Enterobacteriaceae I: Opportunistic Pathogens

1. Enterobacteriaceae I: Opportunistic Pathogens

2. Enterobacteriaceae • Opportunistic Pathogen: • Escherichia • Klebsiella • Proteus • Enterobacter • Serratia • Edwardsiella • Citrobacter • Intestinal Pathogen: • Salmonella • Shigella • Yersinia

3. Enterobacteriaceae: General Characteristics • Gram (-) rod • Catalase(+) • Oxidase(–) • Ferment glucose • Reduce nitrate • Motile by peritrichous flagella (except Klebsiella, Shigella, Yersinia)

4. General Characteristics • Facultative anaerobes • Normal inhabitants of intestinal tract humans, other animals • Enteric pathogen • Some are urinary, respiratory tract pathogen • Differentiate by biochemical test and antigenic structure

5. Lab Culture Media • CBA: • Colonies large, dull gray • ± hemolysis • Selective Media: • Dye, bile salt inhibit G(+) MO • EMB, Mac - selective for Enterobacteriaceae: • SS, HE, XLD - highly selective for enteric pathogens, inhibit NF • Differential: • Lactose fermentation • H2S production

6. Serologic ID: Major Antigens • O - Somatic LPS antigen • Major cell wall antigen • Heat stable polysaccharide • Component of endotoxin • H - Flagellar antigen • Protein • Heat labile • K - Capsule antigen • Polysaccharide • Heat labile • Vi (virulence) antigen in Salmonella typhi • I.D. pathogenic E. coliO157:H7 strain

7. Biochemical Tests • TSI (Triple Sugar Iron Agar) – glucose, lactose, sucrose, H2S indicator • LIA (Lysine Iron Agar) • lysine deaminase – aerobic, slant • Lysine decarboxylase – anaerobic, butt • H2S indicator – anaerobic, butt • Motility – soft (0.4%) agar • 1% Glucose broth (MR-VP) – acid or neutral metabolic end product • Citrate – use as sole C source • Indole – tryptone product • Urea – hydrolysis by urease • PAD – phenylalanine deaminase

8. Virulence Factors • Endotoxin – Gram(-) cell wall: • LPS release upon cell death • Symtoms by Lipid A - fever, leukopenia, cytokine action • Immune and inflammatory response • Vasodilation, capillary leakage • Disseminated Intravascular Coagulation (DIC); blood coagulation, decreased circulation • Hypotension, shock, death • Capsule – prevent phagocytosis, resist serum killing by C’

9. Virulence Factors: Enterotoxin • Movement of water & ions from tissue to bowel resulting in watery diarrhea • Two types of enterotoxin: • Heat-stable (ST) – activate cGMP • Heat-labile (LT) – activate cAMP • Shiga toxin – inhibit protein synthesis: • Enterohemorrhagic strains of E. coli (EHEC) • Cytotoxic, enterotoxic, neurotoxic • Diarrhea, ulceration of G.I. tract • Enteroaggregative toxin - produced by E. coli (EAEC), causes watery diarrhea

10. Virulence Factors • Hemolysin – cytotoxin to RBC’s, leukocytes • Enterochelin - capture iron • Outer membrane proteins – attachment, initiate endocytosis for cell invasion • Adhesions – pili, fimbriae, non-fimbrial factors for host cell (GI, urinary, CNS) attachment; antibodies to adhesions may protect from colonization

11. Sites of Infection • Central Nervous System (CNS) • Escherichia • Lower Respiratory Tract (LRT) • Klebsiella • Enterobacter • Escherichia • Bloodstream • Escherichia • Klebsiella • Enterobacter • Salmonella

12. Sites of Infection • Gastrointestinal (GI) Tract • Salmonella • Shigella • Yersinia • Escherichia • Urinary Tract (UT) • Escherichia • Proteus • Klebsiella • Providencia

13. Escherichia coli • Normal inhabitant G.I. tract, present in water, vegetation • Various forms of gastroenteritis • Major cause of: • Urinary tract infection • Neonatal meningitis • Septicemia • Transmission: • Endogenous spread in susceptible patient • Ingest contaminated food or water • Nosocomial infection

14. Escherichia coli: Biochemical Test ID • May be hemolytic on CBA, more common in pathogenic strains • Key Test: • TSI = A/A + gas • LIA = K/K • IMViC = (++--)

15. Escherichia coli: Infections • Neonatal Meningitis - leading cause, along with Strep Gp. B • Septicemia - GU infection or own GI NF, high mortality • Urinary Tract Infection – leading cause, by own NF: • acute cystitis (bladder infection) • pyelonephritis (kidney infection)

16. Escherichia coli: Gastroenteritis • ETEC (Enterotoxigenic E. coli) – infant diarrhea, Traveler’s diarrhea • EPEC (Enteropathogenic E. coli ) – infant diarrhea • EAEC (Enteroaggregative E. coli) – chronic diarrhea in children • EIEC (Enteroinvasive E. coli) – dysentery in children and adults • EHEC (Enterohemorrhagic E. coli) – hemorrhagic colitis, hemolytic uremic syndrome

17. ETEC (Enterotoxigenic E. coli) • Traveler’s diarrhea • Diarrhea of children in developing countries • These strains rare in U.S. • MO attaches to intestinal mucosa, liberates enterotoxin • Disease characterized: • Watery diarrhea • Nausea • Abdominal cramps • Low-grade fever for 1-5 days • Transmission via contaminated food or water

