Investigation of the Rv3065, Rv2942, Rv1258c, Rv1410c, and Rv2459 efflux pump genes expression among multidrug-resistant Mycobacterium tuberculosis clinical isolates.

ABSTRACT

BACKGROUND: Different resistance mechanisms for multidrug-resistant tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB) have been reported. Although mutations in target genes are the main cause of drug resistance, efflux pumps (Eps) also play an important role in this process. Here, we investigated the overexpression of five putative EP genes plus gene mutations in MDR-TB clinical isolates. METHODS: A total of 27 M. tuberculosis (Mtb) clinical isolates including, 22 MDR and 5 sensitive isolates were analyzed. Minimum inhibitory concentrations (MIC) were determined in the absence and presence of efflux inhibitor. The expression level of 5 EP genes (Rv3065, Rv2942, Rv1258c, Rv1410c, Rv2459) was investigated by quantitative real time PCR (RT-qPCR). DNA sequencing of rpoB, katG, and inhA promoter was done. RESULTS: Among the 22 MDR-TB isolates, 13 (59.1%) showed significant overexpression (>4-fold) for at least one EP gene. The expression levels of 5 genes were significantly higher (P < 0.05) in MDR-TB isolates than sensitive isolates. The Rv3065 (22.7%), and Rv1410c (18.2%) were found to be the most commonly overexpressed EPs. The observed MICs were as follows RIF (2 to >128 µg/ml) and INH (2-32 µg/ml). After efflux pump inhibitor exposure, 10/22 (45.45%) isolates showed a decrease in MIC of INH, and 17/22 (77.27%) isolates showed a decrease in MIC of RIF. Of the isolates that overexpressed, 4 isolates lacked mutation in inhA, rpoB, and katG genes and 10 ones lacked mutation in inhA and katG. CONCLUSION: The results showed that overexpression of EP genes in Mtb isolates, besides target gene mutations can contribute to the development of MDR phenotype.