The VP assay can be applied to (1) identify Enterobacter aerogenes and Escherichia coli, and (2) characterize other bacterial groups, including actinomycetes.

Operation method

Barritt's method

Principle

Certain genera can decarboxylate pyruvic acid to produce neutral acetylmethylmethanol, which by the action of KOH and air converts acetylmethylmethanol (3-hydroxyacetone) to diacetyl, which is catalyzed by α-nairesinol and creatine to produce a reddish compound, which is a positive VP test.

Materials and Instruments

Escherichia coli Enterobacter aerogenes
Glucose peptone water VP test solution Solution A (α-naphthol alcohol solution) Solution B (40% KOH solution)
Petri dish Thermostat

Move

Inoculate the test organisms in the universal medium, incubate at 36±1°C for 4 days, 2.5ml of culture solution first add 5°C α-naphthol (2-na-phthol) pure alcohol solution 0.6ml, then add 40% potassium hydroxide aqueous solution 0.2ml, shake for 2-5min, the positive organisms often show red color immediately, if there is no red color, static at room temperature or in a constant temperature box at 36±1°C, if no red color is still visible within 2h, it can be judged as negative. If no red color appears, put it at room temperature or 36±1°C constant temperature box, if the red color still does not appear in 2h, it can be judged as negative.

Caveat

The experiments need to pay attention to the order of reagent addition, shaking well and left to stand for 10min and then observed.

Common Problems

1.VP is one of four tests in the IMViC series that test for evidence of gut bacteria. The other three tests include: indole test, methyl red test and citric acid test.

2.VP positive organisms include Enterobacteriaceae, Klebsiella, Serratia marcescens, Hafnia alvei, Vibrio cholerae biotype eltor and Vibrio alginolyticus.

3. VP-negative organisms include Citrobacter spp, Shigella, Yersinia, Edwards, Salmonella, Vibrio furnissii, Vibrio fluvialis, Vibrio traumaticus and Vibrio parahaemolyticus.

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