Single-Cell Electrophysiological Measurements Reveal Bacterial Membrane Potential Dynamics during Extracellular Electron Transfer

Abstract Extracellular electron transfer (EET) allows microorganisms to gain energy by linking intracellular reactions to external surfaces ranging from natural minerals to the electrodes of bioelectrochemical renewable energy technologies. In the past two decades, electrochemical techniques have been used to investigate EET in a wide range of microbes, with emphasis on dissimilatory metal-reducing bacteria, such as Shewanella oneidensis MR-1, as model organisms. However, due to the typically bulk nature of these techniques, they are unable to reveal the subpopulation variation in EET or link the observed electrochemical currents to energy gain by individual cells, thus overlooking the potentially complex spatial patterns of activity in bioelectrochemical systems. Here, to address these limitations, we use the cell membrane potential as a bioenergetic indicator of EET by S. oneidensis MR-1 cells. Using a fluorescent membrane potential indicator during in vivo single-cell level fluorescence microscopy in a bioelectrochemical reactor, we demonstrate that membrane potential strongly correlates with the electrode potential, produced current, and position of cells relative to the electrodes. The high spatial and temporal resolution of the reported technique can be used to study the single-cell level dynamics of EET not only on electrode surfaces, but also during respiration of other solid-phase electron acceptors.