Extracellular electron transfer (EET) allows microbes to acquire energy from solid state electron acceptors and donors, such as environmental minerals. This process can also be harnessed at electrode interfaces in bioelectrochemical technologies including microbial fuel cells, microbial electrosynthesis, bioremediation, and wastewater treatment. Improving the performance of these technologies will benefit from a better fundamental understanding of EET in diverse microbial systems. While the mechanisms of outward (i.e. microbe-to-anode) EET is relatively well characterized, specifically in a few metal-reducing bacteria, the reverse process of inward EET from redox-active minerals or cathodes to bacteria remains poorly understood. This knowledge gap stems, at least partly, from the lack of well-established model organisms and general difficulties associated with laboratory studies in existing model systems. Recently, a sulfur oxidizing marine microbe, Thioclava electrotropha ElOx9, was demonstrated to perform electron uptake from cathodes. However, a detailed analysis of the electron uptake pathways has yet to be established, and electrochemical characterization has been limited to aerobic conditions. Here, we report a detailed amperometric and voltammetric characterization of ElOx9 cells coupling cathodic electron uptake to reduction of nitrate as the sole electron acceptor, even in the absence of any added inorganic carbon source. By comparing this cellular activity to spent media controls and using medium exchange experiments, we demonstrate that one of the pathways by which ElOx9 facilitates inward EET is by a direct-contact mechanism through a redox center with a formal potential of −94 mV vs SHE, rather than soluble intermediate electron carriers. In addition to the implications for understanding microbial sulfur oxidation in marine environments, this study highlights the potential for ElOx9 to serve as a convenient and readily culturable model organism for understanding the molecular mechanisms of inward EET.
While typically investigated as a microorganism capable of extracellular electron transfer to minerals or anodes, Shewanella oneidensis MR-1 can also facilitate electron flow from a cathode to terminal electron acceptors, such as fumarate or oxygen, thereby providing a model system for a process that has significant environmental and technological implications. This work demonstrates that cathodic electrons enter the electron transport chain of S. oneidensis when oxygen is used as the terminal electron acceptor. The effect of electron transport chain inhibitors suggested that a proton gradient is generated during cathode oxidation, consistent with the higher cellular ATP levels measured in cathode-respiring cells than in controls. Cathode oxidation also correlated with an increase in the cellular redox (NADH/FMNH2) pool determined with a bioluminescence assay, a proton uncoupler, and a mutant of proton-pumping NADH oxidase complex I. This work suggested that the generation of NADH/FMNH2 under cathodic conditions was linked to reverse electron flow mediated by complex I. A decrease in cathodic electron uptake was observed in various mutant strains, including those lacking the extracellular electron transfer components necessary for anodic-current generation. While no cell growth was observed under these conditions, here we show that cathode oxidation is linked to cellular energy acquisition, resulting in a quantifiable reduction in the cellular decay rate. This work highlights a potential mechanism for cell survival and/or persistence on cathodes, which might extend to environments where growth and division are severely limited.