to increase contrast
fluorochromes
chromophores that fluoresce (absorb and then emit light)
illuminator
the light source on a microscope
immunofluorescence
a technique that uses a fluorescence microscope and antibody-specific fluorochromes to determine the presence of specific pathogens in a specimen
monocular
having a single eyepiece
objective lenses
on a light microscope, the lenses closest to the specimen, typically located at the ends of turrets
ocular lens
on a microscope, the lens closest to the eye (also called an eyepiece)
oil immersion lens
a special objective lens on a microscope designed to be used with immersion oil to improve resolution
phase-contrast microscope
a light microscope that uses an annular stop and annular plate to increase contrast
rheostat
a dimmer switch that controls the intensity of the illuminator on a light microscope
scanning electron microscope (SEM)
a type of electron microscope that bounces electrons off of the specimen, forming an image of the surface
scanning probe microscope
a microscope that uses a probe that travels across the surface of a specimen at a constant distance while the current, which is sensitive to the size of the gap, is measured
scanning tunneling microscope
a microscope that uses a probe that is passed just above the specimen as a constant voltage bias creates the potential for an electric current between the probe and the specimen
stage
the platform of a microscope on which slides are placed
total magnification
in a light microscope is a value calculated by multiplying the magnification of the ocular by the magnification of the objective lenses
transmission electron microscope (TEM)
a type of electron microscope that uses an electron beam, focused with magnets, that passes through a thin specimen
two-photon microscope
a microscope that uses long-wavelength or infrared light to fluoresce fluorochromes in the specimen
x-y mechanical stage knobs
knobs on a microscope that are used to adjust the position of the specimen on the stage surface, generally to center it directly above the light | textbooks/bio/Microbiology/Microbiology_(OpenStax)/02%3A_How_We_See_the_Invisible_World/2.03%3A_Instruments_of_Microscopy.txt |
Learning Objectives
• Differentiate between simple and differential stains
• Describe the unique features of commonly used stains
• Explain the procedures and name clinical applications for Gram, endospore, acid-fast, negative capsule, and flagella staining
In their natural state, most of the cells and microorganisms that we observe under the microscope lack color and contrast. This makes it difficult, if not impossible, to detect important cellular structures and their distinguishing characteristics without artificially treating specimens. We have already alluded to certain techniques involving stains and fluorescent dyes, and in this section we will discuss specific techniques for sample preparation in greater detail. Indeed, numerous methods have been developed to identify specific microbes, cellular structures, DNA sequences, or indicators of infection in tissue samples, under the microscope. Here, we will focus on the most clinically relevant techniques.
Preparing Specimens for Light Microscopy
In clinical settings, light microscopes are the most commonly used microscopes. There are two basic types of preparation used to view specimens with a light microscope: wet mounts and fixed specimens.
The simplest type of preparation is the wet mount, in which the specimen is placed on the slide in a drop of liquid. Some specimens, such as a drop of urine, are already in a liquid form and can be deposited on the slide using a dropper. Solid specimens, such as a skin scraping, can be placed on the slide before adding a drop of liquid to prepare the wet mount. Sometimes the liquid used is simply water, but often stains are added to enhance contrast. Once the liquid has been added to the slide, a coverslip is placed on top and the specimen is ready for examination under the microscope.
The second method of preparing specimens for light microscopy is fixation. The “fixing” of a sample refers to the process of attaching cells to a slide. Fixation is often achieved either by heating (heat fixing) or chemically treating the specimen. In addition to attaching the specimen to the slide, fixation also kills microorganisms in the specimen, stopping their movement and metabolism while preserving the integrity of their cellular components for observation.
To heat-fix a sample, a thin layer of the specimen is spread on the slide (called a smear), and the slide is then briefly heated over a heat source (Figure \(1\)). Chemical fixatives are often preferable to heat for tissue specimens. Chemical agents such as acetic acid, ethanol, methanol, formaldehyde (formalin), and glutaraldehyde can denature proteins, stop biochemical reactions, and stabilize cell structures in tissue samples (Figure \(1\)).
In addition to fixation, staining is almost always applied to color certain features of a specimen before examining it under a light microscope. Stains, or dyes, contain salts made up of a positive ion and a negative ion. Depending on the type of dye, the positive or the negative ion may be the chromophore (the colored ion); the other, uncolored ion is called the counterion. If the chromophore is the positively charged ion, the stain is classified as a basic dye; if the negative ion is the chromophore, the stain is considered an acidic dye.
Dyes are selected for staining based on the chemical properties of the dye and the specimen being observed, which determine how the dye will interact with the specimen. In most cases, it is preferable to use a positive stain, a dye that will be absorbed by the cells or organisms being observed, adding color to objects of interest to make them stand out against the background. However, there are scenarios in which it is advantageous to use a negative stain, which is absorbed by the background but not by the cells or organisms in the specimen. Negative staining produces an outline or silhouette of the organisms against a colorful background (Figure \(2\)).
Because cells typically have negatively charged cell walls, the positive chromophores in basic dyes tend to stick to the cell walls, making them positive stains. Thus, commonly used basic dyes such as basic fuchsin, crystal violet, malachite green, methylene blue, and safranin typically serve as positive stains. On the other hand, the negatively charged chromophores in acidic dyes are repelled by negatively charged cell walls, making them negative stains. Commonly used acidic dyes include acid fuchsin, eosin, and rose bengal. Figure \(10\) provides more detail.
Some staining techniques involve the application of only one dye to the sample; others require more than one dye. In simple staining, a single dye is used to emphasize particular structures in the specimen. A simple stain will generally make all of the organisms in a sample appear to be the same color, even if the sample contains more than one type of organism. In contrast, differential stainingdistinguishes organisms based on their interactions with multiple stains. In other words, two organisms in a differentially stained sample may appear to be different colors. Differential staining techniques commonly used in clinical settings include Gram staining, acid-fast staining, endospore staining, flagella staining, and capsule staining. Figure \(11\) provides more detail on these differential staining techniques.
Exercise \(1\)
1. Explain why it is important to fix a specimen before viewing it under a light microscope.
2. What types of specimens should be chemically fixed as opposed to heat-fixed?
3. Why might an acidic dye react differently with a given specimen than a basic dye?
4. Explain the difference between a positive stain and a negative stain.
5. Explain the difference between simple and differential staining.
Gram Staining
The Gram stain procedure is a differential staining procedure that involves multiple steps. It was developed by Danish microbiologist Hans Christian Gram in 1884 as an effective method to distinguish between bacteria with different types of cell walls, and even today it remains one of the most frequently used staining techniques. The steps of the Gram stain procedure are listed below and illustrated in Figure \(3\).
1. First, crystal violet, a primary stain, is applied to a heat-fixed smear, giving all of the cells a purple color.
2. Next, Gram’s iodine, a mordant, is added. A mordant is a substance used to set or stabilize stains or dyes; in this case, Gram’s iodine acts like a trapping agent that complexes with the crystal violet, making the crystal violet–iodine complex clump and stay contained in thick layers of peptidoglycan in the cell walls.
3. Next, a decolorizing agent is added, usually ethanol or an acetone/ethanol solution. Cells that have thick peptidoglycan layers in their cell walls are much less affected by the decolorizing agent; they generally retain the crystal violet dye and remain purple. However, the decolorizing agent more easily washes the dye out of cells with thinner peptidoglycan layers, making them again colorless.
4. Finally, a secondary counterstain, usually safranin, is added. This stains the decolorized cells pink and is less noticeable in the cells that still contain the crystal violet dye.
The purple, crystal-violet stained cells are referred to as gram-positive cells, while the red, safranin-dyed cells are gram-negative (Figure \(4\)). However, there are several important considerations in interpreting the results of a Gram stain. First, older bacterial cells may have damage to their cell walls that causes them to appear gram-negative even if the species is gram-positive. Thus, it is best to use fresh bacterial cultures for Gram staining. Second, errors such as leaving on decolorizer too long can affect the results. In some cases, most cells will appear gram-positive while a few appear gram-negative (as in Figure \(4\)). This suggests damage to the individual cells or that decolorizer was left on for too long; the cells should still be classified as gram-positive if they are all the same species rather than a mixed culture.
Besides their differing interactions with dyes and decolorizing agents, the chemical differences between gram-positive and gram-negative cells have other implications with clinical relevance. For example, Gram staining can help clinicians classify bacterial pathogens in a sample into categories associated with specific properties. Gram-negative bacteria tend to be more resistant to certain antibiotics than gram-positive bacteria. We will discuss this and other applications of Gram staining in more detail in later chapters.
Exercise \(2\)
1. Explain the role of Gram’s iodine in the Gram stain procedure.
2. Explain the role of alcohol in the Gram stain procedure.
3. What color are gram-positive and gram-negative cells, respectively, after the Gram stain procedure?
Clinical Focus: Part 3
Viewing Cindy’s specimen under the darkfield microscope has provided the technician with some important clues about the identity of the microbe causing her infection. However, more information is needed to make a conclusive diagnosis. The technician decides to make a Gram stain of the specimen. This technique is commonly used as an early step in identifying pathogenic bacteria. After completing the Gram stain procedure, the technician views the slide under the brightfield microscope and sees purple, grape-like clusters of spherical cells (Figure \(5\)).
Exercise \(3\)
1. Are these bacteria gram-positive or gram-negative?
2. What does this reveal about their cell walls?
Acid-Fast Stains
Acid-fast staining is another commonly used, differential staining technique that can be an important diagnostic tool. An acid-fast stain is able to differentiate two types of gram-positive cells: those that have waxy mycolic acids in their cell walls, and those that do not. Two different methods for acid-fast staining are the Ziehl-Neelsen technique and the Kinyoun technique. Both use carbolfuchsin as the primary stain. The waxy, acid-fast cells retain the carbolfuchsin even after a decolorizing agent (an acid-alcohol solution) is applied. A secondary counterstain, methylene blue, is then applied, which renders non–acid-fast cells blue.
The fundamental difference between the two carbolfuchsin-based methods is whether heat is used during the primary staining process. The Ziehl-Neelsen method uses heat to infuse the carbolfuchsin into the acid-fast cells, whereas the Kinyoun method does not use heat. Both techniques are important diagnostic tools because a number of specific diseases are caused by acid-fast bacteria(AFB). If AFB are present in a tissue sample, their red or pink color can be seen clearly against the blue background of the surrounding tissue cells (Figure \(6\)).
Exercise \(4\)
Why are acid-fast stains useful?
Using Microscopy to Diagnose Tuberculosis
Mycobacterium tuberculosis, the bacterium that causes tuberculosis, can be detected in specimens based on the presence of acid-fast bacilli. Often, a smear is prepared from a sample of the patient’s sputum and then stained using the Ziehl-Neelsen technique (Figure \(6\)). If acid-fast bacteria are confirmed, they are generally cultured to make a positive identification. Variations of this approach can be used as a first step in determining whether M. tuberculosis or other acid-fast bacteria are present, though samples from elsewhere in the body (such as urine) may contain other Mycobacterium species.
An alternative approach for determining the presence of M. tuberculosis is immunofluorescence. In this technique, fluorochrome-labeled antibodies bind to M. tuberculosis, if present. Antibody-specific fluorescent dyes can be used to view the mycobacteria with a fluorescence microscope.
Capsule Staining
Certain bacteria and yeasts have a protective outer structure called a capsule. Since the presence of a capsule is directly related to a microbe’s virulence (its ability to cause disease), the ability to determine whether cells in a sample have capsules is an important diagnostic tool. Capsules do not absorb most basic dyes; therefore, a negative staining technique (staining around the cells) is typically used for capsule staining. The dye stains the background but does not penetrate the capsules, which appear like halos around the borders of the cell. The specimen does not need to be heat-fixed prior to negative staining.
One common negative staining technique for identifying encapsulated yeast and bacteria is to add a few drops of India ink or nigrosin to a specimen. Other capsular stains can also be used to negatively stain encapsulated cells (Figure \(7\)). Alternatively, positive and negative staining techniques can be combined to visualize capsules: The positive stain colors the body of the cell, and the negative stain colors the background but not the capsule, leaving halo around each cell.
Exercise \(5\)
How does negative staining help us visualize capsules?
Endospore Staining
Endospores are structures produced within certain bacterial cells that allow them to survive harsh conditions. Gram staining alone cannot be used to visualize endospores, which appear clear when Gram-stained cells are viewed. Endospore staining uses two stains to differentiate endospores from the rest of the cell. The Schaeffer-Fulton method (the most commonly used endospore-staining technique) uses heat to push the primary stain (malachite green) into the endospore. Washing with water decolorizes the cell, but the endospore retains the green stain. The cell is then counterstained pink with safranin. The resulting image reveals the shape and location of endospores, if they are present. The green endospores will appear either within the pink vegetative cells or as separate from the pink cells altogether. If no endospores are present, then only the pink vegetative cells will be visible (Figure \(8\)).
Endospore-staining techniques are important for identifying Bacillus and Clostridium, two genera of endospore-producing bacteria that contain clinically significant species. Among others, B. anthracis(which causes anthrax) has been of particular interest because of concern that its spores could be used as a bioterrorism agent. C. difficile is a particularly important species responsible for the typically hospital-acquired infection known as “C. diff.”
Exercise \(6\)
Is endospore staining an example of positive, negative, or differential staining?
Flagella Staining
Flagella (singular: flagellum) are tail-like cellular structures used for locomotion by some bacteria, archaea, and eukaryotes. Because they are so thin, flagella typically cannot be seen under a light microscope without a specialized flagella staining technique. Flagella staining thickens the flagella by first applying mordant (generally tannic acid, but sometimes potassium alum), which coats the flagella; then the specimen is stained with pararosaniline (most commonly) or basic fuchsin (Figure \(9\)).
Though flagella staining is uncommon in clinical settings, the technique is commonly used by microbiologists, since the location and number of flagella can be useful in classifying and identifying bacteria in a sample. When using this technique, it is important to handle the specimen with great care; flagella are delicate structures that can easily be damaged or pulled off, compromising attempts to accurately locate and count the number of flagella.
Preparing Specimens for Electron Microscopy
Samples to be analyzed using a TEM must have very thin sections. But cells are too soft to cut thinly, even with diamond knives. To cut cells without damage, the cells must be embedded in plastic resin and then dehydrated through a series of soaks in ethanol solutions (50%, 60%, 70%, and so on). The ethanol replaces the water in the cells, and the resin dissolves in ethanol and enters the cell, where it solidifies. Next, thin sections are cut using a specialized device called an ultramicrotome (Figure \(12\)). Finally, samples are fixed to fine copper wire or carbon-fiber grids and stained—not with colored dyes, but with substances like uranyl acetate or osmium tetroxide, which contain electron-dense heavy metal atoms.
When samples are prepared for viewing using an SEM, they must also be dehydrated using an ethanol series. However, they must be even drier than is necessary for a TEM. Critical point drying with inert liquid carbon dioxide under pressure is used to displace the water from the specimen. After drying, the specimens are sputter-coated with metal by knocking atoms off of a palladium target, with energetic particles. Sputter-coating prevents specimens from becoming charged by the SEM’s electron beam.
Exercise \(7\)
1. Why is it important to dehydrate cells before examining them under an electron microscope?
2. Name the device that is used to create thin sections of specimens for electron microscopy.
Using Microscopy to Diagnose Syphilis
The causative agent of syphilis is Treponema pallidum, a flexible, spiral cell (spirochete) that can be very thin (<0.15 μm) and match the refractive index of the medium, making it difficult to view using brightfield microscopy. Additionally, this species has not been successfully cultured in the laboratory on an artificial medium; therefore, diagnosis depends upon successful identification using microscopic techniques and serology (analysis of body fluids, often looking for antibodies to a pathogen). Since fixation and staining would kill the cells, darkfield microscopy is typically used for observing live specimens and viewing their movements. However, other approaches can also be used. For example, the cells can be thickened with silver particles (in tissue sections) and observed using a light microscope. It is also possible to use fluorescence or electron microscopy to view Treponema (Figure \(13\)).
In clinical settings, indirect immunofluorescence is often used to identify Treponema. A primary, unstained antibody attaches directly to the pathogen surface, and secondary antibodies “tagged” with a fluorescent stain attach to the primary antibody. Multiple secondary antibodies can attach to each primary antibody, amplifying the amount of stain attached to each Treponema cell, making them easier to spot (Figure \(14\)).
Preparation and Staining for Other Microscopes
Samples for fluorescence and confocal microscopy are prepared similarly to samples for light microscopy, except that the dyes are fluorochromes. Stains are often diluted in liquid before applying to the slide. Some dyes attach to an antibody to stain specific proteins on specific types of cells (immunofluorescence); others may attach to DNA molecules in a process called fluorescence in situ hybridization (FISH), causing cells to be stained based on whether they have a specific DNA sequence.
Sample preparation for two-photon microscopy is similar to fluorescence microscopy, except for the use of infrared dyes. Specimens for STM need to be on a very clean and atomically smooth surface. They are often mica coated with Au(111). Toluene vapor is a common fixative.
Exercise \(8\)
What is the main difference between preparing a sample for fluorescence microscopy versus light microscopy?
Link to Learning
Cornell University’s Case Studies in Microscopy offers a series of clinical problems based on real-life events. Each case study walks you through a clinical problem using appropriate techniques in microscopy at each step.
Clinical Focus: Resolution
From the results of the Gram stain, the technician now knows that Cindy’s infection is caused by spherical, gram-positive bacteria that form grape-like clusters, which is typical of staphylococcal bacteria. After some additional testing, the technician determines that these bacteria are the medically important species known as Staphylococcus aureus, a common culprit in wound infections. Because some strains of S. aureus are resistant to many antibiotics, skin infections may spread to other areas of the body and become serious, sometimes even resulting in amputations or death if the correct antibiotics are not used.
After testing several antibiotics, the lab is able to identify one that is effective against this particular strain of S. aureus. Cindy’s doctor quickly prescribes the medication and emphasizes the importance of taking the entire course of antibiotics, even if the infection appears to clear up before the last scheduled dose. This reduces the risk that any especially resistant bacteria could survive, causing a second infection or spreading to another person.
Microscopy and Antibiotic Resistance
As the use of antibiotics has proliferated in medicine, as well as agriculture, microbes have evolved to become more resistant. Strains of bacteria such as methicillin-resistant S. aureus (MRSA), which has developed a high level of resistance to many antibiotics, are an increasingly worrying problem, so much so that research is underway to develop new and more diversified antibiotics.
Fluorescence microscopy can be useful in testing the effectiveness of new antibiotics against resistant strains like MRSA. In a test of one new antibiotic derived from a marine bacterium, MC21-A (bromophene), researchers used the fluorescent dye SYTOX Green to stain samples of MRSA. SYTOX Green is often used to distinguish dead cells from living cells, with fluorescence microscopy. Live cells will not absorb the dye, but cells killed by an antibiotic will absorb the dye, since the antibiotic has damaged the bacterial cell membrane. In this particular case, MRSA bacteria that had been exposed to MC21-A did, indeed, appear green under the fluorescence microscope, leading researchers to conclude that it is an effective antibiotic against MRSA.
Of course, some argue that developing new antibiotics will only lead to even more antibiotic-resistant microbes, so-called superbugs that could spawn epidemics before new treatments can be developed. For this reason, many health professionals are beginning to exercise more discretion in prescribing antibiotics. Whereas antibiotics were once routinely prescribed for common illnesses without a definite diagnosis, doctors and hospitals are much more likely to conduct additional testing to determine whether an antibiotic is necessary and appropriate before prescribing.
A sick patient might reasonably object to this stingy approach to prescribing antibiotics. To the patient who simply wants to feel better as quickly as possible, the potential benefits of taking an antibiotic may seem to outweigh any immediate health risks that might occur if the antibiotic is ineffective. But at what point do the risks of widespread antibiotic use supersede the desire to use them in individual cases?
Key Concepts and Summary
• Samples must be properly prepared for microscopy. This may involve staining, fixation, and/or cutting thin sections.
• A variety of staining techniques can be used with light microscopy, including Gram staining, acid-fast staining, capsule staining, endospore staining, and flagella staining.
• Samples for TEM require very thin sections, whereas samples for SEM require sputter-coating.
• Preparation for fluorescence microscopy is similar to that for light microscopy, except that fluorochromes are used. | textbooks/bio/Microbiology/Microbiology_(OpenStax)/02%3A_How_We_See_the_Invisible_World/2.04%3A_Staining_Microscopic_Specimens.txt |
2.1: The Properties of Light
Visible light consists of electromagnetic waves that behave like other waves. Hence, many of the properties of light that are relevant to microscopy can be understood in terms of light’s behavior as a wave. An important property of light waves is the wavelength, or the distance between one peak of a wave and the next peak. The height of each peak (or depth of each trough) is called the amplitude.
Multiple Choice
Which of the following has the highest energy?
1. light with a long wavelength
2. light with an intermediate wavelength
3. light with a short wavelength
4. It is impossible to tell from the information given.
Answer
C
You place a specimen under the microscope and notice that parts of the specimen begin to emit light immediately. These materials can be described as _____________.
1. fluorescent
2. phosphorescent
3. transparent
4. opaque
Answer
A
Fill in the Blank
When you see light bend as it moves from air into water, you are observing _________.
Answer
refraction
Short Answer
Explain how a prism separates white light into different colors.
Critical Thinking
In Figure 2.1.6, which of the following has the lowest energy?
1. visible light
2. X-rays
3. ultraviolet rays
4. infrared rays
2.2: Peering into the Invisible World
Italian scholar Girolamo Fracastoro is regarded as the first person to formally postulate that disease was spread by tiny invisible seminaria. He proposed that these seeds could attach themselves to certain objects that supported their transfer from person to person. However, since the technology for seeing such tiny objects did not yet exist, the existence of the seminaria remained hypothetical for a little over a century—an invisible world waiting to be revealed.
Short Answer
Why is Antonie van Leeuwenhoek’s work much better known than that of Zaccharias Janssen?
Why did the cork cells observed by Robert Hooke appear to be empty, as opposed to being full of other structures?
Multiple Choice
Who was the first to describe “cells” in dead cork tissue?
1. Hans Janssen
2. Zaccharias Janssen
3. Antonie van Leeuwenhoek
4. Robert Hooke
Answer
D
Who is the probable inventor of the compound microscope?
1. Girolamo Fracastoro
2. Zaccharias Janssen
3. Antonie van Leeuwenhoek
4. Robert Hooke
Answer
B
Fill in the Blank
A microscope that uses multiple lenses is called a _________ microscope.
Answer
compound
2.3: Instruments of Microscopy
The 20th century saw the development of microscopes that leveraged nonvisible light, such as fluorescence microscopy, which uses an ultraviolet light source, and electron microscopy, which uses short-wavelength electron beams. These advances led to major improvements in magnification, resolution, and contrast. In this section, we survey the broad range of modern microscopic technology and common applications for each type of microscope.
Multiple Choice
Which would be the best choice for viewing internal structures of a living protist such as a Paramecium?
1. a brightfield microscope with a stain
2. a brightfield microscope without a stain
3. a darkfield microscope
4. a transmission electron microscope
Answer
C
Which type of microscope is especially useful for viewing thick structures such as biofilms?
1. a transmission electron microscope
2. a scanning electron microscopes
3. a phase-contrast microscope
4. a confocal scanning laser microscope
5. an atomic force microscope
Answer
D
Which type of microscope would be the best choice for viewing very small surface structures of a cell?
1. a transmission electron microscope
2. a scanning electron microscope
3. a brightfield microscope
4. a darkfield microscope
5. a phase-contrast microscope
Answer
B
What type of microscope uses an annular stop?
1. a transmission electron microscope
2. a scanning electron microscope
3. a brightfield microscope
4. a darkfield microscope
5. a phase-contrast microscope
Answer
E
What type of microscope uses a cone of light so that light only hits the specimen indirectly, producing a darker image on a brighter background?
1. a transmission electron microscope
2. a scanning electron microscope
3. a brightfield microscope
4. a darkfield microscope
5. a phase-contrast microscope
Answer
D
Fill in the Blank
Chromophores that absorb and then emit light are called __________.
Answer
fluorochromes
In a(n) _______ microscope, a probe located just above the specimen moves up and down in response to forces between the atoms and the tip of the probe.
Answer
atomic force microscope
What is the total magnification of a specimen that is being viewed with a standard ocular lens and a 40⨯ objective lens?
Answer
400⨯
Short Answer
What is the function of the condenser in a brightfield microscope?
Label each component of the brightfield microscope.
Critical Thinking
When focusing a light microscope, why is it best to adjust the focus using the coarse focusing knob before using the fine focusing knob?
You need to identify structures within a cell using a microscope. However, the image appears very blurry even though you have a high magnification. What are some things that you could try to improve the resolution of the image? Describe the most basic factors that affect resolution when you first put the slide onto the stage; then consider more specific factors that could affect resolution for 40⨯ and 100⨯ lenses.
2.4: Staining Microscopic Specimens
In their natural state, most of the cells and microorganisms that we observe under the microscope lack color and contrast. This makes it difficult, if not impossible, to detect important cellular structures and their distinguishing characteristics without artificially treating specimens. We focus on the most clinically relevant techniques developed to identify specific microbes, cellular structures, DNA sequences, or indicators of infection in tissue samples, under the microscope.
Multiple Choice
What mordant is used in Gram staining?
1. crystal violet
2. safranin
3. acid-alcohol
4. iodine
Answer
D
What is one difference between specimen preparation for a transmission electron microscope (TEM) and preparation for a scanning electron microscope (SEM)?
1. Only the TEM specimen requires sputter coating.
2. Only the SEM specimen requires sputter-coating.
3. Only the TEM specimen must be dehydrated.
4. Only the SEM specimen must be dehydrated.
Answer
B
Fill in the Blank
Ziehl-Neelsen staining, a type of _______ staining, is diagnostic for Mycobacterium tuberculosis.
Answer
acid-fast
The _______ is used to differentiate bacterial cells based on the components of their cell walls.
Answer
Gram stain
Short Answer
How could you identify whether a particular bacterial sample contained specimens with mycolic acid-rich cell walls?
Critical Thinking
You use the Gram staining procedure to stain an L-form bacterium (a bacterium that lacks a cell wall). What color will the bacterium be after the staining procedure is finished? | textbooks/bio/Microbiology/Microbiology_(OpenStax)/02%3A_How_We_See_the_Invisible_World/2.E%3A_How_We_See_the_Invisible_World_%28Exercises%29.txt |
Life takes many forms, from giant redwood trees towering hundreds of feet in the air to the tiniest known microbes, which measure only a few billionths of a meter. Humans have long pondered life’s origins and debated the defining characteristics of life, but our understanding of these concepts has changed radically since the invention of the microscope. In the 17th century, observations of microscopic life led to the development of the cell theory: the idea that the fundamental unit of life is the cell, that all organisms contain at least one cell, and that cells only come from other cells.
Despite sharing certain characteristics, cells may vary significantly. The two main types of cells are prokaryotic cells (lacking a nucleus) and eukaryotic cells (containing a well-organized, membrane-bound nucleus). Each type of cell exhibits remarkable variety in structure, function, and metabolic activity. This chapter will focus on the historical discoveries that have shaped our current understanding of microbes, including their origins and their role in human disease. We will then explore the distinguishing structures found in prokaryotic and eukaryotic cells.
• 3.1: Spontaneous Generation
The theory of spontaneous generation states that life arose from nonliving matter. It was a long-held belief dating back to Aristotle and the ancient Greeks. Experimentation by Francesco Redi in the 17th century presented the first significant evidence refuting spontaneous generation by showing that flies must have access to meat for maggots to develop on the meat. Louis Pasteur is credited with conclusively disproving the theory and proposed that “life only comes from life.”
• 3.2: Foundations of Modern Cell Theory
Although cells were first observed in the 1660s by Robert Hooke, cell theory was not well accepted for another 200 years. The work of scientists such as Schleiden, Schwann, Remak, and Virchow contributed to its acceptance. Endosymbiotic theory states that mitochondria and chloroplasts, organelles found in many types of organisms, have their origins in bacteria. Significant structural and genetic information support this theory. The miasma theory was widely accepted until the 19th century.
• 3.3: Unique Characteristics of Prokaryotic Cells
Prokaryotic cells differ from eukaryotic cells in that their genetic material is contained in a nucleoid rather than a membrane-bound nucleus. In addition, prokaryotic cells generally lack membrane-bound organelles. Prokaryotic cells of the same species typically share a similar cell morphology and cellular arrangement. Most prokaryotic cells have a cell wall that helps the organism maintain cellular morphology and protects it against changes in osmotic pressure.
• 3.4: Unique Characteristics of Eukaryotic Cells
Eukaryotic cells are defined by the presence of a nucleus containing the DNA genome and bound by a nuclear membrane (or nuclear envelope) composed of two lipid bilayers that regulate transport of materials into and out of the nucleus through nuclear pores. Eukaryotic cell morphologies vary greatly and may be maintained by various structures, including the cytoskeleton, the cell membrane, and/or the cell wall. The nucleolus in the nucleus of eukaryotic cells is the site of ribosomal synthesis.
• 3.E: The Cell (Exercises)
Thumbnail: A 3D rendering of an animal cell cut in half. (CC -BY-SA 4.0; Zaldua I., Equisoain J.J., Zabalza A., Gonzalez E.M., Marzo A., Public University of Navarre).
03: The Cell
Learning Objectives
• Explain the theory of spontaneous generation and why people once accepted it as an explanation for the existence of certain types of organisms
• Explain how certain individuals (van Helmont, Redi, Needham, Spallanzani, and Pasteur) tried to prove or disprove spontaneous generation
Clinical Focus: Part 1
Barbara is a 19-year-old college student living in the dormitory. In January, she came down with a sore throat, headache, mild fever, chills, and a violent but unproductive (i.e., no mucus) cough. To treat these symptoms, Barbara began taking an over-the-counter cold medication, which did not seem to work. In fact, over the next few days, while some of Barbara’s symptoms began to resolve, her cough and fever persisted, and she felt very tired and weak.
Exercise \(1\)
What types of respiratory disease may be responsible?
Humans have been asking for millennia: Where does new life come from? Religion, philosophy, and science have all wrestled with this question. One of the oldest explanations was the theory of spontaneous generation, which can be traced back to the ancient Greeks and was widely accepted through the Middle Ages.
The Theory of Spontaneous Generation
The Greek philosopher Aristotle (384–322 BC) was one of the earliest recorded scholars to articulate the theory of spontaneous generation, the notion that life can arise from nonliving matter. Aristotle proposed that life arose from nonliving material if the material contained pneuma (“vital heat”). As evidence, he noted several instances of the appearance of animals from environments previously devoid of such animals, such as the seemingly sudden appearance of fish in a new puddle of water.1
This theory persisted into the 17th century, when scientists undertook additional experimentation to support or disprove it. By this time, the proponents of the theory cited how frogs simply seem to appear along the muddy banks of the Nile River in Egypt during the annual flooding. Others observed that mice simply appeared among grain stored in barns with thatched roofs. When the roof leaked and the grain molded, mice appeared. Jan Baptista van Helmont, a 17th century Flemish scientist, proposed that mice could arise from rags and wheat kernels left in an open container for 3 weeks. In reality, such habitats provided ideal food sources and shelter for mouse populations to flourish.
However, one of van Helmont’s contemporaries, Italian physician Francesco Redi (1626–1697), performed an experiment in 1668 that was one of the first to refute the idea that maggots (the larvae of flies) spontaneously generate on meat left out in the open air. He predicted that preventing flies from having direct contact with the meat would also prevent the appearance of maggots. Redi left meat in each of six containers (Figure \(1\)). Two were open to the air, two were covered with gauze, and two were tightly sealed. His hypothesis was supported when maggots developed in the uncovered jars, but no maggots appeared in either the gauze-covered or the tightly sealed jars. He concluded that maggots could only form when flies were allowed to lay eggs in the meat, and that the maggots were the offspring of flies, not the product of spontaneous generation.
In 1745, John Needham (1713–1781) published a report of his own experiments, in which he briefly boiled broth infused with plant or animal matter, hoping to kill all preexisting microbes.2 He then sealed the flasks. After a few days, Needham observed that the broth had become cloudy and a single drop contained numerous microscopic creatures. He argued that the new microbes must have arisen spontaneously. In reality, however, he likely did not boil the broth enough to kill all preexisting microbes.
Lazzaro Spallanzani (1729–1799) did not agree with Needham’s conclusions, however, and performed hundreds of carefully executed experiments using heated broth.3 As in Needham’s experiment, broth in sealed jars and unsealed jars was infused with plant and animal matter. Spallanzani’s results contradicted the findings of Needham: Heated but sealed flasks remained clear, without any signs of spontaneous growth, unless the flasks were subsequently opened to the air. This suggested that microbes were introduced into these flasks from the air. In response to Spallanzani’s findings, Needham argued that life originates from a “life force” that was destroyed during Spallanzani’s extended boiling. Any subsequent sealing of the flasks then prevented new life force from entering and causing spontaneous generation (Figure \(2\)).
