Biofilm Electrode Research Articles

Validation of effective roles of non-electroactive microbes on recalcitrant contaminant degradation in bioelectrochemical systems

Bioelectrochemical systems (BESs) have been widely investigated for recalcitrant waste treatment mainly because of their waste removal effectiveness. Electroactive microbes (EMs) have long been thought to contribute to the high effectiveness by interacting with electrodes via electron chains. However, this work demonstrated the dispensable role of EMs for enhanced recalcitrant contamination degradation in BESs. We revealed enhanced p-fluoronitrobenzene (p-FNB) degradation in a BES by observing a defluorination efficiency that was three times higher than that in biodegradation or electrochemical processes. Such an improvement was achieved by the collaborative roles of electrode biofilms and planktonic microbes, as their individual contributions to p-FNB degradation were found to be similarly stimulated by electricity. However, no bioelectrochemical activity was found in either the electrode biofilms or the planktonic microbes during stimulated p-FNB degradation; because no biocatalytically reductive or oxidative turnovers were observed on cyclic voltammetry curves. The non-involvement of EMs was further proven by the similar microbial community evolution for biofilms and planktonic microbes. In summary, we proposed a mechanism for indirect electrical stimulation of microbial metabolism by electrochemically generating the active mediator p-fluoroaniline (p-FA) and further degradation by a sequential combination of electrochemical p-FNB reduction and biological p-FA oxidation by non-EMs.

Enhanced denitrification by nano ɑ-Fe2O3 induced self-assembled hybrid biofilm on particle electrodes of three-dimensional biofilm electrode reactors

Three-dimensional biofilm electrode reactors (3D-BERs) represent a novel technology for wastewater denitrification. Formation of mature electroactive biofilm on particle electrodes is crucial to realize successful denitrification in 3D-BERs. However, long start-up time and low electroactivity of the biofilm formed on particle electrodes limit the further application of 3D-BERs in wastewater treatment. In this work, self-assembled hybrid biofilms (SAHB) was cultivated on granular activate carbon particle electrodes of the 3D-BER by assembling nano ɑ-Fe2O3 into the biofilm. ɑ-Fe2O3 was selected due to its high affinity to bacterial outer-membrane cytochromes, an important mediator for microbial electron transfer. SAHB formed on particle electrodes were characterized and the denitrification performance of 3D-BERs was also investigated. Results indicate that nano ɑ-Fe2O3 plays positive roles in the start-up of 3D-BER, which captures more microbes into SAHB and constructs thick biofilm on particle electrodes. Special microorganisms with denitrification function related with genera of Hydrogenophaga and Opitutus are distinctively enriched in SAHB. Nano ɑ-Fe2O3 induced SAHB exhibit superior denitrification performance compared to natural biofilm. The average denitrification rate increases from 0.62 mg total nitrogen/L/h for natural biofilm to 1.73 mg total nitrogen/L/h for SAHB, mainly ascribed to accelerated nitrites reduction. Our work provides new technical solution to enhance nitrates removal in 3D-BERs and brings deep insights into application of bio-electrochemical system in wastewater treatment.

Effect of continuous and intermittent electric current on lignin wastewater treatment and microbial community structure in electro-microbial system

In this study, complex structured soluble lignin wastewater was treated by electro-microbial system (EMS) using different direct current (DC) application modes (CR (continuous ON), IR12h (12 h-ON/12 h-OFF) and IR2h (2 h-ON/2 h-OFF)), and physiological characteristics and microbial communities were investigated. Results showed that CR, IR12h and IR2h had higher lignin removals, which were almost two times that of the control reactor (R0′, no current), and IR2h performed best and stably. Furthermore, IR2h exhibited the lowest ohmic resistance (Rs) of electrode biofilms, which could be explained by its higher abundance of electroactive bacteria. In the activated sludge of EMS, the concentration of dehydrogenase activity (DHA) and electronic transport system (ETS) in IR2h were the highest (1.48 and 1.28 times of R0′), which contributed to its high content of adenosine triphosphate (ATP). The viability of activated sludge was not affected by different DC application modes. Phospholipid fatty acids (PLFA) analysis indicated that IR2h had the maximum content of C15:1 anteiso A, C16:0 and C18:0; CR increased the content of C15:0 anteiso and decreased the content of saturated fatty acids. Genus-level results revealed that lignin-degrading bacteria, Pseudoxanthomonas and Mycobacterium, could be enriched in IR2h and CR, respectively.

