Methanogenic Archaea Research Articles

Differences in regulation mechanisms of glutamine synthetases from methanogenic archaea unveiled by structural investigations

Glutamine synthetases (GS) catalyze the ATP-dependent ammonium assimilation, the initial step of nitrogen acquisition that must be under tight control to fit cellular needs. While their catalytic mechanisms and regulations are well-characterized in bacteria and eukaryotes, only limited knowledge exists in archaea. Here, we solved two archaeal GS structures and unveiled unexpected differences in their regulatory mechanisms. GS from Methanothermococcus thermolithotrophicus is inactive in its resting state and switched on by 2-oxoglutarate, a sensor of cellular nitrogen deficiency. The enzyme activation overlays remarkably well with the reported cellular concentration for 2-oxoglutarate. Its binding to an allosteric pocket reconfigures the active site through long-range conformational changes. The homolog from Methermicoccus shengliensis does not harbor the 2-oxoglutarate binding motif and, consequently, is 2-oxoglutarate insensitive. Instead, it is directly feedback-inhibited through glutamine recognition by the catalytic Asp50ʹ-loop, a mechanism common to bacterial homologs, but absent in M. thermolithotrophicus due to residue substitution. Analyses of residue conservation in archaeal GS suggest that both regulations are widespread and not mutually exclusive. While the effectors and their binding sites are surprisingly different, the molecular mechanisms underlying their mode of action on GS activity operate on the same molecular determinants in the active site.

A hybrid photocatalytic system enables direct glucose utilization for methanogenesis

Integration of methanogenic archaea with photocatalysts presents a sustainable solution for solar-driven methanogenesis. However, maximizing CH4 conversion efficiency remains challenging due to the intrinsic energy conservation and strictly restricted substrates of methanogenic archaea. Here, we report a solar-driven biotic-abiotic hybrid (biohybrid) system by incorporating cadmium sulfide (CdS) nanoparticles with a rationally designed methanogenic archaeon Methanosarcina acetivorans C2A, in which the glucose synergist protein and glucose kinase, an energy-efficient route for glucose transport and phosphorylation from Zymomonas mobilis, were implemented to facilitate nonnative substrate glucose for methanogenesis. We demonstrate that the photo-excited electrons facilitate membrane-bound electron transport chain, thereby augmenting the Na+ and H+ ion gradients across membrane to enhance adenosine triphosphate (ATP) synthesis. Additionally, this biohybrid system promotes the metabolism of pyruvate to acetyl coenzyme A (AcCoA) and inhibits the flow of AcCoA to the tricarboxylic acid (TCA) cycle, resulting in a 1.26-fold augmentation in CH4 production from glucose-derived carbon. Our results provide a unique strategy for enhancing methanogenesis through rational biohybrid design and reprogramming, which gives a promising avenue for sustainably manufacturing value-added chemicals.

Fecal Microbiota and Feeding Habitats of Nomadic Indigenous Animals (Deer, Yak, Sheep and Camel) in Baikal Siberia (Russia)

In the vast expanse of Baikal Siberia, indigenous nomadic animal groups have been conserved, grazing on pastures throughout the year. It is believed that the fecal microbiota of these diverse nomadic animal species is unique to each species and closely tied to their feeding environments. We conducted a pioneering comparative analysis of the taxonomic structure and the diversity of fecal microbiota in indigenous nomadic animals inhabiting Baikal Siberia. Our study encompassed 20 deer, 23 yaks, 24 camels, and 29 sheep, using high-throughput 16S rRNA gene profiling. In the fecal microbiota of these animals, we observed a predominant presence of the phyla Bacillota, Bacteroidota, and Verrucomicrobiota, collectively comprising over 88% of the microbial communities. Moreover, these proportions exhibited variations according to the host species. The unculturable Bacillota UCG-005 and UCG-010 are the key groups for all animals. However, at the genus level, distinctive compositions of fecal microbiota were discernible within each animal group. We identified a total of 37 dominant genera across the fecal samples from these four animal species. Principal component analysis (PCA) and cluster analysis demonstrated that the fecal microbiota composition clustered among individuals of the same animal species. Linear discriminant analysis effect size (LEfSe) indicated that camels exhibited higher abundances of the family Akkermansiaceae and the uncultured clostridial lineage UCG-010, while deer featured Lachnospiraceae; sheep had Ruminococcaceae; and yaks displayed Monoglobaceae, Bacteroidaceae, and methanogenic archaea from the family Methanobacteriaceae as distinctive marker taxa. Our studies showed that the studied nomadic animals feed mainly on plants belonging to the families Poaceae, Cyperaceae, Asteraceae, and Rosaceae. Our research indicated that the identity of the host species and, to a lesser degree, their diets and habitats, significantly shape the composition of fecal microbiota in these studied nomadic ruminant animals.

