The mutational signatures of poor treatment outcomes on the drug-susceptible Mycobacterium tuberculosis genome.

Drug resistance is a known risk factor for poor tuberculosis (TB) treatment outcomes, but the contribution of other bacterial factors to poor outcomes in drug-susceptible TB is less well understood. Here, we generate a population-based dataset of drug-susceptible Mycobacterium tuberculosis (MTB) isolates from China to identify factors associated with poor treatment outcomes. We analyzed whole-genome sequencing (WGS) data of MTB strains from 3196 patients, including 3105 patients with good and 91 patients with poor treatment outcomes, and linked genomes to patient epidemiological data. A genome-wide association study (GWAS) was performed to identify bacterial genomic variants associated with poor outcomes. Risk factors identified by logistic regression analysis were used in clinical models to predict treatment outcomes. GWAS identified fourteen MTB fixed mutations associated with poor treatment outcomes, but only 24.2% (22/91) of strains from patients with poor outcomes carried at least one of these mutations. Isolates from patients with poor outcomes showed a higher ratio of reactive oxygen species (ROS)-associated mutations compared to isolates from patients with good outcomes (26.3% vs 22.9%, t-test, p=0.027). Patient age, sex, and duration of diagnostic delay were also independently associated with poor outcomes. Bacterial factors alone had poor power to predict poor outcomes with an AUC of 0.58. The AUC with host factors alone was 0.70, but increased significantly to 0.74 (DeLong's test, p=0.01) when bacterial factors were also included. In conclusion, although we identified MTB genomic mutations that are significantly associated with poor treatment outcomes in drug-susceptible TB cases, their effects appear to be limited.

The potassium transporter KdpA affects persister formation by regulating ATP levels in Mycobacterium marinum.

Mycobacterial persistence mechanisms remain to be fully characterized. Screening a transposon insertion library of Mycobacterium marinum identified kdpA, whose inactivation reduced the fraction of persisters after exposure to rifampicin. kdpA encodes a transmembrane protein that is part of the Kdp-ATPase, an ATP-dependent high-affinity potassium (K+) transport system. We found that kdpA is induced under low K+ conditions and is required for pH homeostasis and growth in media with low concentrations of K+. The inactivation of the Kdp system in a kdpA insertion mutant caused hyperpolarization of the cross-membrane potential, increased proton motive force (PMF) and elevated levels of intracellular ATP. The KdpA mutant phenotype could be complemented with a functional kdpA gene or supplementation with high K+ concentrations. Taken together, our results suggest that the Kdp system is required for ATP homeostasis and persister formation. The results also confirm that ATP-mediated regulation of persister formation is a general mechanism in bacteria, and suggest that K+ transporters could play a role in the regulation of ATP levels and persistence. These findings could have implications for the development of new drugs that could either target persisters or reduce their presence.