Impact of hydraulic retention time and current on the microbial community and denitrification genes in a continuous‐flow biofilm electrode reactor

AbstractBACKGROUNDA biofilm‐electrode reactor (BER) was developed on the basis of hydrogenotrophic denitrification. The BER can be strengthened by adding organic matter, and the nitrate removal will be enhanced by a combination of heterotrophic and autotrophic denitrification. Denitrification in a biological system can be affected by operating parameters, and the microbial community and functional genes can be changed at the same time. In this study, the effect of hydraulic retention time (HRT) and current on the denitrification of a low chemical oxygen demand to nitrogen ratio (COD/N), saline wastewater in a continuous‐flow BER was evaluated and compared to a conventional biofilm reactor (BR).RESULTSThe highest extent of nitrate removal was obtained at the optimum HRT (8 h) and current (10 mA). Low levels of ammonia (0.5–4 mg L−1), produced through dissimilatory nitrate reduction to ammonia (DNRA), were observed. Quantitative real‐time polymerase chain reaction analysis revealed that the proportion of nirS‐type denitrifiers (3.75–8.30%) exceeded nirK‐type denitrifiers (0.24–0.90%) in both BR and BER. The genera of T78, Paracoccus and Azoarcus were identified as the dominant denitrifying bacteria in the BER. The cooperative activity of nitrate, nitrite and nitrous oxide reductase is necessary to reduce nitrate to dinitrogen in the denitrification process. The highest ratio of nosZ/nir genes in the bacterial community was 0.25 when the applied current was 10 mA.CONCLUSIONCompared to other denitrification reactors, the BER is more effective to treat low‐COD/N, saline wastewater with short HRT and low current. © 2018 Society of Chemical Industry

Azo dye as part of co-substrate in a biofilm electrode reactor–microbial fuel cell coupled system and an analysis of the relevant microorganisms

In general, refractory organics were hardly used as co-substrate in bioelectrochemical system. This study established a coupled bioelectrochemical system composed of a biofilm electrode reactor and a microbial fuel cell for using the azo dye X-3B as part of co-substrate. The two units degraded the azo dye X-3B stepwise while using it as part of co-substrate. Our results indicated that the removal efficiency of X-3B increased 28.5% using the coupled system compared with a control system. Moreover, the addition of the co-substrate glucose, which was necessary for MFC electricity generation, was reduced on the premise of stable removal efficiency in the coupled system to prevent resource waste due to using X-3B as part of co-substrate. The intermediate products of X-3B degradation were further explored using gas chromatography–mass spectrometry and a X-3B degradation pathway was proposed at the same time. Microbial communities were analyzed, illustrating that the mechanism of X-3B degradation was dependent on bioelectrochemistry rather than on microbial degradation.

Electrochemical Reduction of Carbon Dioxide to Value-Added Products: The Electrocatalyst and Microbial Electrosynthesis.

The electrochemical reduction of carbon dioxide (CO2 ) to value-added products obtains great attention and investigation worldwide in recent years. The commercialization of this green process relies on the progress of relating high-performance electrocatalysts and their feasibility with proper reactor design. The microbial electrosynthesis (MES) is an alternative route to reduce CO2 with electroactive bio-film electrode as catalyst. This review presents the research status and development of cathode catalysts, particularly focusing on the active sites and development tendency, for highly efficient electrochemical reduction CO2 from personal viewpoint. Some of our results are also presented to exhibit contributions. MES shows a similar process to the typical electrochemical reduction of CO2 . Their combination is an important trend, and the future research in this field is full of challenges and opportunities.

