# Indole test

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{{Short description|Biochemical method to identify bacteria}}
{{About|the biochemical test|the detection of indoles such as LSD|Ehrlich's reagent}}

{{More citations needed|date=November 2022}}
The '''indole test''' is a biochemical test performed on [bacterial species](/source/Bacteria) to determine the ability of the organism to convert [tryptophan](/source/tryptophan) into [indole](/source/indole). This division is performed by a chain of a number of different intracellular [enzyme](/source/enzyme)s, a system generally referred to as "[tryptophanase](/source/tryptophanase)."{{cn|date=November 2022}}

==Biochemistry==
Indole is generated by reductive [deamination](/source/deamination) from tryptophan via the intermediate molecule indolepyruvic acid. [Tryptophanase](/source/Tryptophanase) catalyzes the deamination reaction, during which the [amine](/source/amine) (-NH<sub>2</sub>) group of the tryptophan molecule is removed. Final products of the reaction are indole, [pyruvic acid](/source/pyruvic_acid), [ammonium](/source/ammonium) (NH<sub>4</sub><sup>+</sup>) and energy. [Pyridoxal phosphate](/source/Pyridoxal_phosphate) is required as a [coenzyme](/source/coenzyme). 
center

==Performing a test==
thumb|Indole test positive: appearance of pink layer at top (e.g. ''Escherichia coli'')

Like many biochemical tests on bacteria, results of an indole test are indicated by a change in color following a reaction with an added reagent.

Pure bacterial culture must be grown in sterile tryptophan or peptone broth for 24–48 hours before performing the test. Following incubation, five drops of [Kovács reagent](/source/Kov%C3%A1cs_reagent) ([isoamyl alcohol](/source/isoamyl_alcohol), [para-Dimethylaminobenzaldehyde](/source/para-Dimethylaminobenzaldehyde), concentrated [hydrochloric acid](/source/hydrochloric_acid)) are added to the culture broth.

A positive result is shown by the presence of a red or reddish-violet color in the surface alcohol layer of the broth. A negative result appears yellow. A variable result can also occur, showing an orange color as a result. This is due to the presence of [skatole](/source/skatole), also known as methyl indole or methylated indole, another possible product of tryptophan degradation.

The positive red color forms as a result of a series of reactions. The para-Dimethylaminobenzaldehyde reacts with [indole](/source/indole) present in the medium to form a red rosindole dye. The isoamyl alcohol forms a [complex](/source/Coordination_complex) with rosindole dye, which causes it to [precipitate](/source/Precipitation_(chemistry)). The remaining alcohol and the precipitate then rise to the surface of the medium.

A variation on this test using [Ehrlich's reagent](/source/Ehrlich's_reagent) (using [ethyl alcohol](/source/ethyl_alcohol) in place of isoamyl alcohol, developed by [Paul Ehrlich](/source/Paul_Ehrlich)) is used when performing the test on nonfermenters and [anaerobe](/source/anaerobe)s.

===Indole-Positive Bacteria===
Bacteria that test positive for cleaving indole from tryptophan include: ''[Aeromonas hydrophila](/source/Aeromonas_hydrophila)'', ''[Aeromonas punctata](/source/Aeromonas_punctata)'', ''[Bacillus alvei](/source/Bacillus_alvei)'', ''[Edwardsiella](/source/Edwardsiella_(bacterium))'' sp., ''[Escherichia coli](/source/Escherichia_coli)'', ''[Flavobacterium](/source/Flavobacterium)'' sp., ''[Haemophilus influenzae](/source/Haemophilus_influenzae)'', ''[Klebsiella oxytoca](/source/Klebsiella_oxytoca)'', ''[Proteus](/source/Proteus_(bacterium))'' sp. (not ''P. mirabilis'' and ''P. penneri''), ''[Plesiomonas shigelloides](/source/Plesiomonas_shigelloides)'', ''[Pasteurella multocida](/source/Pasteurella_multocida)'', ''[Pasteurella](/source/Pasteurella) pneumotropica'', ''[Vibrio](/source/Vibrio)'' sp., and ''[Lactobacillus reuteri](/source/Lactobacillus_reuteri)''.

===Indole-Negative Bacteria===
Bacteria which give negative results for the indole test include: ''[Actinobacillus](/source/Actinobacillus)'' spp., ''[Aeromonas salmonicida](/source/Aeromonas_salmonicida)'', ''[Alcaligenes](/source/Alcaligenes)'' sp., most ''[Bacillus](/source/Bacillus)'' sp., ''[Bordetella](/source/Bordetella)'' sp., ''[Enterobacter](/source/Enterobacter)'' sp., most ''[Haemophilus](/source/Haemophilus)'' sp., most ''[Klebsiella](/source/Klebsiella)'' sp., ''[Neisseria](/source/Neisseria)'' sp., ''[Mannheimia haemolytica](/source/Mannheimia_haemolytica)'', ''Pasteurella ureae'', ''[Proteus mirabilis](/source/Proteus_mirabilis)'', ''[P. penneri](/source/Proteus_penneri)'', ''[Pseudomonas](/source/Pseudomonas)'' sp., ''[Salmonella](/source/Salmonella)'' sp., ''[Serratia](/source/Serratia)'' sp., ''[Yersinia](/source/Yersinia)'' sp., and ''[Rhizobium](/source/Rhizobium)'' sp.

The Indole test is one of the four tests of the [IMViC](/source/IMViC) series, which tests for evidence of an enteric bacterium. The other three tests include: the [methyl red test](/source/methyl_red_test) [M], the [Voges–Proskauer test](/source/Voges%E2%80%93Proskauer_test) [V] and the [citrate test](/source/citrate_test) [C].<ref>Bachoon, Dave S., and Wendy A. Dustman. Microbiology Laboratory Manual. Ed. Michael Stranz. Mason, OH: Cengage Learning, 2008. Exercise 15, "Normal Flora of the Intestinal Tract" Print.</ref>

==References==
{{Reflist}}
* MacFaddin, Jean F. "Biochemical Tests for Identification of Medical Bacteria." Williams & Wilkins, 1980, pp 173 – 183.
* [https://web.archive.org/web/20100613045203/http://inst.bact.wisc.edu/inst/index.php?module=Book&func=displayarticle&art_id=119 Example of typical indole reactions]
* {{cite journal | pmid = 10028248 | doi=10.1099/00207713-49-1-67 | volume=49 Pt 1 | title=Taxonomic relationships of the [Pasteurella] haemolytica complex as evaluated by DNA-DNA hybridizations and 16S rRNA sequencing with proposal of Mannheimia haemolytica gen. nov., comb. nov., Mannheimia granulomatis comb. nov., Mannheimia glucosida sp. nov., Mannheimia ruminalis sp. nov. and Mannheimia varigena sp. nov | year=1999 | journal=Int J Syst Bacteriol | pages=67–86  | last1 = Angen | first1 = O | last2 = Mutters | first2 = R | last3 = Caugant | first3 = DA | last4 = Olsen | first4 = JE | last5 = Bisgaard | first5 = M}}

{{Clinical microbiology techniques}}

Category:Bacteriology terminology
Category:Microbiology techniques

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