Acyl homoserine lactone-based regulation strategy for improved methane production in anaerobic digestion of agricultural wastes

The acyl homoserine lactone (AHL)-based regulation strategy presents a considerable potential in improving anaerobic digestion (AD) efficiency of complex substrates. However, the reinforcement mechanisms are still unclear. In this study, the effects of different AHL types (C4-HSL, C7-HSL, C12-HSL) and different adding times (0 d, 5 d, 15 d) on AD performance of agricultural wastes (corn straw and cattle manure) were investigated. The results indicated that all types of AHLs exerted positive effects on AD, and the adding time rather than the AHL type was the key factor affecting AD processes. The C12-5d group obtained the highest accumulated methane production (310.7 ± 2.56 mL/g VS), which increased by 47.71% than that of the Control group. The exogenous AHLs facilitated the hydrolysis and acidification of complex substrates and the utilization of released low-molecular-weight organic acids in AD systems. Furthermore, the role of methanogenic archaea was upregulated and the dominance of bacteria was relatively weakened in AD systems, which favored the establishment of their balanced and symbiotic metabolic relationship. The enriched Methanobacterium strengthened the hydrogenotrophic methanogenesis pathway. This study provides new insights into the regulation strategy of AD for agricultural waste treatment based on AHL addition.

Biocatalysts for biomethanol production: Advancements and future prospects

Biomethanol, a renewable and sustainable alternative to traditional fossil-fuel-derived methanol, has garnered considerable attention as a potential solution to mitigate greenhouse gas emissions and dependence on non-renewable resources. The utilization of biocatalysts in biomethanol production offers a promising avenue to achieve environmentally friendly and economically viable processes. Paper highlights the biocatalytic pathways involved in biomethanol synthesis. Particular emphasis is placed on microbial biocatalysts, such as methanogenic archaea and certain bacteria, which possess the unique capability of converting carbon dioxide and hydrogen into methanol through a series of enzymatic reactions. Additionally, enzyme-based systems derived from various microorganisms and genetically engineered organisms are also discussed as potential biocatalysts for biomethanol synthesis. Paper also delves into the current challenges and limitations faced in harnessing biocatalysts for biomethanol production. These challenges include substrate availability, low conversion rates, enzyme stability, and process scalability. Several strategies to address these issues are highlighted, including metabolic engineering, synthetic biology, and bioprocess optimization techniques. The advantages of utilizing biocatalysts for biomethanol production are outlined. Biocatalytic routes offer the advantage of operating under mild conditions, which reduces energy consumption and minimizes the production of unwanted by-products. Furthermore, the utilization of renewable feedstocks, such as carbon dioxide captured from industrial emissions or waste streams, enhances the sustainability of the process. The final section discusses future prospects and potential research directions in the field of biocatalytic biomethanol production. Advances in biotechnology, omics technologies, and computational modeling are poised to accelerate the discovery and optimization of novel biocatalysts, thereby unlocking the full potential of biomethanol as a sustainable fuel and chemical precursor. The use of biocatalysts for biomethanol production offers an attractive approach to establish a green and circular economy. With ongoing research and technological advancements, the field holds significant promise for reducing carbon emissions and transitioning towards a more sustainable energy landscape. However, to fully realize the potential of biocatalytic biomethanol production, interdisciplinary collaboration and concerted efforts are required to address existing challenges.

