The diversity of microbially mediated redox processes that occur in marine sediments is likely underestimated, especially with respect to the metabolisms that involve solid substrate electron donors or acceptors. Though electrochemical studies that utilize poised potential electrodes as a surrogate for solid substrate or mineral interactions have shed some much needed light on these areas, these studies have traditionally been limited to one redox potential or metabolic condition. This work seeks to uncover the diversity of microbes capable of accepting cathodic electrons from a marine sediment utilizing a range of redox potentials, by coupling electrochemical enrichment approaches to microbial cultivation and isolation techniques. Five lab-scale three-electrode electrochemical systems were constructed, using electrodes that were initially incubated in marine sediment at cathodic or electron-donating voltages (five redox potentials between −400 and −750 mV versus Ag/AgCl) as energy sources for enrichment. Electron uptake was monitored in the laboratory bioreactors and linked to the reduction of supplied terminal electron acceptors (nitrate or sulfate). Enriched communities exhibited differences in community structure dependent on poised redox potential and terminal electron acceptor used. Further cultivation of microbes was conducted using media with reduced iron (Fe0, FeCl2) and sulfur (S0) compounds as electron donors, resulting in the isolation of six electrochemically active strains. The isolates belong to the genera Vallitalea of the Clostridia, Arcobacter of the Epsilonproteobacteria, Desulfovibrio of the Deltaproteobacteria, and Vibrio and Marinobacter of the Gammaproteobacteria. Electrochemical characterization of the isolates with cyclic voltammetry yielded a wide range of midpoint potentials (99.20 to −389.1 mV versus Ag/AgCl), indicating diverse metabolic pathways likely support the observed electron uptake. Our work demonstrates culturing under various electrochemical and geochemical regimes allows for enhanced cultivation of diverse cathode-oxidizing microbes from one environmental system. Understanding the mechanisms of solid substrate oxidation from environmental microbes will further elucidation of the ecological relevance of these electron transfer interactions with implications for microbe-electrode technologies.
Extracellular electron transport (EET) is a microbial process that allows microorganisms to transport electrons to and from insoluble substrates outside of the cell. Although progress has been made in understanding how microbes transfer electrons to insoluble substrates, the process of receiving electrons has largely remained unexplored. We investigated redox potentials favourable for donating electrons to dissolved and insoluble components in Catalina Harbor marine sediment by combining electrochemical techniques with geochemistry and molecular methods. Working electrodes buried in sediment microcosms were poised at seven redox potentials between −300 and −750 mV versus Ag/AgCl using a three‐electrode system. In electrode biofilms recovered after 2‐month incubations, overall community diversity increased with more negative redox potentials. Abundances of known EET‐capable groups (e.g., Alteromonadales and Desulfuromonadales) varied with redox potential. Motility and chemotaxis genes were found in greater abundance in electrode communities, suggesting a possible selective advantage of these pathways for colonization and utilization of the electrode. Our enrichments demonstrated the validity of this approach in capturing groups known, as well as novel groups (e.g., Campylobacterales) that perform EET. The diverse nature of the enriched cathode communities suggest that insoluble substrate oxidation may be a critical, although poorly described microbial metabolic process in marine sediment.
Little is known about the importance and/or mechanisms of biological mineral oxidation in sediments, partially due to the difficulties associated with culturing mineral-oxidizing microbes. We demonstrate that electrochemical enrichment is a feasible approach for isolation of microbes capable of gaining electrons from insoluble minerals. To this end we constructed sediment microcosms and incubated electrodes at various controlled redox potentials. Negative current production was observed in incubations and increased as redox potential decreased (tested −50 to −400 mV vs. Ag/AgCl). Electrode-associated biomass responded to the addition of nitrate and ferric iron as terminal electron acceptors in secondary sediment-free enrichments. Elemental sulfur, elemental iron and amorphous iron sulfide enrichments derived from electrode biomass demonstrated products indicative of sulfur or iron oxidation. The microbes isolated from these enrichments belong to the genera Halomonas, Idiomarina, Marinobacter, and Pseudomonas of the Gammaproteobacteria, and Thalassospira and Thioclava from the Alphaproteobacteria. Chronoamperometry data demonstrates sustained electrode oxidation from these isolates in the absence of alternate electron sources. Cyclic voltammetry demonstrated the variability in dominant electron transfer modes or interactions with electrodes (i.e., biofilm, planktonic or mediator facilitated) and the wide range of midpoint potentials observed for each microbe (from 8 to −295 mV vs. Ag/AgCl). The diversity of extracellular electron transfer mechanisms observed in one sediment and one redox condition, illustrates the potential importance and abundance of these interactions. This approach has promise for increasing our understanding the extent and diversity of microbe mineral interactions, as well as increasing the repository of microbes available for electrochemical applications.