Exercise \(2\)
1. Describe the theory of spontaneous generation and some of the arguments used to support it.
2. Explain how the experiments of Redi and Spallanzani challenged the theory of spontaneous generation.
Disproving Spontaneous Generation
The debate over spontaneous generation continued well into the 19th century, with scientists serving as proponents of both sides. To settle the debate, the Paris Academy of Sciences offered a prize for resolution of the problem. Louis Pasteur, a prominent French chemist who had been studying microbial fermentation and the causes of wine spoilage, accepted the challenge. In 1858, Pasteur filtered air through a gun-cotton filter and, upon microscopic examination of the cotton, found it full of microorganisms, suggesting that the exposure of a broth to air was not introducing a “life force” to the broth but rather airborne microorganisms.
Later, Pasteur made a series of flasks with long, twisted necks (“swan-neck” flasks), in which he boiled broth to sterilize it (Figure \(3\)). His design allowed air inside the flasks to be exchanged with air from the outside, but prevented the introduction of any airborne microorganisms, which would get caught in the twists and bends of the flasks’ necks. If a life force besides the airborne microorganisms were responsible for microbial growth within the sterilized flasks, it would have access to the broth, whereas the microorganisms would not. He correctly predicted that sterilized broth in his swan-neck flasks would remain sterile as long as the swan necks remained intact. However, should the necks be broken, microorganisms would be introduced, contaminating the flasks and allowing microbial growth within the broth.
Pasteur’s set of experiments irrefutably disproved the theory of spontaneous generation and earned him the prestigious Alhumbert Prize from the Paris Academy of Sciences in 1862. In a subsequent lecture in 1864, Pasteur articulated “Omne vivum ex vivo” (“Life only comes from life”). In this lecture, Pasteur recounted his famous swan-neck flask experiment, stating that “…life is a germ and a germ is life. Never will the doctrine of spontaneous generation recover from the mortal blow of this simple experiment.”4 To Pasteur’s credit, it never has.
Exercise \(3\)
1. How did Pasteur’s experimental design allow air, but not microbes, to enter, and why was this important?
2. What was the control group in Pasteur’s experiment and what did it show?
Summary
• The theory of spontaneous generation states that life arose from nonliving matter. It was a long-held belief dating back to Aristotle and the ancient Greeks.
• Experimentation by Francesco Redi in the 17th century presented the first significant evidence refuting spontaneous generation by showing that flies must have access to meat for maggots to develop on the meat. Prominent scientists designed experiments and argued both in support of (John Needham) and against (Lazzaro Spallanzani) spontaneous generation.
• Louis Pasteur is credited with conclusively disproving the theory of spontaneous generation with his famous swan-neck flask experiment. He subsequently proposed that “life only comes from life.”
Footnotes
1. 1 K. Zwier. “Aristotle on Spontaneous Generation.” www.sju.edu/int/academics/cas...R.%20Zwier.pdf
2. 2 E. Capanna. “Lazzaro Spallanzani: At the Roots of Modern Biology.” Journal of Experimental Zoology 285 no. 3 (1999):178–196.
3. 3 R. Mancini, M. Nigro, G. Ippolito. “Lazzaro Spallanzani and His Refutation of the Theory of Spontaneous Generation.” Le Infezioni in Medicina 15 no. 3 (2007):199–206.
4. 4 R. Vallery-Radot. The Life of Pasteur, trans. R.L. Devonshire. New York: McClure, Phillips and Co, 1902, 1:142. | textbooks/bio/Microbiology/Microbiology_(OpenStax)/03%3A_The_Cell/3.01%3A_Spontaneous_Generation.txt |
Learning Objectives
• Explain the key points of cell theory and the individual contributions of Hooke, Schleiden, Schwann, Remak, and Virchow
• Explain the key points of endosymbiotic theory and cite the evidence that supports this concept
• Explain the contributions of Semmelweis, Snow, Pasteur, Lister, and Koch to the development of germ theory
While some scientists were arguing over the theory of spontaneous generation, other scientists were making discoveries leading to a better understanding of what we now call the cell theory. Modern cell theory has two basic tenets:
• All cells only come from other cells (the principle of biogenesis).
• Cells are the fundamental units of organisms.
Today, these tenets are fundamental to our understanding of life on earth. However, modern cell theory grew out of the collective work of many scientists.
The Origins of Cell Theory
The English scientist Robert Hooke first used the term “cells” in 1665 to describe the small chambers within cork that he observed under a microscope of his own design. To Hooke, thin sections of cork resembled “Honey-comb,” or “small Boxes or Bladders of Air.” He noted that each “Cavern, Bubble, or Cell” was distinct from the others (Figure \(1\)). At the time, Hooke was not aware that the cork cells were long dead and, therefore, lacked the internal structures found within living cells.
Despite Hooke’s early description of cells, their significance as the fundamental unit of life was not yet recognized. Nearly 200 years later, in 1838, Matthias Schleiden (1804–1881), a German botanist who made extensive microscopic observations of plant tissues, described them as being composed of cells. Visualizing plant cells was relatively easy because plant cells are clearly separated by their thick cell walls. Schleiden believed that cells formed through crystallization, rather than cell division.
Theodor Schwann (1810–1882), a noted German physiologist, made similar microscopic observations of animal tissue. In 1839, after a conversation with Schleiden, Schwann realized that similarities existed between plant and animal tissues. This laid the foundation for the idea that cells are the fundamental components of plants and animals.
In the 1850s, two Polish scientists living in Germany pushed this idea further, culminating in what we recognize today as the modern cell theory. In 1852, Robert Remak (1815–1865), a prominent neurologist and embryologist, published convincing evidence that cells are derived from other cells as a result of cell division. However, this idea was questioned by many in the scientific community. Three years later, Rudolf Virchow (1821–1902), a well-respected pathologist, published an editorial essay entitled “Cellular Pathology,” which popularized the concept of cell theory using the Latin phrase omnis cellula a cellula (“all cells arise from cells”), which is essentially the second tenet of modern cell theory.1Given the similarity of Virchow’s work to Remak’s, there is some controversy as to which scientist should receive credit for articulating cell theory. See the following Eye on Ethics feature for more about this controversy.
Science and Plagiarism
Rudolf Virchow, a prominent, Polish-born, German scientist, is often remembered as the “Father of Pathology.” Well known for innovative approaches, he was one of the first to determine the causes of various diseases by examining their effects on tissues and organs. He was also among the first to use animals in his research and, as a result of his work, he was the first to name numerous diseases and created many other medical terms. Over the course of his career, he published more than 2,000 papers and headed various important medical facilities, including the Charité – Universitätsmedizin Berlin, a prominent Berlin hospital and medical school. But he is, perhaps, best remembered for his 1855 editorial essay titled “Cellular Pathology,” published in Archiv für Pathologische Anatomie und Physiologie, a journal that Virchow himself cofounded and still exists today.
Despite his significant scientific legacy, there is some controversy regarding this essay, in which Virchow proposed the central tenet of modern cell theory—that all cells arise from other cells. Robert Remak, a former colleague who worked in the same laboratory as Virchow at the University of Berlin, had published the same idea 3 years before. Though it appears Virchow was familiar with Remak’s work, he neglected to credit Remak’s ideas in his essay. When Remak wrote a letter to Virchow pointing out similarities between Virchow’s ideas and his own, Virchow was dismissive. In 1858, in the preface to one of his books, Virchow wrote that his 1855 publication was just an editorial piece, not a scientific paper, and thus there was no need to cite Remak’s work.
By today’s standards, Virchow’s editorial piece would certainly be considered an act of plagiarism, since he presented Remak’s ideas as his own. However, in the 19th century, standards for academic integrity were much less clear. Virchow’s strong reputation, coupled with the fact that Remak was a Jew in a somewhat anti-Semitic political climate, shielded him from any significant repercussions. Today, the process of peer review and the ease of access to the scientific literature help discourage plagiarism. Although scientists are still motivated to publish original ideas that advance scientific knowledge, those who would consider plagiarizing are well aware of the serious consequences.
In academia, plagiarism represents the theft of both individual thought and research—an offense that can destroy reputations and end careers.2 3 4 5
Exercise \(1\)
1. What are the key points of the cell theory?
2. What contributions did Rudolf Virchow and Robert Remak make to the development of the cell theory?
Endosymbiotic Theory
As scientists were making progress toward understanding the role of cells in plant and animal tissues, others were examining the structures within the cells themselves. In 1831, Scottish botanist Robert Brown (1773–1858) was the first to describe observations of nuclei, which he observed in plant cells. Then, in the early 1880s, German botanist Andreas Schimper (1856–1901) was the first to describe the chloroplasts of plant cells, identifying their role in starch formation during photosynthesis and noting that they divided independent of the nucleus.
Based upon the chloroplasts’ ability to reproduce independently, Russian botanist Konstantin Mereschkowski (1855–1921) suggested in 1905 that chloroplasts may have originated from ancestral photosynthetic bacteria living symbiotically inside a eukaryotic cell. He proposed a similar origin for the nucleus of plant cells. This was the first articulation of the endosymbiotic hypothesis, and would explain how eukaryotic cells evolved from ancestral bacteria.
Mereschkowski’s endosymbiotic hypothesis was furthered by American anatomist Ivan Wallin (1883–1969), who began to experimentally examine the similarities between mitochondria, chloroplasts, and bacteria—in other words, to put the endosymbiotic hypothesis to the test using objective investigation. Wallin published a series of papers in the 1920s supporting the endosymbiotic hypothesis, including a 1926 publication co-authored with Mereschkowski. Wallin claimed he could culture mitochondria outside of their eukaryotic host cells. Many scientists dismissed his cultures of mitochondria as resulting from bacterial contamination. Modern genome sequencing work supports the dissenting scientists by showing that much of the genome of mitochondria had been transferred to the host cell’s nucleus, preventing the mitochondria from being able to live on their own.6 7
Wallin’s ideas regarding the endosymbiotic hypothesis were largely ignored for the next 50 years because scientists were unaware that these organelles contained their own DNA. However, with the discovery of mitochondrial and chloroplast DNA in the 1960s, the endosymbiotic hypothesis was resurrected. Lynn Margulis (1938–2011), an American geneticist, published her ideas regarding the endosymbiotic hypothesis of the origins of mitochondria and chloroplasts in 1967.8 In the decade leading up to her publication, advances in microscopy had allowed scientists to differentiate prokaryotic cells from eukaryotic cells. In her publication, Margulis reviewed the literature and argued that the eukaryotic organelles such as mitochondria and chloroplasts are of prokaryotic origin. She presented a growing body of microscopic, genetic, molecular biology, fossil, and geological data to support her claims.
Again, this hypothesis was not initially popular, but mounting genetic evidence due to the advent of DNA sequencing supported the endosymbiotic theory, which is now defined as the theory that mitochondria and chloroplasts arose as a result of prokaryotic cells establishing a symbiotic relationship within a eukaryotic host (Figure \(3\)). With Margulis’ initial endosymbiotic theory gaining wide acceptance, she expanded on the theory in her 1981 book Symbiosis in Cell Evolution. In it, she explains how endosymbiosis is a major driving factor in the evolution of organisms. More recent genetic sequencing and phylogenetic analysis show that mitochondrial DNA and chloroplast DNA are highly related to their bacterial counterparts, both in DNA sequence and chromosome structure. However, mitochondrial DNA and chloroplast DNA are reduced compared with nuclear DNA because many of the genes have moved from the organelles into the host cell’s nucleus. Additionally, mitochondrial and chloroplast ribosomes are structurally similar to bacterial ribosomes, rather than to the eukaryotic ribosomes of their hosts. Last, the binary fission of these organelles strongly resembles the binary fission of bacteria, as compared with mitosis performed by eukaryotic cells. Since Margulis’ original proposal, scientists have observed several examples of bacterial endosymbionts in modern-day eukaryotic cells. Examples include the endosymbiotic bacteria found within the guts of certain insects, such as cockroaches,9 and photosynthetic bacteria-like organelles found in protists.10
Exercise \(2\)
1. What does the modern endosymbiotic theory state?
2. What evidence supports the endosymbiotic theory?
The Germ Theory of Disease
Prior to the discovery of microbes during the 17th century, other theories circulated about the origins of disease. For example, the ancient Greeks proposed the miasma theory, which held that disease originated from particles emanating from decomposing matter, such as that in sewage or cesspits. Such particles infected humans in close proximity to the rotting material. Diseases including the Black Death, which ravaged Europe’s population during the Middle Ages, were thought to have originated in this way.
In 1546, Italian physician Girolamo Fracastoro proposed, in his essay De Contagione et Contagiosis Morbis, that seed-like spores may be transferred between individuals through direct contact, exposure to contaminated clothing, or through the air. We now recognize Fracastoro as an early proponent of the germ theory of disease, which states that diseases may result from microbial infection. However, in the 16th century, Fracastoro’s ideas were not widely accepted and would be largely forgotten until the 19th century.
In 1847, Hungarian obstetrician Ignaz Semmelweis (Figure \(4\)) observed that mothers who gave birth in hospital wards staffed by physicians and medical students were more likely to suffer and die from puerperal fever after childbirth (10%–20% mortality rate) than were mothers in wards staffed by midwives (1% mortality rate). Semmelweis observed medical students performing autopsies and then subsequently carrying out vaginal examinations on living patients without washing their hands in between. He suspected that the students carried disease from the autopsies to the patients they examined. His suspicions were supported by the untimely death of a friend, a physician who contracted a fatal wound infection after a postmortem examination of a woman who had died of a puerperal infection. The dead physician’s wound had been caused by a scalpel used during the examination, and his subsequent illness and death closely paralleled that of the dead patient.
Although Semmelweis did not know the true cause of puerperal fever, he proposed that physicians were somehow transferring the causative agent to their patients. He suggested that the number of puerperal fever cases could be reduced if physicians and medical students simply washed their hands with chlorinated lime water before and after examining every patient. When this practice was implemented, the maternal mortality rate in mothers cared for by physicians dropped to the same 1% mortality rate observed among mothers cared for by midwives. This demonstrated that handwashing was a very effective method for preventing disease transmission. Despite this great success, many discounted Semmelweis’s work at the time, and physicians were slow to adopt the simple procedure of handwashing to prevent infections in their patients because it contradicted established norms for that time period.
Around the same time Semmelweis was promoting handwashing, in 1848, British physician John Snow conducted studies to track the source of cholera outbreaks in London. By tracing the outbreaks to two specific water sources, both of which were contaminated by sewage, Snow ultimately demonstrated that cholera bacteria were transmitted via drinking water. Snow’s work is influential in that it represents the first known epidemiological study, and it resulted in the first known public health response to an epidemic. The work of both Semmelweis and Snow clearly refuted the prevailing miasma theory of the day, showing that disease is not only transmitted through the air but also through contaminated items.
Although the work of Semmelweis and Snow successfully showed the role of sanitation in preventing infectious disease, the cause of disease was not fully understood. The subsequent work of Louis Pasteur, Robert Koch, and Joseph Listerwould further substantiate the germ theory of disease.
While studying the causes of beer and wine spoilage in 1856, Pasteur discovered properties of fermentation by microorganisms. He had demonstrated with his swan-neck flask experiments (link) that airborne microbes, not spontaneous generation, were the cause of food spoilage, and he suggested that if microbes were responsible for food spoilage and fermentation, they could also be responsible for causing infection. This was the foundation for the germ theory of disease.
Meanwhile, British surgeon Joseph Lister (Figure \(5\)) was trying to determine the causes of postsurgical infections. Many physicians did not give credence to the idea that microbes on their hands, on their clothes, or in the air could infect patients’ surgical wounds, despite the fact that 50% of surgical patients, on average, were dying of postsurgical infections.11 Lister, however, was familiar with the work of Semmelweis and Pasteur; therefore, he insisted on handwashing and extreme cleanliness during surgery. In 1867, to further decrease the incidence of postsurgical wound infections, Lister began using carbolic acid (phenol) spray disinfectant/antiseptic during surgery. His extremely successful efforts to reduce postsurgical infection caused his techniques to become a standard medical practice.
A few years later, Robert Koch (Figure \(5\)) proposed a series of postulates (Koch’s postulates) based on the idea that the cause of a specific disease could be attributed to a specific microbe. Using these postulates, Koch and his colleagues were able to definitively identify the causative pathogens of specific diseases, including anthrax, tuberculosis, and cholera. Koch’s “one microbe, one disease” concept was the culmination of the 19th century’s paradigm shift away from miasma theory and toward the germ theory of disease. Koch’s postulates are discussed more thoroughly in How Pathogens Cause Disease.
Exercise \(3\)
1. Compare and contrast the miasma theory of disease with the germ theory of disease.
2. How did Joseph Lister’s work contribute to the debate between the miasma theory and germ theory and how did this increase the success of medical procedures?
Clinical Focus: Part 2
After suffering a fever, congestion, cough, and increasing aches and pains for several days, Barbara suspects that she has a case of the flu. She decides to visit the health center at her university. The PA tells Barbara that her symptoms could be due to a range of diseases, such as influenza, bronchitis, pneumonia, or tuberculosis.
During her physical examination, the PA notes that Barbara’s heart rate is slightly elevated. Using a pulse oximeter, a small device that clips on her finger, he finds that Barbara has hypoxemia—a lower-than-normal level of oxygen in the blood. Using a stethoscope, the PA listens for abnormal sounds made by Barbara’s heart, lungs, and digestive system. As Barbara breathes, the PA hears a crackling sound and notes a slight shortness of breath. He collects a sputum sample, noting the greenish color of the mucus, and orders a chest radiograph, which shows a “shadow” in the left lung. All of these signs are suggestive of pneumonia, a condition in which the lungs fill with mucus (Figure \(6\)).
Exercise \(4\)
What kinds of infectious agents are known to cause pneumonia?
Key Concepts and Summary
• Although cells were first observed in the 1660s by Robert Hooke, cell theory was not well accepted for another 200 years. The work of scientists such as Schleiden, Schwann, Remak, and Virchow contributed to its acceptance.
• Endosymbiotic theory states that mitochondria and chloroplasts, organelles found in many types of organisms, have their origins in bacteria. Significant structural and genetic information support this theory.
• The miasma theory of disease was widely accepted until the 19th century, when it was replaced by the germ theory of disease thanks to the work of Semmelweis, Snow, Pasteur, Lister, and Koch, and others.
Footnotes
1. 1 M. Schultz. “Rudolph Virchow.” Emerging Infectious Diseases 14 no. 9 (2008):1480–1481.
2. 2 B. Kisch. “Forgotten Leaders in Modern Medicine, Valentin, Gouby, Remak, Auerbach.” Transactions of the American Philosophical Society 44 (1954):139–317.
3. 3 H. Harris. The Birth of the Cell. New Haven, CT: Yale University Press, 2000:133.
4. 4 C. Webster (ed.). Biology, Medicine and Society 1840-1940. Cambridge, UK; Cambridge University Press, 1981:118–119.
5. 5 C. Zuchora-Walske. Key Discoveries in Life Science. Minneapolis, MN: Lerner Publishing, 2015:12–13.
6. 6 T. Embley, W. Martin. “Eukaryotic Evolution, Changes, and Challenges.” Nature Vol. 440 (2006):623–630.
7. 7 O.G. Berg, C.G. Kurland. “Why Mitochondrial Genes Are Most Often Found in Nuclei.” Molecular Biology and Evolution 17 no. 6 (2000):951–961.
8. 8 L. Sagan. “On the Origin of Mitosing Cells.” Journal of Theoretical Biology 14 no. 3 (1967):225–274.
9. 9 A.E. Douglas. “The Microbial Dimension in Insect Nutritional Ecology.” Functional Ecology 23 (2009):38–47.
10. 10 J.M. Jaynes, L.P. Vernon. “The Cyanelle of Cyanophora paradoxa: Almost a Cyanobacterial Chloroplast.” Trends in Biochemical Sciences 7 no. 1 (1982):22–24.
11. 11 Alexander, J. Wesley. “The Contributions of Infection Control to a Century of Progress” Annals of Surgery 201:423-428, 1985. | textbooks/bio/Microbiology/Microbiology_(OpenStax)/03%3A_The_Cell/3.02%3A_Foundations_of_Modern_Cell_Theory.txt |
Learning Objectives
• Explain the distinguishing characteristics of prokaryotic cells
• Describe common cell morphologies and cellular arrangements typical of prokaryotic cells and explain how cells maintain their morphology
• Describe internal and external structures of prokaryotic cells in terms of their physical structure, chemical structure, and function
• Compare the distinguishing characteristics of bacterial and archaeal cells
Cell theory states that the cell is the fundamental unit of life. However, cells vary significantly in size, shape, structure, and function. At the simplest level of construction, all cells possess a few fundamental components. These include cytoplasm (a gel-like substance composed of water and dissolved chemicals needed for growth), which is contained within a plasma membrane (also called a cell membrane or cytoplasmic membrane); one or more chromosomes, which contain the genetic blueprints of the cell; and ribosomes, organelles used for the production of proteins.
Beyond these basic components, cells can vary greatly between organisms, and even within the same multicellular organism. The two largest categories of cells—prokaryotic cells and eukaryotic cells—are defined by major differences in several cell structures. Prokaryotic cells lack a nucleus surrounded by a complex nuclear membrane and generally have a single, circular chromosome located in a nucleoid. Eukaryotic cells have a nucleus surrounded by a complex nuclear membrane that contains multiple, rod-shaped chromosomes.1
All plant cells and animal cells are eukaryotic. Some microorganisms are composed of prokaryotic cells, whereas others are composed of eukaryotic cells. Prokaryotic microorganisms are classified within the domains Archaea and Bacteria, whereas eukaryotic organisms are classified within the domain Eukarya.
The structures inside a cell are analogous to the organs inside a human body, with unique structures suited to specific functions. Some of the structures found in prokaryotic cells are similar to those found in some eukaryotic cells; others are unique to prokaryotes. Although there are some exceptions, eukaryotic cells tend to be larger than prokaryotic cells. The comparatively larger size of eukaryotic cells dictates the need to compartmentalize various chemical processes within different areas of the cell, using complex membrane-bound organelles. In contrast, prokaryotic cells generally lack membrane-bound organelles; however, they often contain inclusions that compartmentalize their cytoplasm. Figure \(1\) illustrates structures typically associated with prokaryotic cells. These structures are described in more detail in the next section.
Common Cell Morphologies and Arrangements
Individual cells of a particular prokaryotic organism are typically similar in shape, or cell morphology. Although thousands of prokaryotic organisms have been identified, only a handful of cell morphologies are commonly seen microscopically. Figure \(2\) names and illustrates cell morphologies commonly found in prokaryotic cells. In addition to cellular shape, prokaryotic cells of the same species may group together in certain distinctive arrangements depending on the plane of cell division. Some common arrangements are shown in Figure \(3\).
In most prokaryotic cells, morphology is maintained by the cell wall in combination with cytoskeletal elements. The cell wall is a structure found in most prokaryotes and some eukaryotes; it envelopes the cell membrane, protecting the cell from changes in osmotic pressure (Figure \(4\)). Osmotic pressure occurs because of differences in the concentration of solutes on opposing sides of a semipermeable membrane. Water is able to pass through a semipermeable membrane, but solutes (dissolved molecules like salts, sugars, and other compounds) cannot. When the concentration of solutes is greater on one side of the membrane, water diffuses across the membrane from the side with the lower concentration (more water) to the side with the higher concentration (less water) until the concentrations on both sides become equal. This diffusion of water is called osmosis, and it can cause extreme osmotic pressure on a cell when its external environment changes.
The external environment of a cell can be described as an isotonic, hypertonic, or hypotonic medium. In an isotonic medium, the solute concentrations inside and outside the cell are approximately equal, so there is no net movement of water across the cell membrane. In a hypertonic medium, the solute concentration outside the cell exceeds that inside the cell, so water diffuses out of the cell and into the external medium. In a hypotonic medium, the solute concentration inside the cell exceeds that outside of the cell, so water will move by osmosis into the cell. This causes the cell to swell and potentially lyse, or burst.
The degree to which a particular cell is able to withstand changes in osmotic pressure is called tonicity. Cells that have a cell wall are better able to withstand subtle changes in osmotic pressure and maintain their shape. In hypertonic environments, cells that lack a cell wall can become dehydrated, causing crenation, or shriveling of the cell; the plasma membrane contracts and appears scalloped or notched (Figure \(4\)). By contrast, cells that possess a cell wall undergo plasmolysis rather than crenation. In plasmolysis, the plasma membrane contracts and detaches from the cell wall, and there is a decrease in interior volume, but the cell wall remains intact, thus allowing the cell to maintain some shape and integrity for a period of time (Figure \(5\)). Likewise, cells that lack a cell wall are more prone to lysis in hypotonic environments. The presence of a cell wall allows the cell to maintain its shape and integrity for a longer time before lysing (Figure \(5\)).
Exercise \(1\)
1. Explain the difference between cell morphology and arrangement.
2. What advantages do cell walls provide prokaryotic cells?
The Nucleoid
All cellular life has a DNA genome organized into one or more chromosomes. Prokaryotic chromosomes are typically circular, haploid (unpaired), and not bound by a complex nuclear membrane. Prokaryotic DNA and DNA-associated proteins are concentrated within the nucleoid region of the cell (Figure \(6\)). In general, prokaryotic DNA interacts with nucleoid-associated proteins (NAPs) that assist in the organization and packaging of the chromosome. In bacteria, NAPs function similar to histones, which are the DNA-organizing proteins found in eukaryotic cells. In archaea, the nucleoid is organized by either NAPs or histone-like DNA organizing proteins.
Plasmids
Prokaryotic cells may also contain extrachromosomal DNA, or DNA that is not part of the chromosome. This extrachromosomal DNA is found in plasmids, which are small, circular, double-stranded DNA molecules. Cells that have plasmids often have hundreds of them within a single cell. Plasmids are more commonly found in bacteria; however, plasmids have been found in archaea and eukaryotic organisms. Plasmids often carry genes that confer advantageous traits such as antibiotic resistance; thus, they are important to the survival of the organism. We will discuss plasmids in more detail in Mechanisms of Microbial Genetics.
Ribosomes
All cellular life synthesizes proteins, and organisms in all three domains of life possess ribosomes, structures responsible protein synthesis. However, ribosomes in each of the three domains are structurally different. Ribosomes, themselves, are constructed from proteins, along with ribosomal RNA (rRNA). Prokaryotic ribosomes are found in the cytoplasm. They are called 70S ribosomes because they have a size of 70S (Figure \(7\)), whereas eukaryotic cytoplasmic ribosomes have a size of 80S. (The S stands for Svedberg unit, a measure of sedimentation in an ultracentrifuge, which is based on size, shape, and surface qualities of the structure being analyzed). Although they are the same size, bacterial and archaeal ribosomes have different proteins and rRNA molecules, and the archaeal versions are more similar to their eukaryotic counterparts than to those found in bacteria.
Inclusions
As single-celled organisms living in unstable environments, some prokaryotic cells have the ability to store excess nutrients within cytoplasmic structures called inclusions. Storing nutrients in a polymerized form is advantageous because it reduces the buildup of osmotic pressure that occurs as a cell accumulates solutes. Various types of inclusions store glycogen and starches, which contain carbon that cells can access for energy. Volutin granules, also called metachromatic granules because of their staining characteristics, are inclusions that store polymerized inorganic phosphate that can be used in metabolism and assist in the formation of biofilms. Microbes known to contain volutin granules include the archaea Methanosarcina, the bacterium Corynebacterium diphtheriae, and the unicellular eukaryotic alga Chlamydomonas. Sulfur granules, another type of inclusion, are found in sulfur bacteria of the genus Thiobacillus; these granules store elemental sulfur, which the bacteria use for metabolism.
Occasionally, certain types of inclusions are surrounded by a phospholipid monolayer embedded with protein. Polyhydroxybutyrate (PHB), which can be produced by species of Bacillus and Pseudomonas, is an example of an inclusion that displays this type of monolayer structure. Industrially, PHB has also been used as a source of biodegradable polymers for bioplastics. Several different types of inclusions are shown in Figure \(8\).
Some prokaryotic cells have other types of inclusions that serve purposes other than nutrient storage. For example, some prokaryotic cells produce gas vacuoles, accumulations of small, protein-lined vesicles of gas. These gas vacuoles allow the prokaryotic cells that synthesize them to alter their buoyancy so that they can adjust their location in the water column. Magnetotactic bacteria, such as Magnetospirillum magnetotacticum, contain magnetosomes, which are inclusions of magnetic iron oxide or iron sulfide surrounded by a lipid layer. These allow cells to align along a magnetic field, aiding their movement (Figure \(8\)). Cyanobacteria such as Anabaena cylindrica and bacteria such as Halothiobacillus neapolitanus produce carboxysome inclusions. Carboxysomes are composed of outer shells of thousands of protein subunits. Their interior is filled with ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) and carbonic anhydrase. Both of these compounds are used for carbon metabolism. Some prokaryotic cells also possess carboxysomes that sequester functionally related enzymes in one location. These structures are considered proto-organelles because they compartmentalize important compounds or chemical reactions, much like many eukaryotic organelles.
Endospores
Bacterial cells are generally observed as vegetative cells, but some genera of bacteria have the ability to form endospores, structures that essentially protect the bacterial genome in a dormant state when environmental conditions are unfavorable. Endospores (not to be confused with the reproductive spores formed by fungi) allow some bacterial cells to survive long periods without food or water, as well as exposure to chemicals, extreme temperatures, and even radiation. Table \(1\) compares the characteristics of vegetative cells and endospores.
Table \(1\): Characteristics of Vegetative Cells versus Endospores
Vegetative Cells Endospores
Sensitive to extreme temperatures and radiation Resistant to extreme temperatures and radiation
Gram-positive Do not absorb Gram stain, only special endospore stains (see Staining Microscopic Specimens)
Normal water content and enzymatic activity Dehydrated; no metabolic activity
Capable of active growth and metabolism Dormant; no growth or metabolic activity
The process by which vegetative cells transform into endospores is called sporulation, and it generally begins when nutrients become depleted or environmental conditions become otherwise unfavorable (Figure \(9\)). The process begins with the formation of a septum in the vegetative bacterial cell. The septum divides the cell asymmetrically, separating a DNA forespore from the mother cell. The forespore, which will form the core of the endospore, is essentially a copy of the cell’s chromosomes, and is separated from the mother cell by a second membrane. A cortex gradually forms around the forespore by laying down layers of calcium and dipicolinic acid between membranes. A protein spore coat then forms around the cortex while the DNA of the mother cell disintegrates. Further maturation of the endospore occurs with the formation of an outermost exosporium. The endospore is released upon disintegration of the mother cell, completing sporulation.
Endospores of certain species have been shown to persist in a dormant state for extended periods of time, up to thousands of years.2 However, when living conditions improve, endospores undergo germination, reentering a vegetative state. After germination, the cell becomes metabolically active again and is able to carry out all of its normal functions, including growth and cell division.
Not all bacteria have the ability to form endospores; however, there are a number of clinically significant endospore-forming gram-positive bacteria of the genera Bacillus and Clostridium. These include B. anthracis, the causative agent of anthrax, which produces endospores capable of survive for many decades3; C. tetani (causes tetanus); C. difficile (causes pseudomembranous colitis); C. perfringens (causes gas gangrene); and C. botulinum (causes botulism). Pathogens such as these are particularly difficult to combat because their endospores are so hard to kill. Special sterilization methods for endospore-forming bacteria are discussed in Control of Microbial Growth.
Exercise \(2\)
1. What is an inclusion?
2. What is the function of an endospore?
Plasma Membrane
Structures that enclose the cytoplasm and internal structures of the cell are known collectively as the cell envelope. In prokaryotic cells, the structures of the cell envelope vary depending on the type of cell and organism. Most (but not all) prokaryotic cells have a cell wall, but the makeup of this cell wall varies. All cells (prokaryotic and eukaryotic) have a plasma membrane (also called cytoplasmic membrane or cell membrane) that exhibits selective permeability, allowing some molecules to enter or leave the cell while restricting the passage of others.
The structure of the plasma membrane is often described in terms of the fluid mosaic model, which refers to the ability of membrane components to move fluidly within the plane of the membrane, as well as the mosaic-like composition of the components, which include a diverse array of lipid and protein components (Figure \(10\)). The plasma membrane structure of most bacterial and eukaryotic cell types is a bilayer composed mainly of phospholipids formed with ester linkages and proteins. These phospholipids and proteins have the ability to move laterally within the plane of the membranes as well as between the two phospholipid layers.
Archaeal membranes are fundamentally different from bacterial and eukaryotic membranes in a few significant ways. First, archaeal membrane phospholipids are formed with ether linkages, in contrast to the ester linkages found in bacterial or eukaryotic cell membranes. Second, archaeal phospholipids have branched chains, whereas those of bacterial and eukaryotic cells are straight chained. Finally, although some archaeal membranes can be formed of bilayers like those found in bacteria and eukaryotes, other archaeal plasma membranes are lipid monolayers.
Proteins on the cell’s surface are important for a variety of functions, including cell-to-cell communication, and sensing environmental conditions and pathogenic virulence factors. Membrane proteins and phospholipids may have carbohydrates (sugars) associated with them and are called glycoproteins or glycolipids, respectively. These glycoprotein and glycolipid complexes extend out from the surface of the cell, allowing the cell to interact with the external environment (Figure \(10\)). Glycoproteins and glycolipids in the plasma membrane can vary considerably in chemical composition among archaea, bacteria, and eukaryotes, allowing scientists to use them to characterize unique species.
