1. Ex. 18: IMViC Reactions Objectives ?? Indole production test
Methyl red test
Voges-Proskauer test
Citrate utilization test.
Objectives ??

2. Day 1 Materials needed per team of two students:
2 tryptone broth tubes
2 MR-VP broth tubes
2 Simmons citrate slants
Pure cultures needed per table:
Escherichia coli
Enterobacter aerogenes
Work as a team of two. Each team member will work with one bacterial species

3. Day 2 Materials needed per team of two:
2 clean tubes and two 1 ml transfer pipettes
Materials needed per table:
Small biohazard bag
Kovac’s reagent
Methyl red indicator reagent
VP reagents A and B
Work as a team of two. Each team member will work with one bacterial species

4. Indole Test Checks for Tryptophanase
Medium: 1% Tryptone broth
Test for indole with Kovac’s reagent
dimethylamino-benzaldehyde (DMABA) in
DMABA in Kovac’s reagent combines with indole and forms red band at the top of the medium

5. Table 18-1: Indole Production Results and Interpretation
Symbol
Red in the alcohol layer of Kovac’s reagent
Tryptophan is broken down into indole and pyruvate +
Reagent color is unchanged
Tryptophan is not broken down into indole and pyruvate 
Enterobacter
aerogenes
E. coli

6. Methyl Red and Voges-Proskauer Tests
Physiologically related tests, use same medium (MRVP broth), NB with glucose
MR test checks for large amounts of acid endproducts from glucose fermentation
Do not confuse methyl red indicator with phenol red indicator!!!
VP test checks for production of acetoin, a neutral fermentation end product

7. Table 18-2: Methyl Red Test Results and Interpretation Result
Symbol
Red
Mixed acid fermentation +
No color change
No mixed acid fermentation 
Table 18-3: Voges-Proskauer Test Results and Interpretation
Result
Interpretation
Symbol
Red
Acetoin produced +
No color change
Acetoin is not produced 

8. MRVP after incubation and before adding test reagents

9. MR test After addition of 3 -4 drops of methyl red indicator
Explain the meaning of this outcome

10. VP Test
After addition of 15 drops of VP reagent A, and 5 drops of VP reagent B
E. a.
E. c.
Explain the meaning of this outcome

11. Citrate Utilization Test
Checks if organism has citrate permease
Simmons citrate agar also contains ammonium phosphate as sole N-source and bromothymol blue as indicator.
If bacterium is able to use citrate it will also convert ammonium phosphate to NH3 and NH4OH

12. Table 18-4: Citrate Test Results and Interpretation
Symbol
Blue (even a small amount)
Citrate is utilized +
No growth or weak growth with no color change
Citrate is NOT utilized 

13. the end

14. Could an organism be both MR and VP positive? Explain.
The End