Hydrogenotrophic Methanogenic Pathway Research Articles

Enhancing effect of conductive materials and rumen microorganisms on the anaerobic digestion performance of a real traditional Chinese medicine wastewater

This work evaluated the effect of different concentrations of conductive materials and rumen microorganisms on the anaerobic digestion (AD) performance of real traditional Chinese medicine (TCM) wastewater. The experiments first determined the optimal organic concentration of 0.25 L/L for the AD system and removed suspended solids from real TCM wastewater to reduce the inhibition effect on methanogenesis. Analysis of the modified Gompertz model showed that the addition of activated carbon, biochar, cow manure, and rumen fluid at doses of 12 g/L, 12 g/L, 12 mL/L, and 125 mL/L, respectively, had the greatest effect on CH4 production, increasing the cumulative CH4 yield by 13.5 %, 10.4 %, 26.8 %, and 32.9 %. Through microbial community and metabolism pathway analyses, the conductive materials promoted direct interspecies electron transfer (DIET) between Clostridium and Methanothrix, and the acetoclastic methanogenic pathway dominated by acetyl-CoA synthase. Rumen microorganisms enhanced AD performance by promoting the growth of hydrogenotrophic methanogens and the abundance of genes dominated by formylmethanofuran dehydrogenase in the hydrogenotrophic methanogenic pathway, illustrating the relationship between microbial community and metabolism pathway. Rumen microorganisms increased CH4 production more than conductive materials in real TCM wastewater. This study helps to better understand the internal mechanisms by which different materials enhance AD performance.

Methanogenic degradation of long-chain fatty acids associated with syntrophic microbial dynamics during thermophilic AnMBR treatment of lipid-rich dairy wastes

Anaerobic membrane reactors (AnMBRs) have been applied to effectively treat dairy wastes because they can address the drawbacks of sludge flotation and biomass washout. However, when dairy wastes contain considerable proportion of lipids, any anaerobic systems will likely encounter challenges with long-chain fatty acid (LCFA) inhibition. In this study, a thermophilic AnMBR was operated to treat dairy processing wastewater (DPW) plus ice cream waste (ICW). To evaluate the impact of LCFAs, the lipid-rich ICW was gradually introduced at different levels (0, 5, 10, and 20 %) into the AnMBR, along with a stepwise increase in lipid loading rate (LLR) from 0.22 to 11.24 g lipid/L/d. The results of biomethanation revealed that the AnMBR could be successfully operated when the LLR was maintained at 5.73 g lipid/L/d with a lipid/VS ratio of 50.22 % in the feedstock. However, at ultra-high LLR of 11.24 g lipid/L/d (20 % ICW added), a significant inhibitory effect on methane yields was observed, resulting in a reduction of 56.90 %. Dynamic analysis of the LCFAs profiled that a lower degradation rate was an important factor contributing to LCFA accumulation, and the inhibition concentration of various LCFAs was identified. Based on microbial evolution and co-occurrence network results, potential syntrophic partnerships between LCFA-degrading bacteria and methanogens were proposed. The responsible enzyme genes in the LCFA-degrading, acetoclastic, and hydrogenotrophic methanogenic pathways played crucial roles in LCFA inhibition and degradation. These findings offer practical insights for the efficient methanogenic treatment of lipid-rich wastes.

Unveiling the synergy of volatile fatty acids and ammonia nitrogen in optimizing methane production during dry anaerobic digestion of pig manure and corn straw

Dry anaerobic digestion (AD) is vulnerable to volatile fatty acids (VFA) and ammonia nitrogen. Understanding their synergy is crucial for optimizing strategies. In this study, the digestive performance, microbial dynamics, metabolic pathways, and synergistic relationships were investigated in the dry AD process by adding different concentrations of acetic acid (2, 5, and 8 mg/g) and urea (1, 5 mg/g N). Results showed that acid inhibition was caused by excessive pH reduction due to VFA accumulation. Propionic acid was the primary form of VFA accumulation. The inhibition of VFA accumulation could be alleviated using a low concentration of ammonia nitrogen (1 mg/g N urea) or be strengthened using a high concentration of ammonia nitrogen (5 mg/g N urea). Compared with the control group (82.2 mL/g VS), VFA and 1 mg/g N urea had a synergistic facilitation effect on the methane production (increasing by 1.7 %-18.1 %). This synergistic effect shortened the lag phases, enhanced the activity of Caldicoprobacter and Methanosarcina, increased the expression of genes encoding functional enzymes involved in the acetoclastic-methanogenic pathway. By contrast, VFA and 5 mg/g N urea had a synergistic inhibition effect on methane production (decreasing by 65.7 %–69.8 %). This synergistic effect prolonged lag phases, increased the relative abundance of ammonia-tolerant and acid-tolerant microbial (such as Methanoculleus) and reduced the expression of genes encoding functional enzymes involved in the hydrogenotrophic-methanogenic pathway. These results improve the understanding of methane regulation during dry AD.

