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16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
16  -chapter_19_-_enterobacteriaceae
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16 -chapter_19_-_enterobacteriaceae

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Enterobacteriaceae

Enterobacteriaceae

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  • 1. Chapter 9ENTEROBACTERIACEAE & 19
  • 2. OBJECTIVES Describe the general characteristics of the Enterobacteriaceae family. Discuss the principle, purpose, and interpretation of the following tests:  Citra te  DN a se  L ysine iron a ga r  Methyl R ed  MIO  N itra te  ON PG  Oxida se  Phenyl a nil i n e dea mina se  TSI  Urea se  Voges - Pra sk a uer Distinguish between lactose fermenting and non -lactose fermenting Enterobacteriaceae on the MacConkey, Eosin Methylene Blue, Hektoen, and XLD agars. List the non-motile Enterobacteriaceae. L i s t t h e E n t e r o b a c t e r i a c e a e t h a t p r o d u c e H 2S . Differentiate the genera of Enterobacteriaceae based on biochemical tests. Explain the common modes of transmission and clinical significance of Salmonella, Shigella and E. coli 0157:H7. Differentiate the following species: E.coli and E. coli 0157:H7 Proteus vulgaris and Proteus mirabilis Klebsiella pneumoniae and Klebsiella oxytoca. Discuss the clinical significance of the Enterobacteriaceae in this unit.
  • 3. TAXONOMY Family  Enterobacteriaceae Genera  Escherichia  Serratia  Shigella  Proteus  Edwardsiella  Morganella  Citrobacter  Providencia  Klebsiella  Yersinia  Enterobacter  Salmonella
  • 4. GENERAL CHARACTERISTICS Found in nature  Plants  Soil  Water Intestines of humans Most are normal enteric flora
  • 5. BIOCHEMICAL CHARACTERISTICS Ferment glucose Oxidase negative Reduce nitrate to nitrite Oxidase Test Nitrate Reduction Glucose Fermentation
  • 6. MICROSCOPIC CHARACTERISTICS Gram negative bacilli Most are motile Non-spore forming
  • 7. GROWTH REQUIREMENTS Facultative anaerobes Incubation length  18-24 hours Incubation temperature  35-37°C Incubation atmosphere  Air
  • 8. MACROSCOPIC CHARACTERISTICS Grow well on BAP Grow on enteric media  MAC  HE  XLD  EMB
  • 9. THE SCOOP ON POOP! Enterobacteriaceae are routinely isolated from stool cultures ID work-up  Only performed on those that are pathogens
  • 10. ENTERIC MEDIA
  • 11. BLOOD AGAR PLATE - 1All Enterics grow on BAPAllows for observation of colony “true” morphology
  • 12. BLOOD AGAR PLATE - 2
  • 13. MACCONKEY (MAC) - 1Selective  Gram negativeDifferential  Lactose fermentation  Fermenter  PINK  Non-fermenter  Clear
  • 14. MACCONKEY (MAC) - 2
  • 15. XYLOSE-LYSINE-DESOXYCHOLATE (XLD) - 1Selective  Bile SaltsDifferential  Lactose fermentation  YELLOW  H2S  BLACK
  • 16. XYLOSE-LYSINE-DESOXYCHOLATE (XLD) - 2Lactose Non-Fermenter; H2S Producer Lactose Fermenter
  • 17. EOSIN-METHYLENE BLUE (EMB) - 1 Selective  Gram negative Differential  Lactose fermentation  BLUE-BLACK
  • 18. EOSIN-METHYLENE BLUE (EMB) - 2
  • 19. HEKTOEN ENTERIC (HE) - 1Selective  Gram negativeDifferential  Lactose fermentation  YELLOW/ORANGE  SALMON (Pink)  H2S  BLACK
  • 20. HEKTOEN ENTERIC (HE)
  • 21. BIOCHEMICAL REACTIONS
  • 22. LACTOSE FERMENTATION TESTS Lactose non-fermenters  Cannot break down lactose Lactose fermenters  Have enzymes that allow for transport of lactose into the cell for utilization Slow lactose fermenters  Lack beta-galactosidase permease  But have beta-galactosidase
  • 23. TRIPLE SUGAR IRON AGAR (TSI) - 1 Media composition  Sugars (Glucose, Lactose, Sucrose)  Peptone - nutrient and energy  pH indicator  H 2 S indicator
  • 24. TSI - 2 How to read the tube?  FIRST - Slant reaction  SECOND – Butt reaction  THIRD - H 2 S production  “/” separates the reactions  Circling the butt reaction denotes gas production Rules for test interpretation  Glucose is always the preferred sugar for fermenters  Glucose fermentation results in acid production  Peptone utilization results in alkaline by-products
  • 25. TSI REACTIONS - 1
  • 26. TSI REACTIONS - 2
  • 27. TSI REACTIONS - 3
  • 28. TSI REACTIONS - 4
  • 29. TSI REACTIONS - 5
  • 30. TRIPLE SUGAR IRON AGAR (TSI) Test Your Knowledge
  • 31. O-F TESTS See power point presentation for chapter 21.
