Dylan Huttenlocher

Bedaquiline (BDQ), an ATP synthesis inhibitor, is used to treat drug resistant tuberculosis (TB). Mutations in rv0678, a transcriptional repressor of the efflux pump MmpS5/MmpL5, are associated with bedaquiline resistance. For some patients with multi-drug resistant TB (MDR-TB), despite no prior BDQ treatment history, resistance-associated mutations in rv0678 have been observed. Thus, there is evidence for a selective factor other than BDQ itself for resistance-associated mutations to bedaquiline. Mutations in rv0678 are more prevalent in bedaquiline-naive patients with a history of other TB antibiotic use. As the MmpS5/MmpL5 efflux pump is thought to be involved in the oxidative stress response, it is hypothesized that oxidative stress from prior antibiotic exposure can select for rv0678 mutations. Strains with mutations in rv0678 are needed to determine the link between oxidative stress and bedaquiline resistance. Serial passaging of an attenuated, wild-type strain of Mycobacterium tuberculosis was conducted in media containing increasing concentrations of bedaquiline. Following two serial passages, mutations at fixation levels (>99%) in rv0678 were detected in three independent cultures. A frameshift mutation in the DNA binding domain and a nonsense mutation in the dimerization domain were discovered among these isolates that have not been documented in previous literature. These mutant strains will be used in subsequent studies to determine the role of rv0678 in the oxidative stress response of Mycobacterium tuberculosis and its involvement in bedaquiline resistance.