Anaerobic hydrocarbon biodegradation by alkylotrophic methanogens in deep oil reservoirs.

In subsurface biodegraded oil reservoirs, methanogenic biodegradation of crude oil is a common process. This process was previously assigned to the syntrophy of hydrocarbon-degrading bacteria and methanogenic archaea. Recent studies showed that archaea of the Candidatus Methanoliparum named as alkylotrophic methanogens couple hydrocarbon degradation and methane production in a single archaeon. To assess the geochemical role of Ca. Methanoliparum, we analyzed the chemical and microbial composition and metabolites of 209 samples from 15 subsurface oil reservoirs across China. Gas chromatography-mass spectrometry analysis revealed that 92% of the tested samples were substantially degraded. Molecular analysis showed that 85% of the tested samples contained Ca. Methanoliparum, and 52% of the tested samples harbored multiple alkyl-coenzyme M derivatives, the intercellular metabolites of alkylotrophic archaea. According to metagenomic and metatranscriptomic analyses, Ca. Methanoliparum dominates hydrocarbon degradation in biodegraded samples from the Changqing, Jiangsu, and Shengli (SL) oilfields, and it is persistently present as shown in a 15-year-long sampling effort at the Shengli oilfield. Together, these findings demonstrate that Ca. Methanoliparum is a widely distributed oil degrader in reservoirs of China, suggesting that alkylotrophic methanogenesis by archaea plays a key role in the alteration of oil reservoirs, thereby expanding our understanding of biogeochemical process in the deep biosphere.

Facultative endosymbiosis between cellulolytic protists and methanogenic archaea in the gut of the Formosan termite Coptotermes formosanus.

Anaerobic protists frequently harbour methanogenic archaea, which apparently contribute to the hosts' fermentative metabolism by consuming excess H2. However, the ecological properties of endosymbiotic methanogens remain elusive in many cases. Here we investigated the ecology and genome of the endosymbiotic methanogen of the Cononympha protists in the hindgut of the termite Coptotermes formosanus. Microscopic and 16S rRNA amplicon sequencing analyses revealed that a single species, designated here "Candidatus Methanobrevibacter cononymphae", is associated with both Cononympha leidyi and Cononympha koidzumii and that its infection rate in Cononympha cells varied from 0.0% to 99.8% among termite colonies. Fine-scale network analysis indicated that multiple 16S rRNA sequence variants coexisted within a single host cell and that identical variants were present in both Cononympha species and also on the gut wall. Thus, "Ca. Methanobrevibacter cononymphae" is a facultative endosymbiont, transmitted vertically with frequent exchanges with the gut environment. Indeed, transmission electron microscopy showed escape or uptake of methanogens from/by a Cononympha cell. The genome of "Ca. Methanobrevibacter cononymphae" showed features consistent with its facultative lifestyle: i.e., the genome size (2.7 Mbp) comparable to those of free-living relatives; the pseudogenization of the formate dehydrogenase gene fdhA, unnecessary within the non-formate-producing host cell; the dependence on abundant acetate in the host cell as an essential carbon source; and the presence of a catalase gene, required for colonization on the microoxic gut wall. Our study revealed a versatile endosymbiosis between the methanogen and protists, which may be a strategy responding to changing conditions in the termite gut.

Methanogenic symbionts of anaerobic ciliates are host and habitat specific.

The association between anaerobic ciliates and methanogenic archaea has been recognized for over a century. Nevertheless, knowledge of these associations is limited to a few ciliate species, and so the identification of patterns of host-symbiont specificity has been largely speculative. In this study, we integrated microscopy and genetic identification to survey the methanogenic symbionts of 32 free-living anaerobic ciliate species, mainly from the order Metopida. Based on Sanger and Illumina sequencing of the 16S rRNA gene, our results show that a single methanogenic symbiont population, belonging to Methanobacterium, Methanoregula, or Methanocorpusculum, is dominant in each host strain. Moreover, the host's taxonomy (genus and above) and environment (i.e. endobiotic, marine/brackish, or freshwater) are linked with the methanogen identity at the genus level, demonstrating a strong specificity and fidelity in the association. We also established cultures containing artificially co-occurring anaerobic ciliate species harboring different methanogenic symbionts. This revealed that the host-methanogen relationship is stable over short timescales in cultures without evidence of methanogenic symbiont exchanges, although our intraspecific survey indicated that metopids also tend to replace their methanogens over longer evolutionary timescales. Therefore, anaerobic ciliates have adapted a mixed transmission mode to maintain and replace their methanogenic symbionts, allowing them to thrive in oxygen-depleted environments.