Plasma membranes from different cells types also contain unique phospholipids, which contain fatty acids. As described in Using Biochemistry to Identify Microorganisms, phospholipid-derived fatty acid analysis (PLFA) profiles can be used to identify unique types of cells based on differences in fatty acids. Archaea, bacteria, and eukaryotes each have a unique PFLA profile.
Membrane Transport Mechanisms
One of the most important functions of the plasma membrane is to control the transport of molecules into and out of the cell. Internal conditions must be maintained within a certain range despite any changes in the external environment. The transport of substances across the plasma membrane allows cells to do so.
Cells use various modes of transport across the plasma membrane. For example, molecules moving from a higher concentration to a lower concentration with the concentration gradient are transported by simple diffusion, also known as passive transport (Figure \(11\)). Some small molecules, like carbon dioxide, may cross the membrane bilayer directly by simple diffusion. However, charged molecules, as well as large molecules, need the help of carriers or channels in the membrane. These structures ferry molecules across the membrane, a process known as facilitated diffusion (Figure \(12\)).
Active transport occurs when cells move molecules across their membrane against concentration gradients (Figure \(13\)). A major difference between passive and active transport is that active transport requires adenosine triphosphate (ATP) or other forms of energy to move molecules “uphill.” Therefore, active transport structures are often called “pumps.”
Group translocation also transports substances into bacterial cells. In this case, as a molecule moves into a cell against its concentration gradient, it is chemically modified so that it does not require transport against an unfavorable concentration gradient. A common example of this is the bacterial phosphotransferase system, a series of carriers that phosphorylates (i.e., adds phosphate ions to) glucose or other sugars upon entry into cells. Since the phosphorylation of sugars is required during the early stages of sugar metabolism, the phosphotransferase system is considered to be an energy neutral system.
Photosynthetic Membrane Structures
Some prokaryotic cells, namely cyanobacteria and photosynthetic bacteria, have membrane structures that enable them to perform photosynthesis. These structures consist of an infolding of the plasma membrane that encloses photosynthetic pigments such as green chlorophylls and bacteriochlorophylls. In cyanobacteria, these membrane structures are called thylakoids; in photosynthetic bacteria, they are called chromatophores, lamellae, or chlorosomes.
Cell Wall
The primary function of the cell wall is to protect the cell from harsh conditions in the outside environment. When present, there are notable similarities and differences among the cell walls of archaea, bacteria, and eukaryotes.
The major component of bacterial cell walls is called peptidoglycan (or murein); it is only found in bacteria. Structurally, peptidoglycan resembles a layer of meshwork or fabric (Figure \(14\)). Each layer is composed of long chains of alternating molecules of N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM). The structure of the long chains has significant two-dimensional tensile strength due to the formation of peptide bridges that connect NAG and NAM within each peptidoglycan layer. In gram-negative bacteria, tetrapeptide chains extending from each NAM unit are directly cross-linked, whereas in gram-positive bacteria, these tetrapeptide chains are linked by pentaglycine cross-bridges. Peptidoglycan subunits are made inside of the bacterial cell and then exported and assembled in layers, giving the cell its shape.
Since peptidoglycan is unique to bacteria, many antibiotic drugs are designed to interfere with peptidoglycan synthesis, weakening the cell wall and making bacterial cells more susceptible to the effects of osmotic pressure (see Mechanisms of Antibacterial Drugs). In addition, certain cells of the human immune system are able “recognize” bacterial pathogens by detecting peptidoglycan on the surface of a bacterial cell; these cells then engulf and destroy the bacterial cell, using enzymes such as lysozyme, which breaks down and digests the peptidoglycan in their cell walls (see Pathogen Recognition and Phagocytosis).
The Gram staining protocol (see Staining Microscopic Specimens) is used to differentiate two common types of cell wall structures (Figure \(15\)). Gram-positive cells have a cell wall consisting of many layers of peptidoglycan totaling 30–100 nm in thickness. These peptidoglycan layers are commonly embedded with teichoic acids (TAs), carbohydrate chains that extend through and beyond the peptidoglycan layer.4 TA is thought to stabilize peptidoglycan by increasing its rigidity. TA also plays a role in the ability of pathogenic gram-positive bacteria such as Streptococcus to bind to certain proteins on the surface of host cells, enhancing their ability to cause infection. In addition to peptidoglycan and TAs, bacteria of the family Mycobacteriaceae have an external layer of waxy mycolic acids in their cell wall; as described in Staining Microscopic Specimens, these bacteria are referred to as acid-fast, since acid-fast stains must be used to penetrate the mycolic acid layer for purposes of microscopy (Figure \(16\)).
Gram-negative cells have a much thinner layer of peptidoglycan (no more than about 4 nm thick6) than gram-positive cells, and the overall structure of their cell envelope is more complex. In gram-negative cells, a gel-like matrix occupies the periplasmic space between the cell wall and the plasma membrane, and there is a second lipid bilayer called the outer membrane, which is external to the peptidoglycan layer (Figure \(15\)). This outer membrane is attached to the peptidoglycan by murein lipoprotein. The outer leaflet of the outer membrane contains the molecule lipopolysaccharide (LPS), which functions as an endotoxin in infections involving gram-negative bacteria, contributing to symptoms such as fever, hemorrhaging, and septic shock. Each LPS molecule is composed of Lipid A, a core polysaccharide, and an O side chain that is composed of sugar-like molecules that comprise the external face of the LPS (Figure \(17\)). The composition of the O side chain varies between different species and strains of bacteria. Parts of the O side chain called antigens can be detected using serological or immunological tests to identify specific pathogenic strains like Escherichia coli O157:H7, a deadly strain of bacteria that causes bloody diarrhea and kidney failure.
Archaeal cell wall structure differs from that of bacteria in several significant ways. First, archaeal cell walls do not contain peptidoglycan; instead, they contain a similar polymer called pseudopeptidoglycan (pseudomurein) in which NAM is replaced with a different subunit. Other archaea may have a layer of glycoproteins or polysaccharides that serves as the cell wall instead of pseudopeptidoglycan. Last, as is the case with some bacterial species, there are a few archaea that appear to lack cell walls entirely.
Glycocalyces and S-Layers
Although most prokaryotic cells have cell walls, some may have additional cell envelope structures exterior to the cell wall, such as glycocalyces and S-layers. A glycocalyx is a sugar coat, of which there are two important types: capsules and slime layers. A capsule is an organized layer located outside of the cell wall and usually composed of polysaccharides or proteins (Figure \(18\)). A slime layer is a less tightly organized layer that is only loosely attached to the cell wall and can be more easily washed off. Slime layers may be composed of polysaccharides, glycoproteins, or glycolipids.
Glycocalyces allows cells to adhere to surfaces, aiding in the formation of biofilms (colonies of microbes that form in layers on surfaces). In nature, most microbes live in mixed communities within biofilms, partly because the biofilm affords them some level of protection. Biofilms generally hold water like a sponge, preventing desiccation. They also protect cells from predation and hinder the action of antibiotics and disinfectants. All of these properties are advantageous to the microbes living in a biofilm, but they present challenges in a clinical setting, where the goal is often to eliminate microbes.
The ability to produce a capsule can contribute to a microbe’s pathogenicity (ability to cause disease) because the capsule can make it more difficult for phagocytic cells (such as white blood cells) to engulf and kill the microorganism. Streptococcus pneumoniae, for example, produces a capsule that is well known to aid in this bacterium’s pathogenicity. As explained in Staining Microscopic Specimens, capsules are difficult to stain for microscopy; negative staining techniques are typically used.
An S-layer is another type of cell envelope structure; it is composed of a mixture of structural proteins and glycoproteins. In bacteria, S-layers are found outside the cell wall, but in some archaea, the S-layer serves as the cell wall. The exact function of S-layers is not entirely understood, and they are difficult to study; but available evidence suggests that they may play a variety of functions in different prokaryotic cells, such as helping the cell withstand osmotic pressure and, for certain pathogens, interacting with the host immune system.
Clinical Focus: Part 3
After diagnosing Barbara with pneumonia, the PA writes her a prescription for amoxicillin, a commonly-prescribed type of penicillin derivative. More than a week later, despite taking the full course as directed, Barbara still feels weak and is not fully recovered, although she is still able to get through her daily activities. She returns to the health center for a follow-up visit.
Many types of bacteria, fungi, and viruses can cause pneumonia. Amoxicillin targets the peptidoglycan of bacterial cell walls. Since the amoxicillin has not resolved Barbara’s symptoms, the PA concludes that the causative agent probably lacks peptidoglycan, meaning that the pathogen could be a virus, a fungus, or a bacterium that lacks peptidoglycan. Another possibility is that the pathogen is a bacterium containing peptidoglycan but has developed resistance to amoxicillin.
Exercise \(3\)
1. How can the PA definitively identify the cause of Barbara’s pneumonia?
2. What form of treatment should the PA prescribe, given that the amoxicillin was ineffective?
Filamentous Appendages
Many bacterial cells have protein appendages embedded within their cell envelopes that extend outward, allowing interaction with the environment. These appendages can attach to other surfaces, transfer DNA, or provide movement. Filamentous appendages include fimbriae, pili, and flagella.
Fimbriae and Pili
Fimbriae and pili are structurally similar and, because differentiation between the two is problematic, these terms are often used interchangeably.7 8 The term fimbriae commonly refers to short bristle-like proteins projecting from the cell surface by the hundreds. Fimbriae enable a cell to attach to surfaces and to other cells. For pathogenic bacteria, adherence to host cells is important for colonization, infectivity, and virulence. Adherence to surfaces is also important in biofilm formation.
The term pili (singular: pilus) commonly refers to longer, less numerous protein appendages that aid in attachment to surfaces (Figure \(19\)). A specific type of pilus, called the F pilus or sex pilus, is important in the transfer of DNA between bacterial cells, which occurs between members of the same generation when two cells physically transfer or exchange parts of their respective genomes (see How Asexual Prokaryotes Achieve Genetic Diversity).
Group A Strep
Before the structure and function of the various components of the bacterial cell envelope were well understood, scientists were already using cell envelope characteristics to classify bacteria. In 1933, Rebecca Lancefield proposed a method for serotyping various β-hemolytic strains of Streptococcus species using an agglutination assay, a technique using the clumping of bacteria to detect specific cell-surface antigens. In doing so, Lancefield discovered that one group of S. pyogenes, found in Group A, was associated with a variety of human diseases. She determined that various strains of Group A strep could be distinguished from each other based on variations in specific cell surface proteins that she named M proteins.
Today, more than 80 different strains of Group A strep have been identified based on M proteins. Various strains of Group A strep are associated with a wide variety of human infections, including streptococcal pharyngitis (strep throat), impetigo, toxic shock syndrome, scarlet fever, rheumatic fever, and necrotizing fasciitis. The M protein is an important virulence factor for Group A strep, helping these strains evade the immune system. Changes in M proteins appear to alter the infectivity of a particular strain of Group A strep.
Flagella
Flagella are structures used by cells to move in aqueous environments. Bacterial flagella act like propellers. They are stiff spiral filaments composed of flagellin protein subunits that extend outward from the cell and spin in solution. The basal body is the motor for the flagellum and is embedded in the plasma membrane (Figure \(20\)). A hook region connects the basal body to the filament. Gram-positive and gram-negative bacteria have different basal body configurations due to differences in cell wall structure.
Different types of motile bacteria exhibit different arrangements of flagella (Figure \(21\)). A bacterium with a singular flagellum, typically located at one end of the cell (polar), is said to have a monotrichous flagellum. An example of a monotrichously flagellated bacterial pathogen is Vibrio cholerae, the gram-negative bacterium that causes cholera. Cells with amphitrichous flagella have a flagellum or tufts of flagella at each end. An example is Spirillum minor, the cause of spirillary (Asian) rat-bite fever or sodoku. Cells with lophotrichous flagella have a tuft at one end of the cell. The gram-negative bacillus Pseudomonas aeruginosa, an opportunistic pathogen known for causing many infections, including “swimmer’s ear” and burn wound infections, has lophotrichous flagella. Flagella that cover the entire surface of a bacterial cell are called peritrichous flagella. The gram-negative bacterium E. coli shows a peritrichous arrangement of flagella.
Directional movement depends on the configuration of the flagella. Bacteria can move in response to a variety of environmental signals, including light (phototaxis), magnetic fields (magnetotaxis) using magnetosomes, and, most commonly, chemical gradients (chemotaxis). Purposeful movement toward a chemical attractant, like a food source, or away from a repellent, like a poisonous chemical, is achieved by increasing the length of runs and decreasing the length of tumbles. When running, flagella rotate in a counterclockwise direction, allowing the bacterial cell to move forward. In a peritrichous bacterium, the flagella are all bundled together in a very streamlined way (Figure \(22\)), allowing for efficient movement. When tumbling, flagella are splayed out while rotating in a clockwise direction, creating a looping motion and preventing meaningful forward movement but reorienting the cell toward the direction of the attractant. When an attractant exists, runs and tumbles still occur; however, the length of runs is longer, while the length of the tumbles is reduced, allowing overall movement toward the higher concentration of the attractant. When no chemical gradient exists, the lengths of runs and tumbles are more equal, and overall movement is more random (Figure \(23\)).
Exercise \(4\)
1. What is the peptidoglycan layer and how does it differ between gram-positive and gram-negative bacteria?
2. Compare and contrast monotrichous, amphitrichous, lophotrichous, and peritrichous flagella.
Summary
• Prokaryotic cells differ from eukaryotic cells in that their genetic material is contained in a nucleoid rather than a membrane-bound nucleus. In addition, prokaryotic cells generally lack membrane-bound organelles.
• Prokaryotic cells of the same species typically share a similar cell morphology and cellular arrangement.
• Most prokaryotic cells have a cell wall that helps the organism maintain cellular morphology and protects it against changes in osmotic pressure.
• Outside of the nucleoid, prokaryotic cells may contain extrachromosomal DNA in plasmids.
• Prokaryotic ribosomes that are found in the cytoplasm have a size of 70S.
• Some prokaryotic cells have inclusions that store nutrients or chemicals for other uses.
• Some prokaryotic cells are able to form endospores through sporulation to survive in a dormant state when conditions are unfavorable. Endospores can germinate, transforming back into vegetative cells when conditions improve.
• In prokaryotic cells, the cell envelope includes a plasma membrane and usually a cell wall.
• Bacterial membranes are composed of phospholipids with integral or peripheral proteins. The fatty acid components of these phospholipids are ester-linked and are often used to identify specific types of bacteria. The proteins serve a variety of functions, including transport, cell-to-cell communication, and sensing environmental conditions. Archaeal membranes are distinct in that they are composed of fatty acids that are ether-linked to phospholipids.
• Some molecules can move across the bacterial membrane by simple diffusion, but most large molecules must be actively transported through membrane structures using cellular energy.
• Prokaryotic cell walls may be composed of peptidoglycan (bacteria) or pseudopeptidoglycan (archaea).
• Gram-positive bacterial cells are characterized by a thick peptidoglycan layer, whereas gram-negative bacterial cells are characterized by a thin peptidoglycan layer surrounded by an outer membrane.
• Some prokaryotic cells produce glycocalyx coatings, such as capsules and slime layers, that aid in attachment to surfaces and/or evasion of the host immune system.
• Some prokaryotic cells have fimbriae or pili, filamentous appendages that aid in attachment to surfaces. Pili are also used in the transfer of genetic material between cells.
• Some prokaryotic cells use one or more flagella to move through water. Peritrichous bacteria, which have numerous flagella, use runs and tumbles to move purposefully in the direction of a chemical attractant.
Footnotes
1. 1 Y.-H.M. Chan, W.F. Marshall. “Scaling Properties of Cell and Organelle Size.” Organogenesis 6 no. 2 (2010):88–96.
2. 2 F. Rothfuss, M Bender, R Conrad. “Survival and Activity of Bacteria in a Deep, Aged Lake Sediment (Lake Constance).” Microbial Ecology 33 no. 1 (1997):69–77.
3. 3 R. Sinclair et al. “Persistence of Category A Select Agents in the Environment.” Applied and Environmental Microbiology 74 no. 3 (2008):555–563.
4. 4 T.J. Silhavy, D. Kahne, S. Walker. “The Bacterial Cell Envelope.” Cold Spring Harbor Perspectives in Biology 2 no. 5 (2010):a000414.
5. 5 B. Zuber et al. “Granular Layer in the Periplasmic Space of Gram-Positive Bacteria and Fine Structures of Enterococcus gallinarum and Streptococcus gordonii Septa Revealed by Cryo-Electron Microscopy of Vitreous Sections.” Journal of Bacteriology 188 no. 18 (2006):6652–6660
6. 6 L. Gana, S. Chena, G.J. Jensena. “Molecular Organization of Gram-Negative Peptidoglycan.” Proceedings of the National Academy of Sciences of the United States of America 105 no. 48 (2008):18953–18957.
7. 7 J.A. Garnetta et al. “Structural Insights Into the Biogenesis and Biofilm Formation by the Escherichia coli Common Pilus.” Proceedings of the National Academy of Sciences of the United States of America 109 no. 10 (2012):3950–3955.
8. 8 T. Proft, E.N. Baker. “Pili in Gram-Negative and Gram-Positive Bacteria—Structure, Assembly and Their Role in Disease.” Cellular and Molecular Life Sciences 66 (2009):613. | textbooks/bio/Microbiology/Microbiology_(OpenStax)/03%3A_The_Cell/3.03%3A_Unique_Characteristics_of_Prokaryotic_Cells.txt |
Learning Objectives
• Explain the distinguishing characteristics of eukaryotic cells
• Describe internal and external structures of prokaryotic cells in terms of their physical structure, chemical structure, and function
• Identify and describe structures and organelles unique to eukaryotic cells
• Compare and contrast similar structures found in prokaryotic and eukaryotic cells
Eukaryotic organisms include protozoans, algae, fungi, plants, and animals. Some eukaryotic cells are independent, single-celled microorganisms, whereas others are part of multicellular organisms. The cells of eukaryotic organisms have several distinguishing characteristics. Above all, eukaryotic cells are defined by the presence of a nucleus surrounded by a complex nuclear membrane. Also, eukaryotic cells are characterized by the presence of membrane-bound organelles in the cytoplasm. Organelles such as mitochondria, the endoplasmic reticulum (ER), Golgi apparatus, lysosomes, and peroxisomes are held in place by the cytoskeleton, an internal network that supports transport of intracellular components and helps maintain cell shape (Figure \(1\)). The genome of eukaryotic cells is packaged in multiple, rod-shaped chromosomes as opposed to the single, circular-shaped chromosome that characterizes most prokaryotic cells. Table \(1\) compares the characteristics of eukaryotic cell structures with those of bacteria and archaea.
Table \(1\): Summary of Cell Structures.
Cell Structure Prokaryotes Eukaryotes
Bacteria Archaea
Size ~0.5–1 μM ~0.5–1 μM ~5–20 μM
Surface area-to-volume ratio High High Low
Nucleus No No Yes
Genome characteristics
• Single chromosome
• Circular
• Haploid
• Lacks histones
• Single chromosome
• Circular
• Haploid
• Contains histones
• Multiple chromosomes
• Linear
• Haploid or diploid
• Contains histones
Cell division Binary fission Binary fission Mitosis, meiosis
Membrane lipid composition
• Ester-linked
• Straight-chain fatty acids
• Bilayer
• Ether-linked
• Branched isoprenoids
• Bilayer or monolayer
• Ester-linked
• Straight-chain fatty acids
• Sterols
• Bilayer
Cell wall composition
• Peptidoglycan, or
• None
• Pseudopeptidoglycan, or
• Glycopeptide, or
• Polysaccharide, or
• Protein (S-layer), or
• None
• Cellulose (plants, some algae)
• Chitin (molluscs, insects, crustaceans, and fungi)
• Silica (some algae)
• Most others lack cell walls
Motility structures Rigid spiral flagella composed of flagellin Rigid spiral flagella composed of archaeal flagellins Flexible flagella and cilia composed of microtubules
Membrane-bound organelles No No Yes
Endomembrane system No No Yes (ER, Golgi, lysosomes)
Ribosomes 70S 70S
• 80S in cytoplasm and rough ER
• 70S in mitochondria, chloroplasts
Cell Morphologies
Eukaryotic cells display a wide variety of different cell morphologies. Possible shapes include spheroid, ovoid, cuboidal, cylindrical, flat, lenticular, fusiform, discoidal, crescent, ring stellate, and polygonal (Figure \(2\)). Some eukaryotic cells are irregular in shape, and some are capable of changing shape. The shape of a particular type of eukaryotic cell may be influenced by factors such as its primary function, the organization of its cytoskeleton, the viscosity of its cytoplasm, the rigidity of its cell membrane or cell wall (if it has one), and the physical pressure exerted on it by the surrounding environment and/or adjoining cells.
Exercise \(1\)
Identify two differences between eukaryotic and prokaryotic cell.
Nucleus
Unlike prokaryotic cells, in which DNA is loosely contained in the nucleoid region, eukaryotic cells possess a nucleus, which is surrounded by a complex nuclear membrane that houses the DNA genome (Figure \(3\)). By containing the cell’s DNA, the nucleus ultimately controls all activities of the cell and also serves an essential role in reproduction and heredity. Eukaryotic cells typically have their DNA organized into multiple linear chromosomes. The DNA within the nucleus is highly organized and condensed to fit inside the nucleus, which is accomplished by wrapping the DNA around proteins called histones.
Although most eukaryotic cells have only one nucleus, exceptions exist. For example, protozoans of the genus Paramecium typically have two complete nuclei: a small nucleus that is used for reproduction (micronucleus) and a large nucleus that directs cellular metabolism (macronucleus). Additionally, some fungi transiently form cells with two nuclei, called heterokaryotic cells, during sexual reproduction. Cells whose nuclei divide, but whose cytoplasm does not, are called coenocytes.
The nucleus is bound by a complex nuclear membrane, often called the nuclear envelope, that consists of two distinct lipid bilayers that are contiguous with each other (Figure \(4\)). Despite these connections between the inner and outer membranes, each membrane contains unique lipids and proteins on its inner and outer surfaces. The nuclear envelope contains nuclear pores, which are large, rosette-shaped protein complexes that control the movement of materials into and out of the nucleus. The overall shape of the nucleus is determined by the nuclear lamina, a meshwork of intermediate filaments found just inside the nuclear envelope membranes. Outside the nucleus, additional intermediate filaments form a looser mesh and serve to anchor the nucleus in position within the cell.
Nucleolus
The nucleolus is a dense region within the nucleus where ribosomal RNA (rRNA) biosynthesis occurs. In addition, the nucleolus is also the site where assembly of ribosomes begins. Preribosomal complexes are assembled from rRNA and proteins in the nucleolus; they are then transported out to the cytoplasm, where ribosome assembly is completed (Figure \(5\)).
Ribosomes
Ribosomes found in eukaryotic organelles such as mitochondria or chloroplasts have 70S ribosomes—the same size as prokaryotic ribosomes. However, nonorganelle-associated ribosomes in eukaryotic cells are 80S ribosomes, composed of a 40S small subunit and a 60S large subunit. In terms of size and composition, this makes them distinct from the ribosomes of prokaryotic cells.
The two types of nonorganelle-associated eukaryotic ribosomes are defined by their location in the cell: free ribosomesand membrane-bound ribosomes. Free ribosomes are found in the cytoplasm and serve to synthesize water-soluble proteins; membrane-bound ribosomes are found attached to the rough endoplasmic reticulum and make proteins for insertion into the cell membrane or proteins destined for export from the cell.
The differences between eukaryotic and prokaryotic ribosomes are clinically relevant because certain antibiotic drugs are designed to target one or the other. For example, cycloheximide targets eukaryotic action, whereas chloramphenicol targets prokaryotic ribosomes.1 Since human cells are eukaryotic, they generally are not harmed by antibiotics that destroy the prokaryotic ribosomes in bacteria. However, sometimes negative side effects may occur because mitochondria in human cells contain prokaryotic ribosomes.
Endomembrane System
The endomembrane system, unique to eukaryotic cells, is a series of membranous tubules, sacs, and flattened disks that synthesize many cell components and move materials around within the cell (Figure \(6\)). Because of their larger cell size, eukaryotic cells require this system to transport materials that cannot be dispersed by diffusion alone. The endomembrane system comprises several organelles and connections between them, including the endoplasmic reticulum, Golgi apparatus, lysosomes, and vesicles.
Endoplasmic Reticulum
The endoplasmic reticulum (ER) is an interconnected array of tubules and cisternae (flattened sacs) with a single lipid bilayer (Figure \(7\)). The spaces inside of the cisternae are called lumen of the ER. There are two types of ER, rough endoplasmic reticulum (RER) and smooth endoplasmic reticulum (SER). These two different types of ER are sites for the synthesis of distinctly different types of molecules. RER is studded with ribosomes bound on the cytoplasmic side of the membrane. These ribosomes make proteins destined for the plasma membrane (Figure \(\PageIndex{}\)). Following synthesis, these proteins are inserted into the membrane of the RER. Small sacs of the RER containing these newly synthesized proteins then bud off as transport vesicles and move either to the Golgi apparatus for further processing, directly to the plasma membrane, to the membrane of another organelle, or out of the cell. Transport vesicles are single-lipid, bilayer, membranous spheres with hollow interiors that carry molecules. SER does not have ribosomes and, therefore, appears “smooth.” It is involved in biosynthesis of lipids, carbohydrate metabolism, and detoxification of toxic compounds within the cell.
Golgi Apparatus
The Golgi apparatus was discovered within the endomembrane system in 1898 by Italian scientist Camillo Golgi (1843–1926), who developed a novel staining technique that showed stacked membrane structures within the cells of Plasmodium, the causative agent of malaria. The Golgi apparatus is composed of a series of membranous disks called dictyosomes, each having a single lipid bilayer, that are stacked together (Figure \(8\)).
Enzymes in the Golgi apparatus modify lipids and proteins transported from the ER to the Golgi, often adding carbohydrate components to them, producing glycolipids, glycoproteins, or proteoglycans. Glycolipids and glycoproteins are often inserted into the plasma membrane and are important for signal recognition by other cells or infectious particles. Different types of cells can be distinguished from one another by the structure and arrangement of the glycolipids and glycoproteins contained in their plasma membranes. These glycolipids and glycoproteins commonly also serve as cell surface receptors.
Transport vesicles leaving the ER fuse with a Golgi apparatus on its receiving, or cis, face. The proteins are processed within the Golgi apparatus, and then additional transport vesicles containing the modified proteins and lipids pinch off from the Golgi apparatus on its outgoing, or trans, face. These outgoing vesicles move to and fuse with the plasma membrane or the membrane of other organelles.
Exocytosis is the process by which secretory vesicles (spherical membranous sacs) release their contents to the cell’s exterior (Figure \(8\)). All cells have constitutive secretory pathways in which secretory vesicles transport soluble proteins that are released from the cell continually (constitutively). Certain specialized cells also have regulated secretory pathways, which are used to store soluble proteins in secretory vesicles. Regulated secretion involves substances that are only released in response to certain events or signals. For example, certain cells of the human immune system (e.g., mast cells) secrete histamine in response to the presence of foreign objects or pathogens in the body. Histamine is a compound that triggers various mechanisms used by the immune system to eliminate pathogens.
Lysosomes
In the 1960s, Belgian scientist Christian de Duve (1917–2013) discovered lysosomes, membrane-bound organelles of the endomembrane system that contain digestive enzymes. Certain types of eukaryotic cells use lysosomes to break down various particles, such as food, damaged organelles or cellular debris, microorganisms, or immune complexes. Compartmentalization of the digestive enzymes within the lysosome allows the cell to efficiently digest matter without harming the cytoplasmic components of the cell.
Exercise \(2\)
Name the components of the endomembrane system and describe the function of each component.
Peroxisomes
Christian de Duve is also credited with the discovery of peroxisomes, membrane-bound organelles that are not part of the endomembrane system (Figure \(9\)). Peroxisomes form independently in the cytoplasm from the synthesis of peroxin proteins by free ribosomes and the incorporation of these peroxin proteins into existing peroxisomes. Growing peroxisomes then divide by a process similar to binary fission.
Peroxisomes were first named for their ability to produce hydrogen peroxide, a highly reactive molecule that helps to break down molecules such as uric acid, amino acids, and fatty acids. Peroxisomes also possess the enzyme catalase, which can degrade hydrogen peroxide. Along with the SER, peroxisomes also play a role in lipid biosynthesis. Like lysosomes, the compartmentalization of these degradative molecules within an organelle helps protect the cytoplasmic contents from unwanted damage.
The peroxisomes of certain organisms are specialized to meet their particular functional needs. For example, glyoxysomes are modified peroxisomes of yeasts and plant cells that perform several metabolic functions, including the production of sugar molecules. Similarly, glycosomes are modified peroxisomes made by certain trypanosomes, the pathogenic protozoans that cause Chagas disease and African sleeping sickness.
Cytoskeleton
Eukaryotic cells have an internal cytoskeleton made of microfilaments, intermediate filaments, and microtubules. This matrix of fibers and tubes provides structural support as well as a network over which materials can be transported within the cell and on which organelles can be anchored (Figure \(10\)). For example, the process of exocytosis involves the movement of a vesicle via the cytoskeletal network to the plasma membrane, where it can release its contents.
Microfilaments are composed of two intertwined strands of actin, each composed of actin monomers forming filamentous cables 6 nm in diameter2 (Figure \(11\)). The actin filaments work together with motor proteins, like myosin, to effect muscle contraction in animals or the amoeboid movement of some eukaryotic microbes. In ameboid organisms, actin can be found in two forms: a stiffer, polymerized, gel form and a more fluid, unpolymerized soluble form. Actin in the gel form creates stability in the ectoplasm, the gel-like area of cytoplasm just inside the plasma membrane of ameboid protozoans.
Temporary extensions of the cytoplasmic membrane called pseudopodia (meaning “false feet”) are produced through the forward flow of soluble actin filaments into the pseudopodia, followed by the gel-sol cycling of the actin filaments, resulting in cell motility. Once the cytoplasm extends outward, forming a pseudopodium, the remaining cytoplasm flows up to join the leading edge, thereby creating forward locomotion. Beyond amoeboid movement, microfilaments are also involved in a variety of other processes in eukaryotic cells, including cytoplasmic streaming (the movement or circulation of cytoplasm within the cell), cleavage furrow formation during cell division, and muscle movement in animals (Figure \(11\)). These functions are the result of the dynamic nature of microfilaments, which can polymerize and depolymerize relatively easily in response to cellular signals, and their interactions with molecular motors in different types of eukaryotic cells.
Intermediate filaments (Figure \(12\)) are a diverse group of cytoskeletal filaments that act as cables within the cell. They are termed “intermediate” because their 10-nm diameter is thicker than that of actin but thinner than that of microtubules.3 They are composed of several strands of polymerized subunits that, in turn, are made up of a wide variety of monomers. Intermediate filaments tend to be more permanent in the cell and maintain the position of the nucleus. They also form the nuclear lamina (lining or layer) just inside the nuclear envelope. Additionally, intermediate filaments play a role in anchoring cells together in animal tissues. The intermediate filament protein desmin is found in desmosomes, the protein structures that join muscle cells together and help them resist external physical forces. The intermediate filament protein keratin is a structural protein found in hair, skin, and nails.
Microtubules (Figure \(13\)) are a third type of cytoskeletal fiber composed of tubulin dimers (α tubulin and β tubulin). These form hollow tubes 23 nm in diameter that are used as girders within the cytoskeleton.4 Like microfilaments, microtubules are dynamic and have the ability to rapidly assemble and disassemble. Microtubules also work with motor proteins (such as dynein and kinesin) to move organelles and vesicles around within the cytoplasm. Additionally, microtubules are the main components of eukaryotic flagella and cilia, composing both the filament and the basal body components (Figure \(20\)).
In addition, microtubules are involved in cell division, forming the mitotic spindle that serves to separate chromosomes during mitosis and meiosis. The mitotic spindle is produced by two centrosomes, which are essentially microtubule-organizing centers, at opposite ends of the cell. Each centrosome is composed of a pair of centrioles positioned at right angles to each other, and each centriole is an array of nine parallel microtubules arranged in triplets (Figure \(14\)).
Exercise \(3\)
Compare and contrast the three types of cytoskeletal structures described in this section.
Mitochondria
The large, complex organelles in which aerobic cellular respiration occurs in eukaryotic cells are called mitochondria(Figure \(15\)). The term “mitochondrion” was first coined by German microbiologist Carl Benda in 1898 and was later connected with the process of respiration by Otto Warburg in 1913. Scientists during the 1960s discovered that mitochondria have their own genome and 70S ribosomes. The mitochondrial genome was found to be bacterial, when it was sequenced in 1976. These findings ultimately supported the endosymbiotic theory proposed by Lynn Margulis, which states that mitochondria originally arose through an endosymbiotic event in which a bacterium capable of aerobic cellular respiration was taken up by phagocytosis into a host cell and remained as a viable intracellular component.
