1. Normal Flora and Bacterial Pathogenesis Pin Ling ( 凌 斌 ), Ph.D. Department of Microbiology & Immunology, NCKU ext 5632 lingpin@mail.ncku.edu.tw References: 1. Murray, P. et al., Medical Microbiology (5 th edition) 2. Samuel Baron, Medical Microbiology (4 th edition)

2. Outline Normal Flora (Commensal Microbes) Introduction Significance of the Normal Flora Distribution of the Normal Flora Bacterial Pathogenesis Introduction Host Susceptibility Pathogenic Mechanisms Virulence Factors

3. Transient colonization; Permanent colonization; Disease Colonization vs. infection Colonization: establishment of a site of reproduction of microbes on a person without necessarily resulting in tissue invasion or damage. Infection: growth and multiplication of a microbe in or on the body of the host with or without the production of disease. Outcomes of exposure to a microorganism: Normal Flora and Pathogenesis

4. Introduction of Normal Flora 1.A diverse microbial flora => Human body Area: the skin and mucous membranes Time: shortly after birth until death Number: 10 14 bacteria =>10 13 host cells 2. Normal flora may: a. Aid the host b. Harm the host (in sometimes) c. Exist as commensals (no effect to the host) 3.Viruses and parasites => NOT normal microbial flora Most investigators consider that they are not commensals and do not aid the host.

5. Significance of Normal Flora-I 1.The effect of the normal flora on the host was not well understood until germ-free animals became available. Cesarean Section => Germ-free animals => Isolators w/o detectable pathogens (viruses, bacteria & others) 2.Two interesting observations: a. the germ-free animals lived almost twice as long as their conventionally maintained counterparts. b. the major causes of death were different in the two groups.

6. Germ-free AnimalsRegular Counterparts LifespanTwiceOne Cause of death Intestinal AtoniaInfection Anatomic & Physiological Changes 1. Alimentary lamina propria under- developed 2. No Ab 3. Intestinal epithelial cell renewal rate down half Germ-free Animals vs Counterparts

7. Significance of Normal Flora-II Normal flora may aid the host in several ways: Aid in digestion of food Help the development of mucosa immunity Protect the host from colonization with pathogenic microbes. 10 6 pathogenic microbes GI infection w/ normal flora GI infection w/ reduced flora after Streptomycin treatment 10 pathogenic microbes

8. Normal Flora competing w/ Invading Pathogens Adopted from Samuel Baron “Medical Microbiology”

9. Normal flora may act as opportunistic pathogens Especially in hosts rendered susceptible by: 1. Immuno-suppression (AIDS & SCID) 2. Radiation therapy & Chemotherapy 3. Perforated mucous membranes 4. Rheumatic heart disease…etc.

10. Respiratory tract and head outer ear, eye, mouth, oropharynx, nasopharynx Sterile sites: sinuses, middle ear, brain, lower respiratory tract (trachea, bronchiole, lung) Gastrointestinal tract esophagus, stomach, small intestine, large intestine Genitourinary system anterior urethra, vagina Sterile sites: bladder, cervix, uterus Skin Sites of human body that the normal flora microbes colonize

11. Adopted from Smaul Baron “Medical Microbiology” Distribution of Normal Flora in Human Body

12. 1.Local Environment (pH, temperature, redox potential, O2, H2O, and nutrient levels…). 2.Diet 3.Age 4.Health condition (immune activity…) 5.Antibiotics,…..etc Factors Influencing Normal Flora

13. Outline Normal Flora (Commensal Microbes) Introduction Significance of the Normal Flora Distribution of the Normal Flora Bacterial Pathogenesis Introduction Host Susceptibility Pathogenic Mechanisms Virulence Factors

14. Introduction of Bacterial Pathogenesis 1.Infection: growth and multiplication of a microbe in or on the body of the host with or without the production of disease. 2.The capacity of a bacterium to cause disease reflects its relative pathogenicity. 3. Virulence is the measure of the pathogenicity of an organism. 4. Pathogenesis refers both to the mechanism of infection and to the mechanism by which disease develops.

15. Host Susceptibility 1.Susceptibility to bacterial infections => Host Defenses vs Bacterial Virulence 2.Host Defenses: - Barriers (skin & mucus) – first line - Innate Immune Responses (complement, macrophages & cytokines) – the early stage - Adaptive Immune Responses (Ag-specific B & T cells) – the later stage 3.Host defenses can be comprised by destructing barriers or defective immune response. e.x. Cystic Fibrosis => poor ciliary function => NOT clear mucus efficiently from the respiratory tract => Pseudomonas aeruginosa => serious respiratory distress.

16. Strict pathogens are more virulent and can cause diseases in a normal person. Opportunistic pathogens are typically members of normal flora and cause diseases when they are introduced into unprotected sites, usually occur in people with underlying conditions.

