Modelling anaerobic sulfide removal by sulfide shuttling bacteria.

Sulfide oxidizing bacteria are used in industrial biodesulfurization processes to convert sulfide to sulfur. These bacteria can spatially separate sulfide removal from terminal electron transfer, thereby acting as sulfide shuttles. The mechanisms underlying sulfide shuttling are not yet clear. In this work, newly obtained sulfide removal data were used to develop a new model for anaerobic sulfide removal and this model was shown to be an improvement over two previously published models. The new model describes a fast chemical step and a consecutive slow enzymatic step. The improved model includes the effect of pH, with higher total sulfide removal at increasing pH, as well as partial sulfide removal at higher sulfide concentrations. The two-stage model is supported by recent developments in anaerobic sulfide removal research and contributes to a better understanding of the underlying mechanisms. The model is a step toward accurately modelling anaerobic sulfide removal in industrial systems.

Oxygen-to-ammonium-nitrogen ratio as an indicator for oxygen supply management in microoxic bioanodic ammonium oxidation.

Microbial electrolysis cells (MECs) have been proven effective for oxidizing ammonium (NH4+), where the anode acts as an electron acceptor, reducing the energy input by substituting oxygen (O2). However, O2 has been proved to be essential for achieving high removal rates MECs. Thus, precise control of oxygen supply is crucial for optimizing treatment performance and minimizing energy consumption. Unlike previous studies focusing on dissolved oxygen (DO) levels, this study introduces the O2/NH4+-N ratio as a novel control parameter for balancing oxidation rates and the selectivity of NH4+ oxidation towards dinitrogen gas (N2) under limited oxygen condition. Our results demonstrated that the O2/NH4+-N ratio is a more relevant oxygen supply indicator compared to DO level. Oxygen served as a more favorable electron acceptor than the electrode, increasing NH4+ oxidation rates but also resulting in more oxidized products such as nitrate (NO3-). Additionally, nitrous oxide (N2O) and N2 production were higher with the electrode as the electron acceptor compared to oxygen alone. An O2/NH4+-N ratio of 0.5 was found to be optimal, achieving a balance between product selectivity for N2 (51.4 % ± 4.5 %) and oxidation rates (344.6 ± 14.7 mg-N/L*d), with the columbic efficiency of 30.7 % ± 2.0 %. Microbial community analysis revealed that nitrifiers and denitrifiers were the primary bacteria involved, with oxygen promoting the growth of nitrite-oxidizing bacteria, thus facilitating complete NH4+ oxidation to NO3-. Our study provides new insights and guidelines on the appropriate oxygen dosage, offering strategies into optimizing operational conditions for NH4+ removal using MECs.

Unravelling the mechanisms of organic micropollutant removal in bio-electrochemical systems: Insights into sorption, electrochemical degradation, and biodegradation processes.

Bio-electrochemical systems (BESs) have recently been proposed as an efficient treatment technology to remove organic micropollutants from water treatment plants. In this study, we aimed to differentiate between sorption, electrochemical transport/degradation, and biodegradation. Using electro-active microorganisms and electrodes, we investigated organic micropollutant removal at environmentally relevant concentrations, clarifying the roles of sorption and electrochemical and biological degradation. The role of anodic biofilms on the removal of 10 relevant organic micropollutants was studied by performing separate sorption experiments on carbon-based electrodes (graphite felt, graphite rod, graphite granules, and granular activated carbon) and electrochemical degradation experiments at two different electrode potentials (-0.3 and 0 V). Granular activated carbon showed the highest sorption of micropollutants; applying a potential to graphite felt electrodes increased organic micropollutant removal. Removal efficiencies >80 % were obtained for all micropollutants at high anode potentials (+0.955 V), indicating that the studied compounds were more susceptible to oxidation than to reduction. All organic micropollutants showed removal when under bio-electrochemical conditions, ranging from low (e.g. metformin, 9.3 %) to exceptionally high removal efficiencies (e.g. sulfamethoxazole, 99.5 %). The lower removal observed under bio-electrochemical conditions when compared to only electrochemical conditions indicated that sorption to the electrode is key to guarantee high electrochemical degradation. The detection of transformation products of chloridazon and metformin indicated that (bio)-electrochemical degradation occurred. This study confirms that BES can treat some organic micropollutants through several mechanisms, which merits further investigation.

Energy Efficient Carbon Capture through Electrochemical pH Swing Regeneration of Amine Solution.

