TORTORA FUNKE CASE Microbiology AN INTRODUCTION EIGHTH EDITION

TORTORA • FUNKE • CASE Microbiology AN INTRODUCTION EIGHTH EDITION B. E Pruitt & Jane J. Stein Chapter 15 Microbial Mechanisms of Pathogenicity

Microbial Mechanisms of Pathogenicity • Pathogenicity The ability to cause disease • Virulence The extent of pathogenicity Many properties that determine a microbe’s pathogenicity or virulence are unclear or unknown But, only when a microbe overpowers the hosts defenses, disease results! They need to gain entry, adhere, penetrate and cause damage to cause disease.

Portals of Entry of a Microbe • Need to adhere, penetrate, and then cause damage • Gain access via portal of entry and may a have preferred portal of entry - Streptococcus pneumoniae via GI tract? Small pox via vein? Portals of Entry: • Mucous membranes • Respiratory • GI • Urogenital • conjunctiva • Skin • Tough so rare - Necator americanus - hookworm • Parenteral route • Puncture or injection

Mucous Membranes: Respiratory • Respiratory Tract • microbes inhaled into mouth or nose in droplets of moisture or dust particles • Easiest and most frequently traveled portal of entry • Common cold • Flu • Tuberculosis • Whooping cough • Pneumonia • Measles • Strep Throat • Diphtheria

Mucous membranes: G. I. Tract • Salmonellosis • Salmonella sp. • Shigellosis • Shigella sp. • Cholera • Vibrio cholorea • Ulcers Fecal - Oral Diseases • Helicobacter pylori • • Botulism • Clostridium botulinum These pathogens enter the G. I. Tract at one end and exit at the other end. • Spread by contaminated hands & fingers or contaminated food & water • Poor personal hygiene.

Mucous Membranes of the Genitourinary System - STD’s Gonorrhea Neisseria gonorrhoeae Syphilis Treponema pallidum Chlamydia trachomatis HIV Herpes Simplex II

Mucous Membranes: Conjunctiva • Conjunctiva – • mucous membranes that cover the eyeball and lines the eyelid • Trachoma • Chlamydia trachomatis

2 nd Portal of Entry: Skin • Skin - the largest organ of the body. When unbroken is an effective barrier for most microorganisms. • Some microbes can gain entrance thru openings in the skin: hair follicles and sweat glands

3 rd Portal of Entry: Parenteral Microorganisms are deposited into the tissues below the skin or mucous membranes • Punctures • injections • bites • scratches • surgery • splitting of skin due to swelling or dryness

Preferred Portal of Entry • Just because a pathogen enters your body it does not mean it’s going to cause disease. • pathogens - preferred portal of entry • Small pox via variolation • Streptococcus pneumoniae • if inhaled can cause pneumonia • if enters the G. I. Tract, no disease • Salmonella typhi • if enters the G. I. Tract can cause Typhoid Fever • if on skin, no disease

Numbers of Invading Microbes • ID 50: Infectious dose for 50% of the test population • LD 50: Lethal dose (of a toxin) for 50% of the test population • Example: ID 50 for Vibrio cholerea 108 cells (100, 000 cells) • ID 50 for Inhalation Anthrax - 5, 000 to 10, 000 spores ? ?

ID 50 and LD 50 for Bacillus anthracis Portal of entry Skin ID 50 ? ? ? endospores Inhalation 10, 000 -20, 000 endospores Ingestion 250, 000 -1, 000 endospores

Key traits for a pathogen The ability to: • 1. Adherence • To host surfaces and not be washed off • 2. Avoid phagocytosis • Prevent host defenses from destroying • 3. Penetrate • Get into host and spread • 4. Produce Enzymes • Spread, prevent host defenses and cause damage at or near site of infection • 5. Produce Toxins • Cause damage at distant site

Adherence • Adhesions/ligands bind to receptors on host cells so won’t get flushed off. • Mechanisms to adhere and avoid host defenses: • Glycocalyx Streptococcus mutans Dextran (plaque) • Waxes Mycobacteria • Fimbriae Escherichia coli • M protein Streptococcus pyogenes • Tapered end w/ hooks Treponema pallidum

Figure 15. 1 b-c Adherence. E. coli bacteria (yellow-green) on human urinary bladder cells Bacteria (purple) adhering to human skin

Capsules Prevent phagocytosis and help with attachment (adherence) • Streptococcus pneumoniae • Klebsiella pneumoniae • Haemophilus influenzae • Bacillus anthracis • Streptococcus mutans • Yersinia pestis

