Extracellular Electron Transport Research Articles

Significantly Enhanced Nitrate and Phosphorus Removal by Pyrite/Sawdust Composite-Driven Mixotrophic Denitrification with Boosted Electron Transfer: Comprehensive Evaluation of Water-Gas-Biofilm Phases during a Long-Term Study.

Further reducing total nitrogen (TN) and total phosphorus (TP) in the secondary effluent needs to be realized effectively and in an eco-friendly manner. Herein, four pyrite/sawdust composite-based biofilters were established to treat simulated secondary effluent for 304 days. The results demonstrated that effluent TN and TP concentrations from biofilters under the optimal hydraulic retention time (HRT) of 3.5 h were stable at <2.0 and 0.1 mg/L, respectively, and no significant differences were observed between inoculated sludge sources. The pyrite/sawdust composite-based biofilters had low N2O, CH4, and CO2 emissions, and the effluent's DOM was mainly composed of five fluorescence components. Moreover, mixotrophic denitrifiers (Thiothrix) and sulfate-reducing bacteria (Desulfosporosinus) contributing to microbial nitrogen and sulfur cycles were enriched in the biofilm. Co-occurrence network analysis deciphered that Chlorobaculum and Desulfobacterales were key genera, which formed an obvious sulfur cycle process that strengthened the denitrification capacity. The higher abundances of genes encoding extracellular electron transport (EET) chains/mediators revealed that pyrite not only functioned as an electron conduit to stimulate direct interspecies electron transfer by flagella but also facilitated EET-associated enzymes for denitrification. This study comprehensively evaluates the water-gas-biofilm phases of pyrite/sawdust composite-based biofilters during a long-term study, providing an in-depth understanding of boosted electron transfer in pyrite-based mixotrophic denitrification systems.

Enhanced anaerobic digestion of landfill leachate based on a novel redox mediator: Synergistic mechanism of enhancing extracellular electron transfer

The presence of recalcitrant compounds and toxic substances in landfill leachate results in low anaerobic digestion (AD) efficiency and poor methane production performance. Strengthening the extracellular electron transfer of microorganisms represents an effective approach to promote the AD of landfill leachate. In this study, a novel redox mediator (FeNC-RM) was synthesized through the combination of metal and non-metal doping, intensifying the stable operation of the landfill leachate AD system. Through characterization techniques including X-ray photoelectron spectroscopy and X-ray diffraction, it was observed that FeNC-RM possesses a larger specific surface area, abundant functional groups, and exceptional electrochemical performance. By adding FeNC-RM, the stability and efficiency of the medium-temperature AD system in the treatment of landfill leachate were improved: Obtained a high removal of chemical oxygen demand which was increased by 30 % compared to the control group and enhanced the specific methanogenic activity by 57.5 % compared to the control group. The electron transport system (ETS) activities of the experimental groups with the addition of RMs were all significantly higher than those of the control group, and the ETS activity of the sludge from the FeNC-RM group was 79.6 % higher than that of the control group. Through a comprehensive analysis of the underlying mechanism, it was determined that three aspects synergistically facilitated electron transfer: FeNC-RM, endowed with electrochemical capabilities, assumed a pivotal role in establishing a conduit for electron transport, thus promoting the synthesis of extracellular humic substances and augmenting electron transport activity. Furthermore, the microbial community was optimized by FeNC-RM, thereby fostering a co-nutritional mechanism between hydrolytic acidification bacteria and methanogens, establishing a direct interspecific electron transport pathway. In this study, an “endogenous + exogenous” electron transport model based on the enhanced extracellular electron transport pathway facilitated by FeNC-RM was proposed. This study offered valuable contributions to the advancement of anaerobic treatment technology for landfill leachate.

