Enterobacteriaceae PDF

Enterobacteriaceae ROEL R. GUINTO JR., RMT, MSMT DEPARTMENT OF MEDICAL TECHNOLOGY COLLEGE OF ALLIED HEALTH NATIONAL UNIVERSITY- MANILA General Characteristics: Most medically important of gram-negative bacilli. Most species are normal flora of the GI Tract “Salmonella” “...

Enterobacteriaceae ROEL R. GUINTO JR., RMT, MSMT DEPARTMENT OF MEDICAL TECHNOLOGY COLLEGE OF ALLIED HEALTH NATIONAL UNIVERSITY- MANILA General Characteristics: Most medically important of gram-negative bacilli. Most species are normal flora of the GI Tract “Salmonella” “Shigella” and “Yersinia” are not normal GI flora. Major cause of nosocomial infection. Diseases includes UTIs, gastroenteritis, septicemia, food poisoning, wound infections, peritonitis, pneumonia and meningitis. General Characteristics: Gram (-) Facultative anaerobe All reduces to nitrate to nitrite All catalase (+) Cytochrome oxidase (-) Aerogenic except Shigella Can be differentiated based on “lactose fermentation” **Ferments glucose (4) Serological Characteristics: O (somatic) antigen → A cell wall antigen K (envelope) antigen → Capsular antigen H (flagellar) antigen → Flagellar antigen Vi antigen → Capsular antigen of Salmonella typhi  O-nitrophenyl-beta-D-galactopyranoside (ONPG) Considered to be a very sensitive test for Lactose-Fermentation. An artificial substrate, is incorporated into this test and acts as the substrate for the beta-galactosidase to ascertain the particular enzyme activity which subsequently aids in the identification and differentiation of different organisms. B-galactosidase (+): Yellow → RLF and LLF (They both have B-galactosidase enzyme) Rapid Lactose Fermenters (EKE) A/A (acid/acid) B-galactosidase Lactose permease Organisms: 1. E.coli 2. Klebsiella 3. Enterobacter Late Lactose Fermenters (C-SSSHY) A/A (acid/acid) B-galactosidase Organisms: 1. Citrobacter 2. Salmonella 3. Shigella 4. Serratia 5. Hafnia 6. Yersinia Urease “PPM-C-KEYS” Rapid urease fermenters “PPM” - Proteus - Providencia - Morganella Slow urease fermenters “C-KEYS” - Citrobacter - Klebsiella - Enterobacter except (E. gergoviae) - Yersinia - Serratia Non-motile at 37C “SKY” Shigella Klebsiella Yersinia Pigments Red pigment → Serratia marcescens Yellow pigment → Pantoea and Enterobacter sakazakii  Serratia marcescens Enterobacter sakazakii Indole Test To determine the ability of microorganisms to decompose the amino acid tryptophan to indole Indole Test Results: Methyl Red Test (MR) To determine the ability of microorganism to oxidize glucose with the production and stabilization of high concentrations of acid end products Voges-Proskauer Test (VP) To determine the ability of many microorganisms to produce acetone (acetyl methyl carbinol) during fermentation of glucose. Citrate Utilization Test To determine the ability of a microorganism to utilize citrate as the sole source of carbon and as energy source for the growth and ammonium salt as a sole source of nitrogen.  Escherichia coli IMViC (+,+,-,-) Other name: Colon Bacillus/ Golden Bacillus TSI: A/A with Gas and H2S (-) Mac Conkey: Pink/ red colonies SBA: Beta-hemolytic EMB: Green Metallic Sheen Indole (+) ONPG (+) Methyl red (+) Voges Proskauer (-) Citrate (-) Urease (-) Strains of E. coli  ETEC Enterotoxigenic E.coli Causes traveler’s diarrhea/ transient diarrhea. “Cholera-like” toxin Stool: Watery stool  EPEC Enteropathogenic E.coli Infantile diarrhea Strain: E.coli 0111 Stool: Watery stool with mucus but “without” blood.  EIEC Enteroinvasive E.coli “Shigella-like” diarrhea Stool: Watery with blood, pus (WBC) and mucus  EAEC Enteroaggregative E.coli Acute and chronic diarrhea. Produce adherent biofilm. Stool: Watery with mucus “Stacked-brick appearance”  EHEC Enterohemorrhagic E.coli “Shiga-like” toxin VTEC: Verotoxigenic E.coli → produces verotoxin Stool: watery with excessive blood without WBC. Causes: HUS Anaerogenic E.coli (E.coli Inactive) “Alkalescens dispar” → does not produce gas during fermentation.  