Pleomorphic viruses establish stable relationship with marine hyperthermophilic archaea.

Non-lytic viruses with enveloped pleomorphic virions (family Pleolipoviridae) are ubiquitous in hypersaline environments across the globe and are associated with nearly all major lineages of halophilic archaea. However, their existence in other ecosystems remains largely unknown. Here, we show that evolutionarily-related viruses also infect hyperthermophilic archaea thriving in deep-sea hydrothermal vents. Archaeoglobus veneficus pleomorphic virus 1 (AvPV1), the first virus described for any member of the class Archaeoglobi, encodes a morphogenetic module typical of pleolipoviruses, including the characteristic VP4-like membrane fusion protein. We show that AvPV1 is a non-lytic virus chronically produced in liquid cultures without substantially affecting the growth dynamics of its host with a stable virus-to-host ratio of ~1. Mining of genomic and metagenomic databases revealed broad distribution of AvPV1-like viruses in geographically remote hydrothermal vents. Comparative genomics, coupled with phylogenetic analysis of VP4-like fusogens revealed deep divergence of pleomorphic viruses infecting halophilic, methanogenic, and hyperthermophilic archaea, signifying niche separation and coevolution of the corresponding virus-host pairs. Hence, we propose a new virus family, "Thalassapleoviridae," for classification of the marine hyperthermophilic virus AvPV1 and its relatives. Collectively, our results provide insights into the diversity and evolution of pleomorphic viruses beyond hypersaline environments.

Coupling between 2, 2′, 4, 4′-tetrabromodiphenyl ether (BDE-47) debromination and methanogenesis in anaerobic soil microcosms

Polybrominated diphenyl ethers (PBDEs) are persistent pollutants that may undergo microbial-mediated debromination in anoxic environments, where diverse anaerobic microbes such as methanogenic archaea co-exist. However, current understanding of the relations between PBDE pollution and methanogenic process is far from complete. To address this knowledge gap, a series of anaerobic soil microcosms were established. BDE-47 (2, 2′, 4, 4′-tetrabromodiphenyl ether) was selected as a model pollutant, and electron donors were supplied to stimulate the activity of anaerobes. Debromination and methane production were monitored during the 12 weeks incubation, while obligate organohalide-respiring bacteria (OHRBs), methanogenic, and the total bacterial communities were examined at week 7 and 12. The results demonstrated slow debromination of BDE-47 in all microcosms, with considerable growth of Dehalococcoides and Dehalogenimonas over the incubation observed in most BDE-47 spiked treatments. In addition, the accumulation of intermediate metabolites positively correlated with the abundance of Dehalogenimonas at week 7, suggesting potential role of these OHRBs in debromination. Methanosarcinaceae were identified as the primary methanogenic archaea, and their abundance were correlated with the production of debrominated metabolites at week 7. Furthermore, it was observed for the first time that BDE-47 considerably enhanced methane production and increased the abundance of mcrA genes, highlighting the potential effects of PBDE pollution on climate change. This might be related to the inhibition of reductive N- and S-transforming microbes, as revealed by the quantitative microbial element cycling (QMEC) analysis. Overall, our findings shed light on the intricate interactions between PBDE and methanogenic processes, and contribute to a better understanding of the environmental fate and ecological implication of PBDE under anaerobic settings.

Methanogenic Archaea in the Pediatric Inflammatory Bowel Disease in Relation to Disease Type and Activity.