  • 32. INDOLE Tryptophanase + tryptophan → pyurvic acid + ammonia + indoleindole + 1% paradimethylaminocinnamaldehyde → color change (pink color)
  • 33. UREAUrease + urea (broth or agar) → ammonia + water + CO2 Ammonia produces a pH change in the media = Pink
  • 34. UREASE Tube 1: Control Tube 2: Proteus vulgaris  = Positive Tube 3: E. coli  = Negative
  • 35. CITRATE + = +
  • 36. DNASE Positive Escherichia coli (top): negative-no clearing Serratia marcescens (bottom): positive-clearing
  • 37. DECARBOXYLASE TESTS - ORNITHINE Control Positive Negative Control
  • 38. DECARBOXYLASE TESTS – ARGININE & LYSINE
  • 39. METHYL REDGlucose → Pyruvic acid → Mixed acid fermentation (pH 4.4) Acid end-products detected with methyl red indicator
  • 40. VOGES-PROSKAUERGlucose → Pyruvic acid →Acetoin → DiacetylDiacetyl + KOH + α-naphthol → Red complex
  • 41. ONPG Lactose fermenter have the β- galactosidase enzyme, which hydrolyzes ONPG into galactose and orthonitrophenol = Color change from colorless to yellow Non-lactose fermenters - do not have the beta-galactosidase enzyme to hydrolyze the ONPG = No color change
  • 42. MOTILITY Motility is defined as any growth away from the stab  1 = Non-motile  2 = Motile  3 = Motile
  • 43. MOTILITY/INDOLE/ORNITHINE (MIO) Indole Reaction Motility Reaction Ornithine Reaction
  • 44. PHENYLALANINE DEAMINASE (PDA) Organisms that produce the enzyme deaminase are able to removes the amine group from the amino acid phenylalanine and releases the amine group as free ammonia. As a result of this reaction, phenylpyruvic acid is also produced. Negative After incubation, 10% ferric chloride is added to the media. Positive  If phenylpyruvic acid was produced, it will react with the ferric chloride and turn dark green.  If the medium remains a straw color, the organism is negative for phenylalanine deaminase production.
  • 45. NITRATE Add Zinc dust = = + ? = = ? Nitrate → NitriteNitrite + sulfanilic acid + N,N-Dimethyl-1-naphthlamine → red
  • 46. NITRATEPositive Positive Negative
  • 47. GELATIN HYDROLYSIS Escherichia coli: negative (gelatin is solid) Pseudomonas aeruginosa: positive (gelatin has been liquefied)
  • 48. ANALYTICAL PROFILE INDEX (API)
  • 49. API 20-E
  • 50. Negative TestsPositive Tests
  • 51. OPPORTUNISTIC ENTERICS
  • 52. ESCHERICHIA COLI
  • 53. CLINICAL SIGNIFICANCE Most common human pathogen UTI Septicemia Meningitis Pneumoniae Endocarditis
  • 54. COLONY MORPHOLOGYBAP MACHE XLD EMB
  • 55. MICROSCOPIC EVALUATIONGram negative bacilliBipolar staining
  • 56. BIOCHEMICAL IDENTIFICATIONLactose fermenterGas from GlucoseIMV:C (++--)  Indole – Positive  MR – Positive  VP – Negative  Citrate – Negative  ONPG – Positive  Lysine - Positive
  • 57. E. COLI Serotypes
  • 58. ENTEROHEMMORAGIC E. COLI (EHEC) E. coli 0157:H7  Hemorrhagic colitis  Hemolytic Uremic Syndrome (HUS)  Low platelets  Hemolytic anemia  Kidney failure
  • 59. ENTEROHEMMORAGIC E. COLI (EHEC)MacConkey with Sorbitol  Negative = clear/colorless  Positive = PINK
  • 60. KLEBSIELLA PNEUMONIAE
  • 61. CLINICAL SIGNIFICANCE Most common isolate  Lower respiratory infections in hospitalized patients  Pneumonia  Septicemia  Meningitis  UTI
  • 62. COLONY MORPHOLOGY Very mucoid (wet) looking Due to a polysaccharide capsule
  • 63. BIOCHEMICAL IDENTIFICATION Lactose fermenter Non-motile IMV:C (+-++); (--++)  Indole  K. oxytoca = Positive  K. pneumoniae – Negative  MR = Negative  VP = Positive  Citrate = Positive Urea = Positive DNase = Negative Ornithine = Negative (few + reported)
  • 64. ENTEROBACTER
