I. Normal Human-Microbial Interactions 23.1 Beneficial Human–Microbial Interactions 23.2 Microflora of the Skin

23.1 Beneficial Human–Microbial Interactions Most microorganisms are benign Few contribute to health, and fewer pose direct threats to health Normal microbial flora (“microflora” or “microbiome”) Microorganisms usually found associated with human body tissue

23.1 Beneficial Human–Microbial Interactions Humans are colonized by microorganisms at birth Acquire microbes via birth canal-lactobacillus Trends Mol Med. Author manuscript; available in PMC 2015 Jun 13. Published in final edited form as: Trends Mol Med. 2015 Feb; 21(2): 109–117. Published online 2014 Dec 11. doi:  10.1016/j.molmed.2014.12.002 PMCID: PMC4464665 NIHMSID: NIHMS695563 The infant microbiome development: mom matters Noel T. Mueller,1,2 Elizabeth Bakacs,3 Joan Combellick,4 Zoya Grigoryan,3 and Maria G. Dominguez-Bello3 Author information ► Copyright and License information ► Scientists swab C-section babies with mothers' microbes Newborns were exposed experimentally to vaginal microbes to restore the microbiomes they missed.

23.1 Beneficial Human–Microbial Interactions Continued exposure throughout life Evolutionary perspective??? Clin Exp Immunol. 2010 Apr; 160(1): 1–9. The ‘hygiene hypothesis’ for autoimmune and allergic diseases: an update H Okada, C Kuhn, H Feillet, and J-F Bach Gut. 2005 Mar; 54(3): 317–320. Microbes, immunoregulation, and the gut G A W Rook and L R Brunet ‘old friends hypothesis’

23.2 Microflora of the Skin-a sample habitat The skin surface varies greatly in chemical composition and moisture content Three microenvironments Dry skin Moist skin Sebaceous skin Skin microflora examined by genomic methods 19 phyla detected Each microenvironment shows a unique microbiota (Figure 23.2)

Figure 23.2 Figure 23.2 Normal skin microflora. Others 1% Bacteroidetes 6.3% Gram negative Proteobacteria 16.5% Actinobacteria 51.8% Other Firmicutes 24.4% Flavobacteriales Betaproteobacteria Gram positive Corynebacteria Figure 23.2 Normal skin microflora. Staphylococci Propionibacteria Figure 23.2

Distribution of microflora on surface of human body

II. Pathogenesis 23.6 Pathogenicity and Virulence 23.7 Adherence 23.8 Invasion, Infection, and Virulence Factors 23.9 Exotoxins 23.10 Endotoxins

23.6 Pathogenicity and Virulence Pathogen-a disease-causing microbe Pathogenicity-the ability of a pathogen to inflict damage on the host Virulence is the relative ability of a pathogen to cause disease Measure of pathogenicity variable Pathogens use various strategies to establish virulence (Figure 23.9) Opportunistic pathogen-causes disease only in the absence of normal host resistance

23.6 Pathogenicity and Virulence Measuring virulence Virulence can be estimated from experimental studies of the LD50 (lethal dose50) The amount of an agent that kills 50% of the animals in a test group (Figure 23.10) Highly virulent pathogens show little difference in the number of cells required to kill 100% of the population as compared to 50% of the population

23.6 Pathogenicity and Virulence Attenuation The decrease or loss of virulence Toxicity Organism causes disease by means of a toxin that inhibits host cell function or kills host cells Toxins can travel to sites within host not inhabited by pathogen Exo-, endo-, entero- toxins

23.6 Pathogenicity and Virulence Invasiveness Ability of a pathogen to grow in host tissue at densities that inhibit host function Can cause damage without producing a toxin Many pathogens use a cocktail of adherence, toxins, invasiveness, and other virulence factors to enhance pathogenicity

23.7 Adherence First step in disease Adherence is ability of microbe to stick to a surface Pathogenic bacteria that initiate infection must adhere to epithelial cells through interactions between molecules on the surfaces of the pathogen and the host cell (Figure 23.12)

23.7 Adherence Bacterial adherence can be facilitated by Extracellular macromolecules that are not covalently attached to the bacterial cell surface Examples: slime layer, capsule (Figure 23.13) Fimbriae and pili (Figure 23.14) Examples: pathogenic strains of E. coli, Neisseria gonorrheae, Shigella spp. “adherence factors” or “adhesins”

Figure 23.12 Adherence of pathogens to tissues.

