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.
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.
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
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.
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