Each mitochondrion has two lipid membranes. The outer membrane is a remnant of the original host cell’s membrane structures. The inner membrane was derived from the bacterial plasma membrane. The electron transport chain for aerobic respiration uses integral proteins embedded in the inner membrane. The mitochondrial matrix, corresponding to the location of the original bacterium’s cytoplasm, is the current location of many metabolic enzymes. It also contains mitochondrial DNA and 70S ribosomes. Invaginations of the inner membrane, called cristae, evolved to increase surface area for the location of biochemical reactions. The folding patterns of the cristae differ among various types of eukaryotic cells and are used to distinguish different eukaryotic organisms from each other.
Chloroplasts
Plant cells and algal cells contain chloroplasts, the organelles in which photosynthesis occurs (Figure \(16\)). All chloroplasts have at least three membrane systems: the outer membrane, the inner membrane, and the thylakoid membrane system. Inside the outer and inner membranes is the chloroplast stroma, a gel-like fluid that makes up much of a chloroplast’s volume, and in which the thylakoid system floats. The thylakoid system is a highly dynamic collection of folded membrane sacs. It is where the green photosynthetic pigment chlorophyll is found and the light reactions of photosynthesis occur. In most plant chloroplasts, the thylakoids are arranged in stacks called grana (singular: granum), whereas in some algal chloroplasts, the thylakoids are free floating.
Other organelles similar to mitochondria have arisen in other types of eukaryotes, but their roles differ. Hydrogenosomes are found in some anaerobic eukaryotes and serve as the location of anaerobic hydrogen production. Hydrogenosomes typically lack their own DNA and ribosomes. Kinetoplasts are a variation of the mitochondria found in some eukaryotic pathogens. In these organisms, each cell has a single, long, branched mitochondrion in which kinetoplast DNA, organized as multiple circular pieces of DNA, is found concentrated at one pole of the cell.
Mitochondria-Related Organelles in Protozoan Parasites
Many protozoans, including several protozoan parasites that cause infections in humans, can be identified by their unusual appearance. Distinguishing features may include complex cell morphologies, the presence of unique organelles, or the absence of common organelles. The protozoan parasites Giardia lamblia and Trichomonas vaginalis are two examples.
G. lamblia, a frequent cause of diarrhea in humans and many other animals, is an anaerobic parasite that possesses two nuclei and several flagella. Its Golgi apparatus and endoplasmic reticulum are greatly reduced, and it lacks mitochondria completely. However, it does have organelles known as mitosomes, double-membrane-bound organelles that appear to be severely reduced mitochondria. This has led scientists to believe that G. lamblia’s ancestors once possessed mitochondria that evolved to become mitosomes. T. vaginalis, which causes the sexually transmitted infection vaginitis, is another protozoan parasite that lacks conventional mitochondria. Instead, it possesses hydrogenosomes, mitochondrial-related, double-membrane-bound organelles that produce molecular hydrogen used in cellular metabolism. Scientists believe that hydrogenosomes, like mitosomes, also evolved from mitochondria.5
Plasma Membrane
The plasma membrane of eukaryotic cells is similar in structure to the prokaryotic plasma membrane in that it is composed mainly of phospholipids forming a bilayer with embedded peripheral and integral proteins (Figure \(17\)). These membrane components move within the plane of the membrane according to the fluid mosaic model. However, unlike the prokaryotic membrane, eukaryotic membranes contain sterols, including cholesterol, that alter membrane fluidity. Additionally, many eukaryotic cells contain some specialized lipids, including sphingolipids, which are thought to play a role in maintaining membrane stability as well as being involved in signal transduction pathways and cell-to-cell communication.
Membrane Transport Mechanisms
The processes of simple diffusion, facilitated diffusion, and active transport are used in both eukaryotic and prokaryotic cells. However, eukaryotic cells also have the unique ability to perform various types of endocytosis, the uptake of matter through plasma membrane invagination and vacuole/vesicle formation (Figure \(18\)). A type of endocytosis involving the engulfment of large particles through membrane invagination is called phagocytosis, which means “cell eating.” In phagocytosis, particles (or other cells) are enclosed in a pocket within the membrane, which then pinches off from the membrane to form a vacuole that completely surrounds the particle. Another type of endocytosis is called pinocytosis, which means “cell drinking.” In pinocytosis, small, dissolved materials and liquids are taken into the cell through small vesicles. Saprophytic fungi, for example, obtain their nutrients from dead and decaying matter largely through pinocytosis.
Receptor-mediated endocytosis is a type of endocytosis that is initiated by specific molecules called ligands when they bind to cell surface receptors on the membrane. Receptor-mediated endocytosis is the mechanism that peptide and amine-derived hormones use to enter cells and is also used by various viruses and bacteria for entry into host cells.
The process by which secretory vesicles release their contents to the cell’s exterior is called exocytosis. Vesicles move toward the plasma membrane and then meld with the membrane, ejecting their contents out of the cell. Exocytosis is used by cells to remove waste products and may also be used to release chemical signals that can be taken up by other cells.
Cell Wall
In addition to a plasma membrane, some eukaryotic cells have a cell wall. Cells of fungi, algae, plants, and even some protists have cell walls. Depending upon the type of eukaryotic cell, cell walls can be made of a wide range of materials, including cellulose (fungi and plants); biogenic silica, calcium carbonate, agar, and carrageenan (protists and algae); or chitin (fungi). In general, all cell walls provide structural stability for the cell and protection from environmental stresses such as desiccation, changes in osmotic pressure, and traumatic injury.6
Extracellular Matrix
Cells of animals and some protozoans do not have cell walls to help maintain shape and provide structural stability. Instead, these types of eukaryotic cells produce an extracellular matrix for this purpose. They secrete a sticky mass of carbohydrates and proteins into the spaces between adjacent cells (Figure \(19\)). Some protein components assemble into a basement membrane to which the remaining extracellular matrix components adhere. Proteoglycans typically form the bulky mass of the extracellular matrix while fibrous proteins, like collagen, provide strength. Both proteoglycans and collagen are attached to fibronectin proteins, which, in turn, are attached to integrin proteins. These integrin proteins interact with transmembrane proteins in the plasma membranes of eukaryotic cells that lack cell walls.
In animal cells, the extracellular matrix allows cells within tissues to withstand external stresses and transmits signals from the outside of the cell to the inside. The amount of extracellular matrix is quite extensive in various types of connective tissues, and variations in the extracellular matrix can give different types of tissues their distinct properties. In addition, a host cell’s extracellular matrix is often the site where microbial pathogens attach themselves to establish infection. For example, Streptococcus pyogenes, the bacterium that causes strep throat and various other infections, binds to fibronectin in the extracellular matrix of the cells lining the oropharynx (upper region of the throat).
Flagella and Cilia
Some eukaryotic cells use flagella for locomotion; however, eukaryotic flagella are structurally distinct from those found in prokaryotic cells. Whereas the prokaryotic flagellum is a stiff, rotating structure, a eukaryotic flagellum is more like a flexible whip composed of nine parallel pairs of microtubules surrounding a central pair of microtubules. This arrangement is referred to as a 9+2 array (Figure \(20\)). The parallel microtubules use dynein motor proteins to move relative to each other, causing the flagellum to bend.
Cilia (singular: cilium) are a similar external structure found in some eukaryotic cells. Unique to eukaryotes, cilia are shorter than flagella and often cover the entire surface of a cell; however, they are structurally similar to flagella (a 9+2 array of microtubules) and use the same mechanism for movement. A structure called a basal body is found at the base of each cilium and flagellum. The basal body, which attaches the cilium or flagellum to the cell, is composed of an array of triplet microtubules similar to that of a centriole but embedded in the plasma membrane. Because of their shorter length, cilia use a rapid, flexible, waving motion. In addition to motility, cilia may have other functions such as sweeping particles past or into cells. For example, ciliated protozoans use the sweeping of cilia to move food particles into their mouthparts, and ciliated cells in the mammalian respiratory tract beat in synchrony to sweep mucus and debris up and out of the lungs (Figure \(20\)).
Exercise \(4\)
1. Explain how the cellular envelope of eukaryotic cells compares to that of prokaryotic cells.
2. Explain the difference between eukaryotic and prokaryotic flagella.
Clinical Focus: Resolution
Since amoxicillin has not resolved Barbara’s case of pneumonia, the PA prescribes another antibiotic, azithromycin, which targets bacterial ribosomes rather than peptidoglycan. After taking the azithromycin as directed, Barbara’s symptoms resolve and she finally begins to feel like herself again. Presuming no drug resistance to amoxicillin was involved, and given the effectiveness of azithromycin, the causative agent of Barbara’s pneumonia is most likely Mycoplasma pneumoniae. Even though this bacterium is a prokaryotic cell, it is not inhibited by amoxicillin because it does not have a cell wall and, therefore, does not make peptidoglycan.
Key Concepts and Summary
• Eukaryotic cells are defined by the presence of a nucleus containing the DNA genome and bound by a nuclear membrane (or nuclear envelope) composed of two lipid bilayers that regulate transport of materials into and out of the nucleus through nuclear pores.
• Eukaryotic cell morphologies vary greatly and may be maintained by various structures, including the cytoskeleton, the cell membrane, and/or the cell wall.
• The nucleolus, located in the nucleus of eukaryotic cells, is the site of ribosomal synthesis and the first stages of ribosome assembly.
• Eukaryotic cells contain 80S ribosomes in the rough endoplasmic reticulum (membrane bound-ribosomes) and cytoplasm (free ribosomes). They contain 70s ribosomes in mitochondria and chloroplasts.
• Eukaryotic cells have evolved an endomembrane system, containing membrane-bound organelles involved in transport. These include vesicles, the endoplasmic reticulum, and the Golgi apparatus.
• The smooth endoplasmic reticulum plays a role in lipid biosynthesis, carbohydrate metabolism, and detoxification of toxic compounds. The rough endoplasmic reticulum contains membrane-bound 80S ribosomes that synthesize proteins destined for the cell membrane
• The Golgi apparatus processes proteins and lipids, typically through the addition of sugar molecules, producing glycoproteins or glycolipids, components of the plasma membrane that are used in cell-to-cell communication.
• Lysosomes contain digestive enzymes that break down small particles ingested by endocytosis, large particles or cells ingested by phagocytosis, and damaged intracellular components.
• The cytoskeleton, composed of microfilaments, intermediate filaments, and microtubules, provides structural support in eukaryotic cells and serves as a network for transport of intracellular materials.
• Centrosomes are microtubule-organizing centers important in the formation of the mitotic spindle in mitosis.
• Mitochondria are the site of cellular respiration. They have two membranes: an outer membrane and an inner membrane with cristae. The mitochondrial matrix, within the inner membrane, contains the mitochondrial DNA, 70S ribosomes, and metabolic enzymes.
• The plasma membrane of eukaryotic cells is structurally similar to that found in prokaryotic cells, and membrane components move according to the fluid mosaic model. However, eukaryotic membranes contain sterols, which alter membrane fluidity, as well as glycoproteins and glycolipids, which help the cell recognize other cells and infectious particles.
• In addition to active transport and passive transport, eukaryotic cell membranes can take material into the cell via endocytosis, or expel matter from the cell via exocytosis.
• Cells of fungi, algae, plants, and some protists have a cell wall, whereas cells of animals and some protozoans have a sticky extracellular matrix that provides structural support and mediates cellular signaling.
• Eukaryotic flagella are structurally distinct from prokaryotic flagella but serve a similar purpose (locomotion). Ciliaare structurally similar to eukaryotic flagella, but shorter; they may be used for locomotion, feeding, or movement of extracellular particles.
Footnotes
1. 1 A.E. Barnhill, M.T. Brewer, S.A. Carlson. “Adverse Effects of Antimicrobials via Predictable or Idiosyncratic Inhibition of Host Mitochondrial Components.” Antimicrobial Agents and Chemotherapy 56 no. 8 (2012):4046–4051.
2. 2 Fuchs E, Cleveland DW. “A Structural Scaffolding of Intermediate Filaments in Health and Disease.” Science 279 no. 5350 (1998):514–519.
3. 3 E. Fuchs, D.W. Cleveland. “A Structural Scaffolding of Intermediate Filaments in Health and Disease.” Science 279 no. 5350 (1998):514–519.
4. 4 E. Fuchs, D.W. Cleveland. “A Structural Scaffolding of Intermediate Filaments in Health and Disease.” Science 279 no. 5350 (1998):514–519.
5. 5 N. Yarlett, J.H.P. Hackstein. “Hydrogenosomes: One Organelle, Multiple Origins.” BioScience 55 no. 8 (2005):657–658.
6. 6 M. Dudzick. “Protists.” OpenStax CNX. November 27, 2013. http://cnx.org/contents/f7048bb6-e46...ef291cf7049c@1 | textbooks/bio/Microbiology/Microbiology_(OpenStax)/03%3A_The_Cell/3.04%3A_Unique_Characteristics_of_Eukaryotic_Cells.txt |
3.1: Spontaneous Generation
The theory of spontaneous generation states that life arose from nonliving matter. It was a long-held belief dating back to Aristotle and the ancient Greeks. Experimentation by Francesco Redi in the 17th century presented the first significant evidence refuting spontaneous generation by showing that flies must have access to meat for maggots to develop on the meat. Louis Pasteur is credited with conclusively disproving the theory and proposed that “life only comes from life.”
Multiple Choice
Which of the following individuals argued in favor of the theory of spontaneous generation?
1. Francesco Redi
2. Louis Pasteur
3. John Needham
4. Lazzaro Spallanzani
Answer
C
Which of the following individuals is credited for definitively refuting the theory of spontaneous generation using broth in swan-neck flask?
1. Aristotle
2. Jan Baptista van Helmont
3. John Needham
4. Louis Pasteur
Answer
D
Which of the following experimented with raw meat, maggots, and flies in an attempt to disprove the theory of spontaneous generation.
1. Aristotle
2. Lazzaro Spallanzani
3. Antonie van Leeuwenhoek
4. Francesco Redi
Answer
D
Fill in the Blank
The assertion that “life only comes from life” was stated by Louis Pasteur in regard to his experiments that definitively refuted the theory of ___________.
Answer
spontaneous generation
True/False
Exposure to air is necessary for microbial growth.
Answer
False
Short Answer
Explain in your own words Pasteur’s swan-neck flask experiment.
Explain why the experiments of Needham and Spallanzani yielded in different results even though they used similar methodologies.
Critical Thinking
What would the results of Pasteur’s swan-neck flask experiment have looked like if they supported the theory of spontaneous generation?
3.2: Foundations of Modern Cell Theory
Although cells were first observed in the 1660s by Robert Hooke, cell theory was not well accepted for another 200 years. The work of scientists such as Schleiden, Schwann, Remak, and Virchow contributed to its acceptance. Endosymbiotic theory states that mitochondria and chloroplasts, organelles found in many types of organisms, have their origins in bacteria. Significant structural and genetic information support this theory. The miasma theory was widely accepted until the 19th century.
Multiple Choice
Which of the following individuals did not contribute to the establishment of cell theory?
1. Girolamo Fracastoro
2. Matthias Schleiden
3. Robert Remak
4. Robert Hooke
Answer
A
Whose proposal of the endosymbiotic theory of mitochondrial and chloroplast origin was ultimately accepted by the greater scientific community?
1. Rudolf Virchow
2. Ignaz Semmelweis
3. Lynn Margulis
4. Theodor Schwann
Answer
C
Which of the following developed a set of postulates for determining whether a particular disease is caused by a particular pathogen?
1. John Snow
2. Robert Koch
3. Joseph Lister
4. Louis Pasteur
Answer
B
Fill in the Blank
John Snow is known as the Father of _____________.
Answer
epidemiology
The ____________ theory states that disease may originate from proximity to decomposing matter and is not due to person-to-person contact.
Answer
miasma
The scientist who first described cells was _____________.
Answer
Robert Hooke
Short Answer
How did the explanation of Virchow and Remak for the origin of cells differ from that of Schleiden and Schwann?
What evidence exists that supports the endosymbiotic theory?
What were the differences in mortality rates due to puerperal fever that Ignaz Semmelweis observed? How did he propose to reduce the occurrence of puerperal fever? Did it work?
Critical Thinking
Why are mitochondria and chloroplasts unable to multiply outside of a host cell?
Why was the work of Snow so important in supporting the germ theory?
3.3: Unique Characteristics of Prokaryotic Cells
Prokaryotic cells differ from eukaryotic cells in that their genetic material is contained in a nucleoid rather than a membrane-bound nucleus. In addition, prokaryotic cells generally lack membrane-bound organelles. Prokaryotic cells of the same species typically share a similar cell morphology and cellular arrangement. Most prokaryotic cells have a cell wall that helps the organism maintain cellular morphology and protects it against changes in osmotic pressure.
Multiple Choice
Which of the following terms refers to a prokaryotic cell that is comma shaped?
1. coccus
2. coccobacilli
3. vibrio
4. spirillum
Answer
C
Which bacterial structures are important for adherence to surfaces? (Select all that apply.)
1. endospores
2. cell walls
3. fimbriae
4. capsules
5. flagella
Answer
C, D
Which of the following cell wall components is unique to gram-negative cells?
1. lipopolysaccharide
2. teichoic acid
3. mycolic acid
4. peptidoglycan
Answer
A
Which of the following terms refers to a bacterial cell having a single tuft of flagella at one end?
1. monotrichous
2. amphitrichous
3. peritrichous
4. lophotrichous
Answer
D
Bacterial cell walls are primarily composed of which of the following?
1. phospholipid
2. protein
3. carbohydrate
4. peptidoglycan
Answer
D
True/False
Bacteria have 80S ribosomes each composed of a 60S large subunit and a 40S small subunit.
Answer
False
Fill in the Blank
Prokaryotic cells that are rod-shaped are called _____________.
Answer
bacilli
The type of inclusion containing polymerized inorganic phosphate is called _____________.
Answer
volutin (or metachromatic granule)
Short Answer
What is the direction of water flow for a bacterial cell living in a hypotonic environment? How do cell walls help bacteria living in such environments?
How do bacterial flagella respond to a chemical gradient of an attractant to move toward a higher concentration of the chemical?
Label the parts of the prokaryotic cell.
Critical Thinking
Which of the following slides is a good example of staphylococci?
(credit a: modification of work by U.S. Department of Agriculture; credit b: modification of work by Centers for Disease Control and Prevention; credit c: modification of work by NIAID)
Provide some examples of bacterial structures that might be used as antibiotic targets and explain why.
The causative agent of botulism, a deadly form of food poisoning, is an endospore-forming bacterium called Clostridium botulinim. Why might it be difficult to kill this bacterium in contaminated food?
3.4: Unique Characteristics of Eukaryotic Cells
Eukaryotic cells are defined by the presence of a nucleus containing the DNA genome and bound by a nuclear membrane (or nuclear envelope) composed of two lipid bilayers that regulate transport of materials into and out of the nucleus through nuclear pores. Eukaryotic cell morphologies vary greatly and may be maintained by various structures, including the cytoskeleton, the cell membrane, and/or the cell wall. The nucleolus in the nucleus of eukaryotic cells is the site of ribosomal synthesis.
Multiple Choice
Which of the following organelles is not part of the endomembrane system?
1. endoplasmic reticulum
2. Golgi apparatus
3. lysosome
4. peroxisome
Answer
D
Which type of cytoskeletal fiber is important in the formation of the nuclear lamina?
1. microfilaments
2. intermediate filaments
3. microtubules
4. fibronectin
Answer
B
Sugar groups may be added to proteins in which of the following?
1. smooth endoplasmic reticulum
2. rough endoplasmic reticulum
3. Golgi apparatus
4. lysosome
Answer
C
Which of the following structures of a eukaryotic cell is not likely derived from endosymbiotic bacterium?
1. mitochondrial DNA
2. mitochondrial ribosomes
3. inner membrane
4. outer membrane
Answer
D
Which type of nutrient uptake involves the engulfment of small dissolved molecules into vesicles?
1. active transport
2. pinocytosis
3. receptor-mediated endocytosis
4. facilitated diffusion
Answer
B
Which of the following is not composed of microtubules?
1. desmosomes
2. centrioles
3. eukaryotic flagella
4. eukaryotic cilia
Answer
A
True/False
Mitochondria in eukaryotic cells contain ribosomes that are structurally similar to those found in prokaryotic cells.
Answer
True
Fill in the Blank
Peroxisomes typically produce _____________, a harsh chemical that helps break down molecules.
Answer
hydrogen peroxide
Microfilaments are composed of _____________ monomers.
Answer
actin
Short Answer
What existing evidence supports the theory that mitochondria are of prokaryotic origin?
Why do eukaryotic cells require an endomembrane system?
Name at least two ways that prokaryotic flagella are different from eukaryotic flagella.
Critical Thinking
Label the lettered parts of this eukaryotic cell.
How are peroxisomes more like mitochondria than like the membrane-bound organelles of the endomembrane system? How do they differ from mitochondria?
Why must the functions of both lysosomes and peroxisomes be compartmentalized? | textbooks/bio/Microbiology/Microbiology_(OpenStax)/03%3A_The_Cell/3.E%3A_The_Cell_%28Exercises%29.txt |
Scientists have studied prokaryotes for centuries, but it wasn’t until 1966 that scientist Thomas Brock (1926–) discovered that certain bacteria can live in boiling water. This led many to wonder whether prokaryotes may also live in other extreme environments, such as at the bottom of the ocean, at high altitudes, or inside volcanoes, or even on other planets.
Prokaryotes have an important role in changing, shaping, and sustaining the entire biosphere. They can produce proteins and other substances used by molecular biologists in basic research and in medicine and industry. For example, the bacterium Shewanella lives in the deep sea, where oxygen is scarce. It grows long appendages, which have special sensors used to seek the limited oxygen in its environment. It can also digest toxic waste and generate electricity. Other species of prokaryotes can produce more oxygen than the entire Amazon rainforest, while still others supply plants, animals, and humans with usable forms of nitrogen; and inhabit our body, protecting us from harmful microorganisms and producing some vitally important substances. This chapter will examine the diversity, structure, and function of prokaryotes.
• 4.1: Prokaryote Habitats, Relationships, and Microbiomes
Prokaryotes are unicellular microorganisms whose cells have no nucleus. Prokaryotes can be found everywhere on our planet, even in the most extreme environments. Prokaryotes are very flexible metabolically, so they are able to adjust their feeding to the available natural resources. Prokaryotes live in communities that interact among themselves and with large organisms that they use as hosts (including humans).
• 4.2: Proteobacteria
Proteobacteria is a phylum of gram-negative bacteria and are classified into the classes alpha-, beta-, gamma-, delta- and epsilonproteobacteria, each class having separate orders, families, genera, and species. Alphaproteobacteria are oligotrophs. The taxa chlamydias and rickettsias are obligate intracellular pathogens, feeding on cells of host organisms; they are metabolically inactive outside of the host cell. Some Alphaproteobacteria can convert atmospheric nitrogen to nitrites.
• 4.3: Nonproteobacteria Gram-negative Bacteria and Phototrophic Bacteria
Gram-negative nonproteobacteria include the taxa spirochetes; the Cytophaga, Fusobacterium, Bacteroides group; Planctomycetes; and many representatives of phototrophic bacteria. Spirochetes are motile, spiral bacteria with a long, narrow body; they are difficult or impossible to culture. Several genera of spirochetes contain human pathogens that cause such diseases as syphilis and Lyme disease. Cytophaga, Fusobacterium, and Bacteroides are classified together as a phylum called the CFB group.
• 4.4: Gram-positive Bacteria
Gram-positive bacteria are a very large and diverse group of microorganisms. Understanding their taxonomy and knowing their unique features is important for diagnostics and treatment of infectious diseases. Gram-positive bacteria are classified into high G+C gram-positive and low G+C gram-positive bacteria, based on the prevalence of guanine and cytosine nucleotides in their genome.
• 4.5: Deeply Branching Bacteria
Deeply branching bacteria are phylogenetically the most ancient forms of life, being the closest to the last universal common ancestor. Deeply branching bacteria include many species that thrive in extreme environments that are thought to resemble conditions on earth billions of years ago. Deeply branching bacteria are important for our understanding of evolution; some of them are used in industry
• 4.6: Archaea
Archaea are unicellular, prokaryotic microorganisms that differ from bacteria in their genetics, biochemistry, and ecology. Some archaea are extremophiles, living in environments with extremely high or low temperatures, or extreme salinity. Only archaea are known to produce methane. Methane-producing archaea are called methanogens. Halophilic archaea prefer a concentration of salt close to saturation and perform photosynthesis using bacteriorhodopsin.
• 4.E: Prokaryotic Diversity (Exercises)
Thumbnail: A cladogram linking all major groups of living organisms to the LUCA (the black trunk at the bottom), based on ribosomal RNA sequence data.
04: Prokaryotic Diversity
Learning Objectives
• Identify and describe unique examples of prokaryotes in various habitats on earth
• Identify and describe symbiotic relationships
• Compare normal/commensal/resident microbiota to transient microbiota
• Explain how prokaryotes are classified
Clinical Focus: Part 1
Marsha, a 20-year-old university student, recently returned to the United States from a trip to Nigeria, where she had interned as a medical assistant for an organization working to improve access to laboratory services for tuberculosis testing. When she returned, Marsha began to feel fatigue, which she initially attributed to jet lag. However, the fatigue persisted, and Marsha soon began to experience other bothersome symptoms, such as occasional coughing, night sweats, loss of appetite, and a low-grade fever of 37.4 °C (99.3 °F).
Marsha expected her symptoms would subside in a few days, but instead, they gradually became more severe. About two weeks after returning home, she coughed up some sputum and noticed that it contained blood and small whitish clumps resembling cottage cheese. Her fever spiked to 38.2 °C (100.8 °F), and she began feeling sharp pains in her chest when breathing deeply. Concerned that she seemed to be getting worse, Marsha scheduled an appointment with her physician.
Exercise \(1\)
Could Marsha’s symptoms be related to her overseas travel, even several weeks after returning home?
All living organisms are classified into three domains of life: Archaea, Bacteria, and Eukarya. In this chapter, we will focus on the domains Archaea and Bacteria. Archaea and bacteria are unicellular prokaryotic organisms. Unlike eukaryotes, they have no nuclei or any other membrane-bound organelles.
Prokaryote Habitats and Functions
Prokaryotes are ubiquitous. They can be found everywhere on our planet, even in hot springs, in the Antarctic ice shield, and under extreme pressure two miles under water. One bacterium, Paracoccus denitrificans, has even been shown to survive when scientists removed it from its native environment (soil) and used a centrifuge to subject it to forces of gravity as strong as those found on the surface of Jupiter.
Prokaryotes also are abundant on and within the human body. According to a report by National Institutes of Health, prokaryotes, especially bacteria, outnumber human cells 10:1.1 More recent studies suggest the ratio could be closer to 1:1, but even that ratio means that there are a great number of bacteria within the human body.2 Bacteria thrive in the human mouth, nasal cavity, throat, ears, gastrointestinal tract, and vagina. Large colonies of bacteria can be found on healthy human skin, especially in moist areas (armpits, navel, and areas behind ears). However, even drier areas of the skin are not free from bacteria.
The existence of prokaryotes is very important for the stability and thriving of ecosystems. For example, they are a necessary part of soil formation and stabilization processes through the breakdown of organic matter and development of biofilms. One gram of soil contains up to 10 billion microorganisms (most of them prokaryotic) belonging to about 1,000 species. Many species of bacteria use substances released from plant roots, such as acids and carbohydrates, as nutrients. The bacteria metabolize these plant substances and release the products of bacterial metabolism back to the soil, forming humus and thus increasing the soil’s fertility. In salty lakes such as the Dead Sea (Figure \(1\)), salt-loving halobacteria decompose dead brine shrimp and nourish young brine shrimp and flies with the products of bacterial metabolism.
In addition to living in the ground and the water, prokaryotic microorganisms are abundant in the air, even high in the atmosphere. There may be up to 2,000 different kinds of bacteria in the air, similar to their diversity in the soil.
Prokaryotes can be found everywhere on earth because they are extremely resilient and adaptable. They are often metabolically flexible, which means that they might easily switch from one energy source to another, depending on the availability of the sources, or from one metabolic pathway to another. For example, certain prokaryotic cyanobacteria can switch from a conventional type of lipid metabolism, which includes production of fatty aldehydes, to a different type of lipid metabolism that generates biofuel, such as fatty acids and wax esters. Groundwater bacteria store complex high-energy carbohydrates when grown in pure groundwater, but they metabolize these molecules when the groundwater is enriched with phosphates. Some bacteria get their energy by reducing sulfates into sulfides, but can switch to a different metabolic pathway when necessary, producing acids and free hydrogen ions.
Prokaryotes perform functions vital to life on earth by capturing (or “fixing”) and recycling elements like carbon and nitrogen. Organisms such as animals require organic carbon to grow, but, unlike prokaryotes, they are unable to use inorganic carbon sources like carbon dioxide. Thus, animals rely on prokaryotes to convert carbon dioxide into organic carbon products that they can use. This process of converting carbon dioxide to organic carbon products is called carbon fixation.
Plants and animals also rely heavily on prokaryotes for nitrogen fixation, the conversion of atmospheric nitrogen into ammonia, a compound that some plants can use to form many different biomolecules necessary to their survival. Bacteria in the genus Rhizobium, for example, are nitrogen-fixing bacteria; they live in the roots of legume plants such as clover, alfalfa, and peas (Figure \(2\)). Ammonia produced by Rhizobium helps these plants to survive by enabling them to make building blocks of nucleic acids. In turn, these plants may be eaten by animals—sustaining their growth and survival—or they may die, in which case the products of nitrogen fixation will enrich the soil and be used by other plants.
Another positive function of prokaryotes is in cleaning up the environment. Recently, some researchers focused on the diversity and functions of prokaryotes in manmade environments. They found that some bacteria play a unique role in degrading toxic chemicals that pollute water and soil.3
Despite all of the positive and helpful roles prokaryotes play, some are human pathogens that may cause illness or infection when they enter the body. In addition, some bacteria can contaminate food, causing spoilage or foodborne illness, which makes them subjects of concern in food preparation and safety. Less than 1% of prokaryotes (all of them bacteria) are thought to be human pathogens, but collectively these species are responsible for a large number of the diseases that afflict humans.
Besides pathogens, which have a direct impact on human health, prokaryotes also affect humans in many indirect ways. For example, prokaryotes are now thought to be key players in the processes of climate change. In recent years, as temperatures in the earth’s polar regions have risen, soil that was formerly frozen year-round (permafrost) has begun to thaw. Carbon trapped in the permafrost is gradually released and metabolized by prokaryotes. This produces massive amounts of carbon dioxide and methane, greenhouse gases that escape into the atmosphere and contribute to the greenhouse effect.
Exercise \(2\)
1. In what types of environments can prokaryotes be found?
2. Name some ways that plants and animals rely on prokaryotes.
Symbiotic Relationships
As we have learned, prokaryotic microorganisms can associate with plants and animals. Often, this association results in unique relationships between organisms. For example, bacteria living on the roots or leaves of a plant get nutrients from the plant and, in return, produce substances that protect the plant from pathogens. On the other hand, some bacteria are plant pathogens that use mechanisms of infection similar to bacterial pathogens of animals and humans.
Prokaryotes live in a community, or a group of interacting populations of organisms. A population is a group of individual organisms belonging to the same biological species and limited to a certain geographic area. Populations can have cooperative interactions, which benefit the populations, or competitive interactions, in which one population competes with another for resources. The study of these interactions between populations is called microbial ecology.
Any interaction between different species within a community is called symbiosis. Such interactions fall along a continuum between opposition and cooperation. Interactions in a symbiotic relationship may be beneficial or harmful, or have no effect on one or both of the species involved. Table \(1\) summarizes the main types of symbiotic interactions among prokaryotes.
Table \(1\): Types of Symbiotic Relationships
Type Population A Population B
Mutualism Benefitted Benefitted
Amensalism Harmed Unaffected
Commensalism Benefitted Unaffected
Neutralism Unaffected Unaffected
Parasitism Benefitted Harmed
When two species benefit from each other, the symbiosis is called mutualism (or syntropy, or crossfeeding). For example, humans have a mutualistic relationship with the bacterium Bacteroides thetaiotetraiotamicron, which lives in the intestinal tract. B. thetaiotetraiotamicron digests complex polysaccharide plant materials that human digestive enzymes cannot break down, converting them into monosaccharides that can be absorbed by human cells. Humans also have a mutualistic relationship with certain strains of Escherichia coli, another bacterium found in the gut. E. coli relies on intestinal contents for nutrients, and humans derive certain vitamins from E. coli, particularly vitamin K, which is required for the formation of blood clotting factors. (This is only true for some strains of E. coli, however. Other strains are pathogenic and do not have a mutualistic relationship with humans.)
A type of symbiosis in which one population harms another but remains unaffected itself is called amensalism. In the case of bacteria, some amensalist species produce bactericidal substances that kill other species of bacteria. For example, the bacterium Lucilia sericata produces a protein that destroys Staphylococcus aureus, a bacterium commonly found on the surface of the human skin. Too much handwashing can affect this relationship and lead to S. aureus diseases and transmission.
In another type of symbiosis, called commensalism, one organism benefits while the other is unaffected. This occurs when the bacterium Staphylococcus epidermidis uses the dead cells of the human skin as nutrients. Billions of these bacteria live on our skin, but in most cases (especially when our immune system is healthy), we do not react to them in any way.