Comparison of anaerobic co-digestion of vacuum toilet blackwater and kitchen waste under mesophilic and thermophilic conditions: Reactor performance, microbial response and metabolic pathway

Co-digestion of kitchen waste (KW) and black water (BW) can be considered as an attractive method to efficiently achieve the clean energy from waste. To find the optimal operation parameters for the co-digestion, the effects of different temperatures (35 and 55 °C) and BW:KW ratios on the reactor performances, microbial communities and metabolic pathways were studied. The results showed that the optimum BW:KW ratio was 1:3.6 and 1:4.5 for mesophilic and thermophilic optimal reactors, with methane production of 449.04 mL/g VS and 411.90 mL/g VS, respectively. Microbial communities showed significant differences between the reactors under different temperatures. For bacteria, increasing BW:KW ratio significantly promoted Defluviitoga enrichment (1.1%–9.5%) under thermophilic condition. For Archaea, the increase in BW:KW ratio promoted the enrichment of Methanosaeta (8.6%–56.4%) in the mesophilic reactor and Methanothermobacter (62.0%–89.2%) in the thermophilic reactor. The analysis of the key enzymes showed that, acetoclastic methanogenic pathway performed as the dominant under mesophilic condition, with high abundance of Acetate-CoA ligase (EC:6.2.1.1) and Pyruvate synthase (EC:1.2.7.1). Hydrogenotrophic methanogenic pathway was the main pathway in the thermophilic reactors, with high abundance of Formylmethanofuran dehydrogenase (EC:1.2.99.5).

Isolation of an H2-dependent electron-bifurcating CO2-reducing megacomplex with MvhB polyferredoxin from Methanothermobacter marburgensis.

In the hydrogenotrophic methanogenic pathway, formylmethanofuran dehydrogenase (Fmd) catalyzes the formation of formylmethanofuran through reducing CO2. Heterodisulfide reductase (Hdr) provides two low potential electrons for the Fmd reaction using a flavin-based electron-bifurcating mechanism. [NiFe]-hydrogenase (Mvh) or formate dehydrogenase (Fdh) complexes with Hdr and provides electrons to Hdr from H2 and formate, or the reduced form of F420, respectively. Recently, an Fdh-Hdr complex was purified as a 3-MDa megacomplex that contained Fmd, and its three-dimensional structure was elucidated by cryo-electron microscopy. In contrast, the Mvh-Hdr complex has been characterized only as a complex without Fmd. Here, we report the isolation and characterization of a 1-MDa Mvh-Hdr-Fmd megacomplex from Methanothermobacter marburgensis. After anion-exchange and hydrophobic chromatography was performed, the proteins with Hdr activity eluted in the 1- and 0.5-MDa fractions during size exclusion chromatography. Considering the apparent molecular mass and the protein profile in the fractions, the 1-MDa megacomplex was determined to be a dimeric Mvh-Hdr-Fmd complex. The megacomplex fraction contained a polyferredoxin subunit MvhB, which contains 12 [4Fe-4S]-clusters. MvhB polyferredoxin has never been identified in the previously purified Mvh-Hdr and Fmd preparations, suggesting that MvhB polyferredoxin is stabilized by the binding between Mvh-Hdr and Fmd in the Mvh-Hdr-Fmd complex. The purified Mvh-Hdr-Fmd megacomplex catalyzed electron-bifurcating reduction of [13C]-CO2 to form [13C]-formylmethanofuran in the absence of extrinsic ferredoxin. These results demonstrated that the subunits in the Mvh-Hdr-Fmd megacomplex are electronically connected for the reduction of CO2, which likely involves MvhB polyferredoxin as an electron relay.

