The electrode potential determines the yield coefficients of early-stage Geobacter sulfurreducens biofilm anodes.

Geobacter sulfurreducens is the model for electroactive microorganisms (EAM). EAM can use solid state terminal electron acceptors (TEA) including anodes via extracellular electron transfer (EET). Yield coefficients relate the produced cell number or biomass to the oxidized substrate or the reduced TEA. These data are not yet sufficiently available for EAM growing at anodes. Thus, this study provides information about kinetics as well as yield coefficients of early-stage G. sulfurreducens biofilms using anodes as TEA at the potentials of -200 mV, 0 mV and +200 mV (vs. Ag/AgCl sat. KCl). The selected microorganism was therefore cultivated in single and double chamber batch reactors on graphite or AuPd anodes. Interestingly, whereas the lag time and maximum current density within 12 days of growth differed, the anode potential does not influence the coulombic efficiency and the formal potential of the EET, which remains constant for all the experiments at ~ -300 to -350 mV. We demonstrated for the first time that the anode potential has a strong influence on single cell yield coefficients which ranged from 2.69 × 1012 cells mole--1 at -200 mV and 1.48 × 1012 cells mole--1 at 0 mV to 2.58 × 1011 cells mole--1 at +200 mV in single chamber reactors and from 1.15 × 1012 cells mole--1 at -200 mV to 8.98× 1011 cells mole--1 at 0 mV in double chamber reactors. This data can be useful for optimization and scaling-up of primary microbial electrochemical technologies.

Identification of Clostridium cochlearium as an electroactive microorganism from the mouse gut microbiome.

Microbial electroactivity, the metabolically relevant transfer of electrons between microorganisms and solid conductors, was first discovered for now well characterized model organisms from hypoxic or anaerobic water or sediment samples. Recent findings indicate that the metabolic trait of electroactivity might as well be important within the microbiome of the mammalian gut. Based on a pre-selection from the mouse intestinal bacterial collection five microorganisms originating from diverse parts of the gut were screened for electroactivity. As there is no marker gene for electroactivity, the ability to synthesize cytochromes and metabolize redox-mediators was studied in-silico. Clostridium cochlearium showed highest electroactivity and Lactobacillus reuteri as well as Staphylococcus xylosus show putative electroactivity, as well. The maximum current density of C. cochlearium of 0.53 ±â€¯0.02 mA cm-2 after only 5.2 h of incubation was clearly linked to growth and glucose consumption. Cyclic voltammetric analysis on C. cochlearium revealed a formal potential of the extracellular electron transfer (EET) site of +0.22 ±â€¯0.05 V versus Ag/AgCl sat. KCl (and + 0.42 V versus SHE) and indicates that EET is not based on biofilm formation, but the involvement of either redox-active molecules or planktonic cells. The potential of the gut as habitat for electroactives and their physiological role are discussed.

Microbial recycling cells: First steps into a new type of microbial electrochemical technologies, aimed at recovering nutrients from wastewater.

The aim of this work were to study terracotta-based porous air-water separators (4 mm thickness) in microbial recycling cells (MRCs) fed with cow manure (CM), swine manure (SM) and dairy wastewater (DW). Over 125 days, besides the removal of 60-90% of soluble-COD, considerable fractions of the main macronutrients (C, N, P, K, Fe, Mn, Ca, Mg) were removed from the wastewater and deposited on the terracotta separators as both inorganic salts and biomass deposits. Water evaporation at air-water interface as well as the high cathodic pH (10-12), induced by oxygen reduction to OH-, were the predominant factors leading to precipitation. The separators were saturated of up to 10 g per kg of terracotta of the main macronutrients, with negligible concentrations of the main inorganic contaminants. These materials could be directly reused as nutrients-enriched solid conditioners for agricultural soils.

Bioelectrochemical Nitrogen fixation (e-BNF): Electro-stimulation of enriched biofilm communities drives autotrophic nitrogen and carbon fixation.

A new approach to microbial electrosynthesis is proposed, aimed at producing whole biomass from N2 and inorganic carbon, by electrostimulation of complex microbial communities. On a carbon-based conductor under constant polarization (-0.7 V vs SHE), an electroactive biofilm was enriched with autotrophic nitrogen fixing microorganims and led to biomass synthesis at higher amounts (up to 18 fold), as compared to controls kept at open circuit (OC). After 110 days, the electron transfer had increased by 30-fold, as compared to abiotic conditions. Metagenomics evidenced Nif genes associated with autotrophs (both Archaea and Bacteria) only in polarized biofilms, but not in OC. With this first proof of concept experiment, we propose to call this promising field 'bioelectrochemical nitrogen fixation' (e-BNF): a possible way to 'power' biological nitrogen fixation, organic carbon storage and soil fertility against desertification, and possibly a new tool to study the development of early prokaryotic life in extreme environments.

A study of microbial communities on terracotta separator and on biocathode of air breathing microbial fuel cells.