Carbon capture is widely acknowledged as a promising strategy for achieving negative emissions. Electrochemical carbon capture technologies are considered a viable alternative to conventional temperature swing processes. Among these, employing the hydrogen oxidation and hydrogen evolution reactions as a redox couple, along with an ion exchange membrane, offers an effective means of establishing a pH swing for desorbing CO2 and regenerating the alkaline solvent. However, the practical scalability of this approach is impeded by challenges such as high energy demands resulting from a high pH differential between anodic and cathodic environments and operation with solutions with a low conductivity, required to obtain an acceptable current yield. To address these limitations, this study introduces an innovative anion exchange membrane (AEM)-based electrochemical process for solvent regeneration. Our research demonstrates the advantageous utilization of amines as chemical buffers. Selecting an amine solution with a favorable pKa (∼7 to 10) helps in maintaining bicarbonate as the predominant carbon species within the system, thereby ensuring a high current yield (>80%) across various operational conditions (current, load ratio, and solution concentration). Furthermore, our analysis indicates that the use of amine solutions effectively reduces the overpotential of the hydrogen evolution reaction due to a lower local pH. This results in a minimum energy requirement of 63 kJ/mol at a current density of 20 A/m2 to regenerate the solution (MDEA) while maintaining high (>99%) product (CO2) purity.

Revealing cellular (poly)sulphide storage in electrochemically active sulphide oxidising bacteria using rotating disc electrodes.

Sulphide oxidising bacteria (SOB) have the potential to be used for bioelectrochemical removal, i.e. oxidation, of sulphide from waste streams. In anaerobic conditions, SOB are able to spatially separate sulphide removal and terminal electron transfer to an electrode and act as a sulphide shuttle. However, it is not fully understood how SOB anaerobically remove sulphide and store charge equivalents, and where in this process sulphur is formed. Therefore, the redox behaviour of sulphide shuttling SOB was investigated at haloalkaline conditions using a glassy carbon rotating disc electrode (RDE) and cyclic voltammetry. Voltammograms of SOB in the absence and presence of sulphide were compared to voltammograms of abiotic sulphur species solutions. Polysulphide and sulphide showed different redox behaviour, with distinct potentials for oxidation of > -0.3 V (vs. Ag/AgCl) for polysulphide and > -0.1 V for sulphide. Comparing biotic to abiotic experiments lead to the hypothesis that SOB formed polysulphides during anaerobic sulphide removal, which stayed sorbed to the cells. With this study, further steps were taken in elucidating the mechanisms of sulphide shuttling by SOB.

Mechanisms of extracellular electron transfer in anaerobic methanotrophic archaea.

Anaerobic methanotrophic (ANME) archaea are environmentally important, uncultivated microorganisms that oxidize the potent greenhouse gas methane. During methane oxidation, ANME archaea engage in extracellular electron transfer (EET) with other microbes, metal oxides, and electrodes through unclear mechanisms. Here, we cultivate ANME-2d archaea ('Ca. Methanoperedens') in bioelectrochemical systems and observe strong methane-dependent current (91-93% of total current) associated with high enrichment of 'Ca. Methanoperedens' on the anode (up to 82% of the community), as determined by metagenomics and transmission electron microscopy. Electrochemical and metatranscriptomic analyses suggest that the EET mechanism is similar at various electrode potentials, with the possible involvement of an uncharacterized short-range electron transport protein complex and OmcZ nanowires.

Enhancement of Ammonium Oxidation at Microoxic Bioanodes.

Bioelectrochemical systems (BESs) are considered to be energy-efficient to convert ammonium, which is present in wastewater. The application of BESs as a technology to treat wastewater on an industrial scale is hindered by the slow removal rate and lack of understanding of the underlying ammonium conversion pathways. This study shows ammonium oxidation rates up to 228 ± 0.4 g-N m-3 d-1 under microoxic conditions (dissolved oxygen at 0.02-0.2 mg-O2/L), which is a significant improvement compared to anoxic conditions (120 ± 21 g-N m-3 d-1). We found that this enhancement was related to the formation of hydroxylamine (NH2OH), which is rate limiting in ammonium oxidation by ammonia-oxidizing microorganisms. NH2OH was intermediate in both the absence and presence of oxygen. The dominant end-product of ammonium oxidation was dinitrogen gas, with about 75% conversion efficiency in the presence of a microoxic level of dissolved oxygen and 100% conversion efficiency in the absence of oxygen. This work elucidates the dominant pathways under microoxic and anoxic conditions which is a step toward the application of BESs for ammonium removal in wastewater treatment.