Enzymes to help penetration Many pathogens secrete enzymes that contribute to their pathogenicity: • Increase virulence by use of enzymes • And avoid phagocytosis • Coagulase - Coagulate blood - wall off from host make boil • Kinases - Digest fibrin clot - allow spreading streptokinase and staphylokinase • Hyaluronidase - Hydrolyses hyaluronic acid connective tissue • Collagenase - Hydrolyzes collagen • Hemolysins - lyse RBC’s

Hemolysins Alpha Hemolytic Streptococci - secrete hemolysins that cause the incomplete lysis or RBC’s Beta Hemolytic Streptococci - secrete hemolysins that cause the complete lysis of RBC’s

Leukocidins • Enzymes that attack certain types of WBC’s • 1. Kills WBC’s which prevents phagocytosis • 2. Releases & ruptures lysosomes • lysosomes - contain powerful hydrolytic enzymes which then cause more tissue damage

Enzymes: Necrotizing Factor “Flesh Eating Bacteria” Necrotizing fasciitis causes death (necrosis) to tissue cells

Summary of How Bacterial Pathogens Penetrate Host Defenses • 1. Adherence • 2. Capsule • 3. Enzymes • leukocidins • Hemolysins • Coagulase • Kinases • Hyaluronidase • Collagenase • Necrotizing Factor

Penetration into the Host Cell Figure 15. 2

Pathogens may cause damage to host • Direct damage in the immediate vicinity • • Far removed from site of invasion by toxins • • Grow, destroy healthy cells and/or multiply and clog cells and passageways Toxins spread through blood and lymph By hypersensitivity • The host’s reaction may cause the damage

Toxins Provide properties to spread and cause damage to the host. Compare endotoxins and exotoxins • Endotoxins from inside the cell. Released upon cell lysis. • Exotoxins are secreted out of the cell during cell life. • Toxin Substances that contribute to pathogenicity • Toxigenicity Ability to produce a toxin • Toxemia Presence of toxin the host's blood • Toxoid Inactivated toxin used in a vaccine • Antitoxin Antibodies against a specific toxin

Toxins Portal of Entry Botulinum Shiga toxin Staphylococcal enterotoxin ID 50 0. 03 ng/kg 250 ng/kg 1350 ng/kg

Exotoxins Mostly seen in Gram (+) Bacteria Most gene that code for exotoxins are located on plasmids or phages Figure 15. 4 a

Exotoxin Source Metabolic product Chemistry Fever? Neutralized by antitoxin LD 50 Mostly Gram + By-products of growing cell Protein Water soluble / Heat Liable No Yes Small - Very potent 1 mg of Clostridium botulinum toxin can kill 1 million guinea pigs

Exotoxins - three types • 1. Cytotoxins • kill cells • 2. Neurotoxins • interfere with normal nerve impulses • 3. Enterotoxins • effect cells lining the G. I. Tract Many toxins have A-B subunit toxins or type III toxins • A - active • Causes change in host • B - binding Figure 15. 5

Exotoxins OR • Superantigens or type I toxins • Cause an intense immune response due to release of cytokines from host cells • Fever, nausea, vomiting, diarrhea, shock, death

Exotoxins • Membrane-disrupting toxins or type II toxins • Lyse host’s cells by: • Making protein channels in the plasma membrane (e. g. , leukocidins, hemolysins) • Disrupting phospholipid bilayer Cholera wenterotoxin • Vibrio cholerae • Gram (-) comma shaped rods

Exotoxins • Corynebacterium diphtheriae • Streptococcus pyogenes • Clostridium botulinum • C. tetani • Vibrio cholerae • Staphylococcus aureus Exotoxin Lysogenic conversion A-B toxin type III. Inhibits protein synthesis. + Membrane-disrupting. Type II Erythrogenic. + A-B toxin. Neurotoxin - flaccid paralysis Botox A-B toxin. Neurotoxin - prevents CNS inhibition - spastic paralysis A-B toxin. Enterotoxin. Stimulates c. AMP to cause severe diarrhea Superantigen. Type I. Enterotoxin. + +

Botox • Botulism • Clostridium botulinum • Gram (+), anaerobic, sporeforming rod, found in soil • works at the neuromuscular junction • prevents impulse from nerve cell to muscle cell • results in muscle paralysis • Botulus – latin word for sausage (first known as sausage disease) C. botulinum does not grow in sausage today mainly due to nitrites added. Infant botulism 250 per yr. , most associated with honey due to little microbial flora in G. I.

