Voges-Proskauer test (VP test)
Objective:
- To check whether the microorganism follow 2,3 butanediol production pathway for glucose metabolism or not
Principle of VP test:
The basic principle for Voges-Proskauer test is to determine the ability of some microorganisms to produce a neutral end product 2,3 butanediol from glucose fermentation. Those microorganisms which follows 2,3 butanediol formation pathway give positive VP test and negative MR test. 2,3 butanediol is a neutral end product of glucose fermentation pathway which does not change the pH of media so color of methyl red indicator does not changes and remain yellow. The VP test is given by the intermediate product ie Acetion formed just before butanediol in the pathway. Acetion reacts with VP reagent (α-napthol + KOH) to give pinkish red complex, which indicates positive test.
The reagent α-naptholl is known as VP-I/ Barrit’s reagent A and the reagent 40% KOH is known as VP-II/ Barrit’s reagent B.
Requirements:
- 24 hours Culture of E.coli and Klebsiella spp
- α-napthol
- Potassium hydroxide (40% KOH)
- Absolute alcohol
- Deionized water
Procedure for VP test:
- Prepare MRVP broth or Clark and Lub’s media in test tubes
- Inoculate the broth aseptically 2 loopful of respective bacterial culture
- Label the test tubes with name of organism inoculated
- Incubate the test tubes at 37°C for 48-72 hours.
- Add Barrit’s reagent A (α-napthol) and Barrit’s reagent B (40% KOH) in the ratio 3: 1
- Remove the cotton plug and shake the tubes for aeration.
- Observe the results after 10-15 minutes
Result:
VP positive: Pinkish red color at the surface (Klebsiella spp)
- VP positive organisms are: Klebsiella, Serratia marcescens, Hafnia alvei,Viridans group streptococci (except Streptococcus vestibularis), Listeria, Enterobacter, Vibrio eltor, Vibrio alginolyticus, etc.
VP negative: whitish yellow ( E.coli)
- VP negative organisms are: E. coli, Streptococcus mitis, Citrobacter sp., Shigella, Yersinia, Edwardsiella, Salmonella, Vibrio furnissii, Vibrio fluvialis, Vibrio vulnificus, and Vibrio parahaemolyticus
** Both MR and VP positive organisms: Hafnia spp, Proteus spp, Serratia spp
Precautions:
- The culture must be incubated nit less than 48 hours
- The reagents should be used in suitable amount ie. VP-I and VP-II in ratio 3:1
- Aeration must be maintained by opening cotton plug and shaking the tubes
- Observe the color change carefully.