If neither of the symbiotic organisms is affected in any way, we call this type of symbiosis neutralism. An example of neutralism is the coexistence of metabolically active (vegetating) bacteria and endospores (dormant, metabolically passive bacteria). For example, the bacterium Bacillus anthracis typically forms endospores in soil when conditions are unfavorable. If the soil is warmed and enriched with nutrients, some endospores germinate and remain in symbiosis with other endospores that have not germinated.
A type of symbiosis in which one organism benefits while harming the other is called parasitism. The relationship between humans and many pathogenic prokaryotes can be characterized as parasitic because these organisms invade the body, producing toxic substances or infectious diseases that cause harm. Diseases such as tetanus, diphtheria, pertussis, tuberculosis, and leprosy all arise from interactions between bacteria and humans.
Scientists have coined the term microbiome to refer to all prokaryotic and eukaryotic microorganisms that are associated with a certain organism. Within the human microbiome, there are resident microbiota and transient microbiota. The resident microbiota consists of microorganisms that constantly live in or on our bodies. The term transient microbiota refers to microorganisms that are only temporarily found in the human body, and these may include pathogenic microorganisms. Hygiene and diet can alter both the resident and transient microbiota.
The resident microbiota is amazingly diverse, not only in terms of the variety of species but also in terms of the preference of different microorganisms for different areas of the human body. For example, in the human mouth, there are thousands of commensal or mutualistic species of bacteria. Some of these bacteria prefer to inhabit the surface of the tongue, whereas others prefer the internal surface of the cheeks, and yet others prefer the front or back teeth or gums. The inner surface of the cheek has the least diverse microbiota because of its exposure to oxygen. By contrast, the crypts of the tongue and the spaces between teeth are two sites with limited oxygen exposure, so these sites have more diverse microbiota, including bacteria living in the absence of oxygen (e.g., Bacteroides, Fusobacterium). Differences in the oral microbiota between randomly chosen human individuals are also significant. Studies have shown, for example, that the prevalence of such bacteria as Streptococcus, Haemophilus, Neisseria, and others was dramatically different when compared between individuals.4
There are also significant differences between the microbiota of different sites of the same human body. The inner surface of the cheek has a predominance of Streptococcus, whereas in the throat, the palatine tonsil, and saliva, there are two to three times fewer Streptococcus, and several times more Fusobacterium. In the plaque removed from gums, the predominant bacteria belong to the genus Fusobacterium. However, in the intestine, both Streptococcus and Fusobacterium disappear, and the genus Bacteroides becomes predominant.
Not only can the microbiota vary from one body site to another, the microbiome can also change over time within the same individual. Humans acquire their first inoculations of normal flora during natural birth and shortly after birth. Before birth, there is a rapid increase in the population of Lactobacillus spp. in the vagina, and this population serves as the first colonization of microbiota during natural birth. After birth, additional microbes are acquired from health-care providers, parents, other relatives, and individuals who come in contact with the baby. This process establishes a microbiome that will continue to evolve over the course of the individual’s life as new microbes colonize and are eliminated from the body. For example, it is estimated that within a 9-hour period, the microbiota of the small intestine can change so that half of the microbial inhabitants will be different.5 The importance of the initial Lactobacillus colonization during vaginal child birth is highlighted by studies demonstrating a higher incidence of diseases in individuals born by cesarean section, compared to those born vaginally. Studies have shown that babies born vaginally are predominantly colonized by vaginal lactobacillus, whereas babies born by cesarean section are more frequently colonized by microbes of the normal skin microbiota, including common hospital-acquired pathogens.
Throughout the body, resident microbiotas are important for human health because they occupy niches that might be otherwise taken by pathogenic microorganisms. For instance, Lactobacillus spp. are the dominant bacterial species of the normal vaginal microbiota for most women. lactobacillus produce lactic acid, contributing to the acidity of the vagina and inhibiting the growth of pathogenic yeasts. However, when the population of the resident microbiota is decreased for some reason (e.g., because of taking antibiotics), the pH of the vagina increases, making it a more favorable environment for the growth of yeasts such as Candida albicans. Antibiotic therapy can also disrupt the microbiota of the intestinal tract and respiratory tract, increasing the risk for secondary infections and/or promoting the long-term carriage and shedding of pathogens.
Exercise \(3\)
1. Explain the difference between cooperative and competitive interactions in microbial communities.
2. List the types of symbiosis and explain how each population is affected.
Taxonomy and Systematics
Assigning prokaryotes to a certain species is challenging. They do not reproduce sexually, so it is not possible to classify them according to the presence or absence of interbreeding. Also, they do not have many morphological features. Traditionally, the classification of prokaryotes was based on their shape, staining patterns, and biochemical or physiological differences. More recently, as technology has improved, the nucleotide sequences in genes have become an important criterion of microbial classification.
In 1923, American microbiologist David Hendricks Bergey (1860–1937) published A Manual in Determinative Bacteriology. With this manual, he attempted to summarize the information about the kinds of bacteria known at that time, using Latin binomial classification. Bergey also included the morphological, physiological, and biochemical properties of these organisms. His manual has been updated multiple times to include newer bacteria and their properties. It is a great aid in bacterial taxonomy and methods of characterization of bacteria. A more recent sister publication, the five-volume Bergey’s Manual of Systematic Bacteriology, expands on Bergey’s original manual. It includes a large number of additional species, along with up-to-date descriptions of the taxonomy and biological properties of all named prokaryotic taxa. This publication incorporates the approved names of bacteria as determined by the List of Prokaryotic Names with Standing in Nomenclature (LPSN).
Classification by Staining Patterns
According to their staining patterns, which depend on the properties of their cell walls, bacteria have traditionally been classified into gram-positive, gram-negative, and “atypical,” meaning neither gram-positive nor gram-negative. As explained in Staining Microscopic Specimens, gram-positive bacteria possess a thick peptidoglycan cell wall that retains the primary stain (crystal violet) during the decolorizing step; they remain purple after the gram-stain procedure because the crystal violet dominates the light red/pink color of the secondary counterstain, safranin. In contrast, gram-negative bacteria possess a thin peptidoglycan cell wall that does not prevent the crystal violet from washing away during the decolorizing step; therefore, they appear light red/pink after staining with the safranin. Bacteria that cannot be stained by the standard Gram stain procedure are called atypical bacteria. Included in the atypical category are species of Mycoplasma and Chlamydia, which lack a cell wall and therefore cannot retain the gram-stain reagents. Rickettsiaare also considered atypical because they are too small to be evaluated by the Gram stain.
More recently, scientists have begun to further classify gram-negative and gram-positive bacteria. They have added a special group of deeply branching bacteria based on a combination of physiological, biochemical, and genetic features. They also now further classify gram-negative bacteria into Proteobacteria, Cytophaga-Flavobacterium-Bacteroides (CFB), and spirochetes.
The deeply branching bacteria are thought to be a very early evolutionary form of bacteria (see Deeply Branching Bacteria). They live in hot, acidic, ultraviolet-light-exposed, and anaerobic (deprived of oxygen) conditions. Proteobacteria is a phylum of very diverse groups of gram-negative bacteria; it includes some important human pathogens (e.g., E. coli and Bordetella pertussis). The CFB group of bacteria includes components of the normal human gut microbiota, like Bacteroides. The spirochetes are spiral-shaped bacteria and include the pathogen Treponema pallidum, which causes syphilis. We will characterize these groups of bacteria in more detail later in the chapter.
Based on their prevalence of guanine and cytosine nucleotides, gram-positive bacteria are also classified into low G+C and high G+C gram-positive bacteria. The low G+C gram-positive bacteria have less than 50% of guanine and cytosine nucleotides in their DNA. They include human pathogens, such as those that cause anthrax (Bacillus anthracis), tetanus (Clostridium tetani), and listeriosis (Listeria monocytogenes). High G+C gram-positive bacteria, which have more than 50% guanine and cytosine nucleotides in their DNA, include the bacteria that cause diphtheria (Corynebacterium diphtheriae), tuberculosis (Mycobacterium tuberculosis), and other diseases.
The classifications of prokaryotes are constantly changing as new species are being discovered. We will describe them in more detail, along with the diseases they cause, in later sections and chapters.
Exercise \(4\)
How do scientists classify prokaryotes?
Human Microbiome Project
The Human Microbiome Project was launched by the National Institutes of Health (NIH) in 2008. One main goal of the project is to create a large repository of the gene sequences of important microbes found in humans, helping biologists and clinicians understand the dynamics of the human microbiome and the relationship between the human microbiota and diseases. A network of labs working together has been compiling the data from swabs of several areas of the skin, gut, and mouth from hundreds of individuals.
One of the challenges in understanding the human microbiome has been the difficulty of culturing many of the microbes that inhabit the human body. It has been estimated that we are only able to culture 1% of the bacteria in nature and that we are unable to grow the remaining 99%. To address this challenge, researchers have used metagenomic analysis, which studies genetic material harvested directly from microbial communities, as opposed to that of individual species grown in a culture. This allows researchers to study the genetic material of all microbes in the microbiome, rather than just those that can be cultured.6
One important achievement of the Human Microbiome Project is establishing the first reference database on microorganisms living in and on the human body. Many of the microbes in the microbiome are beneficial, but some are not. It was found, somewhat unexpectedly, that all of us have some serious microbial pathogens in our microbiota. For example, the conjunctiva of the human eye contains 24 genera of bacteria and numerous pathogenic species.7 A healthy human mouth contains a number of species of the genus Streptococcus, including pathogenic species S. pyogenes and S. pneumoniae.8 This raises the question of why certain prokaryotic organisms exist commensally in certain individuals but act as deadly pathogens in others. Also unexpected was the number of organisms that had never been cultured. For example, in one metagenomic study of the human gut microbiota, 174 new species of bacteria were identified.9
Another goal for the near future is to characterize the human microbiota in patients with different diseases and to find out whether there are any relationships between the contents of an individual’s microbiota and risk for or susceptibility to specific diseases. Analyzing the microbiome in a person with a specific disease may reveal new ways to fight diseases.
Summary
• Prokaryotes are unicellular microorganisms whose cells have no nucleus.
• Prokaryotes can be found everywhere on our planet, even in the most extreme environments.
• Prokaryotes are very flexible metabolically, so they are able to adjust their feeding to the available natural resources.
• Prokaryotes live in communities that interact among themselves and with large organisms that they use as hosts (including humans).
• The totality of forms of prokaryotes (particularly bacteria) living on the human body is called the human microbiome, which varies between regions of the body and individuals, and changes over time.
• The totality of forms of prokaryotes (particularly bacteria) living in a certain region of the human body (e.g., mouth, throat, gut, eye, vagina) is called the microbiota of this region.
• Prokaryotes are classified into domains Archaea and Bacteria.
• In recent years, the traditional approaches to classification of prokaryotes have been supplemented by approaches based on molecular genetics.
Footnotes
1. 1 Medical Press. “Mouth Bacteria Can Change Their Diet, Supercomputers Reveal.” August 12, 2014. medicalxpress.com/news/2014-0...rs-reveal.html. Accessed February 24, 2015.
2. 2 A. Abbott. “Scientists Bust Myth That Our Bodies Have More Bacteria Than Human Cells: Decades-Old Assumption about Microbiota Revisited.” Nature. http://www.nature.com/news/scientist...-cells-1.19136. Accessed June 3, 2016.
3. 3 A.M. Kravetz “Unique Bacteria Fights Man-Made Chemical Waste.” 2012. www.livescience.com/25181-bac...s-nsf-bts.html. Accessed March 9, 2015.
4. 4 E.M. Bik et al. “Bacterial Diversity in the Oral Cavity of 10 Healthy Individuals.” The ISME Journal 4 no. 8 (2010):962–974.
5. 5 C.C. Booijink et al. “High Temporal and Intra-Individual Variation Detected in the Human Ileal Microbiota.” Environmental Microbiology 12 no. 12 (2010):3213–3227.
6. 6 National Institutes of Health. “Human Microbiome Project. Overview.” commonfund.nih.gov/hmp/overview. Accessed June 7, 2016.
7. 7 Q. Dong et al. “Diversity of Bacteria at Healthy Human Conjunctiva.” Investigative Ophthalmology & Visual Science 52 no. 8 (2011):5408–5413.
8. 8 F.E. Dewhirst et al. “The Human Oral Microbiome.” Journal of Bacteriology 192 no. 19 (2010):5002–5017.
9. 9 J.C. Lagier et al. “Microbial Culturomics: Paradigm Shift in the Human Gut Microbiome Study.” Clinical Microbiology and Infection 18 no. 12 (2012):1185–1193. | textbooks/bio/Microbiology/Microbiology_(OpenStax)/04%3A_Prokaryotic_Diversity/4.01%3A_Prokaryote_Habitats_Relationships_and_Microbiomes.txt |
Learning Objectives
• Describe the unique features of each class within the phylum Proteobacteria: Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Deltaproteobacteria, and Epsilonproteobacteria
• Give an example of a bacterium in each class of Proteobacteria
In 1987, the American microbiologist Carl Woese (1928–2012) suggested that a large and diverse group of bacteria that he called “purple bacteria and their relatives” should be defined as a separate phylum within the domain Bacteria based on the similarity of the nucleotide sequences in their genome.1 This phylum of gram-negative bacteria subsequently received the name Proteobacteria. It includes many bacteria that are part of the normal human microbiota as well as many pathogens. The Proteobacteria are further divided into five classes: Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Deltaproteobacteria, and Epsilonproteobacteria.
Alphaproteobacteria
The first class of Proteobacteria is the Alphaproteobacteria. The unifying characteristic of this class is that they are oligotrophs, organisms capable of living in low-nutrient environments such as deep oceanic sediments, glacial ice, or deep undersurface soil.
Among the Alphaproteobacteria are two taxa, chlamydias and rickettsias, that are obligate intracellular pathogens, meaning that part of their life cycle must occur inside other cells called host cells. When not growing inside a host cell, Chlamydia and Rickettsia are metabolically inactive outside of the host cell. They cannot synthesize their own adenosine triphosphate (ATP), and, therefore, rely on cells for their energy needs.
Rickettsia spp. include a number of serious human pathogens. For example, R. rickettsii causes Rocky Mountain spotted fever, a life-threatening form of meningoencephalitis (inflammation of the membranes that wrap the brain). R. rickettsii infects ticks and can be transmitted to humans via a bite from an infected tick (Figure \(1\)).
Another species of Rickettsia, R. prowazekii, is spread by lice. It causes epidemic typhus, a severe infectious disease common during warfare and mass migrations of people. R. prowazekii infects human endothelium cells, causing inflammation of the inner lining of blood vessels, high fever, abdominal pain, and sometimes delirium. A relative, R. typhi, causes a less severe disease known as murine or endemic typhus, which is still observed in the southwestern United States during warm seasons.
Chlamydia is another taxon of the Alphaproteobacteria. Members of this genus are extremely resistant to the cellular defenses, giving them the ability to spread from host to host rapidly via elementary bodies. The metabolically and reproductively inactive elementary bodies are the endospore-like form of intracellular bacteria that enter an epithelial cell, where they become active. Figure \(2\) illustrates the life cycle of Chlamydia.
C. trachomatis is a human pathogen that causes trachoma, a disease of the eyes, often leading to blindness. C. trachomatis also causes the sexually transmitted disease lymphogranuloma venereum (LGV). This disease is often mildly symptomatic, manifesting as regional lymph node swelling, or it may be asymptomatic, but it is extremely contagious and is common on college campuses. Table \(1\) summarizes the characteristics of important genera of Alphaproteobacteria.
Table \(1\): Class Alphaproteobacteria
Genus Microscopic Morphology Unique Characteristics
Agrobacterium Gram-negative bacillus Plant pathogen; one species, A. tumefaciens, causes tumors in plants
Bartonella Gram-negative, pleomorphic, flagellated coccobacillus Facultative intracellular bacteria, transmitted by lice and fleas, cause trench fever and cat scratch disease in humans
Brucella Gram-negative, small, flagellated coccobacillus Facultative intracellular bacteria, transmitted by contaminated milk from infected cows, cause brucellosis in cattle and humans
Caulobacter Gram-negative bacillus Used in studies on cellular adaptation and differentiation because of its peculiar life cycle (during cell division, forms “swarm” cells and “stalked” cells)
Chlamydia Gram-negative, coccoid or ovoid bacterium Obligatory intracellular bacteria; some cause chlamydia, trachoma, and pneumonia
Coxiella Small, gram-negative bacillus Obligatory intracellular bacteria; cause Q fever; potential for use as biological weapon
Ehrlichia Very small, gram-negative, coccoid or ovoid bacteria Obligatory intracellular bacteria; can be transported from cell to cell; transmitted by ticks; cause ehrlichiosis (destruction of white blood cells and inflammation) in humans and dogs
Hyphomicrobium Gram-negative bacilli; grows from a stalk Similar to Caulobacter
Methylocystis Gram-negative, coccoid or short bacilli Nitrogen-fixing aerobic bacteria
Rhizobium Gram-negative, rectangular bacilli with rounded ends forming clusters Nitrogen-fixing bacteria that live in soil and form symbiotic relationship with roots of legumes (e.g., clover, alfalfa, and beans)
Rickettsia Gram-negative, highly pleomorphic bacteria (may be cocci, rods, or threads) Obligate intracellular bacteria; transmitted by ticks; may cause Rocky Mountain spotted fever and typhus
Exercise \(1\)
What characteristic do all Alphaproteobacteria share?
Betaproteobacteria
Unlike Alphaproteobacteria, which survive on a minimal amount of nutrients, the class Betaproteobacteria are eutrophs (or copiotrophs), meaning that they require a copious amount of organic nutrients. Betaproteobacteria often grow between aerobic and anaerobic areas (e.g., in mammalian intestines). Some genera include species that are human pathogens, able to cause severe, sometimes life-threatening disease. The genus Neisseria, for example, includes the bacteria N. gonorrhoeae, the causative agent of the STI gonorrhea, and N. meningitides, the causative agent of bacterial meningitis.
Neisseria are cocci that live on mucosal surfaces of the human body. They are fastidious, or difficult to culture, and they require high levels of moisture, nutrient supplements, and carbon dioxide. Also, Neisseria are microaerophilic, meaning that they require low levels of oxygen. For optimal growth and for the purposes of identification, Neisseria spp. are grown on chocolate agar (i.e., agar supplemented by partially hemolyzed red blood cells). Their characteristic pattern of growth in culture is diplococcal: pairs of cells resembling coffee beans (Figure \(3\)).
The pathogen responsible for pertussis (whooping cough) is also a member of Betaproteobacteria. The bacterium Bordetella pertussis, from the order Burkholderiales, produces several toxins that paralyze the movement of cilia in the human respiratory tract and directly damage cells of the respiratory tract, causing a severe cough. Table \(2\) summarizes the characteristics of important genera of Betaproteobacteria.
Table \(2\): Class Betaproteobacteria
Example Genus Microscopic Morphology Unique Characteristics
Bordetella A small, gram-negative coccobacillus Aerobic, very fastidious; B. pertussis causes pertussis (whooping cough)
Burkholderia Gram-negative bacillus Aerobic, aquatic, cause diseases in horses and humans (especially patients with cystic fibrosis); agents of nosocomial infections
Leptothrix Gram-negative, sheathed, filamentous bacillus Aquatic; oxidize iron and manganese; can live in wastewater treatment plants and clog pipes
Neisseria Gram-negative, coffee bean-shaped coccus forming pairs Require moisture and high concentration of carbon dioxide; oxidase positive, grow on chocolate agar; pathogenic species cause gonorrhea and meningitis
Thiobacillus Gram-negative bacillus Thermophilic, acidophilic, strictly aerobic bacteria; oxidize iron and sulfur
Exercise \(2\)
What characteristic do all Betaproteobacteria share?
Clinical Focus: Part 2
When Marsha finally went to the doctor’s office, the physician listened to her breathing through a stethoscope. He heard some crepitation (a crackling sound) in her lungs, so he ordered a chest radiograph and asked the nurse to collect a sputum sample for microbiological evaluation and cytology. The radiologic evaluation found cavities, opacities, and a particular pattern of distribution of abnormal material (Figure \(4\)).
Exercise \(3\)
What are some possible diseases that could be responsible for Marsha’s radiograph results?
Gammaproteobacteria
The most diverse class of gram-negative bacteria is Gammaproteobacteria, and it includes a number of human pathogens. For example, a large and diverse family, Pseudomonaceae, includes the genus Pseudomonas. Within this genus is the species P. aeruginosa, a pathogen responsible for diverse infections in various regions of the body. P. aeruginosa is a strictly aerobic, nonfermenting, highly motile bacterium. It often infects wounds and burns, can be the cause of chronic urinary tract infections, and can be an important cause of respiratory infections in patients with cystic fibrosis or patients on mechanical ventilators. Infections by P. aeruginosa are often difficult to treat because the bacterium is resistant to many antibiotics and has a remarkable ability to form biofilms. Other representatives of Pseudomonas include the fluorescent (glowing) bacterium P. fluorescens and the soil bacteria P. putida, which is known for its ability to degrade xenobiotics (substances not naturally produced or found in living organisms).
The Pasteurellaceae also includes several clinically relevant genera and species. This family includes several bacteria that are human and/or animal pathogens. For example, Pasteurella haemolytica causes severe pneumonia in sheep and goats. P. multocida is a species that can be transmitted from animals to humans through bites, causing infections of the skin and deeper tissues. The genus Haemophilus contains two human pathogens, H. influenzae and H. ducreyi. Despite its name, H. influenzae does not cause influenza (which is a viral disease). H. influenzae can cause both upper and lower respiratory tract infections, including sinusitis, bronchitis, ear infections, and pneumonia. Before the development of effective vaccination, strains of H. influenzae were a leading cause of more invasive diseases, like meningitis in children. H. ducreyi causes the STI known as chancroid.
The order Vibrionales includes the human pathogen Vibrio cholerae. This comma-shaped aquatic bacterium thrives in highly alkaline environments like shallow lagoons and sea ports. A toxin produced by V. cholerae causes hypersecretion of electrolytes and water in the large intestine, leading to profuse watery diarrhea and dehydration. V. parahaemolyticus is also a cause of gastrointestinal disease in humans, whereas V. vulnificus causes serious and potentially life-threatening cellulitis (infection of the skin and deeper tissues) and blood-borne infections. Another representative of Vibrionales, Aliivibrio fischeri, engages in a symbiotic relationship with squid. The squid provides nutrients for the bacteria to grow and the bacteria produce bioluminescence that protects the squid from predators (Figure \(5\)).
The genus Legionella also belongs to the Gammaproteobacteria. L. pneumophila, the pathogen responsible for Legionnaires disease, is an aquatic bacterium that tends to inhabit pools of warm water, such as those found in the tanks of air conditioning units in large buildings (Figure \(6\)). Because the bacteria can spread in aerosols, outbreaks of Legionnaires disease often affect residents of a building in which the water has become contaminated with Legionella. In fact, these bacteria derive their name from the first known outbreak of Legionnaires disease, which occurred in a hotel hosting an American Legion veterans’ association convention in Philadelphia in 1976.
Enterobacteriaceae is a large family of enteric (intestinal) bacteria belonging to the Gammaproteobacteria. They are facultative anaerobes and are able to ferment carbohydrates. Within this family, microbiologists recognize two distinct categories. The first category is called the coliforms, after its prototypical bacterium species, Escherichia coli. Coliforms are able to ferment lactose completely (i.e., with the production of acid and gas). The second category, noncoliforms, either cannot ferment lactose or can only ferment it incompletely (producing either acid or gas, but not both). The noncoliforms include some notable human pathogens, such as Salmonella spp., Shigella spp., and Yersinia pestis.
E. coli has been perhaps the most studied bacterium since it was first described in 1886 by Theodor Escherich (1857–1911). Many strains of E. coli are in mutualistic relationships with humans. However, some strains produce a potentially deadly toxin called Shiga toxin, which perforates cellular membranes in the large intestine, causing bloody diarrhea and peritonitis (inflammation of the inner linings of the abdominal cavity). Other E. coli strains may cause traveler’s diarrhea, a less severe but very widespread disease.
The genus Salmonella, which belongs to the noncoliform group of Enterobacteriaceae, is interesting in that there is still no consensus about how many species it includes. Scientists have reclassified many of the groups they once thought to be species as serotypes (also called serovars), which are strains or variations of the same species of bacteria. Their classification is based on patterns of reactivity by animal antisera against molecules on the surface of the bacterial cells. A number of serotypes of Salmonella can cause salmonellosis, characterized by inflammation of the small and the large intestine, accompanied by fever, vomiting, and diarrhea. The species S. enterobacterica (serovar typhi) causes typhoid fever, with symptoms including fever, abdominal pain, and skin rashes (Figure \(7\)). Table \(3\) summarizes the characteristics of important genera of Gammaproteobacteria.
Table \(3\): Class Gammaproteobacteria
Example Genus Microscopic Morphology Unique Characteristics
Beggiatoa Gram-negative bacteria; disc-shaped or cylindrical Aquatic, live in water with high content of hydrogen disulfide; can cause problems for sewage treatment
Enterobacter Gram-negative bacillus Facultative anaerobe; cause urinary and respiratory tract infections in hospitalized patients; implicated in the pathogenesis of obesity
Erwinia Gram-negative bacillus Plant pathogen causing leaf spots and discoloration; may digest cellulose; prefer relatively low temperatures (25–30 °C)
Escherichia Gram-negative bacillus Facultative anaerobe; inhabit the gastrointestinal tract of warm-blooded animals; some strains are mutualists, producing vitamin K; others, like serotype E. coli O157:H7, are pathogens; E. coli has been a model organism for many studies in genetics and molecular biology
Hemophilus Gram-negative bacillus Pleomorphic, may appear as coccobacillus, aerobe, or facultative anaerobe; grow on blood agar; pathogenic species can cause respiratory infections, chancroid, and other diseases
Klebsiella Gram-negative bacillus; appears rounder and thicker than other members of Enterobacteriaceae Facultative anaerobe, encapsulated, nonmotile; pathogenic species may cause pneumonia, especially in people with alcoholism
Legionella Gram-negative bacillus Fastidious, grow on charcoal-buffered yeast extract; L. pneumophila causes Legionnaires disease
Methylomonas Gram-negative bacillus Use methane as source of carbon and energy
Proteus Gram-negative bacillus (pleomorphic) Common inhabitants of the human gastrointestinal tract; motile; produce urease; opportunistic pathogens; may cause urinary tract infections and sepsis
Pseudomonas Gram-negative bacillus Aerobic; versatile; produce yellow and blue pigments, making them appear green in culture; opportunistic, antibiotic-resistant pathogens may cause wound infections, hospital-acquired infections, and secondary infections in patients with cystic fibrosis
Serratia Gram-negative bacillus Motile; may produce red pigment; opportunistic pathogens responsible for a large number of hospital-acquired infections
Shigella Gram-negative bacillus Nonmotile; dangerously pathogenic; produce Shiga toxin, which can destroy cells of the gastrointestinal tract; can cause dysentery
Vibrio Gram-negative, comma- or curved rod-shaped bacteria Inhabit seawater; flagellated, motile; may produce toxin that causes hypersecretion of water and electrolytes in the gastrointestinal tract; some species may cause serious wound infections
Yersinia Gram-negative bacillus Carried by rodents; human pathogens; Y. pestis causes bubonic plague and pneumonic plague; Y. enterocolitica can be a pathogen causing diarrhea in humans
Exercise \(4\)
List two families of Gammaproteobacteria.
Deltaproteobacteria
The Deltaproteobacteria is a small class of gram-negative Proteobacteria that includes sulfate-reducing bacteria(SRBs), so named because they use sulfate as the final electron acceptor in the electron transport chain. Few SRBs are pathogenic. However, the SRB Desulfovibrio orale is associated with periodontal disease (disease of the gums).
Deltaproteobacteria also includes the genus Bdellovibrio, species of which are parasites of other gram-negative bacteria. Bdellovibrio invades the cells of the host bacterium, positioning itself in the periplasm, the space between the plasma membrane and the cell wall, feeding on the host’s proteins and polysaccharides. The infection is lethal for the host cells.
Another type of Deltaproteobacteria, myxobacteria, lives in the soil, scavenging inorganic compounds. Motile and highly social, they interact with other bacteria within and outside their own group. They can form multicellular, macroscopic “fruiting bodies” (Figure \(8\)), structures that are still being studied by biologists and bacterial ecologists.2These bacteria can also form metabolically inactive myxospores. Table \(4\) summarizes the characteristics of several important genera of Deltaproteobacteria.
Table \(4\): Class Deltaproteobacteria
Genus Microscopic Morphology Unique characteristics
Bdellovibrio Gram-negative, comma-shaped rod Obligate aerobes; motile; parasitic (infecting other bacteria)
Desulfovibrio(formerly Desufuromonas) Gram-negative, comma-shaped rod Reduce sulfur; can be used for removal of toxic and radioactive waste
Myxobacterium Gram-negative, coccoid bacteria forming colonies (swarms) Live in soil; can move by gliding; used as a model organism for studies of intercellular communication (signaling)
Exercise \(5\)
What type of Deltaproteobacteria forms fruiting bodies?
Epsilonproteobacteria
The smallest class of Proteobacteria is Epsilonproteobacteria, which are gram-negative microaerophilic bacteria (meaning they only require small amounts of oxygen in their environment). Two clinically relevant genera of Epsilonproteobacteria are Campylobacter and Helicobacter, both of which include human pathogens. Campylobacter can cause food poisoning that manifests as severe enteritis (inflammation in the small intestine). This condition, caused by the species C. jejuni, is rather common in developed countries, usually because of eating contaminated poultry products. Chickens often harbor C. jejuni in their gastrointestinal tract and feces, and their meat can become contaminated during processing.
Within the genus Helicobacter, the helical, flagellated bacterium H. pylori has been identified as a beneficial member of the stomach microbiota, but it is also the most common cause of chronic gastritis and ulcers of the stomach and duodenum (Figure \(9\)). Studies have also shown that H. pylori is linked to stomach cancer.3 H. pylori is somewhat unusual in its ability to survive in the highly acidic environment of the stomach. It produces urease and other enzymes that modify its environment to make it less acidic. Table \(5\) summarizes the characteristics of the most clinically relevant genera of Epsilonproteobacteria.
Table \(5\): Class Epsilonproteobacteria
Example Genus Microscopic Morphology Unique Characteristics
Campylobacter Gram-negative, spiral-shaped rod Aerobic (microaerophilic); often infects chickens; may infect humans via undercooked meat, causing severe enteritis
Helicobacter Gram-negative, spiral-shaped rod Aerobic (microaerophilic) bacterium; can damage the inner lining of the stomach, causing chronic gastritis, peptic ulcers, and stomach cancer
Exercise \(1\)
Name two Epsilonproteobacteria that cause gastrointestinal disorders.
Summary
• Proteobacteria is a phylum of gram-negative bacteria discovered by Carl Woese in the 1980s based on nucleotide sequence homology.
• Proteobacteria are further classified into the classes alpha-, beta-, gamma-, delta- and epsilonproteobacteria, each class having separate orders, families, genera, and species.
• Alphaproteobacteria are oligotrophs. The taxa chlamydias and rickettsias are obligate intracellular pathogens, feeding on cells of host organisms; they are metabolically inactive outside of the host cell. Some Alphaproteobacteria can convert atmospheric nitrogen to nitrites, making nitrogen usable by other forms of life.
• Betaproteobacteria are eutrophs. They include human pathogens of the genus Neisseria and the species Bordetella pertussis.
• Gammaproteobacteria are the largest and the most diverse group of Proteobacteria. Many are human pathogens that are aerobes or facultative anaerobes. Some Gammaproteobacteria are enteric bacteria that may be coliform or noncoliform. Escherichia coli, a member of Gammaproteobacteria, is perhaps the most studied bacterium.
• Deltaproteobacteria make up a small group able to reduce sulfate or elemental sulfur. Some are scavengers and form myxospores, with multicellular fruiting bodies.
• Epsilonproteobacteria make up the smallest group of Proteobacteria. The genera Campylobacter and Helicobacter are human pathogens.
Footnotes
1. C.R. Woese. “Bacterial Evolution.” Microbiological Review 51 no. 2 (1987):221–271.
2. H. Reichenbach. “Myxobacteria, Producers of Novel Bioactive Substances.” Journal of Industrial Microbiology & Biotechnology 27 no. 3 (2001):149–156.
3. S. Suerbaum, P. Michetti. “Helicobacter pylori infection.” New England Journal of Medicine 347 no. 15 (2002):1175–1186. | textbooks/bio/Microbiology/Microbiology_(OpenStax)/04%3A_Prokaryotic_Diversity/4.02%3A_Proteobacteria.txt |
Learning Objectives
• Describe the unique features of nonproteobacteria gram-negative bacteria
• Give an example of a nonproteobacteria bacterium in each category
• Describe the unique features of phototrophic bacteria
• Identify phototrophic bacteria
The majority of the gram-negative bacteria belong to the phylum Proteobacteria, discussed in the previous section. Those that do not are called the nonproteobacteria. In this section, we will describe three classes of gram-negative nonproteobacteria: the spirochetes, the CFB group, and the Planctomycetes. A diverse group of phototrophic bacteria that includes Proteobacteria and nonproteobacteria will be discussed at the end of this section.
