ABSTRACT
Mycobacterium abscessus is highly resistant to spectinomycin (SPC) thereby making it unavailable for therapeutic use. Sublethal exposure to SPC strongly induces whiB7 and its regulon, and a ΔMab_whiB7 strain is SPC sensitive suggesting that the determinants of SPC resistance are included within its regulon. In the present study we have determined the transcriptomic changes that occur in M. abscessus upon SPC exposure and have evaluated the involvement of 11 genes, that are both strongly SPC induced and whiB7 dependent, in SPC resistance. Of these we show that MAB_2780c can complement SPC sensitivity of ΔMab_whiB7 and that a ΔMab_2780c strain is â¼150 fold more SPC sensitive than wildtype bacteria, but not to tetracycline (TET) or other aminoglycosides. This is in contrast to its homologues, TetV from M. smegmatis and Tap from M. tuberculosis, that confer low-level resistance to TET, SPC and other aminoglycosides. We also show that the addition of the efflux pump inhibitor (EPI), verapamil results in >100-fold decrease in MIC of SPC in bacteria expressing Mab2780c to the levels observed for ΔMab_2780c; moreover a deletion of MAB_2780c results in a decreased efflux of the drug into the cell supernatant. Together our data suggest that Mab2780c is an SPC antiporter. Finally, molecular docking of SPC and TET on models of TetVMs and Mab2780c confirmed our antibacterial susceptibility findings that the Mab2780c pump preferentially effluxes SPC over TET. To our knowledge, this is the first report of an efflux pump that confers high-level drug resistance in M. abscessus. The identification of Mab2780c in SPC resistance opens up prospects for repurposing this relatively well-tolerated antibiotic as a combination therapy with verapamil or its analogs against M. abscessus infections.
