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.
Chemolithoautotrophic Fe(II) oxidizers belonging to Zetaproteobacteria play an important role in multiple biogeochemical cycles in the deep-sea biosphere. However, molecular and mechanistic understanding of Zetaproteobacteria physiology and metabolism remains speculative in the absence of a genetic system. These bacteria are considered recalcitrant to classical genetic techniques owing to their inability to form colonies on a solid medium, and production of copious amounts of iron oxyhydroxides with very few cells during liquid cultivation. Another challenge is their singular life style of Fe(II) dependent autotrophy, which prohibits comparative studies under different metabolic conditions. During my C-DEBI postdoctoral tenure, I successfully developed the first genetic tools and techniques to transform and express foreign genes in Mariprofundus ferrooxydans PV-1 as a model Zetaproteobacteria for genetic studies. I designed, constructed and transformed several different plasmids expressing foreign genes to provide M. ferrooxydans an ability to grow using an alternate metabolism. I successfully modified M. ferrooxydans genetically to grow using glucose as the sole carbon source, instead of carbon dioxide, while using Fe(II) as the energy source. The alternate metabolic ability thus provided to M. ferrooxydans can be leveraged to perform comparative studies in the future to decipher its physiology and metabolism. Furthermore, I developed an undergraduate teaching module for incoming freshmen students, focused on principles of chemolithoautotrophy in Fe(II) oxidizing bacteria.