Synergistic effects of electricity and biofilm on Rhodamine B (RhB) degradation in three-dimensional biofilm electrode reactors (3D-BERs)

Three-dimensional biofilm electrode reactors (3D-BERs) represent an environmentally acceptable and cost effective technology for refractory wastewater treatment. Previous studies on 3D-BERs primarily focused on treatment performance, and little information is available about the microscopic mechanisms of contaminant degradation. The reactions occurred on particle electrodes, the core units of the 3D-BERs served as both electrodes and biofilm carriers, are still unclear. This study comprehensively elucidated the synergistic effects of electricity and biofilm on Rhodamine B (RhB) removal in 3D-BERs from both macroscopic and microcosmic aspects. Continuous-flow 3D-BERs were operated to evaluate overall treatment performance. Batch experiments were conducted to explore the kinetics of RhB degradation as well as the contributions of various physical, chemical, and biological processes to RhB removal. The biofilm formed on the particle electrodes was characterized by imaging and microbial analyses. The results indicated that applying voltage promoted degradation of RhB. Three processes, including electro-adsorption, electrochemical oxidation and electro-biodegradation, were identified to contribute to RhB degradation. Microorganisms in the Rhodanobacter and Thiomonas genera were distinctively enriched under acclimation voltage, which was attributed to the accumulation of intermediates generated by electrochemical oxidation. This study demonstrated that the synergistic effects of electricity and biofilm were dependent on applying voltage, that would be beneficial to comprehensively understand the contaminants removal process in 3D-BERs.

Parylene C-coated PDMS-based microfluidic microbial fuel cells with low oxygen permeability

Oxygen invasion is the main bottleneck in developing microscale microbial fuel cells as an efficient power source. This study reports for the first time the development of a polydimethylsiloxane -based co-laminar microbial fuel cell utilizing a parylene C coating to lower the oxygen permeability. In addition, the surface of the Au electrode is micropillar-structured to reduce the internal resistance of the microbial fuel cell. The performance of this novel microfluidic microbial fuel cell is investigated under various flow rates of electrolytes. The shear stress simulation shows that shear stress, induced by increasing flow rates, strongly impacts the biofilm electrode performance. To the best of our knowledge, the measured peak power density (71.89 ± 5.13 μW cm−2) and maximum current density (182.0 ± 4.82 μA cm−2) with the structured electrode are higher than those of any other reported polydimethylsiloxane-based microscale microbial fuel cells. The proposed microbial fuel cell appears to be a promising power supply that can be easily integrated with portable or implantable biomedical devices.

Azo dye degradation pathway and bacterial community structure in biofilm electrode reactors

In this study, the degradation pathway of the azo dye X-3B was explored in biofilm electrode reactors (BERs). The X-3B and chemical oxygen demand (COD) removal efficiencies were evaluated under different voltages, salinities, and temperatures. The removal efficiencies increased with increasing voltage. Additionally, the BER achieved maximum X-3B removal efficiencies of 66.26% and 75.27% at a NaCl concentration of 0.33 g L−1 and temperature of 32 °C, respectively; it achieved a COD removal efficiency of 75.64% at a NaCl concentration of 0.330 g L−1. Fourier transform infrared spectrometry and gas chromatography–mass spectrometry analysis indicated that the X-3B biodegradation process first involved the interruption of the conjugated double-bond, resulting in aniline, benzodiazepine substance, triazine, and naphthalene ring formation. These compounds were further degraded into lower-molecular-weight products. From this, the degradation pathway of the azo dye X-3B was proposed in BERs. The relative abundances of the microbial community at the phylum and genus levels were affected by temperature, the presence of electrons, and an anaerobic environment in the BERs. To achieve better removal efficiencies, further studies on the functions of the microorganisms are needed.

A continuous flow MFC-CW coupled with a biofilm electrode reactor to simultaneously attenuate sulfamethoxazole and its corresponding resistance genes

