Enterobacteriaceae I: Opportunistic Pathogens

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

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

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

Lab Culture Media CBA: Selective Media: Differential: 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

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 Vi (virulence) antigen in Salmonella typhi I.D. pathogenic E. coli O157:H7 strain

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

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’

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

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

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

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

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

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 = (++--)

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)

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

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

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

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

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

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

Summary: E.coli Gastroenteritis

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

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

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

Proteus: ID NF of GI tract Motile, with swarming 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 (-)

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

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

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

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

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

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

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

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.

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.

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.

Lecture Exam I Thursday, 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!