Mitigating short-circuits through synergistic temperature and hydraulic retention time control for enhancing methane yield in continuous stirred-tank reactors

Temperature and hydraulic retention time (HRT) control are crucial for mitigating the “short-circuit” issue and enhancing methane production in continuous stirred-tank reactors. This study focuses on the combined effects of temperature and HRT on methanation via operating the reactors at different temperatures (35 °C, 45 °C, and 55 °C) and HRT (30, 25, and 20 d) for 116 days. The results demonstrated that the digester conducted at 45 °C and an HRT of 30 d unexpectedly yielded the maximum average methane production of 296.08 ml/g volatile solids (VS). In contrast, the optimal HRT for the 35 °C and 55 °C digesters were 25 and 30 d, with a methane yield of 241.88 ml/g VS and 240.9 ml/g VS, respectively. High-throughput sequencing shows that Firmicutes, Bacteroidetes, Themotogae, Chloroflexi, and Acidobacteria dominated 90–98 % of the total bacteria. Temperature exhibited a significant influence on shaping microbial ecosystems, indicating that temperature, rather than HRT, is the primary factor to consider. Additionally, metagenomic analysis reveals that the levels of genes involved in acetoclastic and hydrogenotrophic methanogenic pathways were enhanced at 45 °C, probably improving methane yield. These findings provide new insights into “short-circuit” mitigation and strategies to strengthen methane generation from biomass.

The Role of Slurry Reflux in a Corn Stalk Continuous Anaerobic Digestion System: Performance and Microbial Community.

Slurry reflux is a low-cost slurry reduction technology, which can solve the problem that a large amount of slurry cannot be completely consumed in a biogas plant. Anaerobic digestion (AD) of corn stalks with slurry reflux and non-reflux was compared and evaluated in continuous anaerobic digestion to clarify the effects of slurry reflux on AD with organic loading rate (OLR) variation. It was found that slurry reflux increased cumulative methane production and improved system stability. The average methane yield of the slurry reflux group was 224.19 mL/gVS, which was 41.35% higher than that of the non-reflux group. High-throughput sequencing results showed that slurry reflux increased the microbial community richness. The dominant microorganisms in the reflux group were in phylum Bacteroidetes, which have the capacity to degrade polymers, and Methanothrix, which is an aceticlastic methanogen. The relative abundances of Bacteroidetes and Methanothrix were 32.41% and 41.75%, respectively. Clostridium III and Saccharofermentans, which are related to syntrophic acetate oxidation and hydrolysis, were increased in relative abundance in the slurry reflux system. The increase of the OLR altered the main methane-producing pathway from the acetoclastic methanogenic pathway to the hydrogenotrophic methanogenic pathway in the AD system, and the slurry reflux can delay this trend. This study provided an effective way for the reduction and utilization of slurry in a biogas plant.

Metabolic effects of Fe0 on simultaneously eliminating excessive acidification and upgrading biogas in mesophilic or thermophilic anaerobic reactor

Excessive acidification and low methane (CH4) percentage are technical challenges restricting anaerobic digestion (AD) of kitchen waste, especially under high organic loading rate (OLR) conditions. Although the contribution of Fe0 powder to excessive acidification elimination and CH4 yield improvement in mesophilic AD (MAD) has been extensively reported, its impact on the thermophilic AD (TAD) process with more frequent excessive acidification problem is rarely studied. Moreover, the difference in microbial interactions and metabolic pathways between MAD and TAD reactors under Fe0 regulation was still unclear. In this study, we compared the performance of Fe0 dosing on excessive acidification elimination and biogas upgrading quality in both MAD and TAD reactors at different OLRs. The results showed that compared with excessive acidification or low CH4 percentage (less than 70%) in control reactors without Fe0 addition, the CH4 yield of MAD and TAD reactors with addition of Fe0 at OLR of 35 gVS/L were 470.88 mLCH4/gVS and 270.40 mLCH4/gVS, with higher CH4 composition of 81.32%–82.23% and 83.68%–88.48% respectively. Microbial diversity analysis showed a sharp distinction between MAD and TAD reactors with addition of Fe0. Analyses of co-occurrence networks and metabolic pathway presented hydrogenotrophic methanogenic pathway, in addition to acetoclastic methanogenesis, was dominant in MAD reactors as results of enhanced interspecies hydrogen transfer between Methanoculleus and some hydrogen producers, especially under high OLR condition. In contrast, syntrophilic acetate oxidization - hydrogenotrophic methanogenesis, which dominated by Methanothermobacter and Alkaliphilus or Pelotomaculum, was the sole methanogenic pathway in TAD reactors. The results will shed lights on the regulation mechanisms of Fe0 in AD of kitchen waste under mesophilic and thermophilic conditions, offering new insight into development of more efficient waste to energy conversion technologies.