MmcA is an electron conduit that facilitates both intracellular and extracellular electron transport in Methanosarcina acetivorans

Methanogens are a diverse group of Archaea that obligately couple energy conservation to the production of methane. Some methanogens encode alternate pathways for energy conservation, like anaerobic respiration, but the biochemical details of this process are unknown. We show that a multiheme c-type cytochrome called MmcA from Methanosarcina acetivorans is important for intracellular electron transport during methanogenesis and can also reduce extracellular electron acceptors like soluble Fe3+ and anthraquinone-2,6-disulfonate. Consistent with these observations, MmcA displays reversible redox features ranging from −100 to −450 mV versus SHE. Additionally, mutants lacking mmcA have significantly slower Fe3+ reduction rates. The mmcA locus is prevalent in members of the Order Methanosarcinales and is a part of a distinct clade of multiheme cytochromes that are closely related to octaheme tetrathionate reductases. Taken together, MmcA might act as an electron conduit that can potentially support a variety of energy conservation strategies that extend beyond methanogenesis.

Expression of filaments of the Geobacter extracellular cytochrome OmcS in Shewanella oneidensis.

The physiological role of Geobacter sulfurreducens extracellular cytochrome filaments is a matter of debate and the development of proposed electronic device applications of cytochrome filaments awaits methods for large-scale cytochrome nanowire production. Functional studies in G. sulfurreducens are stymied by the broad diversity of redox-active proteins on the outer cell surface and the redundancy and plasticity of extracellular electron transport routes. G. sulfurreducens is a poor chassis for producing cytochrome nanowires for electronics because of its slow, low-yield, anaerobic growth. Here we report that filaments of the G. sulfurreducens cytochrome OmcS can be heterologously expressed in Shewanella oneidensis. Multiple lines of evidence demonstrated that a strain of S. oneidensis, expressing the G. sulfurreducens OmcS gene on a plasmid, localized OmcS on the outer cell surface. Atomic force microscopy revealed filaments with the unique morphology of OmcS filaments emanating from cells. Electron transfer to OmcS appeared to require a functional outer-membrane porin-cytochrome conduit. The results suggest that S. oneidensis, which grows rapidly to high culture densities under aerobic conditions, may be suitable for the development of a chassis for producing cytochrome nanowires for electronics applications and may also be a good model microbe for elucidating cytochrome filament function in anaerobic extracellular electron transfer.

Electricity generation by Pseudomonas putida B6-2 in microbial fuel cells using carboxylates and carbohydrate as substrates

Microbial fuel cells (MFCs) employing Pseudomonas putida B6-2 (ATCC BAA-2545) as an exoelectrogen have been developed to harness energy from various conventional substrates, such as acetate, lactate, glucose, and fructose. Owing to its metabolic versatility, P. putida B6-2 demonstrates adaptable growth rates on diverse, cost-effective carbon sources within MFCs, exhibiting distinct energy production characteristics. Notably, the anode chamber's pH rises with carboxylates' (acetate and lactate) consumption and decreases with carbohydrates' (glucose and fructose) utilization. The MFC utilizing fructose as a substrate achieved the highest power density at 411 mW m−2. Initial analysis revealed that P. putida B6-2 forms biofilms covered with nanowires, contributing to bioelectricity generation. These microbial nanowires are likely key players in direct extracellular electron transport through physical contact. This study established a robust foundation for producing valuable compounds and bioenergy from common substrates in bioelectrochemical systems (BESs) utilizing P. putida as an exoelectrogen.

Fe/Co Zeolitic Imidazolate Framework Acted as an Electron Shuttle to Enhance Methanogenesis by Promoting Extracellular Electron Transport

Fe/Co Zeolitic Imidazolate Framework Acted as an Electron Shuttle to Enhance Methanogenesis by Promoting Extracellular Electron Transport

Quantifying cathode biofilm resistance in a cdrAB modified Shewanella oneidensis MR-1 using electrochemical impedance spectroscopy