The inflammatory bowel disease (IBD) is associated with gut microbiota dysbiosis; however, studies on methanogens-especially those focused on children-are extremely limited. The aim of this study was to determine the abundance of total methanogenic archaea and their three subgroups: Methanobrevibacter (Mb.) smithii, Methanosphaera (Ms.) stadtmanae, and Methanomassiliicoccales, in the feces of children with both active and inactive Crohn's disease (CD) and ulcerative colitis (UC). The results of a quantitative real-time PCR were cross-referenced with the disease type (CD vs. UC) and activity assessed with the use of Pediatric Crohn's Disease Activity Index (PCDAI) and Pediatric Ulcerative Colitis Activity Index (PUCAI) indices, and fecal calprotectin (FCP) concentration, and compared with controls. There was a significant decrease in the number of total methanogens in CD and UC compared to controls. The prevalence of total methanogens was also lower in UC compared to controls. Furthermore, patients from the inactive UC group were colonized by a lower number of Mb. smithii, and demonstrated the most pronounced positive correlation between the number of Ms. stadtmanae and the FCP concentration. Our results demonstrate that gut methanogens are related to the type and activity of pediatric IBD.

Effects of Fe–N co-modified biochar on methanogenesis performance, microbial community, and metabolic pathway during anaerobic co-digestion of alternanthera philoxeroides and cow manure

The performance of anaerobic digestion (AD) is susceptible to disturbances in feedstock degradation, intermediates accumulation, and methanogenic archaea activity. To improve the methanogenesis performance of the AD system, Fe–N co-modified biochar was prepared under different pyrolysis temperatures (300,500, and 700 °C). Meanwhile, pristine and Fe-modified biochar were also derived from alternanthera philoxeroides (AP). The aim was to compare the effects of Fe–N co-modification, Fe modification, and pristine biochar on the methanogenic performance and explicit the responding mechanism of the microbial community in anaerobic co-digestion (coAD) of AP and cow manure (CM). The highest cumulative methane production was obtained with the addition of Fe-N-BC500 (260.38 mL/gVS), which was 42.37 % higher than the control, while the acetic acid, propionic acid, and butyric acid concentration of Fe-N-BC were increased by 147.58 %, 44.25 %, and 194.06 % compared with the control, respectively. The co-modified biochar enhanced the abundance of Chloroflexi and Methanosarcina in the AD system. Metabolic pathway analysis revealed that the increased methane production was related to the formation and metabolism of volatile fatty acids and that Fe-N-BC500 enhanced the biosynthesis of coenzyme A and the cell activity of microorganisms, accelerating the degradation of propionic acid and enhancing the hydrogenotrophic methanogenesis pathway. Overall, Fe–N co-modified biochar was proved to be an effective promoter for accelerated methane production during AD.

A unified compendium of prokaryotic and viral genomes from over 300 anaerobic digestion microbiomes

BackgroundThe anaerobic digestion process degrades organic matter into simpler compounds and occurs in strictly anaerobic and microaerophilic environments. The process is carried out by a diverse community of microorganisms where each species has a unique role and it has relevant biotechnological applications since it is used for biogas production. Some aspects of the microbiome, including its interaction with phages, remains still unclear: a better comprehension of the community composition and role of each species is crucial for a cured understanding of the carbon cycle in anaerobic systems and improving biogas production.ResultsThe primary objective of this study was to expand our understanding on the anaerobic digestion microbiome by jointly analyzing its prokaryotic and viral components. By integrating 192 additional datasets into a previous metagenomic database, the binning process generated 11,831 metagenome-assembled genomes from 314 metagenome samples published between 2014 and 2022, belonging to 4,568 non-redundant species based on ANI calculation and quality verification. CRISPR analysis on these genomes identified 76 archaeal genomes with active phage interactions. Moreover, single-nucleotide variants further pointed to archaea as the most critical members of the community. Among the MAGs, two methanogenic archaea, Methanothrix sp. 43zhSC_152 and Methanoculleus sp. 52maCN_3230, had the highest number of SNVs, with the latter having almost double the density of most other MAGs.ConclusionsThis study offers a more comprehensive understanding of microbial community structures that thrive at different temperatures. The findings revealed that the fraction of archaeal species characterized at the genome level and reported in public databases is higher than that of bacteria, although still quite limited. The identification of shared spacers between phages and microbes implies a history of phage-bacterial interactions, and specifically lysogenic infections. A significant number of SNVs were identified, primarily comprising synonymous and nonsynonymous variants. Together, the findings indicate that methanogenic archaea are subject to intense selective pressure and suggest that genomic variants play a critical role in the anaerobic digestion process. Overall, this study provides a more balanced and diverse representation of the anaerobic digestion microbiota in terms of geographic location, temperature range and feedstock utilization.