Klebsiella spp. IMViC (-,-,+,+) Identifying Characteristics: TSI: A/A with gas MAC: pink colonies H2S: (-) MR: (-) Non motile Indole (-) Citrate: (+) Lactose: (+) Klebsiella granulomatis The causative agent of granuloma inguinale (sexually transmitted disease) Identification: - Does not Gram stain or grow on laboratory media. - In clinical specimen, Wright stain or Giemsa stained, “Donovan bodies” may be seen. - Cultured in “human monocytes” - Donovan Bodies → are intracellular pleomorphic bipolar using bacterial cells. Donovan bodies Klebsiella pneumoniae Mac Conkey: Encapsulated, mucoid colonies or tend to string. “Currant jelly-like sputum” Biochemical test IMViC: (-,-,+,+) TSI: A/A with gas Gelatinase: (+) String test: (+) String test K. oxytoca Indole: (+) K. ozonae Atropic rhinitis - inflammation of the mucous membrane of the nose.  Enterobacter spp. IMViC (-,-,+,+) H2S (-) MR: (-) Indole (-) VP (+) **Motile Citrate (+) Lactose: (+)  Enterobacter and Klebsiella has same IMViC reaction. → To differentiate: based on their motility Enterobacter → Motile Klebsiella → Non-motile  LDC: (-) except → E. aerogenes and E. gergoviae (+) Slow urease producer except → E. gergoviae (rapid urease producer) E. cloacae Most common isolated. Arginine: (+) Lysine (-) E. aerogenes 2nd most common isolated Enterobacter. Arginine: (-) Lysine: (+) E. sakazakii (Cronobacter sakazakii) Produces yellow pigment that aids in its presumptive identification. Serratia Causes opportunistic infection in patients undergoing chemotherapy and immunosuppressed patients. S. marcescens → most commonly isolated. Identifying Characteristics: DNase (+) Gelatinase (+) Lipase (+) ONPG (+) → delayed lactose fermenter (some strains)  Hafnia spp. IMViC (-,V,+,-) Identifying Characteristics: Motile Late lactose fermenter ONPG: (+) LDC: (+) ODC: (+) Representative organism: H. alvei (Enterobacter alvei) Hafnia vs Serratia  Notes: KESH → (+) Voges Proskauer, (-) Methyl Red, (+) Gas production (TSI) - Klebsiella - Enterobacter - Serratia - Hafnia  Pantoea IMViC (-,-,+,+) P. agglomerans Nitrate reduction: (-) *Yellow pigment producer LDC: (-) ODC: (-)  Salmonella IMViC (-,+,-,+) Mode of transmission: Ingestion of contaminated food and water. Identifying Characteristics: **H2S (+) Motile Urease (-) Lactose (-) TSI: K/A with gas (+) H2S Infection Enteric/ typhoid fever →Salmonella enterica, S. typhi (80%) and S. paratyphi A/B (10%) Bacteremia → S. cholerasuis Enterocolitis/ gastroenteritis → S. typhimurium Laboratory Assessment (Typhoid) 1st week → Early typhoid fever → Scybala stool: “goat-like dropping” → Specimen: Blood (blood culture) 2nd week → Colonization stage → “Rose spots” → Specimen: Urine (3rd week*) and stool Rose Spots 3rd week Convalescence stage Specimen: serum Widal/ Typhidot test  Shigella IMViC (v,+,-,-) Other name: “Inert bacilli” Lactose: (-) Mac Conkey: Colorless (NLF) Non-motile Causes: Shigellosis/ Bacillary dysentery TSI: K/A (alkaline/acid) H2S: (-) MR (+) **Majority of the biochemical tests are negative Incubation Early stage: 1 to 7 days Diarrheic stage: watery diarrhea (3 days) Dysenteric phase Virulence factor Neurotoxin → Which can cause paralysis and death. Serogroups A: S. dysenteriae → Shiga’s Bacillus B: S. flexneri → Strong bacillus C: S. boydii → New castle Manchester Bacillus D: S. sonnei → Duval’s Bacillus Differentiation of Serogroups Microorganism Catalase ONPG Mannitol fermenter S. dysenteriae - - - S. flexneri + - + S. boydii + - + S. sonnei + + + Shigella vs. Salmonella Salmonella Shigella Motile except S. gallinarum and S. pullorum Non-motile H2S (+) H2S (-) Gas producer Not a gas producer LDC (+) except S. paratyphi A → LDC (-) LDC (-) Proteus Rapid urease producers Typical “swarming motility” H2S (+) TSI but “NOT” in LIA Source of antigen for Weil-Felix Test → Serological test for dx of rickettsial disease P. vulgaris → source of OX2 AND OX19 P. mirabilis → source of OXK (kingsbury stain) P. mirabilis vs. P. vulgaris Microorganism Indole ODC Chloramphenicol susceptibility Test P. mirabilis → Associated with - + (S) wound infection and UTI P. vulgaris + - (R) → Can cause pneumonia and septicemia  Providencia spp. IMViC (+,+,-,+) Lactose (-) Deaminase (+) H2S (-) Motile but not swarming LDC, ODC, ADH (-) Commonly isolated: Microorganism Urease Adonitol Trehalose P. rettgeri + + - P. stuartii - - +  Morganella IMViC (+,+,-,-) M. morganii is the only species. Lactose (-) Mac Conkey: Colorless (NLF) Deaminase (+) H2S (-) LDC (-) Urease (+) ODC (+)  Citrobacter C. freundii is the most common isolated species. → resembles as E. coli → resembles as Salmonella but differs in: ONPG (+) LDC (-) Slow urease producer (C-KEYS) → Citrobacter, Klebsiella, Enterobacter, Yersinia and Serratia Edwardsiella IMViC (+,+,-,-) E. tarda