Intermittent energization improves microbial electrolysis cell-assisted thermophilic anaerobic co-digestion of food waste and spent mushroom substance

Microbial electrolysis cell-assisted thermophilic anaerobic digestion (MEC-TAD) is a promising method to improve anaerobic co-digestion efficiency; however, its application is restricted by high energy consumption. To improve the energy use efficiency of MEC-TAD, this study investigated the effect of different intermittent energization strategies on thermophilic co-digestion performance. Results revealed that an 18 h-ON/6h-OFF energization schedule resulted in the fastest electron transfer rate and the highest methane yield (364.3 mL/g VS). Mechanistic analysis revealed that 18 h-ON/6h-OFF resulted in the enrichment of electroactive microorganisms and increased abundance of enzyme-coding genes associated with energy metabolism (ntp, nuo, atp), electron transfer (pilA, nfrA2, ssuE), and the hydrogenotrophic methanogenic pathway. Finally, energy balance analysis revealed that 18 h-ON/6h-OFF had the highest net energy benefit (2.52 kJ) and energy conversion efficiency (110.76 %). Therefore, intermittent energization of MEC-TAD using an 18 h-ON/6h-OFF schedule can provide improved performance and more energy savings.

Multi-omics joint analysis of the effect of temperature on microbial communities, metabolism, and genetics in full-scale biogas reactors with food waste

Due to the diversity of microbiota and the high complexity of their interactions that mediate biogas production, a detailed understanding of the microbiota is essential for the overall stability and performance of the anaerobic digestion (AD) process. This study evaluated the microbial taxonomy, metabolism, function, and genetic differences in 14 full-scale biogas reactors and laboratory reactors operating under various conditions in China. This is the first known study of the microbial ecology of AD at food waste (FW) at a regional scale based on multi-omics (16S rRNA gene amplicon sequencing, metagenomics, and proteomics). Temperature significantly affected the bacterial and archaeal community structure (R2 = 0.996, P = 0.001; R2 = 0.846, P < 0.002) and total inorganic carbon(TIC) slightly changed the microbial structure (R2 = 0.532, P = 0.005; R2 = 0.349, P = 0.016). The Wood-Ljungdahl coupled with hydrogenotrophic methanogenic pathways were dominant in the thermophilic reactors, where the acs, metF, cooA, mer, mch and ftr genes were 10.1-, 2.8-, 16.2-, 1.74-, 4.15-, 1.04-folds of the mesophilic reactors (P < 0.01). However, acetoclastic and methylotrophic methanogenesis was the primary pathway in the mesophilic reactors, where the ackA, pta, cdh and mta genes were 2.2-, 3.2-, 14.3-, 1.88-folds of the thermophilic group (P < 0.01). Finally, the Wilcoxon rank-sum test was applied to explain the cause of the temperature affecting AD microbial activities. The findings have deepened the understanding of the effect of temperature on AD microbial ecosystems and are expected to guide the construction and management of full-scale FW biogas plants.

Metabolic and ecological controls on the stable carbon isotopic composition of archaeal (isoGDGT and BDGT) and bacterial (brGDGT) lipids in wetlands and lignites