A range of biotechnological applications for converting electricity to fixed metabolic substrates is fuelling the study of cathodic bioelectrochemical systems. Shewanella oneidensis MR-1 has emerged as an important model system for extracellular electron uptake on cathodes, as many of the proteins involved in this process overlap with the organism's previously characterized extracellular electron transport machinery. However, there are still many questions surrounding the mechanics of electron uptake in Shewanella stemming largely from the challenge of quantifying biomass on electrode surfaces. This limits our understanding of the physiologic and kinetic constraints of electron uptake, as well as our ability to make meaningful comparisons across systems. To investigate the relationship between cathodic activity and biomass, we used a Shewanella oneidensis strain genetically modified with cell aggregation protein CdrAB behind a blue light-controlled promotor. Using blue light exposure to control cell deposition, we then investigated the relationship between cathodic activity and biomass. Electrochemical impedance spectroscopy (EIS) confirmed a decrease in biofilm impedance over a range of blue light exposures (i.e., 2 to 8 h). Consistent with previous results, after this timepoint, a drop-in electrochemical activity was observed, and impedances increased. For biofilms within the 2–8 h light exposure range, we observed a trend towards increased biological current consumption by quantifying the difference between pre and post kill currents. Comparing EIS data between pre and post kill experiments supported an increase in impedance post addition of killing agents and a trend towards the lowest biofilm impedances observed in the longest blue light exposed systems. Using an equivalent circuit model to extrapolate specific biofilm parameters we quantified the charger transfer resistance within the biofilm that corresponds to varying biofilm thicknesses and matches previously observed activities. For example, electrochemical activity was highest for the 8 h blue light exposed biofilm condition, with a maximum cathodic biologic current of -5.49 ± 0.85 µA, and a biofilm charge transfer resistance measured at 6909.5 ± 2136.5 Ω. On an individual reactor basis, we correlate this biofilm charge transfer resistance with biologic cathodic current. We observed a linear trend with a correlation score of 0.87 (r2 = 0.773). To the best of our knowledge, this is the first investigation of biofilm physiology on Shewanella cathodes using EIS. Continued efforts in this direction will further our understanding of biofilm-electrode interface during extracellular electron uptake with the goal of enhancing applications to bioelectrochemical systems.

Unraveling response mechanism of electron transport mechanism to uranium-exposure in Bacillus sp. X02

With the development of the uranium mining industry, the remediation of uranium pollution is an urgent environmental problem to be solved. Bioreduction is currently recognized as a U(VI) bioremediation method with good application prospects. However, inadequate reporting of uranium-reducing bacteria and low bioreduction efficiency are the main reasons limiting the application of the bioreduction method. In this paper, we isolated an efficient uranium-reducing bacterium, Bacillus sp. X02, explored the effect of U(VI) on bioreduction efficiency through electron transport mechanisms. The results showed that the maximum removal and reduction rate were 98.35 % and 62.43 %, respectively. Further studies revealed that the efficiency of extracellular electron transport of Bacillus sp. X02 was enhanced under U(VI) exposure. But uranium stress down-regulated the expression of F-type ATPase, Cytochrome c reductase ISP, Cytochrome c oxidase, and Cytochrome bd complex, inhibited the processes of complex III, complex IV, and ATP synthesis in the oxidative phosphorylation electron transport pathway, decreasing the electron transfer and U(VI) reduction capacity of Bacillus sp. X02 ultimately. This work proposed a new electron transfer pathway for U(VI) bioreduction and provided new insights into the effect of uranium on bioreduction efficiency.

Highly conductive riboflavin-based carbon quantum dot–embedded SiO2@MoS2 nanocomposite for enhancing bioelectricity generation through synergistic direct and indirect electron transport

A microbial fuel cell (MFC) is an advanced green battery that has limited application because of its low current density. In the present study, Shewanella oneidensis MR-1, which is an electricity-producing bacterium, was used in an electrochemical reactor as a bacterial model. Mesoporous spherical silica nanoparticles (NPs) loaded with riboflavin (RF) were prepared as the starting material, following which surface modification was conducted to expose the thiol group, which considerably increased the affinity of the NPs to MoS2 and resulted in the formation of a dense MoS2 shell on the surface after calcination. Moreover, the loaded RFs were successfully transformed into RF-based carbon quantum dots (CQDs), which resulted in a substantial increase in conductivity. The CQD-embedded SiO2@MoS2 NPs, which contained redox-active N-doped CQDs and had a metallic MoS2 shell, were able to receive electrons from exoelectrogenic bacteria (charging) and transfer electrons to an indium tin oxide electrode (discharging). Thus, these NPs could act as an electron nanoshuttle and a conductive medium in biofilms for enhancing systematic extracellular electron transport. A 10-fold increase in bioelectricity production than no NPs addition was achieved, which confirmed the applicability of the aforementioned NPs in advanced MFC applications. The NPs prepared in this study, which mimic biological electron shuttles (e.g., RF) in long-distance conduction, can usher in a new era in the development of advanced MFCs.

