Identification of Enterobacteriaceae / Biochemical tests

• Briefly, describe the types of laboratory methods you would use in order to identify the various types of Enterobacteriaceae.

• What biochemical tests are available to test the properties of bacteria? Elaborate on each one.

There are a number of methods available in the laboratory, and only a few are identified here. An enriched broth solution can be used to identify some species. These include: tetrathionate and selenite (for shigella and salmonella). A solid medium with an indicator can also be used, or a more selective solid medium can be used. A solid medium may contain an indicator that can identify whether the specimen ferments lactose or not. If lactose is fermented  then acid is produced (lactic acid) and this turns the indicator pink. The non-lactose fermenters are pathogenic (i.e.: Shigella and Salmonella). The lactose fermenters are rarely pathogenic, and are usually part of the gut flora (i.e.: E.coli, Enterobacter, Citrobacter, Klebsiella). A selective medium can also be used, and these perform two functions: 1) they inhibit the growth of gram positive bacteria so that it will not contaminate the culture, 2) they turn a particular colour if lactic acid is formed. The two main types of selective media are: EMB agar (Eosin methyl blue), where the methyl blue inhibits the gram positive bacteria and if lactic acid is formed – then the medium turns a metallic green colour, MacConkey agar – where the bile salts in medium inhibit gram positive bacteria and if lactic acid is formed the medium turns a purple  black colour. Another identification method: is using a screening media / composite media  triple sugar iron agar – fermentation of glucose, sucrose, lactose, H2S production.

There are a range of biochemical tests available to highlight the biochemical properties of Enterobacteriaceae species – and these properties help in the biochemical classification. The following tests are available in commercial kit form. Some tests are semi-automated, &/or computerised. (Remember: DICKMMUV):

• Decarboxylase production: decarboxylation of arg, lys, orn to form alkaline products / indicator

• Indole production: oxidation of tryptophan to indole

• Citrate utilisation: ability to utilise citrate as the sole carbon source

• KCN: ability to grow in presence of cyanide

• Motility: young broth cultures

• Methyl red: ability to produce <>

• Urease production: ability to split urea in medium to liberate ammonia

• Voges-Proskauer reaction: production of acetyle-methyl cardinol from glucose

Category: Microbiology Notes