Comparing theoretical and practical biomass yields calls for revisiting thermodynamic growth models for electroactive microorganisms.

Research on electroactive microorganisms (EAM) often focuses either on their physiology and the underlying mechanisms of extracellular electron transfer or on their application in microbial electrochemical technologies (MET). Thermodynamic understanding of energy conversions related to growth and activity of EAM has received only a little attention. In this study, we aimed to prove the hypothesized restricted energy harvest of EAM by determining biomass yields by monitoring growth of acetate-fed biofilms presumably enriched in Geobacter, using optical coherence tomography, at three anode potentials and four acetate concentrations. Experiments were concurrently simulated using a refined thermodynamic model for EAM. Neither clear correlations were observed between biomass yield and anode potential nor acetate concentration, albeit the statistical significances are limited, mainly due to the observed experimental variances. The experimental biomass yield based on acetate consumption (YX/ac = 37 ± 9 mgCODbiomass gCODac-1) was higher than estimated by modeling, indicating limitations of existing growth models to predict yields of EAM. In contrast, the modeled biomass yield based on catabolic energy harvest was higher than the biomass yield from experimental data (YX/cat = 25.9 ± 6.8 mgCODbiomass kJ-1), supporting restricted energy harvest of EAM and indicating a role of not considered energy sinks. This calls for an adjusted growth model for EAM, including, e.g., the microbial electrochemical Peltier heat to improve the understanding and modeling of their energy metabolism. Furthermore, the reported biomass yields are important parameters to design strategies for influencing the interactions between EAM and other microorganisms and allowing more realistic feasibility assessments of MET.

Starvation combined with constant anode potential triggers intracellular electron storage in electro-active biofilms.

The accumulation of electrons in the form of Extracellular Polymeric Substances (EPS) and poly-hydroxyalkanoates (PHA) has been studied in anaerobic processes by adjusting the access of microorganisms to the electron donor and final electron acceptor. In Bio-electrochemical systems (BESs), intermittent anode potential regimes have also recently been used to study electron storage in anodic electro-active biofilms (EABfs), but the effect of electron donor feeding mode on electron storage has not been explored. Therefore, in this study, the accumulation of electrons in the form of EPS and PHA was studied as a function of the operating conditions. EABfs were grown under both constant and intermittent anode potential regimes and fed with acetate (electron donor) continuously or in batch. Confocal Laser Scanning Microscopy (CLSM) and Fourier-Transform Infrared Spectroscopy (FTIR) were used to assess electron storage. The range of Coulombic efficiencies, from 25 to 82%, and the biomass yields, between 10 and 20%, indicate that storage could have been an alternative electron consuming process. From image processing, a 0.92 pixel ratio of poly-hydroxybutyrate (PHB) and amount of cells was found in the batch fed EABf grown under a constant anode potential. This storage was linked to the presence of living Geobacter and shows that energy gain and carbon source starvation were the triggers for intracellular electron storage. The highest EPS content (extracellular storage) was observed in the continuously fed EABf under an intermittent anode potential, showing that constant access to electron donor and intermittent access to the electron acceptor leads to the formation of EPS from the excess energy gained. Tailoring operating conditions can thus steer the microbial community and result in a trained EABf to perform a desired biological conversion, which can be beneficial for a more efficient and optimized BES.

Effect of organic amendments obtained from different pretreatment technologies on soil microbial community.

The application of organic amendments (OAs) obtained from biological treatment technologies is a common agricultural practice to increase soil functionality and fertility. OAs and their respective pretreatment processes have been extensively studied. However, comparing the properties of OAs obtained from different pretreatment processes remains challenging. In most cases, the organic residues used to produce OAs exhibit intrinsic variability and differ in origin and composition. In addition, few studies have focused on comparing OAs from different pretreatment processes in the soil microbiome, and the extent to which OAs affect the soil microbial community remains unclear. This limits the design and implementation of effective pretreatments aimed at reusing organic residues and facilitating sustainable agricultural practices. In this study, we used the same model residues to produce OAs to enable meaningful comparisons among compost, digestate, and ferment. These three OAs contained different microbial communities. Compost had higher bacterial but lower fungal alpha diversity than ferment and digestate. Compost-associated microbes were more prevalent in the soil than ferment- and digestate-associated microbes. More than 80% of the bacterial ASVs and fungal OTUs from the compost were detected 3 months after incorporation into the soil. However, the addition of compost had less influence on the resulting soil microbial biomass and community composition than the addition of ferment or digestate. Specific native soil microbes, members from Chloroflexi, Acidobacteria, and Mortierellomycota, were absent after ferment and digestate application. The addition of OAs increased the soil pH, particularly in the compost-amended soil, whereas the addition of digestate enhanced the concentrations of dissolved organic carbon (DOC) and available nutrients (such as ammonium and potassium). These physicochemical variables were key factors that influenced soil microbial communities. This study furthers our understanding of the effective recycling of organic resources for the development of sustainable soils.