Fe/Fe3C nanoparticles in situ-doped with carbon nanofibers embedded in rGO as high-performance anode electrocatalysts of microbial fuel cells

The aerogel structure composed of NCNFs and rGO is conducive to the colonization and extracellular electron transport of microorganisms.

Enhanced performance of biochar-biosensor applied to heavy metals detection in constructed wetlands and biological mechanisms

Biosensors play a very important role in pollutant monitoring, but there are still some problems such as stability, repeatability and low detection limits. Therefore, it is necessary to explore ways to increase the performance of biosensors. In this study, biochar-MFC sensors were constructed for heavy metals monitoring in constructed wetlands (CWs), and the correlation, repeatability and anti-interference were studied. At 1.0 M/L concentrations of heavy metals (Ni, Cu, Zn and Cr), CW with biochar as filler (B-CW) had an output voltage of 0.69 V, extending the detection limit of the sensor. B-CW had advantages in anti-interference and reproducibility. The results showed that B-CW had higher abundance of electrogenic bacteria than CW. Shewanella and Proteobacteria, main electricity-producing bacteria, increased 3-fold. The electrochemically active microorganism Shewanellaceae made up 6.2% of the B-CW microbial population, but was not detected in CW. The abundance of key genes AarC increased by 1295.8% in B-CW. More importantly, the abundance of extracellular electron transport chain related genes (MtrA, OmcA) increased significantly (p < 0.01). Biochar can promote oxidative phosphorylation, thus increasing the sensor's output voltage and detection range. This study is expected to provide support for the engineering application of MFC sensor in CWs.

Genome-Centric Metatranscriptomic Characterization of a Humin-Facilitated Anaerobic Tetrabromobisphenol A-Dehalogenating Consortium.

Tetrabromobisphenol A (TBBPA), a widely used brominated flame retardant in electronics manufacturing, has caused global contamination due to improper e-waste disposal. Its persistence, bioaccumulation, and potential carcinogenicity drive studies of its transformation and underlying (a)biotic interactions. This study achieved an anaerobic enrichment culture capable of reductively dehalogenating TBBPA to the more bioavailable bisphenol A. 16S rRNA gene amplicon sequencing and quantitative PCR confirmed that successive dehalogenation of four bromide ions from TBBPA was coupled with the growth of both Dehalobacter sp. and Dehalococcoides sp. with growth yields of 5.0 ± 0.4 × 108 and 8.6 ± 4.6 × 108 cells per μmol Br- released (N = 3), respectively. TBBPA dehalogenation was facilitated by solid humin and reduced humin, which possessed the highest organic radical signal intensity and reducing groups -NH2, and maintained the highest dehalogenation rate and dehalogenator copies. Genome-centric metatranscriptomic analyses revealed upregulated putative TBBPA-dehalogenating rdhA (reductive dehalogenase) genes with humin amendment, cprA-like Dhb_rdhA1 gene in Dehalobacter species, and Dhc_rdhA1/Dhc_rdhA2 genes in Dehalococcoides species. The upregulated genes of lactate fermentation, de novo corrinoid biosynthesis, and extracellular electron transport in the humin amended treatment also stimulated TBBPA dehalogenation. This study provided a comprehensive understanding of humin-facilitated organohalide respiration.