Gut microbiota at different stages of cardiovascular disease continuum

Aim. To study the relationship of gut microbiota (GM) composition with clinical and paraclinical characteristics of patients with coronary artery disease (CAD) and heart failure with reduced ejection fraction (HFrEF).Material and methods. The study included 64 patients (men, 71,9%) with CAD, 33 (men, 87,9%) with HFrEF and 38 (men, 63,2%) with various cardiovascular risk factors (control group). GM composition was assessed using real-time polymerase chain reaction and next- generation sequencing. To identify correlations of GM composition with clinical and paraclinical characteristics, the Spearman rank correlation coefficient (ρ) was calculated. Generalized linear models (QuasiPoisson, overdispersion) with covariates (age, body mass index) were used to compare groups by phylotype.Results. The relative abundance of Bacillota phylotypes is significantly higher in patients with CAD and HFrEF compared to controls (p<0,001). The Bacillota/Bacteroidota dysbiosis ratio was also significantly higher in patients with CAD and HFrEF than in the control group (p<0,001). According to correlation analysis, the relative representation of methanogenic archaea Methanomassiliicoccaceae, Methanomethylophilaceae (ρ=0,49, p<0,0003) directly correlated with ventricular arrhythmias in patients with HFrEF.Conclusion. Patients with CAD and HFrEF differ significantly from the control group in GM composition at the phylotype level. In the study groups, Bacillota/Bacteroidota dysbiosis ratio was also significantly higher than in the control group. The relative abundance of the archaea Methanomassiliicoccaceae and Methanomethylophilaceae is associated with episodes of ventricular tachycardia/ventricular fibrillation.

Investigating the effect of supplementing different levels of Isochrysis galbana on in vitro rumen fermentation parameters.

This study aimed to investigate the effect of supplementing Isochrysis galbana (I. galbana) at levels of 0 (control), 1, 2, 3, 4, and 5 (g/100 g DM) of the diet on the gas production kinetics, methane production, rumen fermentation parameters, and relative microbial population in vitro. Supplementation of I. galbana at high level (5g/100 g DM) caused a significant decrease in total gas production (p < 0.05). High supplementation rates (4 and 5g/100 g DM) decreased CH4 production relative to the control by 18.4% and 23.2%, respectively. Although rumen ammonia nitrogen (N-NH3) and total volatile fatty acids (VFA) concentrations were affected by dietary treatments, but the VFA profile did not changed. The relative proportion of protozoa and methanogenic archaea as well as Anaerovibrio lipolytica, Prevotella spp., Ruminococcus flavefaciens, and Fibrobacter succinogenes were decreased significantly as a result of microalgae supplementation. However, the relative abundance of Ruminococcus albus, Butyrivibrio fibrisolvens and Selenomonas ruminantium were significantly increased (p < 0.05), related to the control group. As well, the pH was not affected by dietary treatments. It was concluded that I. galbana reduced in vitro CH4 production and methanogenic archaea that its worth to be investigated further in in vivo studies.

Highly diverse-Low abundance methanogenic communities in hypersaline microbial mats of Guerrero Negro B.C.S., assessed through microcosm experiments.