Recently, terracotta has attracted interest as low-cost and biocompatible material to build separators in microbial fuel cells (MFCs). However, the influence of a non-conductive material like terracotta on electroactive microbiological communities remains substantially unexplored. This study aims at describing the microbial pools developed from two different seed inocula (bovine and swine sewage) in terracotta-based air-breathing MFC. A statistical approach on microbiological data confirmed different community enrichment in the MFCs, depending mainly on the inoculum. Terracotta separators impeded the growth of electroactive communities in contact with cathodes (biocathodes), while a thick biofilm was observed on the surface (anolyte-side) of the terracotta separator. Terracotta-free MFCs, set as control experiments, showed a well-developed biocathode, Biocathode-MFCs resulted in 4 to 6-fold higher power densities. All biofilms were analyzed by high-throughput Illumina sequencing applied to 16S rRNA gene. The results showed more abundant (3- to 5-fold) electroactive genera (mainly Geobacter, Pseudomonas, Desulfuromonas and Clostridia MBA03) in terracotta-free biocathodes. Nevertheless, terracotta separators induced only slight changes in anodic microbial communities.

Influences of dissolved oxygen concentration on biocathodic microbial communities in microbial fuel cells.

Dissolved oxygen (DO) at cathodic interface is a critical factor influencing microbial fuel cells (MFC) performance. In this work, three MFCs were operated with cathode under different DO conditions: i) air-breathing (A-MFC); ii) water-submerged (W-MFC) and iii) assisted by photosynthetic microorganisms (P-MFC). A plateau of maximum current was reached at 1.06±0.03mA, 1.48±0.06mA and 1.66±0.04mA, increasing respectively for W-MFC, P-MFC and A-MFC. Electrochemical and microbiological tools (Illumina sequencing, confocal microscopy and biofilm cryosectioning) were used to explore anodic and cathodic biofilm in each MFC type. In all cases, biocathodes improved oxygen reduction reaction (ORR) as compared to abiotic condition and A-MFC was the best performing system. Photosynthetic cultures in the cathodic chamber supplied high DO level, up to 16mgO2L-1, which sustained aerobic microbial community in P-MFC biocathode. Halomonas, Pseudomonas and other microaerophilic genera reached >50% of the total OTUs. The presence of sulfur reducing bacteria (Desulfuromonas) and purple non-sulfur bacteria in A-MFC biocathode suggested that the recirculation of sulfur compounds could shuttle electrons to sustain the reduction of oxygen as final electron acceptor. The low DO concentration limited the cathode in W-MFC. A model of two different possible microbial mechanisms is proposed which can drive predominantly cathodic ORR.

Performance of microbial electrolysis cells with bioanodes grown at different external resistances.

Bioelectrochemical systems need an anode with a high abundance of exoelectrogenic bacteria for an optimal performance. Among all possible operational parameters for an efficient enrichment, the role of external resistance in microbial fuel cell (MFC) has gained a lot of interest since it indirectly poises an anode potential, a key parameter for biofilm distribution and morphology. Thus, this work aims at investigating and discussing whether bioanodes selected at different external resistances under MFC operation present different responses under both MFC and microbial electrolysis cell (MEC) operation. A better MEC performance (i.e. shorter start-up time, higher current intensity and higher H2 production rate) was obtained with an anode from an MFC developed under low external resistance. Quantitative real-time polymerase chain reaction (qPCR) confirmed that a low external resistance provides an MFC anodic biofilm with the highest content of Geobacter because it allows higher current intensity, which is correlated to exoelectrogenic activity. High external resistances such as 1,000 Ω led to a slower start-up time under MEC operation.

Increased performance of hydrogen production in microbial electrolysis cells under alkaline conditions.

This work reports the first successful enrichment and operation of alkaline bioelectrochemical systems (microbial fuel cells, MFC, and microbial electrolysis cells, MEC). Alkaline (pH=9.3) bioelectrochemical hydrogen production presented better performance (+117%) compared to conventional neutral conditions (2.6 vs 1.2 litres of hydrogen gas per litre of reactor per day, LH2·L(-1)REACTOR·d(-1)). Pyrosequencing results of the anodic biofilm showed that while Geobacter was mainly detected under conventional neutral conditions, Geoalkalibacter sp. was highly detected in the alkaline MFC (21%) and MEC (48%). This is the first report of a high enrichment of Geoalkalibacter from an anaerobic mixed culture using alkaline conditions in an MEC. Moreover, Alkalibacter sp. was highly present in the anodic biofilm of the alkaline MFC (37%), which would indicate its potentiality as a new exoelectrogen.

Anode Biofilms of Geoalkalibacter ferrihydriticus Exhibit Electrochemical Signatures of Multiple Electron Transport Pathways.