A continuous flow microbial fuel cell constructed wetland (MFC-CW) coupled with a biofilm electrode reactor (BER) system was constructed to remove sulfamethoxazole (SMX). The BER unit powered by the stacked MFC-CWs was used as a pretreatment unit, and effluent flowed into the MFC-CW for further degradation. The experimental results indicated that the removal rate of 2 or 4 mg/L SMX in a BER unit was nearly 90%, and the total removal rate in the coupled system was over 99%. As the hydraulic retention time (HRT) was reduced from 16 h to 4 h, the SMX removal rate in the BER decreased from 75% to 48%. However, the total removal rate in the coupled system was still over 97%. The maximum SMX removal rate in the MFC-CW, which accounted for 42%–55% of the total removal, was obtained in the anode layer. In addition, the relative abundances of sul genes detected in the systems were in the order of sulI > sulII > sulIII, and significant positive correlations of sul gene copy numbers versus SMX concentration and 16S rRNA gene copy numbers were observed. Furthermore, significant negative correlations were identified between sul genes, 16S rRNA gene copy numbers, and HRT. The abundances of the sul genes in the effluent of the MFC-CW were lower than the abundances observed in the BER effluent. High-throughput sequencing revealed that the microbial community diversity of the BER was affected by running time, power supply forms and HRT. Bio-electricity from the MFC-CW may reduce microbial community diversity and contribute to reduction of the antibiotic resistance gene (ARG) abundance in the BER. Taken together, the BER-MFC-CW coupled system is a potential tool to treat wastewater containing SMX and attenuate corresponding ARG abundance.

Response of the microbial community structure of biofilms to ferric iron in microbial fuel cells

Ferric iron can affect the current generation of microbial electrochemical system (MES); however, how it influences microbial biofilm formation and metabolic activity has not been reported. Here, we describe the response of microbial electrode biofilm communities to insoluble ferric iron (Fe3+) at different concentrations in microbial fuel cells (MFCs). Insoluble ferric iron (200μM) improved electrochemical activity of the MFCs microbial biofilms during start-up and resulted in a higher maximum power density of 0.95W/m2, compared with the control (0.76W/m2), 500μM Fe3+ (0.83W/m2), 1000μM Fe3+ (0.73W/m2), and 2000μM Fe3+ (0.59W/m2) treatments. Illumina Hiseq sequencing of 16S rRNA gene amplicons indicated that the predominant populations in the anode biofilms of the MFCs belonged to Geobacter, with relative abundance of 66–75%. Microbial cathode biofilm communities were more susceptible to Fe3+, as an obvious shift in the cathode biofilm community structures occurred as Fe3+ concentration was increased. The most predominant populations in the MFC cathode biofilms without Fe3+ and with 200μM Fe3+ were affiliated with Thauera (46% and 35%), whereas no absolutely predominant populations were present in the MFC cathode biofilm with 1000μM Fe3+. The results demonstrate that a low concentration of Fe3+ facilitated the power output of MFCs and shaped community structures of the electrode biofilm.

Effects of surface charge, hydrophilicity and hydrophobicity on functional biocathode catalytic efficiency and community structure

The bioelectrotransformation efficiency of various organic matters and corresponding electrode biofilm community formation as well as electron transfer efficiency in bioelectrochemical systems (BESs) with different modified electrodes has been extensively studied on the anode side. However, the effects of cathode interface characteristics towards the BESs bioelectrotransformation performance remain poorly understood. In this study, the nitrobenzene-reducing biocathode catalytic efficiency and community structure in response to different modified electrodes (control: hydrophobic and no charge; –SH: hydrophobic and single negative charge; –NH2: hydrophilic and single positive charge –NH–NH2: hydrophilic and double positive charges) were investigated. The biocathode transformation efficiency of nitrobenzene (NB) to aniline (AN) (ENB-AN) was affected by the nature of electrode interface as well as the biocathode community formation and structure. Cathodes with hydrophilic surface and positive charges have performed well in the bioelectrotransformation experiments, and especially made an outstanding performance when inorganic NaHCO3 was supplied as carbon source and cathode as the sole electron donor. Importantly, the hydrophilic surfaces with positive charges were dominated by the electroactive nitroaromatic reducers (Enterococcus, Desulfovibrio and Klebsiella) with the relative abundance as high as 72.20 ± 1.87% and 74.86 ± 8.71% for –NH2 and –NH–NH2 groups respectively. This could explain the higher ENB-AN in the hydrophilic groups than that of the hydrophobic –SH modified group. This study provides new insights into the effects of electrode interface characteristics on the BESs biocathode performance and offers some suggestions for the future design for the improvement of bioelectroremediation performance.