23.8 Invasion, Infection, and Virulence Factors The initial inoculum of a pathogen is insufficient to cause host damage The pathogen must multiply and colonize the tissue The availability of nutrients is most important in affecting pathogen growth Pathogens may grow locally at the original site or invade throughout the body

23.8 Invasion, Infection, and Virulence Factors Infection: any situation in which a microorganism (not a member of the local flora) is established and growing in a host Bacteremia: the presence of bacteria in the bloodstream Septicemia: bloodborne systemic infection May lead to massive inflammation, septic shock, and death

23.8 Invasion, Infection, and Virulence Factors Virulence factor: any microbial substance that enhances pathogenicity Virulence factors may be: Antibiotic resistance compounds Enzymes that enhance virulence by breaking down or altering host tissue to provide access to nutrients Example: hyaluronidase Enzymes that protect the pathogen by interfering with normal host defense mechanisms Example: coagulase Toxic substances released outside the pathogen: exotoxins Example: neurotoxins

23.8 Invasion, Infection, and Virulence Factors Example of enzyme as virulence factor: streptokinase Produced by Streptococcus pyogenes Strep throat, scarlet fever, impetigo, others Streptokinase is an enzyme released by the pathogen It initiates a cascade that leads to the breakdown of fibrin in blood clots Enhances invasiveness of pathogen Was used as medication to help dissolve blood clots

23.9 Exotoxins Exotoxins Proteins released from the pathogen cell as it grows Three categories based on mechanism of action Cytotoxins AB toxins Superantigen toxins

23.9 Exotoxins Cytotoxins (cytolytic toxins) Work by degrading cytoplasmic membrane integrity, causing cell lysis and death Toxins that lyse red blood cells are called hemolysins (Figure 23.18) Staphylococcal α-toxin induces apoptosis and lyses erythrocytes (Figure 23.19)

Efflux of cytoplasmic components Out Cytoplasmic In membrane RCSB PDB 3ANZ Efflux of cytoplasmic components Out Figure 23.19 Staphylococcal α-toxin. Cytoplasmic membrane In α-Toxin pore Influx of extracellular components

Clostridium perfringens alpha toxin Degrades plasma membrane Leads to cell and tissue death “gas gangrene” Zone of hemolysis

23.9 Exotoxins AB toxins Consist of two subunits, A and B Work by binding to host cell receptor (B subunit) and transferring damaging agent (A subunit) across the cell membrane (Figure 23.20) Examples: diphtheria toxin, tetanus toxin, botulinum toxin

23.9 Exotoxins Clostridium tetani and Clostridium botulinum produce potent AB exotoxins that affect nervous tissue “botulism” food poisoning due to anaerobic production of toxin-often in home canned foods Botulinum toxin or Botox consists of several related AB toxins that are the most potent biological toxins known (Figure 23.21); tetanus toxin is also an AB protein neurotoxin (Figure 23.22)

BTX is the most lethal toxin known Excitation signals from the central nervous system A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A Muscle A A Figure 23.21 The activity of botulinum toxin from Clostridium botulinum. Normal Acetylcholine (A) induces contraction of muscle fibers Botulism Botulinum toxin, , blocks release of A, inhibiting contraction Figure 23.21

23.9 Exotoxins Enterotoxins Exotoxins whose activity affects the small intestine Generally cause massive secretion of fluid into the intestinal lumen, resulting in vomiting and diarrhea Example: cholera toxin (Figure 23.23) Causative agent-Vibrio cholera in contaminated food or water

Figure 23.23 Figure 23.23 The activity of cholera enterotoxin. 1. Normal ion movement, Na+ from lumen to blood, no net Cl– movement 4. Na+ movement blocked, net Cl– movement to lumen Blood Intestinal epithelial cells Lumen of small intestine GM1 5. Massive water movement to the lumen; cholera symptoms 2. Infection and toxin production by V. cholerae Cholera toxin AB form GM1 Vibrio cholerae cell B A 3. Activation of epithelial adenylate cyclase by cholera toxin Figure 23.23 The activity of cholera enterotoxin. A subunits Cholera toxin B subunit Adenylate cyclase ATP Cyclic AMP Figure 23.23

23.9 Exotoxins Superantigens An antigen is a substance that can provoke a response by by the immune system Act by hyperstimulation of immune system Promiscuous binding to immune cell receptors Staphylococcus, Streptococcus S. aureus food poisoning - more than just GI tract

23.10 Endotoxins Endotoxin The lipopolysaccharide portion of the cell envelope of certain gram-negative Bacteria, which is a toxin when solubilized Not secreted but released upon cell breakdown Generally less toxic than exotoxins Neisseria gonorrheae lipo-oligo-saccharides

23.8 Invasion, Infection, and Virulence Factors Salmonella species encode a large number of virulence factors of many types (Figure 23.17) Several genes that direct invasion are clustered together on the chromosome as pathogenicity islands Another Salmonella pathogenicity island contains genes that promote a more systemic disease Salmonella also contains resistance plasmids (R plasmids)

Figure 23.17 Enterotoxin (diarrhea) Siderophores (iron uptake) Injectisome (inv and prg products form complex) Type Ι fimbriae (adherence) Endotoxin in LPS layer (fever) Virulence plasmid SPΙ2 SPΙ1 Anti- phagocytic proteins induced by oxyR Cytotoxin (inhibits host cell protein synthesis; Ca2+ efflux from host cell; adherence) Figure 23.17 Virulence factors in Salmonella. O antigen (inhibits phagocyte killing) Vi capsule antigen; inhibits complement binding Pathogenicity islands on chromosome Flagellum (motility) H antigen (adherence; inhibits phagocyte killing) Figure 23.17