Mycobacterium tuberculosis H2S functions as a sink to modulate central metabolism, bioenergetics, and drug susceptibility

H 2 S is a potent gasotransmitter in eukaryotes and bacteria. Host-derived H 2 S has been shown to profoundly alter M. tuberculosis ( Mtb ) energy metabolism and growth. However, compelling evidence for endogenous production of H 2 S and its role in Mtb physiology is lacking. We show that multidrug-resistant and drug-susceptible clinical Mtb strains produce H 2 S, whereas H 2 S production in non-pathogenic M. smegmatis is barely detectable. We identified Rv3684 (Cds1) as an H 2 S-producing enzyme in Mtb and show that cds1 disruption reduces, but does not eliminate, H 2 S production, suggesting the involvement of multiple genes in H 2 S production. We identified endogenous H 2 S to be an effector molecule that maintains bioenergetic homeostasis by stimulating respiration primarily via cytochrome bd . Importantly, H 2 S plays a key role in central metabolism by modulating the balance between oxidative phosphorylation and glycolysis, and functions as a sink to recycle sulfur atoms back to cysteine to maintain sulfur homeostasis. Lastly, Mtb -generated H 2 S regulates redox homeostasis and susceptibility to anti-TB drugs clofazimine and rifampicin. These findings reveal previously unknown facets of Mtb physiology and have implications for routine laboratory culturing, understanding drug susceptibility, and improved diagnostics.