Spirochetes
Spirochetes are characterized by their long (up to 250 μm), spiral-shaped bodies. Most spirochetes are also very thin, which makes it difficult to examine gram-stained preparations under a conventional brightfield microscope. Darkfield fluorescent microscopy is typically used instead. Spirochetes are also difficult or even impossible to culture. They are highly motile, using their axial filament to propel themselves. The axial filament is similar to a flagellum, but it wraps around the cell and runs inside the cell body of a spirochete in the periplasmic space between the outer membrane and the plasma membrane (Figure \(1\)).
Several genera of spirochetes include human pathogens. For example, the genus Treponema includes a species T. pallidum, which is further classified into four subspecies: T. pallidum pallidum, T. pallidum pertenue, T. pallidum carateum, and T. pallidum endemicum. The subspecies T. pallidum pallidum causes the sexually transmitted infection known as syphilis, the third most prevalent sexually transmitted bacterial infection in the United States, after chlamydia and gonorrhea. The other subspecies of T. pallidum cause tropical infectious diseases of the skin, bones, and joints.
Another genus of spirochete, Borrelia, contains a number of pathogenic species. B. burgdorferi causes Lyme disease, which is transmitted by several genera of ticks (notably Ixodes and Amblyomma) and often produces a “bull’s eye” rash, fever, fatigue, and, sometimes, debilitating arthritis. B. recurrens causes a condition known as relapsing fever.
Exercise \(1\)
Why do scientists typically use darkfield fluorescent microscopy to visualize spirochetes?
Cytophaga, Fusobacterium, and Bacteroides
The gram-negative nonproteobacteria of the genera Cytophaga, Fusobacterium, and Bacteroides are classified together as a phylum and called the CFB group. Although they are phylogenetically diverse, bacteria of the CFB group share some similarities in the sequence of nucleotides in their DNA. They are rod-shaped bacteria adapted to anaerobic environments, such as the tissue of the gums, gut, and rumen of ruminating animals. CFB bacteria are avid fermenters, able to process cellulose in rumen, thus enabling ruminant animals to obtain carbon and energy from grazing.
Cytophaga are motile aquatic bacteria that glide. Fusobacteria inhabit the human mouth and may cause severe infectious diseases. The largest genus of the CFB group is Bacteroides, which includes dozens of species that are prevalent inhabitants of the human large intestine, making up about 30% of the entire gut microbiome (Figure \(2\)). One gram of human feces contains up to 100 billion Bacteroides cells. Most Bacteroides are mutualistic. They benefit from nutrients they find in the gut, and humans benefit from their ability to prevent pathogens from colonizing the large intestine. Indeed, when populations of Bacteroides are reduced in the gut—as often occurs when a patient takes antibiotics—the gut becomes a more favorable environment for pathogenic bacteria and fungi, which can cause secondary infections.
Only a few species of Bacteroides are pathogenic. B. melaninogenicus, for example, can cause wound infections in patients with weakened immune systems.
Exercise \(2\)
Why are Cytophaga, Fusobacterium, and Bacteroides classified together as the CFB group?
Planctomycetes
The Planctomycetes are found in aquatic environments, inhabiting freshwater, saltwater, and brackish water. Planctomycetes are unusual in that they reproduce by budding, meaning that instead of one maternal cell splitting into two equal daughter cells in the process of binary fission, the mother cell forms a bud that detaches from the mother cell and lives as an independent cell. These so-called swarmer cells are motile and not attached to a surface. However, they will soon differentiate into sessile (immobile) cells with an appendage called a holdfast that allows them to attach to surfaces in the water (Figure \(3\)). Only the sessile cells are able to reproduce.
Table \(1\) summarizes the characteristics of some of the most clinically relevant genera of nonproteobacteria. Table \(1\): Nonproteobacteria
Example Genus Microscopic Morphology Unique Characteristics
Bacteroides Gram-negative bacillus Obligate anaerobic bacteria; abundant in the human gastrointestinal tract; usually mutualistic, although some species are opportunistic pathogens
Cytophaga Gram-negative bacillus Motile by gliding; live in soil or water; decompose cellulose; may cause disease in fish
Fusobacterium Gram-negative bacillus with pointed ends Anaerobic; form; biofilms; some species cause disease in humans (periodontitis, ulcers)
Leptospira Spiral-shaped bacterium (spirochetes); gram negative-like (better viewed by darkfield microscopy); very thin Aerobic, abundant in shallow water reservoirs; infect rodents and domestic animals; can be transmitted to humans by infected animals’ urine; may cause severe disease
Sphingobacterium Gram-negative bacillus Oxidase positive; nonmotile; contain high amounts of sphingophospholipids; rarely cause disease in humans
Treponema Gram-negative-like spirochete; very thin; better viewed by darkfield microscopy Motile; do not grow in culture; T. pallidum (subspecies T. pallidum pallidum) causes syphilis
Exercise \(3\)
How do Planctomycetes reproduce?
Phototrophic Bacteria
The phototrophic bacteria are a large and diverse category of bacteria that do not represent a taxon but, rather, a group of bacteria that use sunlight as their primary source of energy. This group contains both Proteobacteria and nonproteobacteria. They use solar energy to synthesize ATP through photosynthesis. When they produce oxygen, they perform oxygenic photosynthesis. When they do not produce oxygen, they perform anoxygenic photosynthesis. With the exception of some cyanobacteria, the majority of phototrophic bacteria perform anoxygenic photosynthesis.
One large group of phototrophic bacteria includes the purple or green bacteria that perform photosynthesis with the help of bacteriochlorophylls, which are green, purple, or blue pigments similar to chlorophyll in plants. Some of these bacteria have a varying amount of red or orange pigments called carotenoids. Their color varies from orange to red to purple to green (Figure \(4\)), and they are able to absorb light of various wavelengths. Traditionally, these bacteria are classified into sulfur and nonsulfur bacteria; they are further differentiated by color.
The sulfur bacteria perform anoxygenic photosynthesis, using sulfites as electron donors and releasing free elemental sulfur. Nonsulfur bacteria use organic substrates, such as succinate and malate, as donors of electrons.
The purple sulfur bacteria oxidize hydrogen sulfide into elemental sulfur and sulfuric acid and get their purple color from the pigments bacteriochlorophylls and carotenoids. Bacteria of the genus Chromatium are purple sulfur Gammaproteobacteria. These microorganisms are strict anaerobes and live in water. They use carbon dioxide as their only source of carbon, but their survival and growth are possible only in the presence of sulfites, which they use as electron donors. Chromatium has been used as a model for studies of bacterial photosynthesis since the 1950s.1
The green sulfur bacteria use sulfide for oxidation and produce large amounts of green bacteriochlorophyll. The genus Chlorobium is a green sulfur bacterium that is implicated in climate change because it produces methane, a greenhouse gas. These bacteria use at least four types of chlorophyll for photosynthesis. The most prevalent of these, bacteriochlorophyll, is stored in special vesicle-like organelles called chlorosomes.
Purple nonsulfur bacteria are similar to purple sulfur bacteria, except that they use hydrogen rather than hydrogen sulfide for oxidation. Among the purple nonsulfur bacteria is the genus Rhodospirillum. These microorganisms are facultative anaerobes, which are actually pink rather than purple, and can metabolize (“fix”) nitrogen. They may be valuable in the field of biotechnology because of their potential ability to produce biological plastic and hydrogen fuel.2
The green nonsulfur bacteria are similar to green sulfur bacteria but they use substrates other than sulfides for oxidation. Chloroflexus is an example of a green nonsulfur bacterium. It often has an orange color when it grows in the dark, but it becomes green when it grows in sunlight. It stores bacteriochlorophyll in chlorosomes, similar to Chlorobium, and performs anoxygenic photosynthesis, using organic sulfites (low concentrations) or molecular hydrogen as electron donors, so it can survive in the dark if oxygen is available. Chloroflexus does not have flagella but can glide, like Cytophaga. It grows at a wide range of temperatures, from 35 °C to 70 °C, thus can be thermophilic.
Another large, diverse group of phototrophic bacteria compose the phylum Cyanobacteria; they get their blue-green color from the chlorophyll contained in their cells (Figure \(5\)). Species of this group perform oxygenic photosynthesis, producing megatons of gaseous oxygen. Scientists hypothesize that cyanobacteria played a critical role in the change of our planet’s anoxic atmosphere 1–2 billion years ago to the oxygen-rich environment we have today.3
Cyanobacteria have other remarkable properties. Amazingly adaptable, they thrive in many habitats, including marine and freshwater environments, soil, and even rocks. They can live at a wide range of temperatures, even in the extreme temperatures of the Antarctic. They can live as unicellular organisms or in colonies, and they can be filamentous, forming sheaths or biofilms. Many of them fix nitrogen, converting molecular nitrogen into nitrites and nitrates that other bacteria, plants, and animals can use. The reactions of nitrogen fixation occur in specialized cells called heterocysts.
Photosynthesis in Cyanobacteria is oxygenic, using the same type of chlorophyll a found in plants and algae as the primary photosynthetic pigment. Cyanobacteria also use phycocyanin and cyanophycin, two secondary photosynthetic pigments that give them their characteristic blue color. They are located in special organelles called phycobilisomes and in folds of the cellular membrane called thylakoids, which are remarkably similar to the photosynthetic apparatus of plants. Scientists hypothesize that plants originated from endosymbiosis of ancestral eukaryotic cells and ancestral photosynthetic bacteria.4 Cyanobacteria are also an interesting object of research in biochemistry,5 with studies investigating their potential as biosorbents6 and products of human nutrition.7
Unfortunately, cyanobacteria can sometimes have a negative impact on human health. Genera such as Microcystis can form harmful cyanobacterial blooms, forming dense mats on bodies of water and producing large quantities of toxins that can harm wildlife and humans. These toxins have been implicated in tumors of the liver and diseases of the nervous system in animals and humans.8
Table \(2\) summarizes the characteristics of important phototrophic bacteria. Table \(2\): Phototrophic Bacteria
Phylum Class Example Genus or Species Common Name Oxygenic or Anoxygenic Sulfur Deposition
Cyanobacteria Cyanophyceae Microcystisaeruginosa Blue-green bacteria Oxygenic None
Chlorobi Chlorobia Chlorobium Green sulfur bacteria Anoxygenic Outside the cell
Chloroflexi(Division) Chloroflexi Chloroflexus Green nonsulfur bacteria Anoxygenic None
Proteobacteria Alphaproteobacteria Rhodospirillum Purple nonsulfur bacteria Anoxygenic None
Betaproteobacteria Rhodocyclus Purple nonsulfur bacteria Anoxygenic None
Gammaproteobacteria Chromatium Purple sulfur bacteria Anoxygenic Inside the cell
Exercise \(4\)
What characteristic makes phototrophic bacteria different from other prokaryotes?
Summary
• Gram-negative nonproteobacteria include the taxa spirochetes; the Cytophaga, Fusobacterium, Bacteroides group; Planctomycetes; and many representatives of phototrophic bacteria.
• Spirochetes are motile, spiral bacteria with a long, narrow body; they are difficult or impossible to culture.
• Several genera of spirochetes contain human pathogens that cause such diseases as syphilis and Lyme disease.
• Cytophaga, Fusobacterium, and Bacteroides are classified together as a phylum called the CFB group. They are rod-shaped anaerobic organoheterotrophs and avid fermenters. Cytophaga are aquatic bacteria with the gliding motility. Fusobacteria inhabit the human mouth and may cause severe infectious diseases. Bacteroides are present in vast numbers in the human gut, most of them being mutualistic but some are pathogenic.
• Planctomycetes are aquatic bacteria that reproduce by budding; they may form large colonies, and develop a holdfast.
• Phototrophic bacteria are not a taxon but, rather, a group categorized by their ability to use the energy of sunlight. They include Proteobacteria and nonproteobacteria, as well as sulfur and nonsulfur bacteria colored purple or green.
• Sulfur bacteria perform anoxygenic photosynthesis, using sulfur compounds as donors of electrons, whereas nonsulfur bacteria use organic compounds (succinate, malate) as donors of electrons.
• Some phototrophic bacteria are able to fix nitrogen, providing the usable forms of nitrogen to other organisms.
• Cyanobacteria are oxygen-producing bacteria thought to have played a critical role in the forming of the earth’s atmosphere.
Footnotes
1. 1 R.C. Fuller et al. “Carbon Metabolism in Chromatium.” Journal of Biological Chemistry 236 (1961):2140–2149.
2. 2 T.T. Selao et al. “Comparative Proteomic Studies in Rhodospirillum rubrum Grown Under Different Nitrogen Conditions.” Journal of Proteome Research 7 no. 8 (2008):3267–3275.
3. 3 A. De los Rios et al. “Ultrastructural and Genetic Characteristics of Endolithic Cyanobacterial Biofilms Colonizing Antarctic Granite Rocks.” FEMS Microbiology Ecology 59 no. 2 (2007):386–395.
4. 4 T. Cavalier-Smith. “Membrane Heredity and Early Chloroplast Evolution.” Trends in Plant Science 5 no. 4 (2000):174–182.
5. 5 S. Zhang, D.A. Bryant. “The Tricarboxylic Acid Cycle in Cyanobacteria.” Science 334 no. 6062 (2011):1551–1553.
6. 6 A. Cain et al. “Cyanobacteria as a Biosorbent for Mercuric Ion.” Bioresource Technology 99 no. 14 (2008):6578–6586.
7. 7 C.S. Ku et al. “Edible Blue-Green Algae Reduce the Production of Pro-Inflammatory Cytokines by Inhibiting NF-κB Pathway in Macrophages and Splenocytes.” Biochimica et Biophysica Acta 1830 no. 4 (2013):2981–2988.
8. 8 I. Stewart et al. Cyanobacterial Poisoning in Livestock, Wild Mammals and Birds – an Overview. Advances in Experimental Medicine and Biology 619 (2008):613–637. | textbooks/bio/Microbiology/Microbiology_(OpenStax)/04%3A_Prokaryotic_Diversity/4.03%3A_Nonproteobacteria_Gram-negative_Bacteria_and_Phototrophic_Bacteria.txt |
Learning Objectives
• Describe the unique features of each category of high G+C and low G+C gram-positive bacteria
• Identify similarities and differences between high G+C and low G+C bacterial groups
• Give an example of a bacterium of high G+C and low G+C group commonly associated with each category
Prokaryotes are identified as gram-positive if they have a multiple layer matrix of peptidoglycan forming the cell wall. Crystal violet, the primary stain of the Gram stain procedure, is readily retained and stabilized within this matrix, causing gram-positive prokaryotes to appear purple under a brightfield microscope after Gram staining. For many years, the retention of Gram stain was one of the main criteria used to classify prokaryotes, even though some prokaryotes did not readily stain with either the primary or secondary stains used in the Gram stain procedure.
Advances in nucleic acid biochemistry have revealed additional characteristics that can be used to classify gram-positive prokaryotes, namely the guanine to cytosine ratios (G+C) in DNA and the composition of 16S rRNA subunits. Microbiologists currently recognize two distinct groups of gram-positive, or weakly staining gram-positive, prokaryotes. The class Actinobacteria comprises the high G+C gram-positive bacteria, which have more than 50% guanine and cytosine nucleotides in their DNA. The class Bacilli comprises low G+C gram-positive bacteria, which have less than 50% of guanine and cytosine nucleotides in their DNA.
Actinobacteria: High G+C Gram-Positive Bacteria
The name Actinobacteria comes from the Greek words for rays and small rod, but Actinobacteria are very diverse. Their microscopic appearance can range from thin filamentous branching rods to coccobacilli. Some Actinobacteria are very large and complex, whereas others are among the smallest independently living organisms. Most Actinobacteria live in the soil, but some are aquatic. The vast majority are aerobic. One distinctive feature of this group is the presence of several different peptidoglycans in the cell wall.
The genus Actinomyces is a much studied representative of Actinobacteria. Actinomyces spp. play an important role in soil ecology, and some species are human pathogens. A number of Actinomyces spp. inhabit the human mouth and are opportunistic pathogens, causing infectious diseases like periodontitis (inflammation of the gums) and oral abscesses. The species A. israelii is an anaerobe notorious for causing endocarditis (inflammation of the inner lining of the heart) (Figure \(1\)).
The genus Mycobacterium is represented by bacilli covered with a mycolic acid coat. This waxy coat protects the bacteria from some antibiotics, prevents them from drying out, and blocks penetration by Gram stain reagents (see Staining Microscopic Specimens). Because of this, a special acid-fast staining procedure is used to visualize these bacteria. The genus Mycobacterium is an important cause of a diverse group of infectious diseases. M. tuberculosis is the causative agent of tuberculosis, a disease that primarily impacts the lungs but can infect other parts of the body as well. It has been estimated that one-third of the world’s population has been infected with M. tuberculosis and millions of new infections occur each year. Treatment of M. tuberculosis is challenging and requires patients to take a combination of drugs for an extended time. Complicating treatment even further is the development and spread of multidrug-resistant strains of this pathogen.
Another pathogenic species, M. leprae, is the cause of Hansen’s disease (leprosy), a chronic disease that impacts peripheral nerves and the integrity of the skin and mucosal surface of the respiratory tract. Loss of pain sensation and the presence of skin lesions increase susceptibility to secondary injuries and infections with other pathogens.
Bacteria in the genus Corynebacterium contain diaminopimelic acid in their cell walls, and microscopically often form palisades, or pairs of rod-shaped cells resembling the letter V. Cells may contain metachromatic granules, intracellular storage of inorganic phosphates that are useful for identification of Corynebacterium. The vast majority of Corynebacterium spp. are nonpathogenic; however, C. diphtheria is the causative agent of diphtheria, a disease that can be fatal, especially in children (Figure \(1\)). C. diphtheria produces a toxin that forms a pseudomembrane in the patient’s throat, causing swelling, difficulty breathing, and other symptoms that can become serious if untreated.
The genus Bifidobacterium consists of filamentous anaerobes, many of which are commonly found in the gastrointestinal tract, vagina, and mouth. In fact, Bifidobacterium spp. constitute a substantial part of the human gut microbiota and are frequently used as probiotics and in yogurt production.
The genus Gardnerella, contains only one species, G. vaginalis. This species is defined as “gram-variable” because its small coccobacilli do not show consistent results when Gram stained (Figure \(1\)). Based on its genome, it is placed into the high G+C gram-positive group. G. vaginalis can cause bacterial vaginosis in women; symptoms are typically mild or even undetectable, but can lead to complications during pregnancy.
Table \(1\) summarizes the characteristics of some important genera of Actinobacteria.
Table \(1\): Actinobacteria: High G+C Gram-Positive
Example Genus Microscopic Morphology Unique Characteristics
Actinomyces Gram-positive bacillus; in colonies, shows fungus-like threads (hyphae) Facultative anaerobes; in soil, decompose organic matter; in the human mouth, may cause gum disease
Arthrobacter Gram-positive bacillus (at the exponential stage of growth) or coccus (in stationary phase) Obligate aerobes; divide by “snapping,” forming V-like pairs of daughter cells; degrade phenol, can be used in bioremediation
Bifidobacterium Gram-positive, filamentous actinobacterium Anaerobes commonly found in human gut microbiota
Corynebacterium Gram-positive bacillus Aerobes or facultative anaerobes; form palisades; grow slowly; require enriched media in culture; C. diphtheriae causes diphtheria
Frankia Gram-positive, fungus-like (filamentous) bacillus Nitrogen-fixing bacteria; live in symbiosis with legumes
Gardnerella Gram-variable coccobacillus Colonize the human vagina, may alter the microbial ecology, thus leading to vaginosis
Micrococcus Gram-positive coccus, form microscopic clusters Ubiquitous in the environment and on the human skin; oxidase-positive (as opposed to morphologically similar S. aureus); some are opportunistic pathogens
Mycobacterium Gram-positive, acid-fast bacillus Slow growing, aerobic, resistant to drying and phagocytosis; covered with a waxy coat made of mycolic acid; M. tuberculosis causes tuberculosis; M. leprae causes leprosy
Nocardia Weakly gram-positive bacillus; forms acid-fast branches May colonize the human gingiva; may cause severe pneumonia and inflammation of the skin
Propionibacterium Gram-positive bacillus Aerotolerant anaerobe; slow-growing; P. acnes reproduces in the human sebaceous glands and may cause or contribute to acne
Rhodococcus Gram-positive bacillus Strict aerobe; used in industry for biodegradation of pollutants; R. fascians is a plant pathogen, and R. equi causes pneumonia in foals
Streptomyces Gram-positive, fungus-like (filamentous) bacillus Very diverse genus (>500 species); aerobic, spore-forming bacteria; scavengers, decomposers found in soil (give the soil its “earthy” odor); used in pharmaceutical industry as antibiotic producers (more than two-thirds of clinically useful antibiotics)
Exercise \(1\)
What is one distinctive feature of Actinobacteria?
Low G+C Gram-positive Bacteria
The low G+C gram-positive bacteria have less than 50% guanine and cytosine in their DNA, and this group of bacteria includes a number of genera of bacteria that are pathogenic.
Clinical Focus: Part 3
Based on her symptoms, Marsha’s doctor suspected that she had a case of tuberculosis. Although less common in the United States, tuberculosis is still extremely common in many parts of the world, including Nigeria. Marsha’s work there in a medical lab likely exposed her to Mycobacterium tuberculosis, the bacterium that causes tuberculosis.
Marsha’s doctor ordered her to stay at home, wear a respiratory mask, and confine herself to one room as much as possible. He also said that Marsha had to take one semester off school. He prescribed isoniazid and rifampin, antibiotics used in a drug cocktail to treat tuberculosis, which Marsha was to take three times a day for at least three months.
Exercise \(2\)
Why did the doctor order Marsha to stay home for three months?
Clostridia
One large and diverse class of low G+C gram-positive bacteria is Clostridia. The best studied genus of this class is Clostridium. These rod-shaped bacteria are generally obligate anaerobes that produce endospores and can be found in anaerobic habitats like soil and aquatic sediments rich in organic nutrients. The endospores may survive for many years.
Clostridium spp. produce more kinds of protein toxins than any other bacterial genus, and several species are human pathogens. C. perfringens is the third most common cause of food poisoning in the United States and is the causative agent of an even more serious disease called gas gangrene. Gas gangrene occurs when C. perfringens endospores enter a wound and germinate, becoming viable bacterial cells and producing a toxin that can cause the necrosis (death) of tissue. C. tetani, which causes tetanus, produces a neurotoxin that is able to enter neurons, travel to regions of the central nervous system where it blocks the inhibition of nerve impulses involved in muscle contractions, and cause a life-threatening spastic paralysis. C. botulinum produces botulinum neurotoxin, the most lethal biological toxin known. Botulinum toxin is responsible for rare but frequently fatal cases of botulism. The toxin blocks the release of acetylcholine in neuromuscular junctions, causing flaccid paralysis. In very small concentrations, botulinum toxin has been used to treat muscle pathologies in humans and in a cosmetic procedure to eliminate wrinkles. C. difficile is a common source of hospital-acquired infections (Figure \(2\)) that can result in serious and even fatal cases of colitis (inflammation of the large intestine). Infections often occur in patients who are immunosuppressed or undergoing antibiotic therapy that alters the normal microbiota of the gastrointestinal tract.
Lactobacillales
The order Lactobacillales comprises low G+C gram-positive bacteria that include both bacilli and cocci in the genera Lactobacillus, Leuconostoc, Enterococcus, and Streptococcus. Bacteria of the latter three genera typically are spherical or ovoid and often form chains.
Streptococcus, the name of which comes from the Greek word for twisted chain, is responsible for many types of infectious diseases in humans. Species from this genus, often referred to as streptococci, are usually classified by serotypes called Lancefield groups, and by their ability to lyse red blood cells when grown on blood agar.
S. pyogenes belongs to the Lancefield group A, β-hemolytic Streptococcus. This species is considered a pyogenic pathogen because of the associated pus production observed with infections it causes (Figure \(3\)). S. pyogenes is the most common cause of bacterial pharyngitis (strep throat); it is also an important cause of various skin infections that can be relatively mild (e.g., impetigo) or life threatening (e.g., necrotizing fasciitis, also known as flesh eating disease), life threatening.
The nonpyogenic (i.e., not associated with pus production) streptococci are a group of streptococcal species that are not a taxon but are grouped together because they inhabit the human mouth. The nonpyogenic streptococci do not belong to any of the Lancefield groups. Most are commensals, but a few, such as S. mutans, are implicated in the development of dental caries.
S. pneumoniae (commonly referred to as pneumococcus), is a Streptococcus species that also does not belong to any Lancefield group. S. pneumoniae cells appear microscopically as diplococci, pairs of cells, rather than the long chains typical of most streptococci. Scientists have known since the 19th century that S. pneumoniae causes pneumonia and other respiratory infections. However, this bacterium can also cause a wide range of other diseases, including meningitis, septicemia, osteomyelitis, and endocarditis, especially in newborns, the elderly, and patients with immunodeficiency.
Bacilli
The name of the class Bacilli suggests that it is made up of bacteria that are bacillus in shape, but it is a morphologically diverse class that includes bacillus-shaped and cocccus-shaped genera. Among the many genera in this class are two that are very important clinically: Bacillus and Staphylococcus.
Bacteria in the genus Bacillus are bacillus in shape and can produce endospores. They include aerobes or facultative anaerobes. A number of Bacillus spp. are used in various industries, including the production of antibiotics (e.g., barnase), enzymes (e.g., alpha-amylase, BamH1 restriction endonuclease), and detergents (e.g., subtilisin).
Two notable pathogens belong to the genus Bacillus. B. anthracis is the pathogen that causes anthrax, a severe disease that affects wild and domesticated animals and can spread from infected animals to humans. Anthrax manifests in humans as charcoal-black ulcers on the skin, severe enterocolitis, pneumonia, and brain damage due to swelling. If untreated, anthrax is lethal. B. cereus, a closely related species, is a pathogen that may cause food poisoning. It is a rod-shaped species that forms chains. Colonies appear milky white with irregular shapes when cultured on blood agar (Figure \(4\)). One other important species is B. thuringiensis. This bacterium produces a number of substances used as insecticides because they are toxic for insects.
The genus Staphylococcus also belongs to the class Bacilli, even though its shape is coccus rather than a bacillus. The name Staphylococcus comes from a Greek word for bunches of grapes, which describes their microscopic appearance in culture (Figure \(5\)). Staphylococcus spp. are facultative anaerobic, halophilic, and nonmotile. The two best-studied species of this genus are S. epidermidis and S. aureus.
S. epidermidis, whose main habitat is the human skin, is thought to be nonpathogenic for humans with healthy immune systems, but in patients with immunodeficiency, it may cause infections in skin wounds and prostheses (e.g., artificial joints, heart valves). S. epidermidis is also an important cause of infections associated with intravenous catheters. This makes it a dangerous pathogen in hospital settings, where many patients may be immunocompromised.
Strains of S. aureus cause a wide variety of infections in humans, including skin infections that produce boils, carbuncles, cellulitis, or impetigo. Certain strains of S. aureus produce a substance called enterotoxin, which can cause severe enteritis, often called staph food poisoning. Some strains of S. aureus produce the toxin responsible for toxic shock syndrome, which can result in cardiovascular collapse and death.
Many strains of S. aureus have developed resistance to antibiotics. Some antibiotic-resistant strains are designated as methicillin-resistant S. aureus (MRSA) and vancomycin-resistant S. aureus (VRSA). These strains are some of the most difficult to treat because they exhibit resistance to nearly all available antibiotics, not just methicillin and vancomycin. Because they are difficult to treat with antibiotics, infections can be lethal. MRSA and VRSA are also contagious, posing a serious threat in hospitals, nursing homes, dialysis facilities, and other places where there are large populations of elderly, bedridden, and/or immunocompromised patients.
Mycoplasmas
Although Mycoplasma spp. do not possess a cell wall and, therefore, are not stained by Gram-stain reagents, this genus is still included with the low G+C gram-positive bacteria. The genus Mycoplasma includes more than 100 species, which share several unique characteristics. They are very small cells, some with a diameter of about 0.2 μm, which is smaller than some large viruses. They have no cell walls and, therefore, are pleomorphic, meaning that they may take on a variety of shapes and can even resemble very small animal cells. Because they lack a characteristic shape, they can be difficult to identify. One species, M. pneumoniae, causes the mild form of pneumonia known as “walking pneumonia” or “atypical pneumonia.” This form of pneumonia is typically less severe than forms caused by other bacteria or viruses.
Table \(2\) summarizes the characteristics of notable genera low G+C Gram-positive bacteria. Table \(2\): Bacilli - Low G+C Gram-Positive Bacteria
Example Genus Microscopic Morphology Unique Characteristics
Bacillus Large, gram-positive bacillus Aerobes or facultative anaerobes; form endospores; B. anthracis causes anthrax in cattle and humans, B. cereus may cause food poisoning
Clostridium Gram-positive bacillus Strict anaerobes; form endospores; all known species are pathogenic, causing tetanus, gas gangrene, botulism, and colitis
Enterococcus Gram-positive coccus; forms microscopic pairs in culture (resembling Streptococcus pneumoniae) Anaerobic aerotolerant bacteria, abundant in the human gut, may cause urinary tract and other infections in the nosocomial environment
Lactobacillus Gram-positive bacillus Facultative anaerobes; ferment sugars into lactic acid; part of the vaginal microbiota; used as probiotics
Leuconostoc Gram-positive coccus; may form microscopic chains in culture Fermenter, used in food industry to produce sauerkraut and kefir
Mycoplasma The smallest bacteria; appear pleomorphic under electron microscope Have no cell wall; classified as low G+C Gram-positive bacteria because of their genome; M. pneumoniae causes “walking” pneumonia
Staphylococcus Gram-positive coccus; forms microscopic clusters in culture that resemble bunches of grapes Tolerate high salt concentration; facultative anaerobes; produce catalase; S. aureus can also produce coagulase and toxins responsible for local (skin) and generalized infections
Streptococcus Gram-positive coccus; forms chains or pairs in culture Diverse genus; classified into groups based on sharing certain antigens; some species cause hemolysis and may produce toxins responsible for human local (throat) and generalized disease
Ureaplasma Similar to Mycoplasma Part of the human vaginal and lower urinary tract microbiota; may cause inflammation, sometimes leading to internal scarring and infertility
Exercise \(3\)
1. Name some ways in which streptococci are classified.
2. Name one pathogenic low G+C gram-positive bacterium and a disease it causes.
Clinical Focus: Resolution
Marsha’s sputum sample was sent to the microbiology lab to confirm the identity of the microorganism causing her infection. The lab also performed antimicrobial susceptibility testing (AST) on the sample to confirm that the physician has prescribed the correct antimicrobial drugs.
Direct microscopic examination of the sputum revealed acid-fast bacteria (AFB) present in Marsha’s sputum. When placed in culture, there were no signs of growth for the first 8 days, suggesting that microorganism was either dead or growing very slowly. Slow growth is a distinctive characteristic of M. tuberculosis.
After four weeks, the lab microbiologist observed distinctive colorless granulated colonies (Figure \(6\)). The colonies contained AFB showing the same microscopic characteristics as those revealed during the direct microscopic examination of Marsha’s sputum. To confirm the identification of the AFB, samples of the colonies were analyzed using nucleic acid hybridization, or direct nucleic acid amplification (NAA) testing. When a bacterium is acid-fast, it is classified in the family Mycobacteriaceae. DNA sequencing of variable genomic regions of the DNA extracted from these bacteria revealed that it was high G+C. This fact served to finalize Marsha’s diagnosis as infection with M. tuberculosis. After nine months of treatment with the drugs prescribed by her doctor, Marsha made a full recovery.
Biopiracy and Bioprospecting
In 1969, an employee of a Swiss pharmaceutical company was vacationing in Norway and decided to collect some soil samples. He took them back to his lab, and the Swiss company subsequently used the fungus Tolypocladium inflatum in those samples to develop cyclosporine A, a drug widely used in patients who undergo tissue or organ transplantation. The Swiss company earns more than \$1 billion a year for production of cyclosporine A, yet Norway receives nothing in return—no payment to the government or benefit for the Norwegian people. Despite the fact the cyclosporine A saves numerous lives, many consider the means by which the soil samples were obtained to be an act of “biopiracy,” essentially a form of theft. Do the ends justify the means in a case like this?
Nature is full of as-yet-undiscovered bacteria and other microorganisms that could one day be used to develop new life-saving drugs or treatments.1 Pharmaceutical and biotechnology companies stand to reap huge profits from such discoveries, but ethical questions remain. To whom do biological resources belong? Should companies who invest (and risk) millions of dollars in research and development be required to share revenue or royalties for the right to access biological resources?
Compensation is not the only issue when it comes to bioprospecting. Some communities and cultures are philosophically opposed to bioprospecting, fearing unforeseen consequences of collecting genetic or biological material. Native Hawaiians, for example, are very protective of their unique biological resources.
For many years, it was unclear what rights government agencies, private corporations, and citizens had when it came to collecting samples of microorganisms from public land. Then, in 1993, the Convention on Biological Diversity granted each nation the rights to any genetic and biological material found on their own land. Scientists can no longer collect samples without a prior arrangement with the land owner for compensation. This convention now ensures that companies act ethically in obtaining the samples they use to create their products.
