Biochemical tests


Written by MicroDok

Biochemical tests are series of experiments that can be used to differentiate bacteria based on their metabolic activity or ability to utilize a given substrate. These tests are generally useful in characterizing bacteria in the microbiology laboratory, and they take advantage of the organism’s physiological, osmotic, and nutritional requirements. The identification of the invading bacterial pathogen in a particular clinical disease condition is critical to the survival of the affected patients. This is because the quick and accurate identification of the bacterial species implicated in the disease condition will automatically lead to a better understanding of the disease process and the subsequent direction of the effective therapy to eliminate it.

A general knowledge of the normal flora that naturally inhabits the human body is indispensable in the diagnosis of bacterial-related diseases, thus the reason to biochemically test suspected bacterial isolates and ensure that treatment is not directed at commensal organisms but only against pathogenic strains. Bacteriological findings including biochemical tests must be able to properly distinguish invasive bacterial pathogens from indigenous microorganisms (i.e. normal floras). Biochemical tests can also help to different even closely related bacterial species. For example, pathogenic S. aureus can be differentiated from non-pathogenic S. aureus in the laboratory because the former (pathogenic S. aureus) produces coagulase enzymes that cause blood plasma to clot while the latter (non-pathogenic S. aureus) does not produce this enzyme, and thus cannot cause blood plasma to clot.

Certain bacterial species have the innate ability to elaborate or express definite enzymes and substances which are used in their own characterization in the microbiology laboratory. Colonies of bacteria growing on an agar plate differ characteristically in their shapes and appearances. Some bacteria also produce specific odour which are also used in their characterization. Bacterial species also differ in their staining properties when exposed to certain dyes, and thus Gram staining and other available staining techniques can as well be used to differentiate one bacterium from another. Bacterial species also differ in their nutritional requirement for growth, thus some selective/differential culture media (e.g. Mannitol salt agar, MacConkey agar) can presumptively help to identify bacteria and even distinguish amongst closely related species.

By growing bacteria on various culture media plates, microbiologists determine the key growth requirements of the respective organism in addition to other criteria which are all dependent on the biochemical and metabolic activity of the bacteria. Biochemical tests assist microbiologists to quickly and accurately identify bacteria from clinical or environmental samples. Many of these tests are commercially available as test kits, and they can help a microbiologist to run as many biochemical tests as possible on a given sample at a relatively short period unlike some conventional biochemical tests that require an overnight incubation before a result can be obtained. Though quick and accurate in identifying bacterial species from either clinical or environmental samples, the series of available commercial systems used for the rapid identification of bacteria are often expensive and difficult to obtain in some developing countries.

Generally, biochemical tests help microbiologists to simply put a name to a particular microorganism, and settle all uncertainty surrounding its identification. Thus, some commonly available conventional biochemical tests used for the identification of bacteria from either clinical or environmental samples in developing countries, some research institutes and even in schools for teaching shall be expanded here, and these tests include coagulase test, indole test, oxidase test, citrate test, catalase test, nitrate reduction test, urease test, triple sugar iron test, motility test, starch hydrolysis test, acid production test, hydrogen sulphide test, blood agar haemolysis test amongst others.

Summary of some biochemical tests and culture media used for identifying bacteria

Gram Positive Bacteria Gram Negative Bacteria
Indole test Voges-Proskauer (VP) test
Coagulase test Urease test
Catalase test Oxidase test
Mannitol Salt Agar (MSA) plates MacConkey agar plates
Blood Agar (BA) plates Simmon’s Citrate Agar (SCA) plates
Optochin sensitivity test Triple Sugar Iron Agar (TSIA)
Bacitracin sensitivity test Sugar utilization test
Starch hydrolysis test Methyl red test
Bile solubility test


Nitrate utilization test (also for Gram positive’s)
DNase test Motility test
CAMP test Eosin Methylene Blue (EMB) Agar

Key: CAMP = Cyclic Adenosine Mono-Phosphate



Cheesbrough M (2006). District Laboratory Practice in Tropical Countries. 2nd Cambridge University Press, UK. Pp. 178-187.

Willey J.M, Sherwood L.M and Woolverton C.J (2008). Harley and Klein’s Microbiology. 7th ed. McGraw-Hill Higher Education, USA.

Woods GL and Washington JA (1995). The Clinician and the Microbiology Laboratory. Mandell GL, Bennett JE, Dolin R (eds): Principles and Practice of Infectious Diseases. 4th ed. Churchill Livingstone, New York.

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