17. Pathological Mechanisms of Bacterial Infections 1.Bacteria-mediated Pathogenesis 2.Host-mediated Pathogenesis 3.Bacterial virulence factors => bacterial factors causing diseases Adopted from Samuel Baron “Medical Microbiology”

18. 1.Transmissibility 2.Adherence to host cells 3. Invasion of host cells and tissue 4.Evasion of the host immune system 5.Toxigenicity A bacterium may cause diseases by 1.Destroying tissue (invasiveness) 2.Producing toxins (toxigenicity) 3.Stimulating overwhelming host immune responses Characteristics of Pathogenic Bacteria

19. Bacterial Virulence Mechanisms

20. Bacterial virulence factors Adhesins Pili (fimbriae) Nonfimbrial adhesins Invasion of host cells Tissue damage Growth byproducts Tissue-degrading enzymes Immunopathogenesis Toxins Exotoxins (cytolytic enzymes and A-B toxins); enterotoxins;A-B toxins superantigens;superantigens endotoxin and other cell wall componentsendotoxin Antiphagocytic factors Intracellular survival Antigenic heterogeneity Antigenic variation Phase variation Iron acquisition Siderophores Receptors for iron-containing molecules Resistance to antibiotics

21. Mechanisms of acquiring bacterial virulence genes

22. Entry into the human body : infection: shedding The most frequent portals of entry are the sites where mucous membranes meet with the skin. Abnormal areas of mucous membranes and skin are also frequent sites of entry. Routes: Ingestion, inhalation, trauma, needles, catheters, arthropod bite, sexual transmission

23. Microbial defenses against host immunologic clearance Encapsulation (Inhibition of phagocytosis and serum bactericidal effect) Antigenic mimicry Antigenic masking Antigenic or phase variation Intracellular multiplication Escape phagosome Inhibition of phagolysosome fusion Resistance to lysosomal enzymes Production of anti-immunoglobulin protease Inhibition of chemotaxis Destruction of phagocytes

24. Mechanisms for escaping phagocytic clearance and intracellular survival

27. Endotoxin-mediated toxicity Fever, leukopenia followed by leukocytosis, activation of complement, thrombocytopenia, disseminated intravasacular coagulation, decreased peripheral circulation and perfusion to major organs (multiple organ system failure), Shock and death. Peptidoglycan, teichoic and lipoteichoic acids of gram- positive bacteria stimulate pyrogenic acute phase responses and produce endotoxin-like toxicity Back

28. Lipid A of lipopolysaccharide is responsible for endotoxin activity Pathogenesis of sepsis (septicemia)

29. Adopted from Samuel Baron “Medical Microbiology”

30. Environmental factors often control the expression of the virulence genes. Common factors: temperature, iron availability, osmolarity, growth phase, pH, specific ions, specific nutrient factors, bacterial cell-density, interaction with host cells. Regulation of bacterial virulence factors

31. The severity of a disease is determined by the inoculum size of the pathogen and the host conditions, such as congenital defect, immunodeficiency states and other disease-related conditions. The symptoms of a disease are determined by the function of the tissue affected.

32. Transmission of infection Carrier: a person or animal with asymptomatic infection that can be transmitted to another person or animal. The clinical symptoms of diseases produced by microbes often promote transmission of the agents. Zoonosis: infectious diseases transmitted between animals and men. Hospital- (nosocomial) vs. community-acquired infections By producing asymptomatic infection or mild disease, rather than death of the host, microorganisms that normally live in people enhance the possibility of transmission from one person to another. Many bacteria are transmitted on hands

33. Adhesion Adherence of bacterium to epithelial or endothelial cells allow them to colonize the tissue. Common adhesins: pili (fimbriae), slime, lipoteichoic acid, surface proteins or lectins. Biofilm, formed on a surface by the bacteria that are bound together within a sticky web of polysaccharide, is a special bacterial adaptation that facilitates colonization on the surgical appliances (e.g., artificial valves or indwelling catheters) and dental plaque. It can protect the bacteria from host defenses and antibiotics. Back

34. The bacteria may invade via the M cells Back

35. The A-B toxins Mode of action Inhibition of protein synthesis Hypersecretion Inhibition of neurotransmitter release A chain has the inhibitory activity against some vital function B chain binds to a receptor and promotes entry of the A chain In many cases the toxin gene is encoded on a plasmid or a lysogenic phage Back

36. SUMMARY 1. Normal flora may aid the host in several ways: Aid in digestion of food Help the development of mucosa immunity Protect the host from colonization with pathogenic microbes 2. Normal flora may act as opportunistic pathogens. 3. Susceptibility to bacterial infections depends on the balance between host defenses and bacterial virulence. 4. Pathogenic mechanisms of bacterial infections include Bacteria-mediated Pathogenesis Host-mediated Pathogenesis

37. Back

39. Transfer of mobile genetic elements within or between species can result in transfer of virulence factors (e.g., pathogenicity islands). Recombination may occur between extrachromosomal DNA and the chromosome to generate new bacterial clone. Different clonal types of a certain species may cause different diseases. Clonality of bacterial pathogens Various virotypes of E. coli