Simultaneous sulfide oxidation and sulfate reduction for intracellular redox homeostasis under highly acidic conditions

Sulfide oxidation and sulfate reduction are core yet counteracting processes in sulfur cycle, typically separated spatially or temporally in microorganisms to avoid energetically futile cycling. Challenging this paradigm, Mycobacterium sp. MAG-M116, a bidirectional sulfur-metabolizing bacterium that simultaneously performs sulfide oxidation and assimilatory sulfate reduction (ASR), was identified in an autotrophic bio-desulfurization system under sulfate-rich and H2S-overloaded conditions. Under aerobic conditions, MAG-M116 mediates sulfide oxidation terminating at elemental sulfur, channeling electrons into the quinone pool to simultaneously drive forward electron transfer for ATP synthesis and reverse electron transfer (RET) for NADPH generation, powering ASR. This bidirectional metabolism enables MAG-M116 to rapidly detoxify sulfide while simultaneously preventing excessive generation of damaging reactive oxygen species (ROS)—a vital adaptation for survival in H2S-overloaded environments. Herein, ASR, acting as a redox homeostat, dynamically captures electrons from leak-prone RET (stimulated by intense sulfide oxidation) to reduce sulfate, thereby mitigating RET electron leakage and resulting in a remarkable 57.5% reduction in ROS production. Consequently, the Mycobacterium genus dominate in this autotrophic system (abundance rising from 3.5% to 99%). These findings redefine intracellular microbial sulfur metabolic network and reveal an energetically futile strategy driven by simultaneous opposing redox reactions, offering a novel paradigm for chemolithoautotrophic redox homeostasis in substrate-excess environments.