The glycerol dialkyl glycerol tetraether (GDGT) lipids of archaea and bacteria have been extensively studied in both modern and ancient wetlands, but their specific biological sources and biogeochemical significance in such settings remain unclear. The stable carbon isotopic composition of GDGTs, and associated tetraether lipids such as butanetriol - DGTs (BDGTs), can help reveal aspects of the carbon metabolism of their source organism. Here we present an in-depth compound-specific analyses of archaeal (isoGDGTs and BDGT-0) and bacterial (brGDGTs) tetraether lipid δ13C values from the Florida Everglades wetland, supplemented by 16S rRNA gene profiling of the archaeal community. We interpret this alongside GDGT δ13C analyses from three additional freshwater wetland sediments, and two ancient wetland deposits (lignites) from the Miocene. Our data suggests that isoGDGT-0 δ13C values, which range from –22.5‰ to −36.5‰ across our wetland sites, could to some degree reflect the relative importance of the acetotrophic and CO2-reducing hydrogenotrophic methanogenic pathways, though definitively delineating between the two is challenging due to (a) potential incorporation of dissolved inorganic carbon (DIC) by CO2 reducers, and (b) the diluting effect of GDGTs derived from seemingly heterotrophic/fermentative Bathyarchaeia, the most abundant archaeal class in the Everglades. In the Everglades, which harbours abundant Methanomassiliicoccales, BDGT-0 is significantly more 13C-depleted than all other lipids, with a δ13C signature at certain depths (∼−40.0 to −45.0‰) that is consistent with an important source from these (likely) methylotrophic methanogens. This adds to a growing body of evidence on the underappreciated role of methylotrophic methanogenesis, largely driven by Methanomassiliicoccales, in the freshwater methane cycle. However, slightly more 13C-enriched values (∼−35.0‰) at certain depths in the same site provide evidence for additional, possibly heterotrophic sources under certain conditions, and BDGT-0 δ13C values in the Miocene lignite (≈ δ13C of TOC) are consistent with a solely heterotrophic source. Where available for measurement, δ13C values of isoGDGTs-1 to -3, averaging –32.7 ± 2.3‰, were significantly more 13C-depleted than isoGDGT-0, suggesting a different mixture of sources, likely including an autotrophic/mixotrophic component derived from archaea belonging to the Crenarchaeota, Nitrososphaeria (formerly Thaumarchaeota) and/or MBG-D/DHVEG-1. Finally, brGDGT δ13C values are generally consistent with their proposed bacterial heterotrophic origin and biosynthetic effects, averaging out at around 4‰ depleted relative to total organic carbon (TOC). Importantly, we find no significant variation between the δ13C values of different brGDGTs across our wetland types. This highlights that spatial and temporal changes in commonly applied brGDGT-based proxies likely do not co-occur with changes in the heterotrophic lifestyle of their producer(s), discounting a possible confounding factor in brGDGT palaeothermometry. Taken together, our results yield insights into wetland archaeal communities and the lipids they produce, and underline the applicability of tetraether δ13C analyses in probing aspects of microbial carbon cycling in modern and ancient wetland sediments.

Revealing the methanogenic pathways for anaerobic digestion of key components in food waste: Performance, microbial community, and implications

Anaerobic digestion (AD) process is widely considered the most sustainable technology for food waste (FW) disposal due to its advantage of biomethane recovery and beneficial environmental consequences. However, the effects of key components in FW (i.e. starchy food, vegetables, fruits, and meats) on AD process and their methanogenic pathways remain unclear. In this study, the biochemical methane potential (BMP) of cooked rice, cabbage, banana peel, pork and local FW was 288, 283, 254, 630, and 476 NmL CH4/g VSadded, with t80 (time required for 80% methane produced) of 3, 9, 3, 11 and 11 days, respectively. Kinetic analysis suggested diverse hydrolysis rates (0.104–0.679 d-1) and specific methane yields (39–119 NmL CH4/g VSadded/d). The relative abundances of key methanogens in the reactors were diverse, leading to the variation in acetoclastic and hydrogenotrophic methanogenic pathways. This study provides fundamental information for the operation of AD systems with different FW compositions.

Genome-centric metagenomics provides new insights into the microbial community and metabolic potential of landfill leachate microbiota

Landfills are important sources of microorganisms associated with anaerobic digestion. However, the knowledge on microbiota along with their functional potential in this special habitat are still lacking. In this study, we recovered 1168 non-redundant metagenome-assembled genomes (MAGs) from nine landfill leachate samples collected from eight cities across China, spanning 42 phyla, 73 classes, 114 orders, 189 families, and 267 genera. Totally, 74.1% of 1168 MAGs could not be classified to any known species and 5.9% of these MAGs belonged to microbial dark matter phyla. Two putative novel classes were discovered from landfill leachate samples. The identification of thousands of novel carbohydrate-active enzymes showed similar richness level compared to the cow rumen microbiota. The methylotrophic methanogenic pathway was speculated to contribute significantly to methane production in the landfill leachate because of its co-occurrence with the acetoclastic and hydrogenotrophic methanogenic pathways. The genetic potential of dissimilatory nitrate reduction to ammonium (DNRA) was observed, implying DNRA may play a role in ammonium generation in landfill leachate. These findings implied that landfill leachate might be a valuable microbial resource repository and filled the previous understanding gaps for both methanogenesis and nitrogen cycling in landfill leachate microbiota. Our study provides a comprehensive genomic catalog and substantially provides unprecedented taxonomic and functional profiles of the landfill leachate microbiota.