Tetanus (Lock Jaw) • Clostridium tetani • Gram (+), spore-forming, anaerobic rod • neurotoxin acts on nerves, resulting in the inhibition of muscle relaxation • Tetanospasmin “spasms” or “Lock Jaw” • 50 cases a yr. in U. S. • 1 million per yr. Worldwide 50% in newborns – because they dress severed umbilical cord with soil, clay or cow dung Tetanospasmin inhibits the release of acetylcholine by interfering with activity of cholinesterase (enzyme that normally breaks down acetylcholine)

Endotoxin Figure 15. 4 b

Endotoxins Source Gram– Metabolic product Present in LPS of outer membrane Chemistry Lipid / Heat Stable Fever? Yes Neutralized by antitoxin No LD 50 Relatively large

Endotoxins - part of the Gram (-) Bacterial cell wall • LPS (Lipopolysaccharides) • O Antigen • Lipid A • Heat Stable (exotoxins are typically heat liable) • Lipid A - Toxin portion of the LPS • responsible for Fever that is associated with many Gram (-) Bacterial infections • Gram (-) cells are “digested” endotoxins are released - fever • Antibiotics • E. coli (0157: H 7) • enterotoxin causes a hemolytic inflammation of the intestines • results in bloody diarrhea

Endotoxins Figure 15. 6

Figure 15. 4 Mechanisms of Exotoxins and Endotoxins. exotoxins endotoxins Exotoxins are proteins produced inside pathogenic bacteria, most commonly grampositive bacteria, as part of their growth and metabolism. The exotoxins are then secreted into the surrounding medium during log phase. Endotoxins are the lipid portions of lipopolysaccharides (LPS) that are part of the outer membrane of the cell wall of gramnegative bacteria (lipid A; see Figure 4. 13 c). The endotoxins are liberated when the bacteria die and the cell wall breaks apart. Cell wall Exotoxin: toxic substances released outside the cell Clostridium botulinum, an example of a grampositive bacterium that produces exotoxins Salmonella typhimurium, an example of a gramnegative bacterium that produces endotoxins Endotoxins: toxins composed of lipids that are part of the cell membrane

Non bacteria pathogens • Viruses • Protozoa • Fungi • Algae • Helminths

Cytopathic Effects of Viruses Table 15. 4

Pathogenic Properties of Fungi • Fungal waste products may cause symptoms • Chronic infections provoke an allergic response • Tichothecene toxins inhibit protein synthesis • Fusarium • Proteases • Candida, Trichophyton • Capsule prevents phagocytosis • Cryptococcus • Ergot toxin • Claviceps • Mycotoxins • Neurotoxins: phalloidin, amanitin • Amanita

Pathogenic Properties of Fungi • Aflatoxin • Aspergillus on peanuts? • Mycotoxins • Neurotoxins: Phalloidin, amanitin • Amanita “death angel” – Liver damage

Pathogenic Properties of Protozoa • Presence of protozoa • Protozoan waste products may cause symptoms • Avoid host defenses by • Growing in phagocytes • Antigenic variation

Pathogenic Properties of Helminths • Use host tissue • Presence of parasite interferes with host function • Parasite's metabolic waste can cause symptoms • Death can cause excessive immune reaction leading to more symptoms

Pathogenic Properties of Algae • Neurotoxins produced by dinoflagellates • Saxitoxin • Paralytic shellfish poisoning

Figure 27. 13 A red tide.

Portals of Exit • Respiratory tract • Coughing, sneezing • Gastrointestinal tract • Feces, saliva • Genitourinary tract • Urine, vaginal secretions • Skin • Blood • Biting arthropods, needles/syringes

Mechanisms of Pathogenicity Figure 15. 9

When the balance between host and microbe is tipped in favor of the microbe, an infection or disease results. Learning these mechanisms of microbial pathogenicity is fundamental to understanding how pathogens are able to overcome the host’s defenses. H 1 N 1 flu virus portals of entry Mucous membranes • Respiratory tract • Gastrointestinal tract • Genitourinary tract • Conjunctiva Skin Parenteral route Number of invading microbes penetration or evasion of host defenses Capsules Cell wall components Enzymes Antigenic variation Invasins Intracellular growth Adherence Mycobacterium intracellulare Clostridium tetani Micrographs are not shown to scale. damage to host cells Siderophores Direct damage Toxins • Exotoxins • Endotoxins Lysogenic conversion Cytopathic effects portals of exit Generally the same as the portals of entry for a given microbe: • Mucous membranes • Skin • Parenteral route