McrA sequencing reveals the role of basophilic methanogens in a cathodic methanogenic community

Cathodic methanogenesis is a promising method for accelerating and stabilising bioenergy recovery in anaerobic processes. The change in composition of microbial (especially methanogenic) communities in response to an applied potential—and especially the associated pH gradient—is critical for achieving this goal, but is not well understood in cathodic biofilms. We found here that the pH-polarised region in the 2 mm surrounding the cathode ranged from 6.9 to 10.1, as determined using a pH microsensor; this substantially affected methane production rate as well as microbial community structure. Miseq sequencing data of a highly conserved region of the mcrA gene revealed a dramatic variation in alpha diversity of methanogens concentrated in electrode biofilms under the applied potential, and confirmed that the dominant microbes at the cathode were hydrogenotrophic methanogens (mostly basophilic Methanobacterium alcaliphilum). These results indicate that regional pH variation in the microenvironment surrounding the electrode is an ecological niche enriched with Methanobacterium.

A system composed of a biofilm electrode reactor and a microbial fuel cell-constructed wetland exhibited efficient sulfamethoxazole removal but induced sul genes

The aim of this work was to study sulfamethoxazole (SMX) removal efficiency and fate of corresponding sul genes in a stacked microbial fuel cell-constructed wetland coupled biofilm electrode reactor system (MFC-CW-BER). Findings showed that two stacked MFC-CWs could provide a relatively stable electricity supply to support the biofilm for SMX removal. Excellent SMX removal (>99.29%) was obtained in the BER-MFC-CW. Compared with the 2000 µg L−1 SMX influent, the relative abundance of the sul genes in biofilm media and effluent was enhanced with continuously high concentrations of SMX (4000 μg L−1). The relative abundances of sul genes in biofilm media and effluent increased as the hydraulic retention time decreased. However, there was no obvious variation in the relative abundance of sul genes in the effluent from MFC-CWs. No effect could be observe of the direct voltage and bioelectricity on the relative abundance of the sul genes in the BER.

Electrochemical study of enhanced nitrate removal in wastewater treatment using biofilm electrode

Bioelectrochemical enhanced nitrate removal in wastewater with high total nitrogen and low organic carbon was electrochemically investigated focusing on the relationship between biochemical and electrochemical nitrogen cycles. Under optimized external voltage of −0.6 V, apparent nitrate removal rate of bioelectrochemical denitrification was 76% higher than normal biofilm denitrification. And with the introduction of biofilm on the electrode, new reduction peak of N2O, much larger current density, and 0.4 V positively shift of on-set potential of nitrate reduction reaction were observed, suggesting a synergy of electrochemical reaction and biological reaction through enhanced electrochemical reduction of intermediate products from biological process. Oxygen reduction reaction could not be avoided during nitrogen electrochemical reduction reaction since their similar reduction potential. But it led to decrease of oxygen concentration and therefore contribute to biological denitrification. Bacteria community tests also supported a dominant bacteria which could denitrify and use external electron.

A 3DBER-S-EC process for simultaneous nitrogen and phosphorus removal from wastewater with low organic carbon content

A new process was proposed by integrating a three-dimensional biofilm electrode reactor with sulfur autotrophic denitrification and electrocoagulation within the same reactor. The results indicated that under the wastewater influent condition of NO3−-N = 30 mg/L, COD = 45 mg/L, total phosphorus (TP) = 1.5 mg/L, hydraulic retention time (HRT) = 8 h, and I = 400 mA, the NO3−-N and TP removal of the proposed process reached 89.8% and 83.0%, respectively. It was observed that the electrocoagulation process improved phosphorus removal, while the simultaneous existence of heterotrophic, hydrogen, sulfur and iron autotrophic denitrifying bacteria led to enhanced and stabilized nitrogen removal. The Sulfuritalea hydrogenivorans sk43H and Sulfuricella denitrificans skB26 were found as the dominant denitrifying bacteria in the electrocoagulation section and the section of biofilm electrode with sulfur filler, respectively. As compared to conventional technologies, the proposed new process can achieve simultaneous, stable and deep nitrogen and phosphorus removal from wastewater treatment plant effluent with low organic carbon content.