Exploring the effect of voltage on biogas production performance and the methanogenic pathway of microbial electrosynthesis

Microbial electrosynthesis (MES) has great potential for energy and resource recovery in wastewater treatment and has therefore gradually become a popular research topic. In this study, MES with an applied voltage (0.6 V, 0.8 V and 1.0 V) significantly improved biogas production (36.0 %, 52.3 %, and 59.1 %, respectively) compared with the control (without electrodes). Analysis of microbial community structure revealed that Clostridium and Methanosaeta had the highest relative abundance in all reactors. The abundances of Acetivibrio and Geobacter dominated the bioanode, and methane production was mainly mediated by the acetoclastic methanogenic pathway. On the biocathode, the abundances of Methanolinea and Methanothermobacter were higher than those of other microbes, and methane was produced primarily through the hydrogenotrophic methanogenic pathway. When the acetoclastic methanogenic pathway in the biocathode was inhibited, MES could efficiently produce methane through other pathways. In addition, MES was more tolerant to the effects of high NH4+-N concentrations than anaerobic digestion (AD). These results suggest that applying voltage to the bioanode and biocathode could better improve the efficiency of methane production.

Meta-proteomics analysis of microbial ecosystem during the anaerobic digestion of chicken manure in biogas production farm

Using meta-proteomics approach for the analysis of biogas production from anaerobic digestion has many advantages that await to be fully uncovered. In this study, we aimed to investigate biogas production from a stable 2-stage chicken manure fermentation system via chemical and biological perspective. The diversity and functional protein changes that occurred from the 1st to 2nd stage were a good indication of exposure on the differential metabolic processes in anaerobic digestion. The highlight of identified functional proteins explained the cause of accumulated ammonia and carbon sources for methane production. Due to the ammonia stress and nutrient limitation, hydrogenotrophic methanogenic pathway was adopted as an indicator of meta-proteomics data involving the key methanogenic substrates (formate and acetate). Unlike traditional meta-genomics analysis, this study identified not only the microorganism's name, but also the enzymes and pathway involved in the generation of methane and carbon dioxide in biogas production of chicken manure.

Explore the difference between the single-chamber and dual-chamber microbial electrosynthesis for biogas production performance

Microbial electrosynthesis (MES) is an advanced technology for efficient treatment of organic wastewater and recovery of new energy, with the advantages and disadvantages of single-chamber and dual-chamber MES reactors being less understood. Therefore, we explored the effects of single-chamber and dual-chamber structures on the methane production performance and microbial community structure of MES. Results indicated that methane concentration and current density of single-chamber MES were higher than those of dual-chamber MES, and the system stability was better, while chemical oxygen demand (COD) removal rate and cumulative methane production were not significantly different. Analysis of microbial community structure showed the abundance of acidogens and H2-producing bacteria was higher in single-chamber MES, while fermentation bacteria and methanogens was lower. The abundance of methanogens of dual-chamber MES (21.74–24.70%) was superior to the single-chamber MES (8.23–10.10%). Moreover, in dual-chamber MES, methane was produced primarily through acetoclastic methanogenic pathway, while in single-chamber MES cathode, methane production was mainly by hydrogenotrophic methanogenic pathway. Information provided will be useful to select suitable reactors and optimize reaction design.

In-situ alkaline enhanced two-stage anaerobic digestion system for waste cooking oil and sewage sludge co-digestion

Anaerobic digestion is a promising way for resource recovery from waste cooking oil (WCO) due to its high bio-methanation potential. In-situ mild alkaline (pH 8) enhanced two-stage continuous stirred tank reactors (ALK-2-CSTRs) were implemented to explore its efficiency in co-digesting WCO and sewage sludge with stepwise increase of WCO in the co-substrates. Results demonstrate that the ALK-2-CSTRs effectively promoted methane yield from the co-substrates via promoting hydrolysis, long chain fatty acids (LCFAs) degradation and protecting methanogens from exposure to high concentration of LCFAs directly. The maximum methane yield of the ALK-2-CSTRs is 39.2% higher than that of a single stage CSTR system at the optimal feed mixture of 45:55 (WCO:SS [VS]). The thermophilic operation applied to the stage-1 of the ALK-2-CSTRs failed to improve the methane yield when the methanogenic performance was stable; while upon WCO overloaded, the elevated temperature mitigated the deterioration of methanogenesis by stimulating the bioconversion of the toxic LCFAs, especially the unsaturated oleic acid. Microbial community analysis reveals the ALK-2-CSTRs stimulated the growth of lipolytic bacteria and hydrogenotrophic methanogens, which suggests the hydrogenotrophic methanogenic pathway was promoted. Cost evaluation demonstrates the economical superiority of the ALK-2-CSTR over the prevailing strategies developed for enhancing methane yield from the co-substrates.

