Key biochemical methods used to distinguish Mycobacterial group

Niacin test: All species of mycobacterium produces Niacin (nicotinic acid) and the Mycobacterium tuberculosis accumulates the most. A positive niacin test provides preliminary evidence that an organism that exhibits a buff-colored, slow-growing rough colony may be M. tuberculosis.

Nitrate Reduction test:  Mycobacterium tuberculosis is a strongly nitrate positive organism. This test is valuable for the identification of M. tuberculosis, M. kansasii, M. szulgai and M. fortuitum.

Rapid growers such as M. fortuitum can be tested within 2 weeks, but slow growers should be tested after 3-4 weeks of luxuriant growth.  Both chemical procedure and commercially available nitrate strips are available. Control strains must be used while performing and interpreting the result.

Catalase test: Semiquantitative catalase test is used for the identification of Mycobacteria. Catalase is an enzyme which splits Hydrogen peroxide to water and oxygen and positive catalase test is indicated by formation of gas bubbles.  Most species of Mycobacteria, except for certain strains of M. tuberculosis complex (Some isoniazid resistant strains) and M. gastri, produce catalase enzyme.

Tween 80 hydrolysis test: Tween 80 hydrolysis test is used to separate the species of Photochromogens, scotochromogens and nonchromogens. Non pathogenic slow growing scotochromogens and nonchromogens produce a lipase that is able to hydrolyze Tween 80 (the detergent polyoxyethylene sorbitan monooleate) into oleic acid and  polyoxyethylated sorbitol, where as pathogenic species do not.

Tellurite reduction test:  The ability of mycobacterial species to reduce tellurite in 3 to 4 days is used to distinguish members of M. avium complex from most other non-chromogenic species. All rapid growers reduce tellurite in 3 days.

Arylsulfatase test: Arylsulfatase enzyme is present in most mycobacteria. The rate by which arylsulfatase enzyme breaks down phenolphthalein disulfate into phenolphthalein (which forms a red color in the presence of sodium bicarbonate) and other salts is used to differentiate certain strains of Mycobacteria. 3 day arylsulfatase test is used to identify potentially pathogenic rapid growers such as M. fortuitum and M. chelonae. Slow growing M. marinum and M. szulgai are positive in the 14 day arylsulfatase test.