Boosting power output in microbial fuel cells with microwave-assisted synthesis of hierarchical bimetallic anodes for enhanced extracellular electron transport

Microbial fuel cells (MFCs) are a promising technology that can generate electricity from various organic waste sources using bacteria as catalysts. However, the low power density of MFCs due to the inefficient extracellular electron transport (EET) remains a major challenge for MFCs. To address this challenge, researchers have been developing new anodes that can enhance the EET efficiency of MFCs. In this study, we report the fabrication of an efficient anode by embedding FeCo bimetallic nanoparticles on carbon cloth (FexCoy@CC) using a microwave-assisted synthesis and calcination process. We find that the Fe1Co1@CC anode demonstrates excellent electrocatalytic activity for electron transfer between bacteria and the anode. Furthermore, in mixed bacterial culture-based MFCs, the Fe1Co1@CC anode shows exceptional electrochemical performances, achieving a high-power density of 3220 mW m−2, a current density of 6.98 A m−2, and a Coulombic efficiency of 16.74 %. The remarkable performance is attributed to the high electronic conductivity provided, the enrichment of electrochemically active Geobacter species, and the EET efficiency enhancement of microbial redox reactions through FeCo bimetallic nanoparticles. This facile and cost-effective method demonstrates great potential for improving the performance of MFCs, which could lead to more efficient and sustainable wastewater treatment and green energy production.

Synergistic effects of trace sulfadiazine and corrosion scales on disinfection by-product formation in bulk water of cast iron pipe

The effects of trace sulfadiazine (SDZ) and cast-iron corrosion scales on the disinfection by-product (DBP) formation in drinking water distribution systems (DWDSs) were investigated. The results show that under the synergistic effect of trace SDZ (10 μg/L) and magnetite (Fe3O4), higher DBP concentration occurred in the bulk water with the transmission and distribution of the drinking water. Microbial metabolism-related substances, one of the important DBP precursors, increased under the SDZ/Fe3O4 condition. It was found that Fe3O4 induced a faster microbial extracellular electron transport (EET) pathway, resulting in a higher microbial regrowth activity. On the other hand, the rate of chlorine consumption was quite high, and the enhanced microbial EET based on Fe3O4 eliminated the need for microorganisms to secrete excessive extracellular polymeric substances (EPS). More importantly, EPS could be continuously secreted due to the higher microbial activity. Finally, high reactivity between EPS and chlorine disinfectant resulted in the continuous formation of DBPs, higher chlorine consumption, and lower EPS content. Therefore, more attention should be paid to the trace antibiotics polluted water sources and cast-iron corrosion scale composition in the future. This study reveals the synergistic effects of trace antibiotics and corrosion scales on the DBP formation in DWDSs, which has important theoretical significance for the DBP control of tap water.

Dual role of birnessite on the modulation of acid production and reinforcement of interspecific electron transfer in anaerobic digestion

Achieving efficient anaerobic digestion of highly loaded substrates is one of the most challenging problems in the field of waste resourcing. Here, the addition of birnessite (2.0 g/L) to kitchen wastewater increased the acetate and final methane yields by 40.53 and 99.18 %, respectively, while reducing the yields of propionate and butyrate by 38.17 and 48.86 %, respectively. There were two main pathways for birnessite to enhance anaerobic digestion, one of which is to act as an electron acceptor, by inducing an alteration in the ratio of reduced-state coenzyme I in the microorganism, allowing the acid production process to proceed towards deeper oxidation. Another pathway enhances the interspecific electron transfer between bacteria and archaea and improves methane yield by optimizing the metabolic relationship. The Kyoto Encyclopedia of Genes and Genomes (KEGG) functional predictions suggest that the extracellular electron transport pathway of the microorganism is enhanced with the addition of birnessite and that its intracellular metabolic pathway is biased towards the nicotinamide adenine dinucleotide (NADH) generation pathway. This work demonstrated that anaerobic digestion facilitation by metallic minerals was not monolithic; that is, different properties of the minerals were employed to intensify the different stages of anaerobic digestion and obtain an amplification cascade.

Bacterial extracellular electron transfer components are spin selective.