Nutrient recovery and pollutant removal during renewable fuel production: opportunities and challenges.

Stimulated by the desire to achieve a Net Zero energy economy, the demand for renewable fuels is growing rapidly. The production of toxic waste streams that accompanies the transition from fossil fuels to renewable fuels is often overlooked. These waste streams include, among others, thiols and ammonia, and benzene, toluene, and xylene (BTX). When suitable treatment technologies are available, these compounds can be converted to valuable nutrients. In this opinion article, we provide an overview of expected waste streams and their characteristics. We indicate future challenges for associated waste streams, such as the lag in developing resource recovery technologies. Furthermore, we discuss unexploited opportunities to recover valuable nutrients from these waste streams.

Anaerobic sulphide removal by haloalkaline sulphide oxidising bacteria.

Sulphide is a toxic and corrosive compound and requires removal from waste streams. Recent discoveries show that sulphide oxidising bacteria (SOB) from modern desulphurisation plants are able to spatially separate sulphide removal and oxygen reduction when exposed to intermittent anaerobic and aerobic environments. Here, SOB act as electron shuttles between electron donor and acceptor. The underlying mechanisms for electron shuttling are of yet unknown. To investigate the anaerobic sulphide removal of SOB, batch experiments and mathematical models were applied. The sulphide removal capacity decreased at increasing biomass concentrations. At 0.6 mgN/L SOB could remove up to 8 mgS/mgN in 30 min. It was found that biological activity determines sulphide removal, alongside chemical processes. Anaerobic oxidation of electron carriers was determined to only explain 0.1% of charge storage, where irreversible cleavage of long chain polysulphides could explain full sulphide storage. Different sulphide removal and intracellular storage processes are postulated.

Maximum thickness of non-buffer limited electro-active biofilms decreases at higher anode potentials.

The accumulation of protons in electro-active biofilms (EABfs) has been reported as a critical parameter determining produced currents at the anode since the very beginning of the studies on Bio-electrochemical systems (BESs). Even though the knowledge gained on the influence of this parameter on the produced currents, its influence on EABfs growth is frequently overlooked. In this study, we quantified EABfs thicknesses in real-time and related them to the produced current at three buffer concentrations, two anode potentials and two acetate concentrations. The thickest EABfs (80 µm) and higher produced currents (2.5 A.m-2) were measured when a 50 mM buffer concentration was used. By combining the measured EABfs thicknesses with the pH in the anolyte, a simple model was developed to identify buffer limitations. Buffer limited EABfs with thicknesses of 15 and 42 µm were identified at -0.3 V vs Ag/AgCl when 10 and 50 mM buffer concentrations were used, respectively. At -0.2 V vs Ag/AgCl, the thicknesses of buffer limited EABfs decreased to 13 and 20 µm, respectively. The model also estimated buffer and acetate diffusion rates in EABfs and allowed to determine the boundary between a buffer and acetate limited EABfs. The diffusion rates reported in this study and the definition of the boundary between buffer and acetate limited EABfs provide a powerful tool to avoid limitations, leading to higher produced currents at the anode.

The effect of anode potential on electrogenesis, methanogenesis and sulfidogenesis in a simulated sewer condition.

Methane emissions from the sewer system are considered to be a non-negligible source of aggravating the greenhouse effect. Meanwhile, the sewer system has long been plagued by sulfide-induced corrosion problems. This study explored the possibility of using a bioelectrochemical system to intensify the competition between electroactive bacteria, methanogens and sulfate-reducing bacteria, thereby reducing the production of methane and sulfide. Dual-chamber bioelectrochemical reactors were constructed and operated in fed-batch mode with the coexistence of Electroactive bacteria, Methanogenic archaea and Sulfate-reducing bacteria. Acetate was supplied as the sole carbon source. The results indicated that electrogenesis induced by the anode potentials of -0.42 V and -0.2 V (vs. Ag/AgCl) had advantages over methanogenesis and sulfidogenesis in consuming acetate. The stimulated electrogenesis by anode potentials resulted in a decrease in pH. Methane production was suppressed in the reactors with anode potentials of -0.42 and -0.2 V compared to open circuit controls. In contrast to methane, the capacity for sulfide production was facilitated in the reactors with the anode potentials of -0.42 V and -0.2 V compared to open circuit controls. 16s rRNA gene analysis showed that Geobacter was the most abundant genus on the anode biofilm in the anode potential-controlled reactor, while acetoclastic methanogens dominated in open circuit controls. Methanosaeta and Methanosarcina were the most abundant methanogens in open circuit controls. Collectively, our study demonstrates that the use of electrodes with anode potential control can help to control methane emissions, but could not yet prevent sulfide production, which requires further research.