  • 65. CLINICAL SIGNIFICANCE Opportunistic pathogens  E. aerogenes  E. cloacae – bacteremia and wound infections Meningitis Endocarditis Urinary tract infection Lower respiratory tract infections
  • 66. BIOCHEMICAL IDENTIFICATION Lactose = Positive Motile IMV:C (--++)  Indole = Negative  MR = Negative  VP = Positive  Citrate = Positive Ornithine = Positive
  • 67. SERRATIA SPECIES
  • 68. CLINICAL SIGNIFICANCE Nosocomial  UTI  Respiratory infections  Septicemia  Endocarditis
  • 69. COLONY MORPHOLOGYRed pigmentation  S. rubidea  S. marcescensSmells like peeled potatoes  S. odorifera
  • 70. BIOCHEMICAL IDENTIFICATION Lactose = Negative IMV:C (-+++)  Indole = Negative  MR = Positive  VP = Positive  Citrate = Positive
  • 71. PROTEUS SPECIES
  • 72. CLINICAL SIGNIFICANCE Nosocomial infections UTIs Wound infections Septicemia Pneumonia
  • 73. COLONY MORPHOLOGY Swarm on BAP Smells like burnt chocolate
  • 74. BIOCHEMICAL IDENTIFICATION Lactose negative PDA - Positive Urease – Positive H 2 S – Positive (most)
  • 75. SPECIES DIFFERENTIATION P. mirabilis  Indole = Negative  Ornithine = Positive P. vulgaris  Indole = Positive  Ornithine = Negative
  • 76. MORGANELLA SPECIES
  • 77. CLINICAL SIGNIFICANCE Nosocomial infections UTIs Post-operative infections Septicemia in the immunocompromised
  • 78. BIOCHEMICAL IDENTIFICATION Lactose – Negative PDA – Positive Urease – Positive Ornithine – Positive Indole - Positive
  • 79. PROVIDENCIA SPECIES
  • 80. CLINICAL SIGNIFICANCE Nosocomial infections Wound infections Skin infections Septicemia
  • 81. BIOCHEMICAL IDENTIFICATION Lactose – Negative Indole – Positive Citrate – Positive PDA – Positive MR – Positive VP- Negative
  • 82. EDWARDSIELLA TARDA
  • 83. CLINICAL SIGNIFICANCE Rare causes of GI infections Hepatic abscesses Meningitis Wound infections
  • 84. BIOCHEMICAL IDENTIFICATIONNon-lactose fermenterGas production from glucoseIMVI:C (++--)  Indole = Positive  MR = Positive  VP = Negative  Citrate = NegativeH 2 S – positiveLysine – PositiveOrnithine – Positive
  • 85. CITROBACTER FREUNDII
  • 86. CLINICAL SIGNIFICANCE UTIs Respiratory infections Septicemia Wound infections
  • 87. BIOCHEMICAL IDENTIFICATION Lactose – Positive Gas production from glucose Lysine – Negative Urea – Positive (70%) Citrate = Positive H 2 S - Positive
  • 88. SalmonellaENTERIC PATHOGENS Shigella Yersinia
  • 89. SALMONELLA SPECIES
  • 90. CLINICAL SIGNIFICANCE Salmonella typhi  Not human normal flora  Humans are the only reservoirs  Ingestion of contaminated food products  Fecal-oral transmission  Typhoid fever
  • 91. BIOCHEMICAL IDENTIFICATION Lactose = Negative H 2 S = Positive Gas production from glucose Lysine = Positive Urease, Indole & Citrate = Negative
  • 92. SHIGELLA SPECIES
  • 93. CLINICAL SIGNIFICANCE Shegellosis or bacterial dysentery  Transmitted by ingesting contaminated food or water  Fever  Abdominal cramping and pain  Scant stools with blood, mucous and pus  Few organisms are capable of causing disease
  • 94. BIOCHEMICAL IDENTIFICATION Non-motile Lactose = negative Anaerogenic MR = Positive Ornithine = Positive
  • 95. YERSINIA SPECIES
  • 96. CLINICAL SIGNIFICANCE Yersinia entercolitica  Entercolitis  Mimics acute appendicitis Yersinia pestis  Bubonic plague  Pneumonic plague
  • 97. COLONY MORPHOLOGY
  • 98. BIOCHEMICAL IDENTIFICATION Lactose – negative Motile at 22°C but not at 37°C
  • 99. SUMMARY TABLES
  • 100. ID THE BUG!
  • 101. ID THE BUG - 1
  • 102. ID THE BUG - 1 - ANSWERIMViC = ++-- E. coli
  • 103. ID THE BUG - 2
  • 104. ID THE BUG - 2 - ANSWERIMViC = --++ K. pneumoniae
  • 105. WHAT’S NEXT? Review Mahon, Chapter 19 Complete AUMoodle Review

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