Methanogenic communities of hypersaline microbial mats of Guerrero Negro, Baja California Sur, Mexico, have been recognized to be dominated by methylotrophic methanogens. However, recent studies of environmental samples have evidenced the presence of hydrogenotrophic and methyl-reducing methanogenic members, although at low relative abundances. Physical and geochemical conditions that stimulate the development of these groups in hypersaline environments, remains elusive. Thus, in this study the taxonomic diversity of methanogenic archaea of two sites of Exportadora de Sal S.A was assessed by mcrA gene high throughput sequencing from microcosm experiments with different substrates (both competitive and non-competitive). Results confirmed the dominance of the order Methanosarcinales in all treatments, but an increase in the abundance of Methanomassiliiccocales was also observed, mainly in the treatment without substrate addition. Moreover, incubations supplemented with hydrogen and carbon dioxide, as well as the mixture of hydrogen, carbon dioxide and trimethylamine, managed to stimulate the richness and abundance of other than Methanosarcinales methanogenic archaea. Several OTUs that were not assigned to known methanogens resulted phylogenetically distributed into at least nine orders. Environmental samples revealed a wide diversity of methanogenic archaea of low relative abundance that had not been previously reported for this environment, suggesting that the importance and diversity of methanogens in hypersaline ecosystems may have been overlooked. This work also provided insights into how different taxonomic groups responded to the evaluated incubation conditions.

Enhancing Anaerobic Biodegradation of Phenanthrene in Polluted Soil by Bioaugmentation and Biostimulation: Focus on the Distribution of Phenanthrene and Microbial Community Analysis

The remediation of polycyclic aromatic hydrocarbon (PAH)-contaminated soils has received much attention in recent years, and most of the contaminated sites are in anaerobic environments, such as deep soils and flooded soils. We simulated the natural flooded soil environment, selected phenanthrene (PHE) as a model PAH contaminant, and designed batch experiments run for 63 days to comprehensively investigate the effects of the combined addition of anaerobic sludge and granular biochar on microbial community and function and the anaerobic biodegradation of PHE. Firstly, the residue, distribution, and removal of PHE in the flooded soil environment were quantified for each group. Secondly, the effects of bioaugmentation of soil indigenous microorganisms by the addition of anaerobic activated sludge and biostimulation of biochar on the removal of PHE from the soil were analyzed against each other. Lastly, the changes in the structure of the microbial community under the effect of bioaugmentation and biostimulation were illustrated by sequencing analyses. The results of this study showed that the removal efficiency of PHE reached 72.0% after the addition of anaerobic activated sludge. The incorporation of anaerobic activated sludge and biochar resulted in a 25.3% increase in PHE removal compared to a single soil, suggesting that the combination of bioaugmentation and biostimulation can have a synergistic effect on the anaerobic biodegradation of PHE in contaminated soils. The results of sequencing analysis further indicated that the introduction of an exogenous microbial community changed the dominant genera associated with PHE degradation and introduced methanogenic archaea, which enriched the metabolic pathways of the carbon cycle in the system. On this basis, the addition of biochar resulted in higher anaerobic microbial community diversity, functional dominant species were enriched, and the direct interspecies electron transfer (DIET) process between electroactive bacteria (Bacteroides, f_Geobacteraceae) and Methanosaeta was facilitated, which accelerated the degradation of PHE by anaerobic microbial communities. The results of this study provide regulatory tools and basic data support for enhanced bioremediation of PAHs in flooded soils.

Leaching of biochar at different stages of anaerobic digestion: Leaching behaviors, leachate chemodiversity and effects

Biochar addition is recognized as a promising way to increase the potential of anaerobic digestion. However, few studies have investigated the interaction between biochar leaching in complex, practical anaerobic digestion systems and the negative effects that biochar leachate may have on the anaerobic digestion process. The traditional measurement methods are relatively simple, but may underestimate the complexity of dissolved organic matter in biochar leachate. In this study, the leaching behaviors of biochar and the influence of biochar leachate on anaerobic digestion were studied at the molecular level by applying ultra-performance liquid chromatography online coupled with a hybrid quadrupole orbitrap mass spectrometer (UPLC Orbitrap MS/MS) test analysis and various molecular information-based analytical methods such as van Krevelen diagrams and by conducting a series of anaerobic digestion experiments. Dissolved organic matter leaching was higher in buffered saline solution and methanol than in ultrapure water and the dissolved organic matter mainly consisted of lipids and proteins. In the lactic acid solution, a large amount of dissolved organic matter was leached from the biochar, dominated by low molecular weight dissolved organic matter such as CHO and CHON, and the lower the pH, the more was leached, suggesting that lactic acid may have an inhibitory effect on anaerobic digestion. The maximum methane production amount and microbial abundance especially key methanogenic archaea such as Methanosaeta in the sample with biochar leachate addition were lower than those for the other samples. This meant that the dissolved organic matter in the biochar leachate might inhibit the growth of microorganisms and adversely affect the acetic acid methanogenesis process. The results from this study showed that a greater number of substances with more complex structures were leached out than had been detected in previous studies. When applying biochar to anaerobic digestion in the future, it is recommended that biochar is leached first to remove toxic and detrimental substances to prevent possible inhibitory effects.

