Voges-Proskauer Test

 

What is the purpose of the test? 

 

This test determines whether the microbe produces 2,3-butanediol as a fermentation product from glucose.

 

 

How is 2,3-butanediol production determined? 

 

2,3-butanediol is the end product of a long fermentation pathway, but it is not easily detected. So, the test detects one of the pathway intermediates instead: acetylmethylcarbinol (acetoin). The production of 2,3-butanediol is thus indirectly detected when the pathway intermediate acetoin reacts with reagents to turn red.

 

 

What medium is used? 

 

The medium used is MRVP broth, a nutrient broth medium with 0.5% glucose added. This medium is also used for the methyl red test, which determines whether acidic fermentation products result from growth.

 

 

How is the test performed? 

 

An inoculum from a pure culture is transferred aseptically to a sterile tube of MRVP broth. The inoculated tube is incubated at 35-37 C for 24 hours.

 

 

What reagents are added?

 

After incubation, five drops of Barritt's A are added and the tube is shaken gently to mix the ingredients. Then, five drops of Barritt's B are added. The tube is then slanted to allow maximum oxygen exposure to the reaction mixture and allowed to stand for up to 30 minutes. Development of a red color indicates a positive test.

 

 

To perform this test in VirtualUnknown™ Microbiology, complete the following steps:

 

Inoculation of Medium

 

1. Select the MRVP broth medium.  

2. Start your Bunsen burner.  

3. Select the inoculating loop tool.  

4. Flame your inoculating loop to sterilize it.  

5. Remove the caps from your test tubes.  

6. Flame the mouths of your test tubes.  

7. Use the sterile inoculating tool to pick up an inoculum from the culture tube of the unknown bacterium.

8. Immediately transfer the inoculum into the fresh, sterile medium.  

9. Flame the mouths of your tubes once again.  

10. Replace the caps on the test tubes.  

11. Re-flame the inoculating tool.  

 

Incubation of the Inoculated Medium

 

12. Place the inoculated tube into the 35-37 C incubator.  

13. Press the New Day button to move forward 24 hours.  

14. Incubate for the appropriate length of time. This test is properly interpreted after 24 hours.  

15. Retrieve desired incubated culture from the incubator.  

 

Addition of Reagents

 

16. Next, you must add Barritt's A reagent followed by Barritt's B reagent to the culture to enact chemical changes that are necessary to complete this test. Select the dropper tool and then highlight Barritt's A from the Reagents list.  

17. Remove the cap from the tube.  

18. Place the end of the dropper into the tube and use a left mouse click to add the Barritt’s A reagent to the culture.  

19. Select the Barritt's B reagent from the reagent list.  Place the end of the dropper into the tube and add the Barritt's B reagent to the culture.  

 

Determination of Test Results

 

20. Observe test result. For this test, addition of the appropriate reagents makes the culture turn red if the test is positive. If negative, no color change is observed.  

22. Record test result.  Dispose of the culture.