Quorum quenching altered microbial diversity and activity of anaerobic membrane bioreactor (AnMBR) and enhanced methane generation

A facultative bacterium Microbacterium sp. (QQ strain) was found significantly mitigated membrane biofouling and also increased methane production. It was found genera Nitrospira, norank-c-Bacterodetes vadinHA17, Trichococcus and family Anaerolineaceae were likely responsible for membrane biofouling. The presence of QQ strain increased the total abundance of fermentative and acetogenic genera by 0.61% and 379.61%, respectively, but had a minor effect on the abundance of methanogens. The increased methane production was likely due to the strengthened methanogenic activity and more available substrates. Homo-acetogenic Treponema was enriched (9.01%) in the presence of QQ strain suggesting that apart from hydrogenotrophic methanogenic pathway, extra CH4 could be also produced from the additional acetate synthesized via homo-acetogenic pathway. This study advances knowledge about the effects of QQ strain on microbial communities, microbiota biofouling behavior and anaerobic fermentation process in AnMBRs.

Long-term evaluation of bioaugmentation to alleviate ammonia inhibition during anaerobic digestion: Process monitoring, microbial community response, and methanogenic pathway modeling

The effect of different bioaugmentation strategies on anaerobic digestion related to the alleviation of ammonia inhibition was investigated in a long-term operation. The long-term operation confirmed that bioaugmentation is a stable method. A 35% increase in methane production (MP) was observed in bottles bioaugmented with Methanosarcina barkeri (MSB) or Syntrophaceticu schinkii (SS) + Methanobrevibacter smithii (MBS), and a 49% increment was obtained from the bottles bioaugmented with Methanosaeta harundinacea (MSH) + SS + MBS. Results suggest that the enhancement in both aceticlastic and hydrogenotrophic methanogenic pathways should be considered, and bioaugmentation strain should be properly selected to achieve a synergistic effect. The microbial community analysis indicated Methanosarcina spp. was the dominant archaea. Combined with specific methanogenic activity and carbon isotope fractionation analysis, it was suggested that Methanosarcina spp. performed differently in methanogenic pathways in different bottles. The abundance of COG and total enzymes in the bottles with high MP (MSB and MSH + SS + MBS) was higher than that in the other bottles. The ratio of the functional enzyme tetrahydromethanopterin S-methyltransferase subunit H (EC 2.1.1.86) to formylmethanofuran dehydrogenase subunit E (EC 1.2.99.5) and the relative abundance of enolase (EC 1.2.1.2) confirmed the aceticlastic methanogenic pathway of MSB in Group 1 and the pathway enhancement balance in MSH + SS + MBS. The modified Anaerobic Digestion Model No.1 including syntrophic acetate oxidation was used for simulation with R2 > 0.96. Simulated contribution rate data indicated that the hydrogenotrophic methanogenic pathway was dominant while both two pathways got strengthened, which is consistent with the findings of the microbial analysis.

A Mini-Review on In situ Biogas Upgrading Technologies via Enhanced Hydrogenotrophic Methanogenesis to Improve the Quality of Biogas From Anaerobic Digesters

In the context of accelerating development of renewable and sustainable energy, biogas upgrading has attracted significant attention in order to yield high quality biomethane which can be utilized as an alternative to natural gas. Biogas upgrading techniques have been revolutionized in the past decade with the emergence of new strategies such as enhanced hydrogenotrophic methanogenesis. This review comprehensively summarizes the latest in situ biogas upgrading technologies via enhanced hydrogenotrophic methanogenesis in order to facilitate their development and commercial application: (1) hydrogen injection into anaerobic digesters for in situ enhancement of hydrogenotrophic methanogenic pathways, (2) membrane electrolysis-assisted CO2 extraction and bioelectrochemical removal of CO2, and (3) additives (e.g. ash, zero-valent iron and biochar). The key findings and technical limitations are summarized and critically analyzed for each in situ biogas upgrading technique. Additionally, potential industrial applications of these in situ biogas upgrading strategies in anaerobic digesters are also discussed, before concluding with their future development.