Metal-reducing bacteria have adapted the ability to respire extracellular solid surfaces instead of soluble oxidants. This process requires an electron transport pathway that spans from the inner membrane, across the periplasm, through the outer membrane, and to an external surface. Multiheme cytochromes are the primary machinery for moving electrons through this pathway. Recent studies show that the chiral-induced spin selectivity (CISS) effect is observable in some of these proteins extracted from the model metal-reducing bacteria, Shewanella oneidensis MR-1. It was hypothesized that the CISS effect facilitates efficient electron transport in these proteins by coupling electron velocity to spin, thus reducing the probability of backscattering. However, these studies focused exclusively on the cell surface electron conduits, and thus, CISS has not been investigated in upstream electron transfer components such as the membrane-associated MtrA, or periplasmic proteins such as small tetraheme cytochrome (STC). By using conductive probe atomic force microscopy measurements of protein monolayers adsorbed onto ferromagnetic substrates, we show that electron transport is spin selective in both MtrA and STC. Moreover, we have determined the spin polarization of MtrA to be ∼77% and STC to be ∼35%. This disparity in spin polarizations could indicate that spin selectivity is length dependent in heme proteins, given that MtrA is approximately two times longer than STC. Most significantly, our study indicates that spin-dependent interactions affect the entire extracellular electron transport pathway.

Humic acid-dependent respiratory growth of Methanosarcina acetivorans involves pyrroloquinoline quinone.

Although microbial humus respiration plays a critical role in organic matter decomposition and biogeochemical cycling of elements in diverse anoxic environments, the role of methane-producing species (methanogens) is not well defined. Here we report that a major fraction of humus, humic acid reduction enhanced the growth of Methanosarcina acetivorans above that attributed to methanogenesis when utilizing the energy sources methanol or acetate, results which showed both respiratory and fermentative modes of energy conservation. Growth characteristics with methanol were the same for an identically cultured mutant deleted for the gene encoding a multi-heme cytochrome c (MmcA), results indicating MmcA is not essential for respiratory electron transport to humic acid. Transcriptomic analyses revealed that growth with humic acid promoted the upregulation of genes annotated as cell surface pyrroloquinoline quinone (PQQ)-binding proteins. Furthermore, PQQ isolated from the membrane fraction was more abundant in humic acid-respiring cells, and the addition of PQQ improved efficiency of the extracellular electron transport. Given that the PQQ-binding proteins are widely distributed in methanogens, the findings extend current understanding of microbial humus respiration in the context of global methane dynamics.

Mechanism of extracellular electron transport and reactive oxygen mediated Sb(III) oxidation by Klebsiella aerogenes HC10

Microbial oxidation and the mechanism of Sb(III) are key governing elements in biogeochemical cycling. A novel Sb oxidizing bacterium, K. aerogenes HC10, was attracted early and revealed that extracellular metabolites were the main fractions driving Sb oxidation. However, linkages between the extracellular metabolite driven Sb oxidation process and mechanism remain elusive. Here, model phenolic and quinone compounds, i.e., anthraquinone-2,6-disulfonate (AQDS) and hydroquinone (HYD), representing extracellular oxidants secreted by K. aerogenes HC10, were chosen to further study the Sb(III) oxidation mechanism. N2 purging and free radical quenching showed that oxygen-induced oxidation accounted for 36.78% of Sb(III) in the metabolite reaction system, while hydroxyl free radicals (·OH) accounted for 15.52%. ·OH and H2O2 are the main driving factors for Sb oxidation. Radical quenching, methanol purification and electron paramagnetic resonance (EPR) analysis revealed that ·OH, superoxide radical (O2•−) and semiquinone (SQ−•) were reactive intermediates of the phenolic induced oxidation process. Phenolic-induced ROS are one of the main oxidants in metabolites. Cyclic voltammetry (CV) showed that electron transfer of quinone also mediated Sb(III) oxidation. Part of Sb(V) was scavenged by the formation of the secondary Sb(V)-bearing mineral mopungite [NaSb(OH)6] in the incubation system. Our study demonstrates the microbial role of oxidation detoxification and mineralization of Sb and provides scientific references for the biochemical remediation of Sb-contaminated soil.

