Susceptibility of MTB to p-Nitrobenzoic acid (PNB)

PNB p-Nitrobenzoic acid susceptibility of MTB

Principle:

The inability to grow in the presence of PNB is one of the key elements in the phenotypic differentiation of tubercle bacilli from other mycobacterial species and is part of the identification process for M. tuberculosis.

M. tuberculosis and other tubercle bacilli will not grow on culture medium containing PNB, 500 µg/ml; other mycobacterial species, with the exception of  M.gastri and some strains of M. kansasii and M. marinum, will grow in the presence of PNB.

The test must be carried out on pure cultures; otherwise, it will yield false results.

Procedure:

•      Weigh out 0.5 gm PNB and dissolve in the minimum amount of dimethylformamide (~15ml).
•       Add to 1 litre of L-J fluid, distribute and inspissate once for 50 minutes at 85 degrees. 
•       Store in a cold room.
•       Inoculate with the neat bacterial suspension one slope of LJ medium and one slope of p-nitrobenzioc acid (PNB) at a concentration of 500 µg/ml and incubate at 37⁰C for each set. 
•      Read on the 28th day.
•      PNB should not be kept for reading on the 42nd day. 
•      It is critical to inoculate with a neat suspension prepared for DST, and reading should be only on the 28th day. 

Results and Interpretation:

 M. tuberculosis does not grow on the PNB medium. All other mycobacteria are resistant to PNB.

Resource

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

Assessment