Rifampin-Resistant RNA Polymerase Mutants of Chlamydia trachomatis Remain Susceptible to the Ansamycin Rifalazil

Stable, homotypic mutants of Chlamydia trachomatis for which MICs of rifampin and rifalazil are elevated were isolated by serial passage at sub-MIC concentrations of these compounds. An alternative approach, in which Chlamydia cells were incubated and subsequently passaged three times, all in the presence of the selective agent at concentrations above the MIC, appeared to be a more effective means of selecting for mutants. In every instance where an elevation in the MIC occurred, one or more mutations in the rpoB gene, encoding the rifampin binding site, were detected. With one exception, all rpoB mutants that contained a single mutation conferred lower levels of resistance than mutants containing multiple mutations. Some rpoB mutations conferred very high levels of resistance to rifampin, up to 512 mug/ml. In all cases, mutants remained susceptible to concentrations of rifalazil at or below 0.064 microg/ml. Thus, rifalazil, a compound that is extremely potent against Chlamydia wild-type cells (MIC of 0.00025 microg/ml), may also protect against the selection of mutants at physiologically achievable concentrations.