Electrochemical Investigation of a Microbial Solar Cell Reveals a Nonphotosynthetic Biocathode Catalyst

Microbial solar cells (MSCs) are microbial fuel cells (MFCs) that generate their own oxidant and/or fuel through photosynthetic reactions. Here, we present electrochemical analyses and biofilm 16S rRNA gene profiling of biocathodes of sediment/seawaterbased MSCs inoculated from the biocathode of a previously described sediment/seawater-based MSC. Electrochemical analyses indicate that for these second-generation MSC biocathodes, catalytic activity diminishes over time if illumination is provided during growth, whereas it remains relatively stable if growth occurs in the dark. For both illuminated and dark MSC biocathodes, cyclic voltammetry reveals a catalytic-current–potential dependency consistent with heterogeneous electron transfer mediated by an insoluble microbial redox cofactor, which was conserved following enrichment of the dark MSC biocathode using a three-electrode configuration. 16S rRNA gene profiling showed Gammaproteobacteria, most closely related to Marinobacter spp., predominated in the enriched biocathode. The enriched biocathode biofilm is easily cultured on graphite cathodes, forms a multimicrobe-thick biofilm (up to 8.2 μm), and does not lose catalytic activity after exchanges of the reactor medium. Moreover, the consortium can be grown on cathodes with only inorganic carbon provided as the carbon source, which may be exploited for proposed bioelectrochemical systems for electrosynthesis of organic carbon from carbon dioxide. These results support a scheme where two distinct communities of organisms develop within MSC biocathodes: one that is photosynthetically active and one that catalyzes reduction of O2 by the cathode, where the former partially inhibits the latter. The relationship between the two communities must be further explored to fully realize the potential for MSC applications

Metaproteomic evidence of changes in protein expression following a change in electrode potential in a robust biocathode microbiome

Microorganisms that respire electrodes may be exploited for biotechnology applications if key pathways for extracellular electron transfer (EET) can be identified and manipulated through bioengineering. To determine whether expression of proposed Biocathode-MCL EET proteins are changed by modulating electrode potential without disrupting the relative distribution of microbial constituents, metaproteomic and 16S rRNA gene expression analyses were performed after switching from an optimal to suboptimal potential based on an expected decrease in electrode respiration. Five hundred and seventy-nine unique proteins were identified across both potentials, the majority of which were assigned to three previously defined Biocathode-MCL metagenomic clusters: a Marinobacter sp., a member of the family Chromatiaceae, and a Labrenzia sp. Statistical analysis of spectral counts using the Fisher's exact test identified 16 proteins associated with the optimal potential, five of which are predicted electron transfer proteins. The majority of proteins associated with the suboptimal potential were involved in protein turnover/turnover, motility, and membrane transport. Unipept and 16S rRNA gene expression analyses indicated that the taxonomic profile of the microbiome did not change after 52 hours at the suboptimal potential. These findings show that protein expression is sensitive to the electrode potential without inducing shifts in community composition, a feature that may be exploited for engineering Biocathode-MCL

A Previously Uncharacterized, Nonphotosynthetic Member of the Chromatiaceae Is the Primary CO_2-Fixing Constituent in a Self-Regenerating Biocathode

Biocathode extracellular electron transfer (EET) may be exploited for biotechnology applications, including microbially mediated O_2 reduction in microbial fuel cells and microbial electrosynthesis. However, biocathode mechanistic studies needed to improve or engineer functionality have been limited to a few select species that form sparse, homogeneous biofilms characterized by little or no growth. Attempts to cultivate isolates from biocathode environmental enrichments often fail due to a lack of some advantage provided by life in a consortium, highlighting the need to study and understand biocathode consortia in situ. Here, we present metagenomic and metaproteomic characterization of a previously described biocathode biofilm (+310 mV versus a standard hydrogen electrode [SHE]) enriched from seawater, reducing O_2, and presumably fixing CO_2 for biomass generation. Metagenomics identified 16 distinct cluster genomes, 15 of which could be assigned at the family or genus level and whose abundance was roughly divided between Alpha- and Gammaproteobacteria. A total of 644 proteins were identified from shotgun metaproteomics and have been deposited in the the ProteomeXchange with identifier PXD001045. Cluster genomes were used to assign the taxonomic identities of 599 proteins, with Marinobacter, Chromatiaceae, and Labrenzia the most represented. RubisCO and phosphoribulokinase, along with 9 other Calvin-Benson-Bassham cycle proteins, were identified from Chromatiaceae. In addition, proteins similar to those predicted for iron oxidation pathways of known iron-oxidizing bacteria were observed for Chromatiaceae. These findings represent the first description of putative EET and CO_2 fixation mechanisms for a self-regenerating, self-sustaining multispecies biocathode, providing potential targets for functional engineering, as well as new insights into biocathode EET pathways using proteomics

Microbial Electrochemical Energy Storage and Recovery in a Combined Electrotrophic and Electrogenic Biofilm

Here we report enrichment from a marine-derived inoculum of a nonphotosynthetic electroactive biofilm that is capable of both consuming electricity (electrotrophy) and producing electricity (electrogenesis) from a single electrode. With the alternation of the electrode potential between −0.4 and 0.0 VSHE every 10 min, alternating anodic and cathodic currents increased in lock step (maximum current density of ±1.4 ± 0.4 A/m2 in both modes, Coulombic efficiency of ∼98% per charge−discharge cycle), which is consistent with alternating between generation and consumption of energy storage compounds by the biofilm. Cyclic voltammetry exhibited a single sigmoid-shaped feature spanning anodic and cathodic limiting currents centered at −0.15 VSHE, a phenomenon not observed to date for an electroactive biofilm, and square wave voltammetry exhibited reversible peaks at −0.15 and −0.05 VSHE, suggesting the same redox cofactor(s) facilitates electron transport at the biofilm−electrode interface in both modes. Hydrogen and carbon monoxide, known energy and/or carbon sources for cellular metabolism, but no volatile fatty acids, were detected in reactors. Cells and cell clusters were spread across the electrode surface, as seen by confocal microscopy. These results suggest that a single microbial electrochemical biofilm can alternate between storing energy and generating power, furthering the potential applicability of bioelectrochemical systems

Biocathode MCL Curated Metagenome

This file contains the curated metagenome assembled from a combination of eight biological replicates of Illumina short read sequencing data, two closed genomes and one plasmid sequence derived from PacBio RSII sequencing of DNA isolated from two pure cultures, and one closed genome with a plasmid assembled from PacBio metagenomic data.  The genome bins and unbinned contigs matching the three complete genome sequences and two plasmids were removed from the short read metagenome assembly and replaced by the closed sequences.  Raw data, parent metagenome assembly, and isolate sequences are linked through NCBI Bioproject PRJNA244670 at https://www.ncbi.nlm.nih.gov