IMViC Tests | Procedure and Principle | Microbiology

The IMViC tests are four biochemical assays designed to differentiate and identify members of the Enterobacteriaceae family, particularly coliforms. The acronym stands for Indole test, Methyl Red test, Voges-Proskauer test, and Citrate utilization test. These tests analyze the biochemical properties of unknown bacteria for characterization purposes. To conduct these tests effectively, inoculate a bacterial culture in an appropriate medium and incubate it at 37 degrees Celsius for 24 to 48 hours.

The indole test detects a bacterium's ability to produce indole from the amino acid tryptophan. To conduct the test, inoculate tryptophan broth with bacterial culture and incubate for 24 hours at 37°C. After incubation, add Kovac's reagent, which reacts with any produced indole to yield a red color if positive. A red coloration indicates that the bacteria have successfully degraded tryptophan into pyruvic acid and ammonium through the action of streptophanase enzyme.

The methyl red test assesses a bacterium's ability to produce and sustain stable acids from glucose fermentation. To conduct the test, inoculate glucose phosphate broth or MRVP medium with a sample and incubate for 24 hours at 37°C. After incubation, add methyl red indicator; if the bacteria have produced sufficient acid, the pH will drop, turning the solution red—indicating a positive result.

The Butylene Glycol Pathway Test identifies bacteria's ability to produce acetone, an intermediate in the pathway. A negative result is indicated by yellow color, while a positive test shows red after adding Alpha naphthol and potassium hydroxide post-incubation. Bacteria ferment glucose into pyruvate before producing acetone and carbon dioxide; if acid oil is present, it reacts with potassium hydroxide to yield diacetyl which turns pinkish-red when combined with guanidine from peptones. The Citrate Utilization Test assesses whether bacteria can use citrate as their sole carbon source through the production of citrate permease enzymes. In this test, streaking on Simmons citrate agar slant followed by incubation reveals results: a green-to-blue color change indicates successful utilization of citrate due to increased pH from sodium carbonate formation.