Detrimental impact of the Geobacter metallireducens type VI secretion system on direct interspecies electron transfer.

Direct interspecies electron transfer (DIET) is important in anaerobic communities of environmental and practical significance. Other than the need for close physical contact for electrical connections, the interactions of DIET partners are poorly understood. Type VI secretion systems (T6SSs) typically kill competitive microbes. Surprisingly, Geobacter metallireducens highly expressed T6SS genes when DIET-based co-cultures were initiated with Geobacter sulfurreducens. T6SS gene expression was lower when the electron shuttle anthraquinone-2,6-disulfonate was added to alleviate the need for interspecies contact. Disruption of hcp, the G. metallireducens gene for the main T6SS needle-tube protein subunit, and the most highly upregulated gene in DIET-grown cells eliminated the long lag periods required for the initiation of DIET. The mutation did not aid DIET in the presence of granular-activated carbon (GAC), consistent with the fact that DIET partners do not make physical contact when electrically connected through conductive materials. The hcp-deficient mutant also established DIET quicker with Methanosarcina barkeri. However, the mutant also reduced Fe(III) oxide faster than the wild-type strain, a phenotype not expected from the loss of the T6SS. Quantitative PCR revealed greater gene transcript abundance for key components of extracellular electron transfer in the hcp-deficient mutant versus the wild-type strain, potentially accounting for the faster Fe(III) oxide reduction and impact on DIET. The results highlight that interspecies interactions beyond electrical connections may influence DIET effectiveness. The unexpected increase in the expression of genes for extracellular electron transport components when hcp was deleted emphasizes the complexities in evaluating the electromicrobiology of highly adaptable Geobacter species. IMPORTANCE Direct interspecies electron transfer is an alternative to the much more intensively studied process of interspecies H2 transfer as a mechanism for microbes to share electrons during the cooperative metabolism of energy sources. DIET is an important process in anaerobic soils and sediments generating methane, a significant greenhouse gas. Facilitating DIET can accelerate and stabilize the conversion of organic wastes to methane biofuel in anaerobic digesters. Therefore, a better understanding of the factors controlling how fast DIET partnerships are established is expected to lead to new strategies for promoting this bioenergy process. The finding that when co-cultured with G. sulfurreducens, G. metallireducens initially expressed a type VI secretion system, a behavior not conducive to interspecies cooperation, illustrates the complexity of establishing syntrophic relationships.

Life strategies for Aminicenantia in subseafloor oceanic crust.

After decades studying the microbial "deep biosphere" in subseafloor oceanic crust, the growth and life strategies in this anoxic, low energy habitat remain poorly described. Using both single cell genomics and metagenomics, we reveal the life strategies of two distinct lineages of uncultivated Aminicenantia bacteria from the basaltic subseafloor oceanic crust of the eastern flank of the Juan de Fuca Ridge. Both lineages appear adapted to scavenge organic carbon, as each have genetic potential to catabolize amino acids and fatty acids, aligning with previous Aminicenantia reports. Given the organic carbon limitation in this habitat, seawater recharge and necromass may be important carbon sources for heterotrophic microorganisms inhabiting the ocean crust. Both lineages generate ATP via several mechanisms including substrate-level phosphorylation, anaerobic respiration, and electron bifurcation driving an Rnf ion translocation membrane complex. Genomic comparisons suggest these Aminicenantia transfer electrons extracellularly, perhaps to iron or sulfur oxides consistent with mineralogy of this site. One lineage, called JdFR-78, has small genomes that are basal to the Aminicenantia class and potentially use "primordial" siroheme biosynthetic intermediates for heme synthesis, suggesting this lineage retain characteristics of early evolved life. Lineage JdFR-78 contains CRISPR-Cas defenses to evade viruses, while other lineages contain prophage that may help prevent super-infection or no detectable viral defenses. Overall, genomic evidence points to Aminicenantia being well adapted to oceanic crust environments by taking advantage of simple organic molecules and extracellular electron transport.