Evaluating the potential of different bioaugmented strains to enhance methane production during thermophilic anaerobic digestion of food waste

Bioaugmentation technology for improving the performance of thermophilic anaerobic digestion (TAD) of food waste (FW) treatment is gaining more attention. In this study, four thermophilic strains (Ureibacillus suwonensis E11, Clostridium thermopalmarium HK1, Bacillus thermoamylovorans Y25 and Caldibacillus thermoamylovorans QK5) were inoculated in the TAD of FW system, and the biochemical methane potential (BMP) batch study was conducted to assess the potential of different bioaugmented strains to enhance methane production. The results showed that the cumulative methane production in groups inoculated with E11, HK1, Y25 and QK5 improved by 2.05%, 14.54%, 19.79% and 9.17%, respectively, compared with the control group with no inoculation. Moreover, microbial community composition analysis indicated that the relative abundance of the main hydrolytic bacteria and/or methanogenic archaea was increased after bioaugmentation, and the four strains successfully became representative bacterial biomarkers in each group. The four strains enhanced methane production by strengthening starch, sucrose, galactose, pyruvate and methane metabolism functions. Further, the correlation networks demonstrated that the representative bacterial genera had positive correlations with the differential metabolic functions in each bioaugmentation group. This study provides new insights into the TAD of FW with bioaugmented strains.

Microbial coalbed gas and its geological formation process in the Jimsar region of the southern Junggar Basin, NW China

The southern Junggar Basin is abundant in coalbed gas (CBG) resources, with the Jimsar region in the southern part of the basin emerging as a potential CBG development demonstration area. However, further development of the CBG in this region is restricted by the lack of understanding of the genesis and gas reservoir formation mechanisms of the CBG. This study systematically collected CBG samples and associated water samples from the Jimsar region. Stable isotope, hydrogeochemical, and 16S ribosomal ribonucleic acid (rRNA) sequencing studies were carried out. The results demonstrate that the CBG is the primary microbial gas, and CO2 reduction is its formation pathway. Hydrogeochemical analysis shows that the coalbed water receives recharge from meteoric or surface water. The water environment is relatively active (open system). The total dissolved organic carbon (TDOC) in the coalbed water is primarily from coal, and the supply and consumption of the total dissolved nitrogen (TDN) may be relatively balanced. The dissolution of CO2 is the main source of the dissolved inorganic carbon (DIC). Furthermore, the positive δ13CDIC values and high alkalinity indicate microbial methanogenesis. Microbial community structure analysis revealed that the archaea are hydrogenotrophic methanogens in the Jimsar region. The microbial basis for the production of microbial gas is provided by the existence of hydrolytic, acidogenic, and hydrogen-producing acetogenic bacteria as well as methanogenic archaea. Natural gas genetic identification diagrams and detection of methane-oxidizing bacteria (Methylomicrobium) reveal that the CBG may be affected by slight oxidation. In addition, the methanogenic effect is much greater than the methane oxidation. Finally, based on these results and the regional tectonic background, the CBG enrichment and accumulation mode in the Jimsar region was established. This mode can be divided into three geological stages: burial and coalification, surface water recharge and microbial gas generation after uplift and denudation, and CBG accumulation controlled by reverse faults and hydrodynamic sealing. This study enriches the geological theory of CBG accumulation in the Jimsar region and provides guidance for the subsequent exploration and development of the CBG.