Efflux pumps positively contribute to rifampin resistance in rpoB mutant Mycobacterium tuberculosis.

BACKGROUND: While several recent studies have documented the importance of efflux pumps as mediators of rifampin (RIF) resistance, it remains uncertain which efflux pumps play major roles in rifampin-resistant Mycobacterium tuberculosis strains harboring rpoB gene mutations. METHODS: In this study, minimum inhibitory concentration (MIC) values for RIF were calculated and the expression of 13 efflux pump genes was evaluated across 35 clinical rifampicin-resistant M. tuberculosis isolates carrying the rpoB mutation before and after efflux pump inhibitor treatment. RESULTS: Rv0677c and Rv0191 were identified as the efflux pump genes that were most frequently overexpressed, and treatment with the inhibitor verapamil was sufficient to synergistically enhance the antibacterial effects of RIF and downregulate efflux pump gene expression. Greater numbers of overexpressed efflux pump genes were associated with a more significant decrease in the MIC value for RIF following verapamil treatment. Levels of RIF resistance for clinical isolates with the rpoB codon 445 mutation were also found to be significantly less susceptible to the effects of verapamil as compared to the resistance of strains with the codon 450 and 170 mutations. CONCLUSIONS: These results suggest that levels of RIF resistance in clinical RIF-resistant M. tuberculosis isolates are ultimately determined by a combination of efflux pump activity and rpoB gene mutations.