18. EPEC (Enteropathogenic E. coli ) • Bundle forming pili attachment intestinal mucosa • Effacement of microvilli, exact pathogenesis unclear • Diarrhea, large amounts mucous, no blood or pus, vomiting, malaise, low grade fever • Outbreaks in nurseries and day care centers

19. EAEC (Enteroaggregative E. coli) • Bacteria associated autoagglutinins cause aggregation in “stacked-brick” arrangement at cell surface of mucus biofilm • Process mediated by bundle-forming pili (BFP) • MO attach and liberate cytotoxin • Symptoms include watery diarrhea, vomiting, occasional abdominal pain, dehydration >14 days • Chronic diarrhea and growth retardation of children in developing countries

20. EIEC (Enteroinvasive E. coli) • MO attaches to intestinal mucosa; direct penetration, invasion of intestinal cells, destruction of intestinal mucosa • Lateral movement of MO from one cell to adjacent cells • Symptoms include fever, severe abdominal cramps, malaise, watery diarrhea, followed by scanty stools containing blood, mucus, leukocytes and pus • Person to person transmission via fecal-oral route

21. EHEC (Enterohemorrhagic E. coli) • Attach to intestinal mucosa, liberates Shiga toxin, also termed Verotoxin (cytopathic to Vero monkey kidney cell culture) • Watery diarrhea, progress to bloody diarrhea without pus, crampy abdominal pain, ±low-grade fever • May progress to hemolytic uremic syndrome (HUS): • Anemia, kidney failure • Highly fatal - infants (<5 yrs.), elderly • By E. coli O157:H7 serotype • Epidemics traced to consumption of undercooked beef, unpasteurized juice

22. Summary: E.coli Gastroenteritis

23. E. coli: Treatment and Control • Susceptible to variety of antibiotics • Drug resistant strains increasing, need susceptibility testing • Hospital infection control important • Education to improve hygiene, hand washing

24. Klebsiella: ID • NF of GI tract, potential pathogen in other sites • TSI = A/A + gas • LIA = K/K • Urea (+) • Citrate (+) • MR(-), VP(+) • Motility (–)

25. Klebsiella: Infections • Virulence factors: • Capsule • Adhesions • Iron capturing ability • Clinical significance: • Pneumonia - mostly immunocompromised hosts; permanent lung damage frequent occurrence (rare in other bacterial pneumonia) • Major cause of nosocomial infection – septicemia, meningitis

26. Proteus: ID • NF of GI tract • Motile, withswarming on CBA • TSI = K/A + H2S (similar to Salmonella) • LIA = R/A (lysine deamination) • PAD = (+) (phenylalanine deamination) • Urea = strongly (+) for Proteus • Indole – only P. mirabilis is (-)

27. Proteus: Infections • Virulence factors • Urease – ammonia produced may damage epithelial cells of UT • Clinical Significance • UT infections; also pneumonia, septicemia, wound infections

28. Enterobacter: ID &Infections • NF of GI tract • TSI = A/A • LIA = K/K • IMViC = (--++) • Clinical significance: • Nosocomial infections • Bacteremia in burn patients

29. Serratia: ID & Infections • Free-living saprophyte • TSI = K/A • LIA = K/K • Citrate (+) • Urea (±) • Has been found in RT and UT infections • Resistant to many antibiotics

30. Edwardsiella: ID & Infections • TSI = K/A + gas, H2S • LIA = K/K +H2S • Urea(–) • Citrate(–) • Indole(+) • Clinical significance – causes GI disease in tropical and subtropical countries

31. Citrobacter: ID & Infections • TSI = K/A or A/A both + gas, H2S • LIA = K/A + H2S • Citrate (+) • Urea usually (+) • Opportunistic pathogen causing urinary tract or respiratory tract infections; occasionally wound infections, osteomyelitis, endocarditis, meningitis

32. Lab: Enterobacteriaceae I Group Unknowns TSI LIA • Escherichia A/A K/K • Shigella K/A K/A • Enterobacter A/A K/K • Klebsiella A/A K/K

33. Class Assignment • Textbook Reading: Chapter 19 Enterobacteriaceae • Key Terms • Learning Assessment Questions

34. Case Study 3 -Enterobacteriaceae • A 25-year-old previously healthy woman came to the emergency room for the evaluation of bloody diarrhea and diffuse abdominal pain of 24 hours’ duration. • She complained of nausea and had vomited twice. • She reported no history of inflammatory bowel disease, previous diarrhea, or contact with other people with diarrhea.

35. Case Study 3 • The symptoms began 24 hours after she had eaten an undercooked hamburger at a local fast food restaurant. • Rectal examination revealed watery stool with gross blood. • Sigmoidoscopy showed diffuse mucosal erythema and petechiae with a modest exudation but no ulceration or pseudomembranes.

36. Case Study 3 - Questions • 1. Name four genera of Enterobacteriaceae that can cause gastrointestinal disease. Name two genera that can cause hemolytic colitis. • 2. What virulence factor mediates this disease? • 3. Name the five groups of E. coli that can cause gastroenteritis. What is characteristic of each group of organisms? • 4. Differentiate between disease caused by S. typhi and that caused by S. sonnei.

37. Lecture Exam IThursday, Feb. 2, 2012 • Introduction thru Enterobacteriaceae • Lecture, Reading (Chap. 14, 15, 17, 19), Key Terms, Learning assessment Questions, Case Study 1,2,3 • Exam Format: • Multiple Choice • Terms • True/False Statements • Short Essay • Review, Review, Review!