New agar medium for testing susceptibility of Mycobacterium tuberculosis to pyrazinamide

A new agar medium to perform pyrazinamide (PZA) susceptibility testing with Mycobacterium tuberculosis has been developed. This medium has an acidic pH of 6.0 instead of the usual for agar media, pH 6.8, to provide optimal conditions for PZA activity, and it also differs from conventional Middlebrook 7H10/7H11 agar in that animal serum (fetal or calf bovine or fetal equine serum) is used instead of oleic acid-albumin-dextrose-catalase to support good growth of M. tuberculosis at the low pH of 6.0. A critical concentration of 900 or 1,200 microg of PZA/ml in this medium made it possible to differentiate between PZA-susceptible and PZA-resistant clinical isolates. This agar medium has the following advantages compared to a liquid medium: it allows determination of the actual proportion of PZA-resistant bacteria in the isolate and it is simple and inexpensive. In addition, it has the potential of being used for a direct susceptibility test with PZA, but this approach will require further confirmation. Further studies to develop critical concentrations of other drugs for this low-pH medium, as well as to investigate the possibility of cultivation in regular (non-CO(2)) incubators, are in progress.