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06.The Count

F agar is a very reliable medium lab 6 page 27 the

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F Agar is a very Reliable Medium.
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Lab 6 Page 27 The IMViC The IMViC (rhymes with Rhythmic) isn’t a Single Test but rather is a Series of Tests designed to definitively distinguish E. coli from Enterobacter . The Name is an Acronym (G= top letters) for Indole, Methyl Red, Voges-Proskauer, and Citrate, with an “i” added as a convenient Vowel. Why does the IMViC go to such Lengths to distinguish these two Bacteria? This happens to be particularly important in Water Testing. The Presence of Enterobacter in a Water Sample is no Big Deal. Enterobacter can live in the Gut of Warm Blooded Animals but it’s also a normal Inhabitant of the Soil. But E. coli cannot live outside of the Gut of a Warm Blooded Animal for very long, so its Presence is considered Positive for Fecal Contamination. E. coli itself generally isn’t all that nasty a Bacterium (although some Strains have gone over to The Dark Side) but the Presence of E. coli implies the Presence of undeniable Nasty Pathogens like Hepatitis Virus. The Results from IMViCs for E. coli and Enterobacter are neatly Reciprocal: Indole MR VP Citrate E. coli + + - - Enterobacter - - + + Indole (Atlas Page 74) “Does this Bacterium have the Enzyme Tryptophanase?” Bacteria with the Enzyme Tryptophanase can convert the Amino Acid Tryptophan to Indole, Ammonia, and Pyruvic Acid. The Pyruvic Acid can be shunted into the TCA Cycle, the Ammonia can be used in the Synthesis of Amino Acids. The Indole is actually just a By-Product of this Reaction. The Presence of Indole is detected by adding Kovac’s Reagent, which consists of p-Dimethylaminobenzaldehyde (DMBA), HCl, and Amyl Alcohol. The DMBA reacts with Indole to form a Cherry Red Rosindole Dye. Since this Dye will remain in the Amyl Alcohol -- which is insoluble in Water -- a Positive Indole Test results in a Red Layer of Rosindole in the Amyl Alcohol Layer at the Top of the Tube. All you’ll do in Today’s Lab is inoculate separate Tubes of Tryptone Broth with your E. coli and/or your Enterobacter . Next Time you’ll add Kovac’s Reagent. E. coli is Indole Positive and will produce a Red Layer of Rosindol Dye in the Amyl Alcohol Layer at the Top of the Tube. Enterobacter is Indole Negative and will not produce any Dye in the Amyl Alcohol at the Top of the Tube so this Layer will remain Yellowish.
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Lab 6 Page 28 Methyl Red/Voges-Proskauer The Methyl Red/Voges-Proskauer Tests together determine whether Enteric Bacteria are Mixed Acid Fermenters of Glucose or Butanediol Fermenters of Glucose. You’ll be inoculating a Single Tube of MR-VP Medium with each Bacterium in Today’s Lab. This Medium contains Tryptone, Glucose, and a Phosphate Buffer. In the next Lab we’ll split the Contents of this Tube into separate Tubes and perform separate Methyl Red and Voges-Proskauer Tests. Methyl Red (Atlas Page 82) “Is this Bacterium a Mixed-Acid Fermenter?” Mixed Acid Fermenters of Glucose will produce Carbon Dioxide, Hydrogen, and Ethanol, as well as large Amounts of stable Acids, particularly Acetic Acid, Formic Acid, Lactic Acid, and Succinic Acid. After a 48 Hour Incubation in MR-VP Medium, we’ll add a Methyl Red pH Indicator.
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F Agar is a very Reliable Medium Lab 6 Page 27 The IMViC...

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