Results of developing a special agar medium method for pyrazinamide susceptibility testing of the Mycobacterium tuberculosis strains in National Research Institute of Tuberculosis & Lung Diseases, Tehran, Iran.

Pyrazinamide (PZA) is one of the four front- line antituberculosis agents, which is very effective in eradicating the organism from the body by killing the active and non-active types of Mycobacterium tuberculosis in the acidic pH within macrophages, as well as in shortening the treatment course. Unlike the other antituberculosis agents, PZA is active in acidic pH, while the favorable pH for growth of the organism is 6. 8. This fact makes the susbptibility testing of the organism to PZA very difficult. With an appropriate and stable buffering system, pH of the 7H10 agar base medium was adjusted to be equal with 6. Moreover, the growth-inhibiting agents were omitted from the composition of the 7H10 agar base medium and it was highly enriched with special animal serum supplements, so that the organism could grow at a pH of 6 like the same way it would grow at the pH of 6. 8. Using the Henderson-Hassle Bausch's equation, which determines the enzyme-substrate activity at different pH levels, individual critical concentrations of PZA were determined and added to the medium, and the results were traced using the proportional method. Standard PZA-susceptible and PZA-resistant strains were used as controls along with the specific strains under investigation. As additional control groups, the specific strains under study and the standard PZA-susceptible and PZA-resistant strains were also cultivated in PZA-free mediums. In addition to PZA-susceptibility testing, susceptibility of the specific strains under study to the other front-line antituberculosis agents was also assessed (rifampin, isoniazid, etambotul, and streptomycin). Approximately 6 ٪ of the clinically isolated strains were identified as PZA-resistant and about 1 ٪ of the strains were resistant only to PZA. Since the low pH of the cultivation medium was continuously controlled in order not to interfere with growth of the organism, and the Henderson-Hassle Bausch's equation is very accurate in determination of the enzyme-substrate activity, our study results are reliable. An additional advantage of this method is feasibility of actual susceptibility or resistance determination based on different concentrations of the drug. It is also feasible to directly determine the susceptibility or resistance to PZA simultaneously with the primary sample culture.