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Introduction of Indole Test

Indole test positive bacteria as shown above picture and it is useful for both aerobic and anaerobic gram negative rods and really very useful in routine identification of bacteria in Microbiology.

Principle

The ability of bacteria to split indole form the amino acid tryptophan is due to the presence of enzyme, tryptophanase.  Indole,  if present  combines with aldehyde  in the reagent to produce a pink to red -violet quinoidal compound if using bezaldehyde reagent or blue to green when using cinnamaldehyde reagent.

Requirements for Test

• Fresh growth of gram negative rods (test organism)
• Sulphide indole motility (SIM) medium/ Motility indole urea (MIU) medium/ peptone water
• Kovac’s reagent
• Incubator
• Inoculating  wire  and Bunsen burner

Quality Control Strains

•  Positive control-E. coli (ATCC25922)
• Negative Control-Pseudomonas aeruginosa (ATCC27853)

 Test Procedure

1. Inoculate liquid tube medium or stab agar medium with colony.
2. Incubate for 18 to 24 hours in  BOD incubator.
3. Add 3 drops of Kovac’s reagent down the side of the tube  and observe color change at meniscus.
4. If the test is negative, repeat the test after additional 24 hours of incubation , if desired.

Result Interpretation

• Development of a brown-red to purple-red color:  Test  Positive (presence of indole)
• Colorless or slightly yellow:Negative test
• Tests positive (left)
• Test negative (Right) as shown above picture.

Indole Test Positive Organisms are-

• Escherichia coli
• Klebsiella oxytoca
• Proteus vulgaris
• Citrobacter koseri
• Morgenella morganii
• Vibrio cholerae
• Providencia species
• Aeromonas species
• Plesiomonas species
• Pasteurella species
• Cardiobacterium hominis
• Propionibacterium acenes

While Indole Test Negative 0rganisms are-

• Klebsiella pneumoniae
• Enterobacter species
• Proteus mirabilis
• Citrobacter freundii

Limitation of Indole Test

• Do not use media that contain dyes (e.g. Eosin methylene blue, MacConkey agar).

Bibliography

1. Cowan & Steel’s Manual for identification of Medical Bacteria. Editors: G.I. Barron & R.K. Felthani, 3rd ed 1993, Publisher Cambridge University press.
2. Bailey & Scott’s Diagnostic Microbiology. Editors: Bettey A. Forbes, Daniel F. Sahm & Alice S. Weissfeld, 12th ed 2007, Publisher Elsevier.
3. Clinical Microbiology Procedure Hand book, Chief in editor H.D. Isenberg, Albert Einstein College of Medicine, New York, Publisher ASM (American Society for Microbiology), Washington DC.
4. Colour Atlas and Text book of Diagnostic Microbiology. Editors: Koneman E.W., Allen D.D., Dowell V.R. Jr and Sommers H.M.
5. Jawetz, Melnick and Adelberg’s Medical Microbiology. Editors: Geo. F. Brook, Janet S. Butel & Stephen A. Morse, 21st ed 1998, Publisher Appleton & Lance, Co Stamford Connecticut.
6. Mackie and Mc Cartney Practical Medical Microbiology. Editors: J.G. Colle, A.G. Fraser, B.P. Marmion, A. Simmous, 4th ed, Publisher Churchill Living Stone, New York, Melborne, Sans Franscisco 1996.
7.  Text book of Diagnostic Microbiology. Editors: Connie R. Mahon, Donald G. Lehman & George Manuselis, 3rd edition2007, Publisher Elsevier.