The capacity of Mycobacterium tuberculosis (Mtb) to tolerate multiple antibiotics represents a major problem in tuberculosis (TB) management. Heterogeneity in Mtb populations is one of the factors that drives antibiotic tolerance during infection. However, the mechanisms underpinning this variation in bacterial population remain poorly understood. Here, we show that phagosomal acidification alters the redox physiology of Mtb to generate a population of replicating bacteria that display drug tolerance during infection. RNA sequencing of this redox-altered population revealed the involvement of iron-sulfur (Fe-S) cluster biogenesis, hydrogen sulfide (H2S) gas, and drug efflux pumps in antibiotic tolerance. The fraction of the pH- and redox-dependent tolerant population increased when Mtb infected macrophages with actively replicating HIV-1, suggesting that redox heterogeneity could contribute to high rates of TB therapy failure during HIV-TB coinfection. Pharmacological inhibition of phagosomal acidification by the antimalarial drug chloroquine (CQ) eradicated drug-tolerant Mtb, ameliorated lung pathology, and reduced postchemotherapeutic relapse in in vivo models. The pharmacological profile of CQ (Cmax and AUClast) exhibited no major drug-drug interaction when coadministered with first line anti-TB drugs in mice. Our data establish a link between phagosomal pH, redox metabolism, and drug tolerance in replicating Mtb and suggest repositioning of CQ to shorten TB therapy and achieve a relapse-free cure.

SummaryAn increasing number of studies have established hydrogen sulfide (H2S) gas as a major cytoprotectant and redox modulator. Following its discovery, H2S has been found to have pleiotropic effects on physiology and human health. H2S acts as a gasotransmitter and exerts its influence on gastrointestinal, neuronal, cardiovascular, respiratory, renal, and hepatic systems. Recent discoveries have clearly indicated the importance of H2S in regulating vasorelaxation, angiogenesis, apoptosis, ageing, and metabolism. Contrary to studies in higher organisms, the role of H2S in the pathophysiology of infectious agents such as bacteria and viruses has been less studied. Bacterial and viral infections are often accompanied by changes in the redox physiology of both the host and the pathogen. Emerging studies indicate that bacterial-derived H2S constitutes a defense system against antibiotics and oxidative stress. The H2S signaling pathway also seems to interfere with redox-based events affected on infection with viruses. This review aims to summarize recent advances on the emerging role of H2S gas in the bacterial physiology and viral infections. Such studies have opened up new research avenues exploiting H2S as a potential therapeutic intervention.

Methionine and cysteine metabolisms are important for the survival and pathogenesis of Mycobacterium tuberculosis ( Mtb ). The transsulfuration pathway converts methionine to cysteine and represents an important link between antioxidant and methylation metabolism in diverse organisms. Using a combination of biochemistry and cryo–electron microscopy, we characterized the first enzyme of the transsulfuration pathway, cystathionine β-synthase ( Mtb Cbs) in Mtb . We demonstrated that Mtb Cbs is a heme-less, pyridoxal-5′-phosphate–containing enzyme, allosterically activated by S -adenosylmethionine (SAM). The atomic model of Mtb Cbs in its native and SAM-bound conformations revealed a unique mode of SAM-dependent allosteric activation. Further, SAM stabilized Mtb Cbs by sterically occluding proteasomal degradation, which was crucial for supporting methionine and redox metabolism in Mtb . Genetic deficiency of Mtb Cbs reduced Mtb survival upon homocysteine overload in vitro, inside macrophages, and in mice coinfected with HIV. Thus, the Mtb Cbs-SAM axis constitutes an important mechanism of coordinating sulfur metabolism in Mtb .