is the common isolate spp. → Resembles as Salmonella spp. → H2S (+) → ONPG (-); Salmonella (+) Edwardsiella vs. Salmonella Organism Indole Citrate Edwardsiella tarda + - Salmonella - + Edwardsiella vs. E. coli Edwardsiella biochemical results: H2S (+) → E.coli (-) Lactose (-) → E.coli (+) (Rapid lactose fermenter EKE) E.coli, Kleb, Enterobacter LDC (+) ODC (+) ADH (-) Yersinia (3) Pathogenic species: 1. Y. pestis 2. Y. enterocolitica 3. Y. psedotuberculosis Yersinia pestis Causes plague. The bacteria transmitted by “fleas” Considered as a potential bioterrorism agent. Rodents/ rats: natural reservoir Yersinia enterocolitica Causes enterocolitis in humans It is acquired by drinking contaminated water or by eating contaminated meat. “Bull’s eye colonies” ONPG (+) → delayed lactose fermenter Sucrose (+) MAC: Colorless at 18 hours TSI: Acid/ Acid (A/A) Yersinia pseudotuberculosis is a rare cause of lymphadenitis in children. It has been isolated in avian animals such as turkeys and pigeons. Identifying Characteristics (Yersinia) MAC: small coccobacilli that produces “small pinpoint” colonies. Suspected if TSI is yellow/ red (acid/alkaline or A/K) All are motile in 37’C except Yersinia pestis. Selective Media for Yersinia CIN Agar (Cefsoludin Irgasan Novobiocin Agar) → selective media also for a. Aeromonas b. Plesiomonas spp. Contains: Mannitol Peptones Yeast Neutral red indicator Crystal Violet  Microorganisms Disease Sucrose Rhamnose Sorbitol Y. pestis Plague - - - Y. enterocolitica Enterocolitis + - + Y. pseudotuberculosis Lymphadenitis - + - Haemophilus Most spp. are normal upper respiratory tract flora. Haemophilus spp. esp. H. aphrophilus are considered member of HACEK (Haemophilus, Actinobacillus, Actinomycetemcomitans, Cardiobacterium hominis, Eikenella corrodens and Kingella) group. → Fastidious → Causes “endocarditis” Pleomorphic Gram (-) negative coccobacilli ranging from small to filamentous. Growth requirement for Haemophilus spp. Hemin (X factor) → Which released from hemoglobin → via cell lysis (destruction of red cells). NAD (V factor) → heat labile compound → S. aureus also produces this type of compound. (Haemophilus Satellitism occurs) Culture Media for Haemophilus spp. Haemophilus does “NOT” grow on SBA because of the presence of enzyme NADase in the agar (NADase inactivates NAD) but “will grow” on HORSE OR RABBIT BLOOD AGAR which contain no NADase. Chocolate agar plate (CAP) → Is routinely used for cultures. Grows at 35-37’C with 5-10% CO2 and is susceptible (S) to drying and temperature changes  Colony Morphology → smooth, round, flat opaque and tan on CAP. Haemophilus on Chcocolate Agar Plate (CAP) Haemophilus Satellitism Haemophilus can grow around colonies of S. aureus growing on SBA plate (NOTE: Haemophilus cannot grow on SBA because it contains NADase which inactivates NAD which is essential for the growth of Haemophilus). S. aureus releases NAD on BAP/SBA that’s why Haemophilus will grow near the S. aureus, forming tiny clear pinpoint colonies. Characteristics of Haemophilus Non-motile Catalase (+) Oxidase (+) Haemophilus influenzae Formerly called Pfeiffer 's bacillus or Bacillus influenzae. Major cause of meningitis in “children” Haemophilus influenzae type b (Hib) vaccine → has greatly reduced childhood meningitis and other invasive diseases caused by this serotype. H. influenzae isolates should be tested for beta-lactamase. Specimen of choice: Blood Sputum CSF Eye swabs Latex agglutination Specific detection of Hib capsular antigen. Haemophilus aegyptius Causes “Pink eye”, a very contagious conjunctivitis. Similar to H. influenzae with the exception of being sucrose (+) Haemophilus influenzae biogroup aegyptius Causes conjunctivitis followed by invasive disease called as “Brazillian purpuric fever”. Haemophilus ducreyi Causes genital ulcers, a sexually transmitted disease (STD). Produces chancroid and buboes (swollen lymph nodes). CAP with vancomycin is used to inhibit normal flora and contaminants. Chancroid A bacterial sexually transmitted infection characterized by painful sores on the genitalia.  Microorganisms X factor V factor Horse Blood ALA* H. influenzae + + - - H. ducreyi + - - - H. aegytius + + - - H. aprophilus - - - + H. haemolyticus + + + - H. parainfluenzae - + - + H. paraphrophilus - + - + Aminolevulinic acid (ALA) is converted to porphyrin + = positive; - = negative