CLS05 - Biochemical test media for GNB

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  1. Carb fermentation (setup, purpose)
    • Fermentation media and durham tube (upside down tube in medium)
    • medium contains peptones, ONE carb, and a pH indicator (colormetric)
    • durham tube determines gas production
    • red is neg, yellow is pos, gas is present or not
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  2. Citrate utilization (diff, purpose, protocol)
    • +: Klebsiella, Salmonella enteritidis, citrobacter, Enterobacter, Serratia
    • -: E. coli, Salmonella typhi, Shigella, Yersinia enterocolitica
    • IDs which bacteria can utilize citrate as carbon source and inorganic ammonium salts as nitrogen source
    • Synthetic medium (slant) w/ bromthymol blue pH indicator
    • *positive result is growth REGARDLESS of color change (color change to blue indicates citrate breakdown)
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  3. Gelatin hydrolysis (diff, purpose, protocol)
    • +: Proteus vulgaris AND P. mirabilis (and Serratia odorifera biotype 2)
    • -: everything else
    • Determines ability of bacteria to produce gelatinase (liquify gelatin)
    • stab inoculate in gelatin deep OR add 4-5 drops of broth culture
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  4. Indole test (diff, purpose, protocol)
    • +: E. coli, Proteus vulgaris
    • -: other enterics, Proteus mirabilis
    • IDs bacteria that produce tryptophanase by detecting end product
    • add Kovac's reagant (entero) or Ehrlich's reagant (anaerobes/nonfermentors) - produces red color
    • Test often combined with other biochemical test (SIM or MIO agar)
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  5. MR/VP (diff, purpose, mechanism, protocol)
    • Methyl red/Voges-Proskauer test
    • diff between many gram- rods (E. coli MR+/VP- | Enterobacter MR-/VP+)
    • Determines metabolic pathway for glucose ferm (mixed acid = acidic pH, acetoin = neutral pH)
    • MR coloration - 4.4 red, 7 orange, 9 yellow
    • VP coloration - pink-red complex
    • protocol: innoculate broth, incubate, then aliquot.
    • add methyl red to tube 1, look for immediate change (red is +, no change is -)
    • add a-naphthol to tube 2, shake well, observe for 5 minutes (red = +, no change is -)
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  6. Moeller Decarboxylase medium (diff, purpose, mechanism, protocol)
    • Arginine - E. cloacae+, E. aerogenes-
    • Lysine - E. aerogenes+, E. cloacae-
    • Ornithine - Enerobacter+, Klebsiella-
    • Differentiates by measuring ability to decarboxylate an amino acid, raising pH (indicator becomes purple)
    • *anaerobic process, requires mineral oil overlay
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  7. Motility medium (purpose, protocol)
    • Differentiates bacteria on ability to move through semi-solid medium
    • stab innoculate into agar, check growth daily
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  8. MUG test (diff, purpose)
    • Diff between commensal E. coli (fluorescence) and O157:H7 (no fluorescence)
    • Looks for β-D-glucuronidase enzyme, end product in medium fluroesces blue under UV
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  9. Nitrate reduction (purpose, mechanism, protocol)
    • Determines if nitrate is final e- acceptor in aerobic resp
    • All enterics use nitrate reductase, some use nitrite reductase 
    • Reagant A + Reagant B = pink color if Nitrite is present
    • no color could be no nitrite OR nitrite further reduced
    • Zinc dust (after) = pink color means zinc reduced nitrate (∴ bacteria doesn't reduce nitrate)
    • if no color change then bacteria reduces nitrate AND nitrite
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  10. ONPG test (diff, QC, purpose, protocol)
    • O-nitrophenyl-B-D-galactopyranoside
    • diff between late-lactose-fermentors and non-lactose fermenters
    • *ONPG is like lactose
    • Shigella is + (color change) and salmonella is -
    • place ONPG disc in suspension and incubate, look for color change
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  11. Oxidase test (diff, purpose, protocol)
    • Diff between gram- nonfermentors (pos) and gram- enterics (neg)
    • Tests for cytochrome oxidase
    • innoculate filter paper, add drop of oxidase reagent. Purple = +, no color = -
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  12. Oxidation-fermentation tubes (diff, purpose, protocol)
    • Diff between G- nonfermentors (P. aeruginosa = oxidizer / E. coli = fermentor / Alcaligenes faecalis = asaccharolytic)
    • 2 tubes are used - one left open to O2, one overlaid with mineral oil
    • Stab innoculated
    • both yellow = fermentation
    • aerobic yellow, oil overlay red = oxidation
    • both red = no utilization of glucose
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  13. Phenylalanine deamination (diff, purpose, protocol)
    • +: Proteus, Morganella, Providencia
    • -: everything else
    • Determines ability to oxidatively deaminate phenylalanine
    • 1 drop broth on slant, leave caps loose, add 4-5 drops FeCl3 (Green = + / no color change = -)
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  14. TSI agar (diff, purpose, protocol, important results to know)
    • Triple Sugar Iron agar
    • Diff G- based on ferm of glu, lac, and suc AND H2S production
    • Tube uses 2 areas - aerobic slant and anaerobic butt (stab, then streak as you remove needle)
    • results are given "slant/butt" yellow = acid, red = alkaline, black = H2S (can only happen in acid), gas = gap at bottom of slant
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  15. TSI important results to know (E. coli, S. typhi, Shigella, Plesiomonas, Pseudomonas, Alcaligenes, Salmonella serotypes)
    • E. coli: A/(A)
    • S. typhi: K/A H2S
    • Shigella: K/A
    • Plesiomonas: K/A
    • Pseudomonas: K/K (K/NC)
    • Alcaligenes: K/K (K/NC)
    • Salmonella serotypes: A/(A) H2S
  16. Urea hydrolysis (diff, purpose, protocol)
    • + (rapid): Proteus
    • + (weak): Klebsiella pneumoniae, some Enterobacter sp
    • enzyme used to break down urea
    • slant or broth
    • pink = +, colorless = -
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Card Set Information

Author:
victimsofadown
ID:
318375
Filename:
CLS05 - Biochemical test media for GNB
Updated:
2016-04-04 01:58:37
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CLS05 Biochemical test media GNB
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CLS05
Description:
CLS05 - Biochemical test media for GNB
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