Thriving under alkaliphilic conditions, Geoalkalibacter ferrihydriticus (Glk. ferrihydriticus) provides new applications in treating alkaline waste streams as well as a possible new model organism for microbial electrochemistry. We investigated the electrochemical response of biofilms of the alkaliphilic anode-respiring bacterium (ARB) Glk. ferrihydriticus voltammetry (CV), electrochemical impedance spectroscopy (EIS), and chronoamperometry. We observed there to be at least four dominant electron transfer pathways, with their contribution to the overall current produced dependent on the set anode potential. These pathways appear to be manifested at midpoint potentials of approximately -0.14 V, -0.2 V, -0.24 V, and -0.27 V vs standard hydrogen electrode. The individual contributions of the pathways change upon equilibration from a set anode potential to another anode potential. Additionally, the contribution of each pathway to the overall current produced is reversible when the anode potential is changed back to the original set potential. The pathways involved in anode respiration in Glk. ferrihydriticus biofilms follow a similar, but more complicated, pattern as compared to those in the model ARB, Geobacter sulfurreducens. This greater diversity of electron transport pathways in Glk. ferrihydriticus could be related to its wider metabolic capability (e.g., higher pH and larger set of possible substrates, among others).

Microbial community analysis in a long-term membrane-less microbial electrolysis cell with hydrogen and methane production.

A single-chamber microbial electrolysis cell (MEC) aiming at hydrogen production with acetate as sole carbon source failed due to methanogenesis build-up despite the significant amount of 2-bromoethanesulfonate (BES) dosage, 50 mM. Specific batch experiments and a thorough microbial community analysis, pyrosequencing and qPCR, of cathode, anode and medium were performed to understand these observations. The experimental data rebuts different hypothesis and shows that methanogenesis at high BES concentration was likely due to the capacity of some Archaea (hydrogen-oxidizing genus Methanobrevibacter) to resist high BES concentration up to 200 mM. Methanobrevibacter, of the Methanobacteriales order, represented almost the 98% of the total Archaea in the cathode whereas Geobacter was highly abundant in the anode (72% of bacteria). Moreover, at higher BES concentration (up to 200 mM), methanogenesis activity decreased resulting in an increase of homoacetogenic activity, which challenged the performance of the MEC for H2 production.

2-Bromoethanesulfonate degradation in bioelectrochemical systems.

2-Bromoethanesulfonate (BES) is the most reported chemical inhibitor for methanogenesis in laboratory-scale bioelectrochemical systems. However, there is doubt about BES's long-term effectiveness in microbial fuel cells (MFCs). We observed BES degradation in MFCs, whereas not in microbial electrolysis cells (MECs). Our results suggest that BES degradation is only possible under aerobic conditions (such as in MFCs) when some oxygen diffuses through the cathode. Experimental BES degradation was linked to the release of bromide (Br(-)) into the medium, with an average recovery of 67 ± 16%. Microbial analysis of the cathodic biomass distribution revealed the presence of Pseudomonas and Alcaligenes genera, which are able to use sulfonates as carbon or sulfur sources under aerobic conditions.

Hydrogen production in single chamber microbial electrolysis cells with different complex substrates.

The use of synthetic wastewater containing carbon sources of different complexity (glycerol, milk and starch) was evaluated in single chamber microbial electrolysis cell (MEC) for hydrogen production. The growth of an anodic syntrophic consortium between fermentative and anode respiring bacteria was operationally enhanced and increased the opportunities of these complex substrates to be treated with this technology. During inoculation, current intensities achieved in single chamber microbial fuel cells were 50, 62.5, and 9 A m⁻³ for glycerol, milk and starch respectively. Both current intensities and coulombic efficiencies were higher than other values reported in previous works. The simultaneous degradation of the three complex substrates favored power production and COD removal. After three months in MEC operation, hydrogen production was only sustained with milk as a single substrate and with the simultaneous degradation of the three substrates. The later had the best results in terms of current intensity (150 A m⁻³), hydrogen production (0.94 m³ m⁻³ d⁻¹) and cathodic gas recovery (91%) at an applied voltage of 0.8 V. Glycerol and starch as substrates in MEC could not avoid the complete proliferation of hydrogen scavengers, even under low hydrogen retention time conditions induced by continuous nitrogen sparging.

Operational aspects, pH transition and microbial shifts of a H2S desulfurizing biotrickling filter with random packing material.

Pall rings, a common random packing material, were used in the biotrickling filtration of biogas with high H2S. Assessment of 600d of operation covered the reactor start-up, the operation at neutral pH and the transition from neutral to acid pH. During the start-up period, operational parameters such as the aeration rate and the trickling liquid velocity were optimized. During the steady-state operation at neutral pH, the performance of the random packing material was investigated by reducing the gas contact time at both constant and increasing H2S loads. The random packing material showed similar elimination capacities and removal efficiencies in comparison with previous studies with a structured packing material, indicating that Pall rings are suitable for biogas desulfurization in biotrickling filters. The diversity of Eubacteria and the structure of the community were investigated before and after the pH transition using the bacterial tag-encoded FLX amplicon pyrosequencing. The pH transition to acid pH drastically reduced the microbial diversity and produced a progressive specialization of the sulfur-oxidizing bacteria community without any detrimental effect on the overall desulfurizing capacity of the reactor. During acidic pH operation, a persistent accumulation of elemental sulfur was found.