Effects of salinity and COD/N on denitrification and bacterial community in dicyclic-type electrode based biofilm reactor

A dicyclic-type electrode based biofilm electrode reactor (BER) was developed for advanced nitrate removal from saline municipal wastewater. The denitrification efficiency was evaluated with a synthetic feed (NO3−-N, 20 mg L−1) under different salinity and COD to nitrogen ratios (COD/N). As the salinity increased from 0% to 1.0%, the denitrification performance of both the traditional biofilm reactor (BR) and BER was inhibited; however, the BER showed better adaptation and ability to recover. The BER achieved a high nitrate removal efficiency (≥90%) at a salinity of 1.0% and a low COD/N of 2.5 (theoretical stoichiometric 2.86 ignoring microbial growth). The abundance of Methylotenera mobilis in BR and Clostridium sticklandii in BER was higher than in the initial sludge sample used as inoculum. Likewise, the abundance of napA, nirS and nosZ genes increased as the COD/N further decreased. Under high salinity stress, the BER had a higher denitrification efficiency and the consumption of the organic carbon source (i.e., methanol) was reduced compared to BR. The cooperation between heterotrophic and autotrophic denitrifiers in the BER system provides a more efficient and feasible solution for nitrate removal from saline municipal wastewater.

Anoxic biodegradation of triclosan and the removal of its antimicrobial effect in microbial fuel cells

Triclosan (TCS) is an emerging organic contaminant in the environment. Here, the anoxic bio-degradation of TCS in microbial fuel cells (MFCs) was explored. It was found that anoxic biodegradation of TCS could be achieved in MFC, and the removal rate of TCS was accelerated after reactor acclimation. After 7 months of operation, 10mg/L TCS could be removed within 8days in MFCs. Fluorescence microscopy results revealed that the microbe cells in the reactors were intact, and the microbes were in active state. Flow cytometry test showed that the proliferation of inoculated microbe was higher in MFC effluent than that in TCS solution. These data indicate that the biotoxicity of TCS has been largely eliminated after the treatment. The microbial community shift during the TCS degradation process was investigated as well. Species such as Geothrix, Corynebacterium, Sulfobacillus, GOUTA19, Geobacter, Acidithiobacillus and Acinetobacter, which were capable for the degradation of benzene-related and dechlorination of chlorine-containing chemicals, were flourished in the electrode biofilm. They may participate in the biodegradation of TCS. This work provides a new perspective for the anoxic biodegradation of recalcitrant organics, and can be useful for the in-situ bioremediation of environmental pollutants with the removal of their biotoxicity.

Energy recovery evaluation in an up flow microbial electrolysis coupled anaerobic digestion (ME-AD) reactor: Role of electrode positions and hydraulic retention times

Microbial catalysed electrochemical systems are intensively used in basic and applied research as a sustainable platform for harnessing energy and generating value added bio-products. Recently more emphasis is being laid on enhancement of AD (Anaerobic Digestion) by integrating ME (Microbial Electrolysis) with it to convert CO2 (Carbon dioxide) directly to CH4 (Methane). This research attempts to shed light on the effects of electrode positioning and arrangement along with hydraulic retention time (HRT), on CH4 generation, and organic removal in novel microbial electrolysis coupled anaerobic digestion (ME-AD) reactors. However, the positioning and placement of electrodes in a ME-AD reactor have not yet been evaluated. Four reactors (S1, S2, S3 and S4) are designed with different electrode arrangements and run in four different HRTs (12, 18, 24 and 36 h) with beer brewery wastewater as the substrate for treatment. The reactors with electrodes arranged at the bottom are better in performance than the reactors with electrodes placed at the top. They have maximum COD, TOC and Carbohydrate removal efficiencies of 92.1%, 64.2% and 98.9% respectively, high methane production rate (MPR) and methane yield (MY) with 304.5 mLCH4/Lreactor/day and 275.8 mL/gCOD respectively and a maximum current generation of 10 mA, all at 36 h HRT. Electrode placement also has crucial roles to play in microbial community prevalence on the electrode biofilm of the reactors. Methanogens and electrogens are well enriched on the electrode biofilms of the reactors with bottom positioned electrodes, revealing the basis behind their maximum organic removal, methane production efficiencies and current generation. This study demonstrates that the optimization of appropriate electrode position and placement in ME-AD reactors is crucial for their performance and development.

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...