Summary
• Gram-positive bacteria are a very large and diverse group of microorganisms. Understanding their taxonomy and knowing their unique features is important for diagnostics and treatment of infectious diseases.
• Gram-positive bacteria are classified into high G+C gram-positive and low G+C gram-positive bacteria, based on the prevalence of guanine and cytosine nucleotides in their genome
• Actinobacteria is the taxonomic name of the class of high G+C gram-positive bacteria. This class includes the genera Actinomyces, Arthrobacter, Corynebacterium, Frankia, Gardnerella, Micrococcus, Mycobacterium, Nocardia, Propionibacterium, Rhodococcus, and Streptomyces. Some representatives of these genera are used in industry; others are human or animal pathogens.
• Examples of high G+C gram-positive bacteria that are human pathogens include Mycobacterium tuberculosis, which causes tuberculosis; M. leprae, which causes leprosy (Hansen’s disease); and Corynebacteriumdiphtheriae, which causes diphtheria.
• Clostridia spp. are low G+C gram-positive bacteria that are generally obligate anaerobes and can form endospores. Pathogens in this genus include C. perfringens (gas gangrene), C. tetani (tetanus), and C. botulinum (botulism).
• Lactobacillales include the genera Enterococcus, Lactobacillus, Leuconostoc, and Streptococcus. Streptococcus is responsible for many human diseases, including pharyngitis (strep throat), scarlet fever, rheumatic fever, glomerulonephritis, pneumonia, and other respiratory infections.
• Bacilli is a taxonomic class of low G+C gram-positive bacteria that include rod-shaped and coccus-shaped species, including the genera Bacillus and Staphylococcus. B. anthracis causes anthrax, B. cereus may cause opportunistic infections of the gastrointestinal tract, and S. aureus strains can cause a wide range of infections and diseases, many of which are highly resistant to antibiotics.
• Mycoplasma spp. are very small, pleomorphic low G+C gram-positive bacteria that lack cell walls. M. pneumoniae causes atypical pneumonia.
Footnotes
1. 1 J. Andre. Bioethics as Practice. Chapel Hill, NC: University of North Carolina Press, 2002. | textbooks/bio/Microbiology/Microbiology_(OpenStax)/04%3A_Prokaryotic_Diversity/4.04%3A_Gram-positive_Bacteria.txt |
Learning Objectives
• Describe the unique features of deeply branching bacteria
• Give examples of significant deeply branching bacteria
On a phylogenetic tree (see A Systematic Approach), the trunk or root of the tree represents a common ancient evolutionary ancestor, often called the last universal common ancestor (LUCA), and the branches are its evolutionary descendants. Scientists consider the deeply branching bacteria, such as the genus Acetothermus, to be the first of these non-LUCA forms of life produced by evolution some 3.5 billion years ago. When placed on the phylogenetic tree, they stem from the common root of life, deep and close to the LUCA root—hence the name “deeply branching” (Figure \(1\)).
The deeply branching bacteria may provide clues regarding the structure and function of ancient and now extinct forms of life. We can hypothesize that ancient bacteria, like the deeply branching bacteria that still exist, were thermophiles or hyperthermophiles, meaning that they thrived at very high temperatures. Acetothermus paucivorans, a gram-negative anaerobic bacterium discovered in 1988 in sewage sludge, is a thermophile growing at an optimal temperature of 58 °C.1 Scientists have determined it to be the deepest branching bacterium, or the closest evolutionary relative of the LUCA (Figure \(1\)).
The class Aquificae includes deeply branching bacteria that are adapted to the harshest conditions on our planet, resembling the conditions thought to dominate the earth when life first appeared. Bacteria from the genus Aquifex are hyperthermophiles, living in hot springs at a temperature higher than 90 °C. The species A. pyrophilus thrives near underwater volcanoes and thermal ocean vents, where the temperature of water (under high pressure) can reach 138 °C. Aquifex bacteria use inorganic substances as nutrients. For example, A. pyrophilus can reduce oxygen, and it is able to reduce nitrogen in anaerobic conditions. They also show a remarkable resistance to ultraviolet light and ionizing radiation. Taken together, these observations support the hypothesis that the ancient ancestors of deeply branching bacteria began evolving more than 3 billion years ago, when the earth was hot and lacked an atmosphere, exposing the bacteria to nonionizing and ionizing radiation.
The class Thermotogae is represented mostly by hyperthermophilic, as well as some mesophilic (preferring moderate temperatures), anaerobic gram-negative bacteria whose cells are wrapped in a peculiar sheath-like outer membrane called a toga. The thin layer of peptidoglycan in their cell wall has an unusual structure; it contains diaminopimelic acid and D-lysine. These bacteria are able to use a variety of organic substrates and produce molecular hydrogen, which can be used in industry. The class contains several genera, of which the best known is the genus Thermotoga. One species of this genus, T. maritima, lives near the thermal ocean vents and thrives in temperatures of 90 °C; another species, T. subterranea, lives in underground oil reservoirs.
Finally, the deeply branching bacterium Deinococcus radiodurans belongs to a genus whose name is derived from a Greek word meaning terrible berry. Nicknamed “Conan the Bacterium,” D. radiodurans is considered a polyextremophile because of its ability to survive under the many different kinds of extreme conditions—extreme heat, drought, vacuum, acidity, and radiation. It owes its name to its ability to withstand doses of ionizing radiation that kill all other known bacteria; this special ability is attributed to some unique mechanisms of DNA repair.
Summary
• Deeply branching bacteria are phylogenetically the most ancient forms of life, being the closest to the last universal common ancestor.
• Deeply branching bacteria include many species that thrive in extreme environments that are thought to resemble conditions on earth billions of years ago
• Deeply branching bacteria are important for our understanding of evolution; some of them are used in industry
Footnotes
1. 1 G. Dietrich et al. “Acetothermus paucivorans, gen. nov., sp. Nov., a Strictly Anaerobic, Thermophilic Bacterium From Sewage Sludge, Fermenting Hexoses to Acetate, CO2, and H2.” Systematic and Applied Microbiology 10 no. 2 (1988):174–179. | textbooks/bio/Microbiology/Microbiology_(OpenStax)/04%3A_Prokaryotic_Diversity/4.05%3A_Deeply_Branching_Bacteria.txt |
Learning Objectives
• Describe the unique features of each category of Archaea
• Explain why archaea might not be associated with human microbiomes or pathology
• Give common examples of archaea commonly associated with unique environmental habitats
Like organisms in the domain Bacteria, organisms of the domain Archaea are all unicellular organisms. However, archaea differ structurally from bacteria in several significant ways, as discussed in Unique Characteristics of Prokaryotic Cells. To summarize:
• The archaeal cell membrane is composed of ether linkages with branched isoprene chains (as opposed to the bacterial cell membrane, which has ester linkages with unbranched fatty acids).
• Archaeal cell walls lack peptidoglycan, but some contain a structurally similar substance called pseudopeptidoglycan or pseudomurein.
• The genomes of Archaea are larger and more complex than those of bacteria.
Domain Archaea is as diverse as domain Bacteria, and its representatives can be found in any habitat. Some archaea are mesophiles, and many are extremophiles, preferring extreme hot or cold, extreme salinity, or other conditions that are hostile to most other forms of life on earth. Their metabolism is adapted to the harsh environments, and they can perform methanogenesis, for example, which bacteria and eukaryotes cannot.
The size and complexity of the archaeal genome makes it difficult to classify. Most taxonomists agree that within the Archaea, there are currently five major phyla: Crenarchaeota, Euryarchaeota, Korarchaeota, Nanoarchaeota, and Thaumarchaeota. There are likely many other archaeal groups that have not yet been systematically studied and classified.
With few exceptions, archaea are not present in the human microbiota, and none are currently known to be associated with infectious diseases in humans, animals, plants, or microorganisms. However, many play important roles in the environment and may thus have an indirect impact on human health.
Crenarchaeota
Crenarchaeota is a class of Archaea that is extremely diverse, containing genera and species that differ vastly in their morphology and requirements for growth. All Crenarchaeota are aquatic organisms, and they are thought to be the most abundant microorganisms in the oceans. Most, but not all, Crenarchaeota are hyperthermophiles; some of them (notably, the genus Pyrolobus) are able to grow at temperatures up to 113 °C.1
Archaea of the genus Sulfolobus (Figure \(1\)) are thermophiles that prefer temperatures around 70–80°C and acidophiles that prefer a pH of 2–3.2 Sulfolobus can live in aerobic or anaerobic environments. In the presence of oxygen, Sulfolobus spp. use metabolic processes similar to those of heterotrophs. In anaerobic environments, they oxidize sulfur to produce sulfuric acid, which is stored in granules. Sulfolobus spp. are used in biotechnology for the production of thermostable and acid-resistant proteins called affitins.3 Affitins can bind and neutralize various antigens (molecules found in toxins or infectious agents that provoke an immune response from the body).
Another genus, Thermoproteus, is represented by strictly anaerobic organisms with an optimal growth temperature of 85 °C. They have flagella and, therefore, are motile. Thermoproteus has a cellular membrane in which lipids form a monolayer rather than a bilayer, which is typical for archaea. Its metabolism is autotrophic. To synthesize ATP, Thermoproteus spp. reduce sulfur or molecular hydrogen and use carbon dioxide or carbon monoxide as a source of carbon. Thermoproteus is thought to be the deepest-branching genus of Archaea, and thus is a living example of some of our planet’s earliest forms of life.
Exercise \(1\)
What types of environments do Crenarchaeota prefer?
Euryarchaeota
The phylum Euryarchaeota includes several distinct classes. Species in the classes Methanobacteria, Methanococci, and Methanomicrobia represent Archaea that can be generally described as methanogens. Methanogens are unique in that they can reduce carbon dioxide in the presence of hydrogen, producing methane. They can live in the most extreme environments and can reproduce at temperatures varying from below freezing to boiling. Methanogens have been found in hot springs as well as deep under ice in Greenland. Some scientists have even hypothesized that methanogens may inhabit the planet Mars because the mixture of gases produced by methanogens resembles the makeup of the Martian atmosphere.4
Methanogens are thought to contribute to the formation of anoxic sediments by producing hydrogen sulfide, making “marsh gas.” They also produce gases in ruminants and humans. Some genera of methanogens, notably Methanosarcina, can grow and produce methane in the presence of oxygen, although the vast majority are strict anaerobes.
The class Halobacteria (which was named before scientists recognized the distinction between Archaea and Bacteria) includes halophilic (“salt-loving”) archaea. Halobacteria require a very high concentrations of sodium chloride in their aquatic environment. The required concentration is close to saturation, at 36%; such environments include the Dead Sea as well as some salty lakes in Antarctica and south-central Asia. One remarkable feature of these organisms is that they perform photosynthesis using the protein bacteriorhodopsin, which gives them, and the bodies of water they inhabit, a beautiful purple color (Figure \(2\)).
Notable species of Halobacteria include Halobacterium salinarum, which may be the oldest living organism on earth; scientists have isolated its DNA from fossils that are 250 million years old.5 Another species, Haloferax volcanii, shows a very sophisticated system of ion exchange, which enables it to balance the concentration of salts at high temperatures
Exercise \(2\)
Where do Halobacteria live?
Finding a Link Between Archaea and Disease
Archaea are not known to cause any disease in humans, animals, plants, bacteria, or in other archaea. Although this makes sense for the extremophiles, not all archaea live in extreme environments. Many genera and species of Archaea are mesophiles, so they can live in human and animal microbiomes, although they rarely do. As we have learned, some methanogens exist in the human gastrointestinal tract. Yet we have no reliable evidence pointing to any archaean as the causative agent of any human disease.
Still, scientists have attempted to find links between human disease and archaea. For example, in 2004, Lepp et al. presented evidence that an archaean called Methanobrevibacter oralis inhabits the gums of patients with periodontal disease. The authors suggested that the activity of these methanogens causes the disease.6 However, it was subsequently shown that there was no causal relationship between M. oralis and periodontitis. It seems more likely that periodontal disease causes an enlargement of anaerobic regions in the mouth that are subsequently populated by M. oralis.7
There remains no good answer as to why archaea do not seem to be pathogenic, but scientists continue to speculate and hope to find the answer.
Summary
• Archaea are unicellular, prokaryotic microorganisms that differ from bacteria in their genetics, biochemistry, and ecology.
• Some archaea are extremophiles, living in environments with extremely high or low temperatures, or extreme salinity.
• Only archaea are known to produce methane. Methane-producing archaea are called methanogens.
• Halophilic archaea prefer a concentration of salt close to saturation and perform photosynthesis using bacteriorhodopsin.
• Some archaea, based on fossil evidence, are among the oldest organisms on earth.
• Archaea do not live in great numbers in human microbiomes and are not known to cause disease.
Footnotes
1. E. Blochl et al.“Pyrolobus fumani, gen. and sp. nov., represents a novel group of Archaea, extending the upper temperature limit for life to 113°C.” Extremophiles 1 (1997):14–21.
2. T.D. Brock et al. “Sulfolobus: A New Genus of Sulfur-Oxidizing Bacteria Living at Low pH and High Temperature.” Archiv für Mikrobiologie 84 no. 1 (1972):54–68.
3. S. Pacheco et al. “Affinity Transfer to the Archaeal Extremophilic Sac7d Protein by Insertion of a CDR.” Protein Engineering Design and Selection 27 no. 10 (2014):431-438.
4. R.R. Britt “Crater Critters: Where Mars Microbes Might Lurk.” www.space.com/1880-crater-cri...obes-lurk.html. Accessed April 7, 2015.
5. H. Vreeland et al. “Fatty acid and DA Analyses of Permian Bacterium Isolated From Ancient Salt Crystals Reveal Differences With Their Modern Relatives.” Extremophiles 10 (2006):71–78.
6. P.W. Lepp et al. “Methanogenic Archaea and Human Gum Disease.” Proceedings of the National Academies of Science of the United States of America 101 no. 16 (2004):6176–6181.
7. R.I. Aminov. “Role of Archaea in Human Disease.” Frontiers in Cellular and Infection Microbiology 3 (2013):42. | textbooks/bio/Microbiology/Microbiology_(OpenStax)/04%3A_Prokaryotic_Diversity/4.06%3A_Archaea.txt |
4.1: Prokaryote Habitats, Relationships, and Microbiomes
Prokaryotes are unicellular microorganisms whose cells have no nucleus. Prokaryotes can be found everywhere on our planet, even in the most extreme environments. Prokaryotes are very flexible metabolically, so they are able to adjust their feeding to the available natural resources. Prokaryotes live in communities that interact among themselves and with large organisms that they use as hosts (including humans).
Multiple Choice
The term prokaryotes refers to which of the following?
1. very small organisms
2. unicellular organisms that have no nucleus
3. multicellular organisms
4. cells that resemble animal cells more than plant cells
Answer
B
The term microbiota refers to which of the following?
1. all microorganisms of the same species
2. all of the microorganisms involved in a symbiotic relationship
3. all microorganisms in a certain region of the human body
4. all microorganisms in a certain geographic region
Answer
C
Which of the following refers to the type of interaction between two prokaryotic populations in which one population benefits and the other is not affected?
1. mutualism
2. commensalism
3. parasitism
4. neutralism
Answer
B
True/False
Among prokaryotes, there are some that can live in every environment on earth.
Answer
True
Fill in the Blank
When prokaryotes live as interacting communities in which one population benefits to the harm of the other, the type of symbiosis is called ________.
Answer
parasitism
The domain ________ does not include prokaryotes.
Answer
Eukarya
Pathogenic bacteria that are part of the transient microbiota can sometimes be eliminated by ________ therapy.
Answer
antibiotic
Nitrogen-fixing bacteria provide other organisms with usable nitrogen in the form of ________.
Answer
ammonia
Short Answer
Compare commensalism and amensalism.
Give an example of the changes of human microbiota that result from medical intervention.
4.2: Proteobacteria
Proteobacteria is a phylum of gram-negative bacteria and are classified into the classes alpha-, beta-, gamma-, delta- and epsilonproteobacteria, each class having separate orders, families, genera, and species. Alphaproteobacteria are oligotrophs. The taxa chlamydias and rickettsias are obligate intracellular pathogens, feeding on cells of host organisms; they are metabolically inactive outside of the host cell. Some Alphaproteobacteria can convert atmospheric nitrogen to nitrites.
Multiple Choice
Which of the following describes Proteobacteria in domain Bacteria?
1. phylum
2. class
3. species
4. genus
Answer
A
All Alphaproteobacteria are which of the following?
1. oligotrophs
2. intracellular
3. pathogenic
4. all of the above
5. none of the above
Answer
A
Class Betaproteobacteria includes all but which of the following genera?
1. Neisseria.
2. Bordetella.
3. Leptothrix.
4. Campylobacter.
Answer
D
Haemophilus influenzae is a common cause of which of the following?
1. influenza
2. dysentery
3. upper respiratory tract infections
4. hemophilia
Answer
C
Fill in the Blank
Rickettsias are ________ intracellular bacteria.
Answer
obligate
The species ________, which belongs to Epsilonproteobacteria, causes peptic ulcers of the stomach and duodenum.
Answer
Helicobacter pylori
The genus Salmonella belongs to the class ________ and includes pathogens that cause salmonellosis and typhoid fever.
Answer
Gammaproteobacteria
Short Answer
What is the metabolic difference between coliforms and noncoliforms? Which category contains several species of intestinal pathogens?
Why are Mycoplasma and Chlamydia classified as obligate intracellular pathogens?
Critical Thinking
The cell shown is found in the human stomach and is now known to cause peptic ulcers. What is the name of this bacterium?
(credit: American Society for Microbiology)
4.3: Nonproteobacteria Gram-negative Bacteria and Phototrophic Bacteria
Gram-negative nonproteobacteria include the taxa spirochetes; the Cytophaga, Fusobacterium, Bacteroides group; Planctomycetes; and many representatives of phototrophic bacteria. Spirochetes are motile, spiral bacteria with a long, narrow body; they are difficult or impossible to culture. Several genera of spirochetes contain human pathogens that cause such diseases as syphilis and Lyme disease. Cytophaga, Fusobacterium, and Bacteroides are classified together as a phylum called the CFB group.
Multiple Choice
Which of the following is the organelle that spirochetes use to propel themselves?
1. plasma membrane
2. axial filament
3. pilum
4. fimbria
Answer
B
Which of the following bacteria are the most prevalent in the human gut?
1. cyanobacteria
2. staphylococci
3. Borrelia
4. Bacteroides
Answer
D
Which of the following refers to photosynthesis performed by bacteria with the use of water as the donor of electrons?
1. oxygenic
2. anoxygenic
3. heterotrophic
4. phototrophic
Answer
A
Fill in the Blank
The bacterium that causes syphilis is called ________.
Answer
Treponema pallidum pallidum
Bacteria in the genus Rhodospirillum that use hydrogen for oxidation and fix nitrogen are ________ bacteria.
Answer
purple nonsulfur
Short Answer
Explain the term CFB group and name the genera that this group includes.
Name and briefly describe the bacterium that causes Lyme disease.
Characterize the phylum Cyanobacteria.
4.4: Gram-positive Bacteria
Gram-positive bacteria are a very large and diverse group of microorganisms. Understanding their taxonomy and knowing their unique features is important for diagnostics and treatment of infectious diseases. Gram-positive bacteria are classified into high G+C gram-positive and low G+C gram-positive bacteria, based on the prevalence of guanine and cytosine nucleotides in their genome.
Multiple Choice
Which of the following bacterial species is classified as high G+C gram-positive?
1. Corynebacterium diphtheriae
2. Staphylococcus aureus
3. Bacillus anthracis
4. Streptococcus pneumonia
Answer
A
Fill in the Blank
Streptococcus is the ________ of bacteria that is responsible for many human diseases.
Answer
genus
One species of Streptococcus, S. pyogenes, is a classified as a ________ pathogen due to the characteristic production of pus in infections it causes.
Answer
pyogenic
Propionibacterium belongs to ________ G+C gram-positive bacteria. One of its species is used in the food industry and another causes acne.
Answer
high
Short Answer
Name and describe two types of S. aureus that show multiple antibiotic resistance.
Critical Thinking
The microscopic growth pattern shown is characteristic of which genus of bacteria?
(credit: modification of work by Janice Haney Carr/Centers for Disease Control and Prevention)
4.5: Deeply Branching Bacteria
Deeply branching bacteria are phylogenetically the most ancient forms of life, being the closest to the last universal common ancestor. Deeply branching bacteria include many species that thrive in extreme environments that are thought to resemble conditions on earth billions of years ago. Deeply branching bacteria are important for our understanding of evolution; some of them are used in industry.
Multiple Choice
The term “deeply branching” refers to which of the following?
1. the cellular shape of deeply branching bacteria
2. the position in the evolutionary tree of deeply branching bacteria
3. the ability of deeply branching bacteria to live in deep ocean waters
4. the pattern of growth in culture of deeply branching bacteria
Answer
B
Which of these deeply branching bacteria is considered a polyextremophile?
1. Aquifex pyrophilus
2. Deinococcus radiodurans
3. Staphylococcus aureus
4. Mycobacterium tuberculosis
Answer
B
Fill in the Blank
The length of the branches of the evolutionary tree characterizes the evolutionary ________ between organisms.
Answer
distance
The deeply branching bacteria are thought to be the form of life closest to the last universal ________ ________.
Answer
common ancestor
Many of the deeply branching bacteria are aquatic and hyperthermophilic, found near underwater volcanoes and thermal ocean ________.
Answer
vents
The deeply branching bacterium Deinococcus radiodurans is able to survive exposure to high doses of ________.
Answer
ionizing radiation
Short Answer
Briefly describe the significance of deeply branching bacteria for basic science and for industry.
What is thought to account for the unique radiation resistance of D. radiodurans?
4.6: Archaea
Archaea are unicellular, prokaryotic microorganisms that differ from bacteria in their genetics, biochemistry, and ecology. Some archaea are extremophiles, living in environments with extremely high or low temperatures, or extreme salinity. Only archaea are known to produce methane. Methane-producing archaea are called methanogens. Halophilic archaea prefer a concentration of salt close to saturation and perform photosynthesis using bacteriorhodopsin.
Multiple Choice
Archaea and Bacteria are most similar in terms of their ________.
1. genetics
2. cell wall structure
3. ecology
4. unicellular structure
Answer
D
Which of the following is true of archaea that produce methane?
1. They reduce carbon dioxide in the presence of nitrogen.
2. They live in the most extreme environments.
3. They are always anaerobes.
4. They have been discovered on Mars.
Answer
B
Fill in the Blank
________ is a genus of Archaea. Its optimal environmental temperature ranges from 70 °C to 80 °C, and its optimal pH is 2–3. It oxidizes sulfur and produces sulfuric acid.
Answer
Sulfolobus
________ was once thought to be the cause of periodontal disease, but, more recently, the causal relationship between this archaean and the disease was not confirmed.
Answer
Methanobrevibacter oralis
Short Answer
What accounts for the purple color in salt ponds inhabited by halophilic archaea?
What evidence supports the hypothesis that some archaea live on Mars?
Critical Thinking
What is the connection between this methane bog and archaea?
(credit: Chad Skeers) | textbooks/bio/Microbiology/Microbiology_(OpenStax)/04%3A_Prokaryotic_Diversity/4.E%3A_Prokaryotic_Diversity_%28Exercises%29.txt |
Although bacteria and viruses account for a large number of the infectious diseases that afflict humans, many serious illnesses are caused by eukaryotic organisms. One example is malaria, which is caused by Plasmodium, a eukaryotic organism transmitted through mosquito bites. Malaria is a major cause of morbidity (illness) and mortality (death) that threatens 3.4 billion people worldwide.1 In severe cases, organ failure and blood or metabolic abnormalities contribute to medical emergencies and sometimes death. Even after initial recovery, relapses may occur years later. In countries where malaria is endemic, the disease represents a major public health challenge that can place a tremendous strain on developing economies.
Worldwide, major efforts are underway to reduce malaria infections. Efforts include the distribution of insecticide-treated bed nets and the spraying of pesticides. Researchers are also making progress in their efforts to develop effective vaccines.2 The President’s Malaria Initiative, started in 2005, supports prevention and treatment. The Bill and Melinda Gates Foundation has a large initiative to eliminate malaria. Despite these efforts, malaria continues to cause long-term morbidity (such as intellectual disabilities in children) and mortality (especially in children younger than 5 years), so we still have far to go.
• 5.1: Unicellular Eukaryotic Microorganisms
Protists are a diverse, polyphyletic group of eukaryotic organisms. Protists may be unicellular or multicellular. They vary in how they get their nutrition, morphology, method of locomotion, and mode of reproduction. Important structures of protists include contractile vacuoles, cilia, flagella, pellicles, and pseudopodia; some lack organelles such as mitochondria. Taxonomy of protists is changing rapidly as relationships are reassessed using newer techniques.
• 5.2: Parasitic Helminths
Helminth parasites are included within the study of microbiology because they are often identified by looking for microscopic eggs and larvae. The two major groups of helminth parasites are the roundworms (Nematoda) and the flatworms (Platyhelminthes). Nematodes are common intestinal parasites often transmitted through undercooked foods, although they are also found in other environments. Platyhelminths include tapeworms and flukes, which are often transmitted through undercooked meat.
• 5.3: Fungi
The fungi include diverse saprotrophic eukaryotic organisms with chitin cell walls. Fungi can be unicellular or multicellular; some (like yeast) and fungal spores are microscopic, whereas some are large and conspicuous. Reproductive types are important in distinguishing fungal groups. Medically important species exist in the four fungal groups Zygomycota, Ascomycota, Basidiomycota, and Microsporidia.
• 5.4: Algae
Algae are a diverse group of photosynthetic eukaryotic protists. Algae may be unicellular or multicellular. Large, multicellular algae are called seaweeds but are not plants and lack plant-like tissues and organs. Although algae have little pathogenicity, they may be associated with toxic algal blooms that can harm aquatic wildlife and contaminate seafood with toxins that cause paralysis.
• 5.5: Lichens
Lichens are a symbiotic association between a fungus and an algae or a cyanobacterium. The symbiotic association found in lichens is currently considered to be a controlled parasitism, in which the fungus benefits and the algae or cyanobacterium is harmed. Lichens are slow growing and can live for centuries in a variety of habitats. Lichens are environmentally important, helping to create soil, providing food, and acting as indicators of air pollution.
• 5.E: The Eukaryotes of Microbiology (Exercises)
Footnotes
1. 1 Centers for Disease Control and Prevention. “Impact of Malaria.” September 22, 2015. http://www.cdc.gov/malaria/malaria_w...de/impact.html. Accessed January 18, 2016.
2. 2 RTS, S Clinical Trials Partnership. “Efficacy and safety of RTS,S/AS01 malaria vaccine with or without a booster dose in infants and children in Africa: final results of a phase 3, individually randomised, controlled trial.” The Lancet 23 April 2015. DOI: http://dx.doi.org/10.1016/S0140-6736(15)60721-8.
Thumbnail: Mold growing on a clementine. (CC SA-BY 3.0; NotFromUtrecht).
05: The Eukaryotes of Microbiology
Learning Objectives
• Summarize the general characteristics of unicellular eukaryotic parasites
• Describe the general life cycles and modes of reproduction in unicellular eukaryotic parasites
• Identify challenges associated with classifying unicellular eukaryotes
• Explain the taxonomic scheme used for unicellular eukaryotes
• Give examples of infections caused by unicellular eukaryotes
Clinical Focus: Part 1
Upon arriving home from school, 7-year-old Sarah complains that a large spot on her arm will not stop itching. She keeps scratching at it, drawing the attention of her parents. Looking more closely, they see that it is a red circular spot with a raised red edge (Figure \(1\)). The next day, Sarah’s parents take her to their doctor, who examines the spot using a Wood’s lamp. A Wood’s lamp produces ultraviolet light that causes the spot on Sarah’s arm to fluoresce, which confirms what the doctor already suspected: Sarah has a case of ringworm.
Sarah’s mother is mortified to hear that her daughter has a “worm.” How could this happen?
Exercise \(1\)
What are some likely ways that Sarah might have contracted ringworm?
Eukaryotic microbes are an extraordinarily diverse group, including species with a wide range of life cycles, morphological specializations, and nutritional needs. Although more diseases are caused by viruses and bacteria than by microscopic eukaryotes, these eukaryotes are responsible for some diseases of great public health importance. For example, the protozoal disease malaria was responsible for 584,000 deaths worldwide (primarily children in Africa) in 2013, according to the World Health Organization (WHO). The protist parasite Giardia causes a diarrheal illness (giardiasis) that is easily transmitted through contaminated water supplies. In the United States, Giardia is the most common human intestinal parasite (Figure \(2\)). Although it may seem surprising, parasitic worms are included within the study of microbiology because identification depends on observation of microscopic adult worms or eggs. Even in developed countries, these worms are important parasites of humans and of domestic animals. There are fewer fungal pathogens, but these are important causes of illness, as well. On the other hand, fungi have been important in producing antimicrobial substances such as penicillin. In this chapter, we will examine characteristics of protists, worms, and fungi while considering their roles in causing disease.
Characteristics of Protists
The word protist is a historical term that is now used informally to refer to a diverse group of microscopic eukaryotic organisms. It is not considered a formal taxonomic term because the organisms it describes do not have a shared evolutionary origin. Historically, the protists were informally grouped into the “animal-like” protozoans, the “plant-like” algae, and the “fungus-like” protists such as water molds. These three groups of protists differ greatly in terms of their basic characteristics. For example, algae are photosynthetic organisms that can be unicellular or multicellular. Protozoa, on the other hand, are nonphotosynthetic, motile organisms that are always unicellular. Other informal terms may also be used to describe various groups of protists. For example, microorganisms that drift or float in water, moved by currents, are referred to as plankton. Types of plankton include zooplankton, which are motile and nonphotosynthetic, and phytoplankton, which are photosynthetic.
Protozoans inhabit a wide variety of habitats, both aquatic and terrestrial. Many are free-living, while others are parasitic, carrying out a life cycle within a host or hosts and potentially causing illness. There are also beneficial symbionts that provide metabolic services to their hosts. During the feeding and growth part of their life cycle, they are called trophozoites; these feed on small particulate food sources such as bacteria. While some types of protozoa exist exclusively in the trophozoite form, others can develop from trophozoite to an encapsulated cyst stage when environmental conditions are too harsh for the trophozoite. A cyst is a cell with a protective wall, and the process by which a trophozoite becomes a cyst is called encystment. When conditions become more favorable, these cysts are triggered by environmental cues to become active again through excystment.
One protozoan genus capable of encystment is Eimeria, which includes some human and animal pathogens. Figure \(3\) illustrates the life cycle of Eimeria.
Protozoans have a variety of reproductive mechanisms. Some protozoans reproduce asexually and others reproduce sexually; still others are capable of both sexual and asexual reproduction. In protozoans, asexual reproduction occurs by binary fission, budding, or schizogony. In schizogony, the nucleus of a cell divides multiple times before the cell divides into many smaller cells. The products of schizogony are called merozoites and they are stored in structures known as schizonts. Protozoans may also reproduce sexually, which increases genetic diversity and can lead to complex life cycles. Protozoans can produce haploid gametes that fuse through syngamy. However, they can also exchange genetic material by joining to exchange DNA in a process called conjugation. This is a different process than the conjugation that occurs in bacteria. The term protist conjugation refers to a true form of eukaryotic sexual reproduction between two cells of different mating types. It is found in ciliates, a group of protozoans, and is described later in this subsection.
All protozoans have a plasma membrane, or plasmalemma, and some have bands of protein just inside the membrane that add rigidity, forming a structure called the pellicle. Some protists, including protozoans, have distinct layers of cytoplasm under the membrane. In these protists, the outer gel layer (with microfilaments of actin) is called the ectoplasm. Inside this layer is a sol (fluid) region of cytoplasm called the endoplasm. These structures contribute to complex cell shapes in some protozoans, whereas others (such as amoebas) have more flexible shapes (Figure \(4\)).
Different groups of protozoans have specialized feeding structures. They may have a specialized structure for taking in food through phagocytosis, called a cytostome, and a specialized structure for the exocytosis of wastes called a cytoproct. Oral grooves leading to cytostomes are lined with hair-like cilia to sweep in food particles. Protozoans are heterotrophic. Protozoans that are holozoic ingest whole food particles through phagocytosis. Forms that are saprozoic ingest small, soluble food molecules.
Many protists have whip-like flagella or hair-like cilia made of microtubules that can be used for locomotion (Figure \(4\)). Other protists use cytoplasmic extensions known as pseudopodia (“false feet”) to attach the cell to a surface; they then allow cytoplasm to flow into the extension, thus moving themselves forward.
Protozoans have a variety of unique organelles and sometimes lack organelles found in other cells. Some have contractile vacuoles, organelles that can be used to move water out of the cell for osmotic regulation (salt and water balance) (Figure \(4\)). Mitochondria may be absent in parasites or altered to kinetoplastids (modified mitochondria) or hydrogenosomes (see Unique Characteristics of Eukaryotic Cells for more discussion of these structures).
Exercise \(2\)
What is the sequence of events in reproduction by schizogony and what are the cells produced called?
