Study Guide, Exam 1 2021

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MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide Below are questions covering the lectures through September 7 th . You should be able to answer these questions for the exam. The questions on the exam are a subset of the questions in this study guide, although slight variations of the questions may be used. 1. Pathogenesis Define pathogenesis. The development of a disease, chain of events leading to that disease, molecular mechanism leading to disease. The infection process can be divided into four stages, what are they? 1. Invasion 2. Multiplication 3. Spread 4. Production of Disease Pathogenic bacteria often enter our bodies at mucosae found in several body sites. Name three such sites. Mouth, eyes, and nose. Mucosae of humans secrete chemicals to combat infectious microbes. Name three proteins secreted by mucosae that combat infectious microbes and describe the antimicrobial activity of each of the three proteins. 1. Lysozyme secreted in sweat and tears. Its enzymatic activity attacks cell walls of bacteria 2. Collectins are proteins in blood, lymph fluid, and tissues that bind microbial surface carbohydrates. Growth inhibition, aggregation, or tagging for phagocytosis. 3. Mucins + colloid bind microbes, enzymes attack microbes, antibodies bind micorbes. 1

MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide Name two chemicals made in mucosae that are not proteins that combat infectious microbes and describe how they work. 1. Defensins are antimicrobial peptides found in secretions of mucosae and skin. They insert into microbial membranes, forming pores, allowing efflux of ions and nutrients, killing the cell. 2. Cathelicidins are antibacterial peptides found in mucosal secretions. They disrupt microbial membranes killing the cells. They also signal and activate many parts of the host immune system. Describe the kind and shape of cells that line the respiratory tract. Epithelial cells: Pseudo-stratified columnar epithelium Mucus is secreted by epithelial cells of the gut, respiratory tract, and urogenital tract. Describe what is mucus is and how it combats infectious microbes. Slimy and sticky, it entraps microbes so they can be expelled. Colloid (suspension) of water plus mucins (proteins) binds microbes, enzymes attack microbes, antibodies bind microbes. What is a physical barrier mechanism unique to the respiratory tract that can help expel microbes? Cilia Describe three ways pathogens spread inside an infected host (do not discuss enzymes used in spreading). 1. Blood System: Blood vessels, Lymphatic vessels, Tropism 2. Cell-to-cell contact 3. Spread within body cavities When an infected host has a disease, what has happened to the host and what are some ways that the pathogens have caused this? Toxigenesis Intracellular pathogens 2

MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide Physical blockage 2. Host Immunity Name five kinds of mature cells that function in the innate immune system. 1. Neutrophils (PMN) 2. Monocytes 3. Macrophages 4. Dendritic Cells 5. Eosinophils Describe four major steps in the process of phagocytosis. 1. Bind 2. Internalize 3. Kill 4. Degrade pathogen Describe one way that macrophages are activated by cytokines and one way they are activated without cytokines. Cytokines: Without Cytokines: What are four physiological features that happen in activated macrophages? 1. Increased killing activity against microbes 2. they become enlarged 3. They are more motile 4. Express more MHCII proteins 3

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MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide What are three different functions of complement proteins? 1. Opsonization; 2. Signaling; 3. Cell lysis. What is the function of Fc-receptors in the innate immune system? Fc-receptors on immune cells bind antibodies that have opsonized microbes Antibodies bind microbe  macrophage Fc Receptor bind antibody/microbe  Phagocytosis What does PAMP mean? Give three examples. Pathogen-associated molecular patterns (PAMPs) Includes; Carbohydrates (LPS, mannose), nucleic acids (bacterial or viral DNA or RNA), Lipoproteins. Toll-like receptors (TLRs) recognize PAMPs. Where are TLRs located? Exterior of the host cytoplasmic membrane, endosomes (phagosome) Name three PAMPs that Toll-like receptors bind to. 1. Lipopolysaccharide (LPS)  TLR4 2. 3. NOD1 and NOD2 recognize intracellular pathogens by binding to what bacterial components? Bacterial Peptidoglycan  NOD1 = meso-DAP & NOD2 = muramyl dipeptide After binding to bacterial components, what responses do NOD1 and NOD2 elicit? NOD1 & NOD2 have caspase activation and recruitment domains (CARDs) that control a signaling pathway that activates NF-kB and MAP kinase pathways. NF-kB and MAP Kinase activate inflammatory response. 4

MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide Besides secreting cytokines, what is the primary function of cytolytic T- lymphocytes (CTLs)? Cytotoxic T lymphocytes are CD8+ T cells that recognize when a specific host cell is infected with intracellular microbes. What type of immune cell activates B cells and what do activated B cells produce to directly attack microbes? CD4+ T cells that activate B cells via B-cell receptors. Activated B cells produce antibodies. What are the two major types of T cells (as discussed in class)? CD4+ T cells & CD8+ T cells After a neutrophil phagocytizes a bacterial cell, what does it do to attack that cell, including molecular components involved, and how fast does this occur? Neutrophils kill cells by oxidative bursts (like macrophages), making superoxide anion O2-, which can also be converted to H2O2, and then to hypochlorous acid (HOCl), which reacts with amines forming stable microbicidal N-chloramines. Describe four steps in the process for identifying a virus-infected cell by our immune system. 1. 2. 3. 4. What are two ways that antibodies help eliminate invading bacteria? Bind to specific antigens on surface of microbes to cause opsonization and increased phagocytosis by cells of the innate immunity system. What are the four generalized functions of cytokines? 5

MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide 1. Differentiation of immature immune cells into mature immune cells. 2. Activation of some immune cells, increasing their ability to fight pathogens. 3. Induce secretion of additional cytokines. 4. Act as signals (chemokines) that allow migration of immune cells to sites of infection. What is immune memory? Adaptive immunity continues to produce CTLs and antibodies that will attack the same microbes if they try to invade the host a second time, with more rapid clearance. The basis of vaccination. What cellular immune responses occur in SCID mice infected with Listeria and what cellular immune responses are absent? Macrophages, PMNs, and NK cells are active in SCID mice. CD4+ T cells and CD8+ T cells are absent. What is the outcome of SCID mice infected with Listeria ? Infection is controlled but bacteria persist and there is no sterilization. What cells display the MHC I system and what cells display the MHC II system? MHC I: Displayed by most cells MHC II: Displayed by antigen presenting cells (Macrophages & dendritic cells) How is the MHC II system restricted to carry only antigens for its own pathway and not for the MHC I system? Peptides bind to groove of complex by displacing CLIP Describe the process that delivers antigens to the MHC I system. Intracellular antigens, such as virus or tumor antigens, are processed into peptides by the proteasome. Peptides are transported into the ER by TAP 6

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MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide where they are loaded into the groove of the MHC class I complex, which is composed of a heavy chain Beta2m. MHC class I complexes present antigens on the cell surface to CD8+ T cells. Describe the process that delivers antigens to the MHC II system. Antigens from extracellular sources, such as bacterial antigens, are processed by endolysosomal enzymes into peptides. These peptides bind to the groove of the MHC class II complex by displacing the class II- associated invariant chain peptide (CLIP), which is derived from the MHC class II-associated invariant chain (Ii). The MHC class II complex presents antigens to CD4+ T cells. Describe the structure of TAP and its function. Transporter associated with Antigen Processing complex (TAP) transports peptides generated by proteasome into the ER. What molecules on T cells recognize the MHC I::antigen complex? CD8+ What molecules on T cells recognize the MHC II::antigen complex? CD4+ What is the result of those recognitions? Activated B cells 3. Antigenic Variation, Invasins and Evasins What is the definition of an invasin and how do invasins differ from exotoxins? Invasins are enzymes that act locally to damage host cells and facilitate spread of the pathogen Invasins act at short range while toxins may act at remote sites. Invasins may not actually kill host cells while toxins are often cytotoxic. Toxins are typically more specific and potent. 7

MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide Bacteria secrete many substances during the invasion of hosts called “invasins”. One class of invasins consists of spreading factors. Name three spreading factors discussed in class and their biochemical activities. 1. Hyaluronidase 2. Collagenase 3. Neruaminidase Another class of invasions consists of those that help evade complement- dependent killing. Describe two such factors and their biochemical activities. 1. Elastase is a protease that degrades complement components. 2. Capsules protect bacteria from complement activation and killing. Capsules contain sialic acid which mimics host structures. Staphylococcus aureus uses the enzyme coagulase to protect itself from the immune system when infecting blood. What does coagulase do and how does this protect S. aureus from the immune system? Coagulase is an enzyme that converts fibrinogen to fibrin, which causes blood clotting, this provides an antigenic disguise when it clots fibrin on the bacteria cell surface How do some pathogenic bacteria to use sialic acid to avoid the host immune system? Sialic acid allows a pathogenic bacteria to disguise its own antigenic surface components How does a soluble antibody decoy help an invading microbe? Soluble antigens such as LPS are able to combine with and neutralize antibodies. Antibodies bind soluble molecules instead of microbial cells. Some pathogens cause immunosuppression in the infected host. Describe one of the mechanisms discussed in class. LcrV protein is secreted by Yersinia. LcrV signals host cells to secrete IL- 10. IL-10 cytokine causes immunosuppression. 8

MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide Some bacteria have special mechanisms for antigenic variation . What advantage does this provide the pathogen? The ability for a microbe to alter the antigenic character of its surface components, to express several antigenic variants of a cellular component. Neisseria gonorrhoeae can express numerous antigenic variants of pilin, which is the major protein of the type IV pili that function in adherence to various mammalian cell types. The type IV pilin protein is encoded by the pilE1 gene (sometimes also a pilE2 gene). Describe what information is in a pilS gene, in particular how it differs from the information in pilE . pilS genes are pseudogenes (silent genes), they have variants of pilin protein coding sequences but that are not expressed. pile are complete genes and express pilin protein. Describe the process for changing the information present in a pilE gene. Copy of a silent gene can carry antigenic variants of pilin protein that can replace parts of an expressed gene, which changes the sequence of the expressed pilin protein How frequently does antigenic variation for pilin occur and roughly how much variation is theoretically possible? Change in pilin expression occurs once in 10 2 to 10 3 generations, hundreds of thousands of pilin antigenic variants are possible. The Opa protein (protein II) is encoded in eleven different copies of the opa gene. Sometimes an Opa protein is made from a particular gene and sometimes it is not. What determines whether a particular Opa protein is made or not and what is the molecular mechanism controlling this? Translation of opa mRNA into functional Opa protein depends on number of CTCTT repeats (ranging from 3-13) that occur in opening reading frame at the beginning (5' end) of each mRNA, in the leader region. Salmonella bacteria produce antigenic variants of its flagella. Describe the structure of the operon with the H2 gene, including the function of each gene product. 9

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MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide What the expression of the H1 gene? Inversion of the promoter for the H2/rh1 operon indirectly allows for the expression of the H1 gene. Describe the molecule event that occurs when a Salmonella bacterium switches from expressing H2 to not expressing H2. Inversion prevents transcription of the H2 structural gene by the H2 promoter. The inversion also prevents the expression of rh1. Without rh1 repressor, H1 gene is no longer repressed and is thus transcribed from the H1 promoter. 4. Extracellular Matrix and Host Cell Surfaces What are two functions that the ECM contributes to? Mechanical: tensile and compressive strength and elasticity Protection: buffering against extracellular change and retention of water What is one protein that forms fibrils in the ECM? Collagen What is one non-protein that forms fibrils in the ECM? Hyaluronan What ECM component does Staphylococcus aureus bind to which helps it cause arthritis? Explain why. Binds to collagen (Type II) in cartilage What are the two primary proteins that function in creating elasticity of lungs, skin, and blood vessels? Highly cross-linked elastin protein and fibrillin 10

MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide What molecular interactions are involved when Neisseria bacteria use Opa to bind host cells? OpaA proteins on Neisseria cells bind to GAG which then binds to Vn. Vn can bind to host cell with its RGB motif thus allowing Bacteria to bind indirectly to the host cells How does fibrinogen function in blood clotting? Fibrinogen is a soluble plasma glycoprotein that when cleaved by thrombin during blood-clotting cascade forms fibrin. Fibrin alongside other adhesion proteins constitutes the majority of the initial ECM for sealing a wound site. Staphylococcus aureus binds fibrinogen. How does this contribute to the endovascular infections caused by these bacteria? S. aureus binds fibrinogen which constitutes a bridge to endothelial cells because fibrinogen is able to bind to these cells. What are the major steps involved (as described in your handouts) in removal of blood clots after healing? Degrading of fibrin (fibrinolysis) 1. Plasminogen is incorporated into the fibrin clot 2. Host cell produces urokinase type and tissue-type (t-PA) plasminogen activators that do proteolysis on plasminogen protein to form plasmin. 3. Plasmin digests fibrin causing fibrinolysis What are two different mechanisms that microbes have to prevent blood clotting or to remove blood clots? Yersinia pestis has an invasion that acts as a bacterial tissue plasminogen activator, the production of plasmin disrupts blood clotting. Borrelia burgdorferi (Lyme disease) induces monocytes to produce host urokinase-type plasminogen activator. What are the benefits for the pathogens when they prevent blood clotting or remove blood clots? 11

MICR 470/670, Microbial Pathogenesis, Exam 1 Study Guide When a blood clot is prevented or removed it allows promotes the invasion and spread of pathogens. What are three different mechanisms that pathogens have to bind to host integrins? 1. Ligand Mimicry 2. Ancillary Recognition 3. Bridging and Masking What is integrin mimicry? Some microbes are capable of expressing proteins that look like integrins, this allows that bacteria to bind to the ECM. 12

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