A new and rapid testing method for drug susceptibility of Mycobacterium leprae using RT-PCR

Due to the uncultivable nature of Mycobacterium leprae in vitro, the fast, easy, and accurate measurement of the antimicrobial drug susceptibility of this microbe has been difficult. Conventional methods for such testing are subjective, cumbersome, and expensive in some cases. Here, the utility of a reverse transcriptase-PCR (RT-PCR)-based assay for testing was examined and compared with a Buddmeyer-type radiorespirometric assay. The susceptibility of M. leprae to rifampin was determined by probing the presence of M. leprae-specific 18 kDa gene mRNA in M. leprae-infected IC-21 macrophage cells after drag treatment. The results showed that, as the rifampin concentration was increased, the 360-bp cDNA products generated by the RT-PCR-based assay decreased in a dose-dependent manner as in the drug susceptibility observed in the Buddmeyer-type assay. The drag susceptibility testing of M. leprae by the RT-PCR based assay was found to be not only faster but also nearly 10⁴-fold more sensitive than the Buddmeyer-type assay. Moreover, it was also found that, unlike the RT-PCR based assay, the same testing by a DNA-PCR resulted in no differences in the 360-bp signal, regardless of the rifampin concentrations used. Accordingly, these results demonstrate that the drug susceptibility of M. leprae can be determined effectively by an RT-PCR-based assay, thereby providing a new, fast, and sensitive testing method.