Taxonomy of Protists
The protists are a polyphyletic group, meaning they lack a shared evolutionary origin. Since the current taxonomy is based on evolutionary history (as determined by biochemistry, morphology, and genetics), protists are scattered across many different taxonomic groups within the domain Eukarya. Eukarya is currently divided into six supergroups that are further divided into subgroups, as illustrated in (Figure \(5\)). In this section, we will primarily be concerned with the supergroups Amoebozoa, Excavata, and Chromalveolata; these supergroups include many protozoans of clinical significance. The supergroups Opisthokonta and Rhizaria also include some protozoans, but few of clinical significance. In addition to protozoans, Opisthokonta also includes animals and fungi, some of which we will discuss in Parasitic Helminths and Fungi. Some examples of the Archaeplastida will be discussed in Algae. Figure \(6\) and Figure \(7\) summarize the characteristics of each supergroup and subgroup and list representatives of each.
Exercise \(3\)
Which supergroups contain the clinically significant protists?
Amoebozoa
The supergroup Amoebozoa includes protozoans that use amoeboid movement. Actin microfilaments produce pseudopodia, into which the remainder of the protoplasm flows, thereby moving the organism. The genus Entamoebaincludes commensal or parasitic species, including the medically important E. histolytica, which is transmitted by cysts in feces and is the primary cause of amoebic dysentery. The notorious “brain-eating amoeba,” Naegleria fowleri, is also classified within the Amoebozoa. This deadly parasite is found in warm, fresh water and causes primary amoebic meningoencephalitis (PAM). Another member of this group is Acanthamoeba, which can cause keratitis (corneal inflammation) and blindness.
The Eumycetozoa are an unusual group of organisms called slime molds, which have previously been classified as animals, fungi, and plants (Figure \(8\)). Slime molds can be divided into two types: cellular slime molds and plasmodial slime molds. The cellular slime molds exist as individual amoeboid cells that periodically aggregate into a mobile slug. The aggregate then forms a fruiting body that produces haploid spores. Plasmodial slime molds exist as large, multinucleate amoeboid cells that form reproductive stalks to produce spores that divide into gametes. One cellular slime mold, Dictyostelium discoideum, has been an important study organism for understanding cell differentiation, because it has both single-celled and multicelled life stages, with the cells showing some degree of differentiation in the multicelled form. Figure \(9\)and Figure \(10\) illustrate the life cycles of cellular and plasmodial slime molds, respectively.
Chromalveolata
The supergroup Chromalveolata is united by similar origins of its members’ plastids and includes the apicomplexans, ciliates, diatoms, and dinoflagellates, among other groups (we will cover the diatoms and dinoflagellates in Algae). The apicomplexans are intra- or extracellular parasites that have an apical complex at one end of the cell. The apical complex is a concentration of organelles, vacuoles, and microtubules that allows the parasite to enter host cells (Figure \(11\)). Apicomplexans have complex life cycles that include an infective sporozoite that undergoes schizogony to make many merozoites (see the example in Figure \(3\)). Many are capable of infecting a variety of animal cells, from insects to livestock to humans, and their life cycles often depend on transmission between multiple hosts. The genus Plasmodium is an example of this group.
Other apicomplexans are also medically important. Cryptosporidium parvum causes intestinal symptoms and can cause epidemic diarrhea when the cysts contaminate drinking water. Theileria (Babesia) microti, transmitted by the tick Ixodes scapularis, causes recurring fever that can be fatal and is becoming a common transfusion-transmitted pathogen in the United States (Theileria and Babesia are closely related genera and there is some debate about the best classification). Finally, Toxoplasma gondii causes toxoplasmosis and can be transmitted from cat feces, unwashed fruit and vegetables, or from undercooked meat. Because toxoplasmosis can be associated with serious birth defects, pregnant women need to be aware of this risk and use caution if they are exposed to the feces of potentially infected cats. A national survey found the frequency of individuals with antibodies for toxoplasmosis (and thus who presumably have a current latent infection) in the United States to be 11%. Rates are much higher in other countries, including some developed countries.1 There is also evidence and a good deal of theorizing that the parasite may be responsible for altering infected humans’ behavior and personality traits.2
The ciliates (Ciliaphora), also within the Chromalveolata, are a large, very diverse group characterized by the presence of cilia on their cell surface. Although the cilia may be used for locomotion, they are often used for feeding, as well, and some forms are nonmotile. Balantidium coli (Figure \(12\)) is the only parasitic ciliate that affects humans by causing intestinal illness, although it rarely causes serious medical issues except in the immunocompromised (those having a weakened immune system). Perhaps the most familiar ciliate is Paramecium, a motile organism with a clearly visible cytostomeand cytoproct that is often studied in biology laboratories (Figure \(13\)). Another ciliate, Stentor, is sessile and uses its cilia for feeding (Figure \(14\)). Generally, these organisms have a micronucleus that is diploid, somatic, and used for sexual reproduction by conjugation. They also have a macronucleus that is derived from the micronucleus; the macronucleus becomes polyploid (multiple sets of duplicate chromosomes), and has a reduced set of metabolic genes.
Ciliates are able to reproduce through conjugation, in which two cells attach to each other. In each cell, the diploid micronuclei undergo meiosis, producing eight haploid nuclei each. Then, all but one of the haploid micronuclei and the macronucleus disintegrate; the remaining (haploid) micronucleus undergoes mitosis. The two cells then exchange one micronucleus each, which fuses with the remaining micronucleus present to form a new, genetically different, diploid micronucleus. The diploid micronucleus undergoes two mitotic divisions, so each cell has four micronuclei, and two of the four combine to form a new macronucleus. The chromosomes in the macronucleus then replicate repeatedly, the macronucleus reaches its polyploid state, and the two cells separate. The two cells are now genetically different from each other and from their previous versions.
Öomycetes have similarities to fungi and were once classified with them. They are also called water molds. However, they differ from fungi in several important ways. Öomycetes have cell walls of cellulose (unlike the chitinous cell walls of fungi) and they are generally diploid, whereas the dominant life forms of fungi are typically haploid. Phytophthora, the plant pathogen found in the soil that caused the Irish potato famine, is classified within this group (Figure \(15\)).
Link to Learning
This video shows the feeding of Stentor.
Excavata
The third and final supergroup to be considered in this section is the Excavata, which includes primitive eukaryotes and many parasites with limited metabolic abilities. These organisms have complex cell shapes and structures, often including a depression on the surface of the cell called an excavate. The group Excavata includes the subgroups Fornicata, Parabasalia, and Euglenozoa. The Fornicata lack mitochondria but have flagella. This group includes Giardia lamblia (also known as G. intestinalis or G. duodenalis), a widespread pathogen that causes diarrheal illness and can be spread through cysts from feces that contaminate water supplies (Figure \(2\)). Parabasalia are frequent animal endosymbionts; they live in the guts of animals like termites and cockroaches. They have basal bodies and modified mitochondria (kinetoplastids). They also have a large, complex cell structure with an undulating membrane and often have many flagella. The trichomonads (a subgroup of the Parabasalia) include pathogens such as Trichomonas vaginalis, which causes the human sexually transmitted disease trichomoniasis. Trichomoniasis often does not cause symptoms in men, but men are able to transmit the infection. In women, it causes vaginal discomfort and discharge and may cause complications in pregnancy if left untreated.
The Euglenozoa are common in the environment and include photosynthetic and nonphotosynthetic species. Members of the genus Euglena are typically not pathogenic. Their cells have two flagella, a pellicle, a stigma (eyespot) to sense light, and chloroplasts for photosynthesis (Figure \(16\)). The pellicle of Euglena is made of a series of protein bands surrounding the cell; it supports the cell membrane and gives the cell shape.
The Euglenozoa also include the trypanosomes, which are parasitic pathogens. The genus Trypanosoma includes T. brucei, which causes African trypanosomiasis (African sleeping sickness and T. cruzi, which causes American trypanosomiasis (Chagas disease). These tropical diseases are spread by insect bites. In African sleeping sickness, T. brucei colonizes the blood and the brain after being transmitted via the bite of a tsetse fly (Glossina spp.) (Figure \(17\)). The early symptoms include confusion, difficulty sleeping, and lack of coordination. Left untreated, it is fatal.
Chagas’ disease originated and is most common in Latin America. The disease is transmitted by Triatoma spp., insects often called “kissing bugs,” and affects either the heart tissue or tissues of the digestive system. Untreated cases can eventually lead to heart failure or significant digestive or neurological disorders.
The genus Leishmania includes trypanosomes that cause disfiguring skin disease and sometimes systemic illness as well.
Neglected Parasites
The Centers for Disease Control and Prevention (CDC) is responsible for identifying public health priorities in the United States and developing strategies to address areas of concern. As part of this mandate, the CDC has officially identified five parasitic diseases it considers to have been neglected (i.e., not adequately studied). These neglected parasitic infections (NPIs) include toxoplasmosis, Chagas disease, toxocariasis (a nematode infection transmitted primarily by infected dogs), cysticercosis (a disease caused by a tissue infection of the tapeworm Taenia solium), and trichomoniasis (a sexually transmitted disease caused by the parabasalid Trichomonas vaginalis).
The decision to name these specific diseases as NPIs means that the CDC will devote resources toward improving awareness and developing better diagnostic testing and treatment through studies of available data. The CDC may also advise on treatment of these diseases and assist in the distribution of medications that might otherwise be difficult to obtain.3
Of course, the CDC does not have unlimited resources, so by prioritizing these five diseases, it is effectively deprioritizing others. Given that many Americans have never heard of many of these NPIs, it is fair to ask what criteria the CDC used in prioritizing diseases. According to the CDC, the factors considered were the number of people infected, the severity of the illness, and whether the illness can be treated or prevented. Although several of these NPIs may seem to be more common outside the United States, the CDC argues that many cases in the United States likely go undiagnosed and untreated because so little is known about these diseases.4
What criteria should be considered when prioritizing diseases for purposes of funding or research? Are those identified by the CDC reasonable? What other factors could be considered? Should government agencies like the CDC have the same criteria as private pharmaceutical research labs? What are the ethical implications of deprioritizing other potentially neglected parasitic diseases such as leishmaniasis?
Key Concepts and Summary
• Protists are a diverse, polyphyletic group of eukaryotic organisms.
• Protists may be unicellular or multicellular. They vary in how they get their nutrition, morphology, method of locomotion, and mode of reproduction.
• Important structures of protists include contractile vacuoles, cilia, flagella, pellicles, and pseudopodia; some lack organelles such as mitochondria.
• Taxonomy of protists is changing rapidly as relationships are reassessed using newer techniques.
• The protists include important pathogens and parasites.
Footnotes
1. 1 J. Flegr et al. “Toxoplasmosis—A Global Threat. Correlation of Latent Toxoplasmosis With Specific Disease Burden in a Set of 88 Countries.” PloS ONE 9 no. 3 (2014):e90203.
2. 2 J. Flegr. “Effects of Toxoplasma on Human Behavior.” Schizophrenia Bull 33, no. 3 (2007):757–760.
3. 3 Centers for Disease Control and Prevention. “Neglected Parasitic Infections (NPIs) in the United States.” http://www.cdc.gov/parasites/npi/. Last updated July 10, 2014.
4. 4 Centers for Disease Control and Prevention. “Fact Sheet: Neglected Parasitic Infections in the United States.” www.cdc.gov/parasites/resourc..._factsheet.pdf | textbooks/bio/Microbiology/Microbiology_(OpenStax)/05%3A_The_Eukaryotes_of_Microbiology/5.01%3A_Unicellular_Eukaryotic_Microorganisms.txt |
Learning Objectives
• Explain why we include the study of parasitic worms within the discipline of microbiology
• Compare the basic morphology of the major groups of parasitic helminthes
• Describe the characteristics of parasitic nematodes, and give an example of infective eggs and infective larvae
• Describe the characteristics of parasitic trematodes and cestodes, and give examples of each
• Identify examples of the primary causes of infections due to nematodes, trematodes, and cestodes
• Classify parasitic worms according to major groups
Parasitic helminths are animals that are often included within the study of microbiology because many species of these worms are identified by their microscopic eggs and larvae. There are two major groups of parasitic helminths: the roundworms (Nematoda) and flatworms (Platyhelminthes). Of the many species that exist in these groups, about half are parasitic and some are important human pathogens. As animals, they are multicellular and have organ systems. However, the parasitic species often have limited digestive tracts, nervous systems, and locomotor abilities. Parasitic forms may have complex reproductive cycles with several different life stages and more than one type of host. Some are monoecious, having both male and female reproductive organs in a single individual, while others are dioecious, each having either male or female reproductive organs.
Nematoda (Roundworms)
Phylum Nematoda (the roundworms) is a diverse group containing more than 15,000 species, of which several are important human parasites (Figure \(1\)). These unsegmented worms have a full digestive system even when parasitic. Some are common intestinal parasites, and their eggs can sometimes be identified in feces or around the anus of infected individuals. Ascaris lumbricoides is the largest nematode intestinal parasite found in humans; females may reach lengths greater than 1 meter. A. lumbricoides is also very widespread, even in developed nations, although it is now a relatively uncommon problem in the United States. It may cause symptoms ranging from relatively mild (such as a cough and mild abdominal pain) to severe (such as intestinal blockage and impaired growth).
Of all nematode infections in the United States, pinworm (caused by Enterobius vermicularis) is the most common. Pinworm causes sleeplessness and itching around the anus, where the female worms lay their eggs during the night. Toxocara canis and T. cati are nematodes found in dogs and cats, respectively, that can be transmitted to humans, causing toxocariasis. Antibodies to these parasites have been found in approximately 13.9% of the U.S. population, suggesting that exposure is common.1 Infection can cause larval migrans, which can result in vision loss and eye inflammation, or fever, fatigue, coughing, and abdominal pain, depending on whether the organism infects the eye or the viscera. Another common nematode infection is hookworm, which is caused by Necator americanus (the New World or North American hookworm) and Ancylostoma duodenale (the Old World hookworm). Symptoms of hookworm infection can include abdominal pain, diarrhea, loss of appetite, weight loss, fatigue, and anemia.
Trichinellosis, also called trichinosis, caused by Trichinella spiralis, is contracted by consuming undercooked meat, which releases the larvae and allows them to encyst in muscles. Infection can cause fever, muscle pains, and digestive system problems; severe infections can lead to lack of coordination, breathing and heart problems, and even death. Finally, heartworm in dogs and other animals is caused by the nematode Dirofilaria immitis, which is transmitted by mosquitoes. Symptoms include fatigue and cough; when left untreated, death may result.
Clinical Focus: Part 2
The physician explains to Sarah’s mother that ringworm can be transferred between people through touch. “It’s common in school children, because they often come in close contact with each other, but anyone can become infected,” he adds. “Because you can transfer it through objects, locker rooms and public pools are also a potential source of infection. It’s very common among wrestlers and athletes in other contact sports.”
Looking very uncomfortable, Sarah says to her mother “I want this worm out of me.”
The doctor laughs and says, “Sarah, you’re in luck because ringworm is just a name; it is not an actual worm. You have nothing wriggling around under your skin.”
“Then what is it?” asks Sarah.
Exercise \(1\)
1. What type of pathogen causes ringworm?
2. What is the most common nematode infection in the United States?
Platyhelminths (Flatworms)
Phylum Platyhelminthes (the platyhelminths) are flatworms. This group includes the flukes, tapeworms, and the turbellarians, which include planarians. The flukes and tapeworms are medically important parasites (Figure \(2\)).
The flukes (trematodes) are nonsegmented flatworms that have an oral sucker (Figure \(3\)) (and sometimes a second ventral sucker) and attach to the inner walls of intestines, lungs, large blood vessels, or the liver. Trematodes have complex life cycles, often with multiple hosts. Several important examples are the liver flukes (Clonorchis and Opisthorchis), the intestinal fluke (Fasciolopsis buski), and the oriental lung fluke (Paragonimus westermani). Schistosomiasis is a serious parasitic disease, considered second in the scale of its impact on human populations only to malaria. The parasites Schistosoma mansoni, S. haematobium, and S. japonicum, which are found in freshwater snails, are responsible for schistosomiasis (Figure \(4\)). Immature forms burrow through the skin into the blood. They migrate to the lungs, then to the liver and, later, other organs. Symptoms include anemia, malnutrition, fever, abdominal pain, fluid buildup, and sometimes death.
The other medically important group of platyhelminths are commonly known as tapeworms (cestodes) and are segmented flatworms that may have suckers or hooks at the scolex (head region) (Figure \(3\)). Tapeworms use these suckers or hooks to attach to the wall of the small intestine. The body of the worm is made up of segments called proglottids that contain reproductive structures; these detach when the gametes are fertilized, releasing gravid proglottids with eggs. Tapeworms often have an intermediate host that consumes the eggs, which then hatch into a larval form called an oncosphere. The oncosphere migrates to a particular tissue or organ in the intermediate host, where it forms cysticerci. After being eaten by the definitive host, the cysticerci develop into adult tapeworms in the host's digestive system (Figure \(5\)). Taenia saginata (the beef tapeworm) and T. solium (the pork tapeworm) enter humans through ingestion of undercooked, contaminated meat. The adult worms develop and reside in the intestine, but the larval stage may migrate and be found in other body locations such as skeletal and smooth muscle. The beef tapeworm is relatively benign, although it can cause digestive problems and, occasionally, allergic reactions. The pork tapeworm can cause more serious problems when the larvae leave the intestine and colonize other tissues, including those of the central nervous system. Diphylobothrium latum is the largest human tapeworm and can be ingested in undercooked fish. It can grow to a length of 15 meters. Echinococcus granulosus, the dog tapeworm, can parasitize humans and uses dogs as an important host.
Exercise \(2\)
What group of medically important flatworms is segmented and what group is unsegmented?
Food for Worms?
For residents of temperate, developed countries, it may be difficult to imagine just how common helminth infections are in the human population. In fact, they are quite common and even occur frequently in the United States. Worldwide, approximately 807–1,221 million people are infected with Ascaris lumbricoides (perhaps one-sixth of the human population) and far more are infected if all nematode species are considered.2 Rates of infection are relatively high even in industrialized nations. Approximately 604–795 million people are infected with whipworm (Trichuris) worldwide (Trichuris can also infect dogs), and 576–740 million people are infected with hookworm (Necator americanus and Ancylostoma duodenale).3 Toxocara, a nematode parasite of dogs and cats, is also able to infect humans. It is widespread in the United States, with about 10,000 symptomatic cases annually. However, one study found 14% of the population (more than 40 million Americans) was seropositive, meaning they had been exposed to the parasite at one time. More than 200 million people have schistosomiasis worldwide. Most of the World Health Organization (WHO) neglected tropical diseases are helminths. In some cases, helminths may cause subclinical illnesses, meaning the symptoms are so mild that that they go unnoticed. In other cases, the effects may be more severe or chronic, leading to fluid accumulation and organ damage. With so many people affected, these parasites constitute a major global public health concern.
Eradicating the Guinea Worm
Dracunculiasis, or Guinea worm disease, is caused by a nematode called Dracunculus medinensis. When people consume contaminated water, water fleas (small crustaceans) containing the nematode larvae may be ingested. These larvae migrate out of the intestine, mate, and move through the body until females eventually emerge (generally through the feet). While Guinea worm disease is rarely fatal, it is extremely painful and can be accompanied by secondary infections and edema (Figure \(6\)).
An eradication campaign led by WHO, the CDC, the United Nations Children’s Fund (UNICEF), and the Carter Center (founded by former U.S. president Jimmy Carter) has been extremely successful in reducing cases of dracunculiasis. This has been possible because diagnosis is straightforward, there is an inexpensive method of control, there is no animal reservoir, the water fleas are not airborne (they are restricted to still water), the disease is geographically limited, and there has been a commitment from the governments involved. Additionally, no vaccines or medication are required for treatment and prevention. In 1986, 3.5 million people were estimated to be affected. After the eradication campaign, which included helping people in affected areas learn to filter water with cloth, only four countries continue to report the disease (Chad, Mali, South Sudan, and Ethiopia) with a total of 126 cases reported to WHO in 2014.
Key Concepts and Summary
• Helminth parasites are included within the study of microbiology because they are often identified by looking for microscopic eggs and larvae.
• The two major groups of helminth parasites are the roundworms (Nematoda) and the flatworms (Platyhelminthes).
• Nematodes are common intestinal parasites often transmitted through undercooked foods, although they are also found in other environments.
• Platyhelminths include tapeworms and flukes, which are often transmitted through undercooked meat.
Footnotes
1. 1 Won K, Kruszon-Moran D, Schantz P, Jones J. “National seroprevalence and risk factors for zoonotic Toxocara spp. infection.” In: Abstracts of the 56th American Society of Tropical Medicine and Hygiene; Philadelphia, Pennsylvania; 2007 Nov 4-8.
2. 2 Fenwick, A. “The global burden of neglected tropical diseases.” Public health 126 no.3 (Mar 2012): 233–6.
3. 3 de Silva, N., et. al. (2003). “Soil-transmitted helminth infections: updating the global picture”. Trends in Parasitology 19 (December 2003): 547–51.
4. 4 World Health Organization. “South Sudan Reports Zero Cases of Guinea-Worm Disease for Seventh Consecutive Month.” 2016. http://www.who.int/dracunculiasis/no...ive_months/en/. Accessed May 2, 2016. | textbooks/bio/Microbiology/Microbiology_(OpenStax)/05%3A_The_Eukaryotes_of_Microbiology/5.02%3A_Parasitic_Helminths.txt |
Learning Objectives
• Explain why the study of fungi such as yeast and molds is within the discipline of microbiology
• Describe the unique characteristics of fungi
• Describe examples of asexual and sexual reproduction of fungi
• Compare the major groups of fungi in this chapter, and give examples of each
• Identify examples of the primary causes of infections due to yeasts and molds
• Identify examples of toxin-producing fungi
• Classify fungal organisms according to major groups
The fungi comprise a diverse group of organisms that are heterotrophic and typically saprozoic. In addition to the well-known macroscopic fungi (such as mushrooms and molds), many unicellular yeasts and spores of macroscopic fungi are microscopic. For this reason, fungi are included within the field of microbiology.
Fungi are important to humans in a variety of ways. Both microscopic and macroscopic fungi have medical relevance, with some pathogenic species that can cause mycoses (illnesses caused by fungi). Some pathogenic fungi are opportunistic, meaning that they mainly cause infections when the host’s immune defenses are compromised and do not normally cause illness in healthy individuals. Fungi are important in other ways. They act as decomposers in the environment, and they are critical for the production of certain foods such as cheeses. Fungi are also major sources of antibiotics, such as penicillin from the fungus Penicillium.
Characteristics of Fungi
Fungi have well-defined characteristics that set them apart from other organisms. Most multicellular fungal bodies, commonly called molds, are made up of filaments called hyphae. Hyphae can form a tangled network called a mycelium and form the thallus (body) of fleshy fungi. Hyphae that have walls between the cells are called septate hyphae; hyphae that lack walls and cell membranes between the cells are called nonseptate or coenocytic hyphae). (Figure \(1\)).
In contrast to molds, yeasts are unicellular fungi. The budding yeasts reproduce asexually by budding off a smaller daughter cell; the resulting cells may sometimes stick together as a short chain or pseudohypha (Figure \(1\)). Candida albicans is a common yeast that forms pseudohyphae; it is associated with various infections in humans, including vaginal yeast infections, oral thrush, and candidiasis of the skin.
Some fungi are dimorphic, having more than one appearance during their life cycle. These dimorphic fungi may be able to appear as yeasts or molds, which can be important for infectivity. They are capable of changing their appearance in response to environmental changes such as nutrient availability or fluctuations in temperature, growing as a mold, for example, at 25 °C (77 °F), and as yeast cells at 37 °C (98.6 °F). This ability helps dimorphic fungi to survive in diverse environments. Histoplasma capsulatum, the pathogen that causes histoplasmosis, a lung infection, is an example of a dimorphic fungus (Figure \(2\)).
There are notable unique features in fungal cell walls and membranes. Fungal cell walls contain chitin, as opposed to the cellulose found in the cell walls of plants and many protists. Additionally, whereas animals have cholesterol in their cell membranes, fungal cell membranes have different sterols called ergosterols. Ergosterols are often exploited as targets for antifungal drugs.
Fungal life cycles are unique and complex. Fungi reproduce sexually either through cross- or self-fertilization. Haploid fungi form hyphae that have gametes at the tips. Two different mating types (represented as “+ type” and “– type”) are involved. The cytoplasms of the + and – type gametes fuse (in an event called plasmogamy), producing a cell with two distinct nuclei (a dikaryotic cell). Later, the nuclei fuse (in an event called karyogamy) to create a diploid zygote. The zygote undergoes meiosis to form spores that germinate to start the haploid stage, which eventually creates more haploid mycelia (Figure \(3\)). Depending on the taxonomic group, these sexually produced spores are known as zygospores (in Zygomycota), ascospores (in Ascomycota), or basidiospores (in Basidiomycota) (Figure \(4\)).
Fungi may also exhibit asexual reproduction by mitosis, mitosis with budding, fragmentation of hyphae, and formation of asexual spores by mitosis. These spores are specialized cells that, depending on the organism, may have unique characteristics for survival, reproduction, and dispersal. Fungi exhibit several types of asexual spores and these can be important in classification.
Exercise \(1\)
Is a dimorphic fungus a yeast or a mold? Explain.
Fungal Diversity
The fungi are very diverse, comprising seven major groups. Not all of the seven groups contain pathogens. Some of these groups are generally associated with plants and include plant pathogens. For example, Urediniomycetes and Ustilagomycetes include the plant rusts and smuts, respectively. These form reddish or dark masses, respectively, on plants as rusts (red) or smuts (dark). Some species have substantial economic impact because of their ability to reduce crop yields. Glomeromycota includes the mycorrhizal fungi, important symbionts with plant roots that can promote plant growth by acting like an extended root system. The Glomeromycota are obligate symbionts, meaning that they can only survive when associated with plant roots; the fungi receive carbohydrates from the plant and the plant benefits from the increased ability to take up nutrients and minerals from the soil. The Chytridiomycetes (chytrids) are small fungi, but are extremely ecologically important. Chytrids are generally aquatic and have flagellated, motile gametes; specific types are implicated in amphibian declines around the world. Because of their medical importance, we will focus on Zygomycota, Ascomycota, Basidiomycota, and Microsporidia. Figure \(9\) summarizes the characteristics of these medically important groups of fungi.
The Zygomycota (zygomycetes) are mainly saprophytes with coenocytic hyphae and haploid nuclei. They use sporangiospores for asexual reproduction. The group name comes from the zygospores that they use for sexual reproduction (Figure \(3\)), which have hard walls formed from the fusion of reproductive cells from two individuals. Zygomycetes are important for food science and as crop pathogens. One example is Rhizopus stolonifer (Figure \(4\)), an important bread mold that also causes rice seedling blight. Mucor is a genus of fungi that can potentially cause necrotizing infections in humans, although most species are intolerant of temperatures found in mammalian bodies (Figure \(4\)).
The Ascomycota include fungi that are used as food (edible mushrooms, morels, and truffles), others that are common causes of food spoilage (bread molds and plant pathogens), and still others that are human pathogens. Ascomycota may have septate hyphae and cup-shaped fruiting bodies called ascocarps. Some genera of Ascomycota use sexually produced ascospores as well as asexual spores called conidia, but sexual phases have not been discovered or described for others. Some produce an ascus containing ascospores within an ascocarp (Figure \(5\)).
Examples of the Ascomycota include several bread molds and minor pathogens, as well as species capable of causing more serious mycoses. Species in the genus Aspergillus are important causes of allergy and infection, and are useful in research and in the production of certain fermented alcoholic beverages such as Japanese sake. The fungus Aspergillus flavus, a contaminant of nuts and stored grains, produces an aflatoxin that is both a toxin and the most potent known natural carcinogen. Neurospora crassa is of particular use in genetics research because the spores produced by meiosis are kept inside the ascus in a row that reflects the cell divisions that produced them, giving a direct view of segregation and assortment of genes (Figure \(6\)). Penicillium produces the antibiotic penicillin (Figure \(5\)).
Many species of ascomycetes are medically important. A large number of species in the genera Trichophyton, Microsporum, and Epidermophyton are dermatophytes, pathogenic fungi capable of causing skin infections such as athlete’s foot, jock itch, and ringworm. Blastomyces dermatitidis is a dimorphic fungus that can cause blastomycosis, a respiratory infection that, if left untreated, can become disseminated to other body sites, sometimes leading to death. Another important respiratory pathogen is the dimorphic fungus Histoplasma capsulatum (Figure \(2\)), which is associated with birds and bats in the Ohio and Mississippi river valleys. Coccidioides immitis causes the serious lung disease Valley fever. Candida albicans, the most common cause of vaginal and other yeast infections, is also an ascomycete fungus; it is a part of the normal microbiota of the skin, intestine, genital tract, and ear (Figure \(5\)). Ascomycetes also cause plant diseases, including ergot infections, Dutch elm disease, and powdery mildews.
Saccharomyces yeasts, including the baker’s yeast S. cerevisiae, are unicellular ascomycetes with haploid and diploid stages (Figure \(7\)). This and other Saccharomyces species are used for brewing beer.
The Basidiomycota (basidiomycetes) are fungi that have basidia (club-shaped structures) that produce basidiospores(spores produced through budding) within fruiting bodies called basidiocarps (Figure \(8\)). They are important as decomposers and as food. This group includes rusts, stinkhorns, puffballs, and mushrooms. Several species are of particular importance. Cryptococcus neoformans, a fungus commonly found as a yeast in the environment, can cause serious lung infections when inhaled by individuals with weakened immune systems. The edible meadow mushroom, Agricus campestris, is a basidiomycete, as is the poisonous mushroom Amanita phalloides, known as the death cap. The deadly toxins produced by A. phalloides have been used to study transcription.
Finally, the Microsporidia are unicellular fungi that are obligate intracellular parasites. They lack mitochondria, peroxisomes, and centrioles, but their spores release a unique polar tubule that pierces the host cell membrane to allow the fungus to gain entry into the cell. A number of microsporidia are human pathogens, and infections with microsporidia are called microsporidiosis. One pathogenic species is Enterocystozoan bieneusi, which can cause symptoms such as diarrhea, cholecystitis (inflammation of the gall bladder), and in rare cases, respiratory illness.
Exercise \(2\)
Which group of fungi appears to be associated with the greatest number of human diseases?
Eukaryotic Pathogens in Eukaryotic Hosts
When we think about antimicrobial medications, antibiotics such as penicillin often come to mind. Penicillin and related antibiotics interfere with the synthesis of peptidoglycan cell walls, which effectively targets bacterial cells. These antibiotics are useful because humans (like all eukaryotes) do not have peptidoglycan cell walls.
Developing medications that are effective against eukaryotic cells but not harmful to human cells is more difficult. Despite huge morphological differences, the cells of humans, fungi, and protists are similar in terms of their ribosomes, cytoskeletons, and cell membranes. As a result, it is more challenging to develop medications that target protozoans and fungi in the same way that antibiotics target prokaryotes.
Fungicides have relatively limited modes of action. Because fungi have ergosterols(instead of cholesterol) in their cell membranes, the different enzymes involved in sterol production can be a target of some medications. The azole and morpholine fungicidesinterfere with the synthesis of membrane sterols. These are used widely in agriculture (fenpropimorph) and clinically (e.g., miconazole). Some antifungal medications target the chitin cell walls of fungi. Despite the success of these compounds in targeting fungi, antifungal medications for systemic infections still tend to have more toxic side effects than antibiotics for bacteria.
Clinical Focus: Part 3
Sarah is relieved the ringworm is not an actual worm, but wants to know what it really is. The physician explains that ringworm is a fungus. He tells her that she will not see mushrooms popping out of her skin, because this fungus is more like the invisible part of a mushroom that hides in the soil. He reassures her that they are going to get the fungus out of her too.
The doctor cleans and then carefully scrapes the lesion to place a specimen on a slide. By looking at it under a microscope, the physician is able to confirm that a fungal infection is responsible for Sarah’s lesion. In Figure \(10\), it is possible to see macro- and microconidia in Trichophyton rubrum. Cell walls are also visible. Even if the pathogen resembled a helminth under the microscope, the presence of cell walls would rule out the possibility because animal cells lack cell walls.
The doctor prescribes an antifungal cream for Sarah’s mother to apply to the ringworm. Sarah’s mother asks, “What should we do if it doesn’t go away?”
Exercise \(3\)
Can all forms of ringworm be treated with the same antifungal medication?
Key Concepts and Summary
• The fungi include diverse saprotrophic eukaryotic organisms with chitin cell walls
• Fungi can be unicellular or multicellular; some (like yeast) and fungal spores are microscopic, whereas some are large and conspicuous
• Reproductive types are important in distinguishing fungal groups
• Medically important species exist in the four fungal groups Zygomycota, Ascomycota, Basidiomycota, and Microsporidia
• Members of Zygomycota, Ascomycota, and Basidiomycota produce deadly toxins
• Important differences in fungal cells, such as ergosterols in fungal membranes, can be targets for antifungal medications, but similarities between human and fungal cells make it difficult to find targets for medications and these medications often have toxic adverse effects | textbooks/bio/Microbiology/Microbiology_(OpenStax)/05%3A_The_Eukaryotes_of_Microbiology/5.03%3A_Fungi.txt |
Subsets and Splits
No community queries yet
The top public SQL queries from the community will appear here once available.