A simple method for routine measurement of organosulfur compounds in complex liquid and gaseous matrices.

The measurement of VOSCs in complex matrices is challenging due to their volatile and reactive nature. A straightforward method using headspace chromatography was developed for routine analyses of organosulfur compounds in a high saline liquid matrix with a pH of 8.4. Direct sample acidification with a 1M acetate buffer (pH 3.6) showed an increased response for methanethiol, ethanethiol, propanethiol, dimethyl sulfide, dimethyl disulfide and diethyl disulfide. A good quadratic fit (R2 <0.995) was obtained for each compound over a calibration range of 5 µM-S until 125 µM-S (µmol sulfur/L). Gas standards were measured using the same chromatographic conditions over a calibration range of 0.08 µM-S until 1.85 µM-S (R2 <0.999). Gas standards could also be used to calibrate the liquid phase with a response ratio of 105.2% for ET, 107% for DMS, 105.7% for PT, 108.9% for DMDS and 106% for DEDS. This alternative calibration strategy reduced the preparation time and does not rely on liquid standards, which were unstable over time. This method was used to determine Henry constants for the organosulfur compounds both in demineralized water and the high saline liquid matrix and to analyze samples from a bio electrochemical experiment that treated methanethiol. This new method allows for routine analysis of samples originating from natural gas desulfurization plants and can potentially also be used to analyze organosulfur compounds in other complex waste streams.

Opportunities for visual techniques to determine characteristics and limitations of electro-active biofilms.

Optimization of bio-electrochemical systems (BESs) relies on a better understanding of electro-active biofilms (EABfs). These microbial communities are studied with a range of techniques, including electrochemical, visual and chemical techniques. Even though each of these techniques provides very valuable and wide-ranging information about EABfs, such as performance, morphology and biofilm composition, they are often destructive. Therefore, the information obtained from EABfs development and characterization studies are limited to a single characterization of EABfs and often limited to one time point that determines the end of the experiment. Despite being scarcer and not as commonly reported as destructive techniques, non-destructive visual techniques can be used to supplement EABfs characterization by adding in-situ information of EABfs functioning and its development throughout time. This opens the door to EABfs monitoring studies that can complement the information obtained with destructive techniques. In this review, we provide an overview of visual techniques and discuss the opportunities for combination with the established electrochemical techniques to study EABfs. By providing an overview of suitable visual techniques and discussing practical examples of combination of visual with electrochemical methods, this review aims at serving as a source of inspiration for future studies in the field of BESs.

Methane-Dependent Extracellular Electron Transfer at the Bioanode by the Anaerobic Archaeal Methanotroph "Candidatus Methanoperedens".

Anaerobic methanotrophic (ANME) archaea have recently been reported to be capable of using insoluble extracellular electron acceptors via extracellular electron transfer (EET). In this study, we investigated EET by a microbial community dominated by "Candidatus Methanoperedens" archaea at the anode of a bioelectrochemical system (BES) poised at 0 V vs. standard hydrogen electrode (SHE), in this way measuring current as a direct proxy of EET by this community. After inoculation of the BES, the maximum current density was 274 mA m-2 (stable current up to 39 mA m-2). Concomitant conversion of 13CH4 into 13CO2 demonstrated that current production was methane-dependent, with 38% of the current attributed directly to methane supply. Based on the current production and methane uptake in a closed system, the Coulombic efficiency was about 17%. Polarization curves demonstrated that the current was limited by microbial activity at potentials above 0 V. The metatranscriptome of the inoculum was mined for the expression of c-type cytochromes potentially used for EET, which led to the identification of several multiheme c-type cytochrome-encoding genes among the most abundant transcripts in "Ca. Methanoperedens." Our study provides strong indications of EET in ANME archaea and describes a system in which ANME-mediated EET can be investigated under laboratory conditions, which provides new research opportunities for mechanistic studies and possibly the generation of axenic ANME cultures.