Niacin production test for MTB

Principle:

Although all mycobacteria produce niacin, comparative studies have shown that M. tuberculosis accumulates the largest amount of nicotinic acid, and its detection is useful for its definitive diagnosis. Niacin-negative M .tuberculosis strains are very rare, while very few other mycobacterial species yield positive niacin tests. 

Cultures grown on egg medium containing Asparagine yield the most consistent results in the niacin test, and LJ medium is therefore recommended. A culture must be at least three to four weeks old and must have sufficient growth of at least 100 colonies. 

Controls:

Check the reagents by testing extract from an uninoculated tube of medium (negative control) and use an extract from a culture of M. tuberculosis H37Rv as a positive control.

Reagents: 

1) O-tolidine - 1.5%: Mix 1.5 gm of O-tolidine in 100 ml of ethanol. Keep in an amber bottle and store in the dark in the refrigerator. Prepare fresh weekly.

 2) Cyanogen bromide solution, approx. 10%.: Prepare a saturated aqueous solution of cyanogen bromide (approx. 10%). Store at 4^oC in the refrigerator.

Procedure:

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Results and Interpretation:

Pink colour                 = positive result

White precipitation     = negative result

Add 2-3 ml of 4%NaOH to all test tubes and discard them.

Precautions: 

• Cyanogen bromide is a severe lachrymator and toxic if inhaled. Work in a well-ventilated fume hood when preparing the solution and in a biological safety cabinet while testing cultures. Alternatively, commercially available niacin strips could also be used. 

• In acid solutions, cyanogen bromide hydrolyses to hydrocyanic acid, which is extremely toxic. Discard all reaction tubes into a disinfectant solution made alkaline by the addition of sodium hydroxide.

Resource

Revised National TB Control Programme Training Manual for Mycobacterium tuberculosis Culture & Drug susceptibility testing

Assessment