EfpA is required for regrowth of Mycobacterium tuberculosis following isoniazid exposure

Efflux of antibiotics is an important survival strategy in bacteria. Mycobacterium tuberculosis has approximately sixty efflux pumps, but little is known about the role of each pump or the substrates they efflux. The putative efflux pump, EfpA, is a member of the major facilitator superfamily and has been shown to be essential by saturation transposon mutagenesis studies. It has been implicated in the efflux of isoniazid (INH), which is a first-line drug used to treat tuberculosis (TB). This is supported by evidence from transcriptional profiling showing that efpA is induced in response to INH exposure. However, its roles in the physiology and adaptation of M. tuberculosis to antibiotics have yet to be determined. In this study, we describe the repression of efpA in M. tuberculosis, using CRISPR interference (CRISPRi) to knockdown the expression of this essential gene and the direct effect of this on the ability of M. tuberculosis to survive exposure to INH over a 45-day time course. We determined that wild-type levels of efpA were required for recovery of M. tuberculosis following INH exposure and that, after 45 days of INH exposure, only a few viable colonies were recoverable from efpA-repressed M. tuberculosis. We conclude that EfpA is required for recovery of M. tuberculosis following INH exposure, which could reduce the efficacy of INH in vivo, and that EfpA may have a role in the development of resistance during drug therapy.