BDQ and CFZ resistant mutants of Mycobacterium tuberculosis. Emergence of non-canonical genomic variants following in vitro exposure of clinical Mycobacterium tuberculosis strains to bedaquiline or clofazimine

Resistance to bedaquiline and clofazimine occurs primarily through variants in Rv0678, a gene encoding a repressor protein, which affects the expression of the mmpS5/mmpL5 efflux pump proteins. Despite the shared effect of both these drugs on efflux, little else is known about other pathways affected. We hypothesized that the use of progenitor strains for generation of either bedaquiline- or clofazimine-resistant mutants could provide further insight into their mechanism of action. We performed whole genome sequencing and determined minimal inhibitory concentrations for both drugs at baseline and after mutant induction. This study reiterates the role of Rv0678 variants in bedaquiline and clofazimine resistance. We identified a number of genetic pathways that appear to be activated following induction of resistance. The shared genes affected for both bedaquiline and clofazimine include glpK, nuoG and uvrD1. The genes with variants in the bedaquiline resistant mutants included atpE, fadE28, truA, mmpL5, glnH and pks8, while those affected in clofazimine resistant mutants only included ppsD, fbiA, fbiD and Rv0988. These results highlight the complexity of bedaquiline and clofazimine resistance acquisition and cross resistance in Mycobacterium tuberculosis and show the importance of epistatic mechanisms as a means of responding to drug exposure.