Methanooceanicella nereidis gen. nov., sp. nov., the first oceanic Methanocellaceae methanogen, isolated from potential methane hydrate bearing area offshore southwestern Taiwan.

A novel mesophilic, hydrogenotrophic methanogen, strain CWC-04T, was obtained from a sediment sample extracted from a gravity core retrieved at station 22 within the KP-9 area off the southwestern coast of Taiwan during the ORIII-1368 cruise in 2009. Cells of strain CWC-04T were rod-shaped, 1.4-2.9 µm long by 0.5-0.6 µm wide, and occurred singly. Strain CWC-04Tutilized formate, H2/CO2, 2-propanol/CO2 or 2-butanol/CO2 as catabolic substrates. The optimal growth conditions were 42 °C, 0.17 M NaCl and pH 5.35. The genomic DNA G+C content calculated from the genome sequence of strain CWC-04T was 46.19 mol%. Phylogenetic analysis of 16S rRNA gene revealed that strain CWC-04T is affiliated with the genus Methanocella. The 16S rRNA gene sequences similarities within strains Methanocella arvoryzae MRE50T, Methanocella paludicola SANAET and Methanocella conradii HZ254T were 93.7, 93.0 and 91.3 %, respectively. In addition, the optical density of CWC-04T culture dropped abruptly upon entering the late-log growth phase, with virus-like particles (150 nm in diameter) being observed on and around the cells. This observation suggests that strain CWC-04T harbours a lytic virus. Based on these phenotypic, phylogenetic and genomic results, we propose that strain CWC-04T represents a novel species of a novel genus in the family Methanocellaceae, for which the name Methanooceanicella nereidis gen. nov., sp. nov. is proposed. The type strain is CWC-04T (=BCRC AR10050T=NBRC 113165T).

The Selenoproteome as a Dynamic Response Mechanism to Oxidative Stress in Hydrogenotrophic Methanogenic Communities.

Methanogenesis is a critical process in the carbon cycle that is applied industrially in anaerobic digestion and biogas production. While naturally occurring in diverse environments, methanogenesis requires anaerobic and reduced conditions, although varying degrees of oxygen tolerance have been described. Microaeration is suggested as the next step to increase methane production and improve hydrolysis in digestion processes; therefore, a deeper understanding of the methanogenic response to oxygen stress is needed. To explore the drivers of oxygen tolerance in methanogenesis, two parallel enrichments were performed under the addition of H2/CO2 in an environment without reducing agents and in a redox-buffered environment by adding redox mediator 9,10-anthraquinone-2,7-disulfonate disodium. The cellular response to oxidative conditions is mapped using proteomic analysis. The resulting community showed remarkable tolerance to high-redox environments and was unperturbed in its methane production. Next to the expression of pathways to mitigate reactive oxygen species, the higher redox potential environment showed an increased presence of selenocysteine and selenium-associated pathways. By including sulfur-to-selenium mass shifts in a proteomic database search, we provide the first evidence of the dynamic and large-scale incorporation of selenocysteine as a response to oxidative stress in hydrogenotrophic methanogenesis and the presence of a dynamic selenoproteome.

Tartrate fermentation with H2 production by a new member of Sporomusaceae enriched from rice paddy soil.

In rice paddies, soil and plant-derived organic matter are degraded anaerobically to methane (CH4), a powerful greenhouse gas. The highest rate of methane emission occurs during the reproductive stage of the plant when mostly dicarboxylic acids are exudated by the roots. The emission of methane at this stage depends largely on the cooperative interaction between dicarboxylic acid-fermenting bacteria and methanogenic archaea in the rhizosphere. The fermentation of tartrate, one of the major acids exudated, has been scarcely explored in rice paddy soils. In this work, we characterized an anaerobic consortium from rice paddy soil composed of four bacterial strains, whose principal member (LT8) can ferment tartrate, producing H2 and acetate. Tartrate fermentation was accelerated by co-inoculation with a hydrogenotrophic methanogen. The assembled genome of LT8 possesses a Na+-dependent oxaloacetate decarboxylase and shows that this bacterium likely invests part of the H2 produced to reduce NAD(P)+ to assimilate C from tartrate. The phylogenetic analysis of the 16S rRNA gene, the genome-based classification as well as the average amino acid identity (AAI) indicated that LT8 belongs to a new genus within the Sporomusaceae family. LT8 shares a few common features with its closest relatives, for which tartrate degradation has not been described. LT8 is limited to a few environments but is more common in rice paddy soils, where it might contribute to methane emissions from root exudates.IMPORTANCEThis is the first report of the metabolic characterization of a new anaerobic bacterium able to degrade tartrate, a compound frequently associated with plants, but rare as a microbial metabolite. Tartrate fermentation by this bacterium can be coupled to methanogenesis in the rice rhizosphere where tartrate is mainly produced at the reproductive stage of the plant, when the maximum methane rate emission occurs. The interaction between secondary fermentative bacteria, such as LT8, and methanogens could represent a fundamental step in exploring mitigation strategies for methane emissions from rice fields. Possible strategies could include controlling the activity of these secondary fermentative bacteria or selecting plants whose exudates are more difficult to ferment.

Expanded phylogeny of extremely halophilic archaea shows multiple independent adaptations to hypersaline environments.

Extremely halophilic archaea (Haloarchaea, Nanohaloarchaeota, Methanonatronarchaeia and Halarchaeoplasmatales) thrive in saturating salt concentrations where they must maintain osmotic equilibrium with their environment. The evolutionary history of adaptations enabling salt tolerance remains poorly understood, in particular because the phylogeny of several lineages is conflicting. Here we present a resolved phylogeny of extremely halophilic archaea obtained using improved taxon sampling and state-of-the-art phylogenetic approaches designed to cope with the strong compositional biases of their proteomes. We describe two uncultured lineages, Afararchaeaceae and Asbonarchaeaceae, which break the long branches at the base of Haloarchaea and Nanohaloarchaeota, respectively. We obtained 13 metagenome-assembled genomes (MAGs) of these archaea from metagenomes of hypersaline aquatic systems of the Danakil Depression (Ethiopia). Our phylogenomic analyses including these taxa show that at least four independent adaptations to extreme halophily occurred during archaeal evolution. Gene-tree/species-tree reconciliation suggests that gene duplication and horizontal gene transfer played an important role in this process, for example, by spreading key genes (such as those encoding potassium transporters) across extremely halophilic lineages.

Genome-based classification of the family Haloferacaceae and description of five novel species of Halobaculum.

The taxonomic status of some species of Halobellus, Haloferax, Halogranum, and Haloplanus within the family Haloferacaceae was elucidated by phylogenetic, phylogenomic, and comparative genomic analyses. The relative species of each genus should constitute a single species based on the overall genome-related indexes proposed for species demarcation. The cutoff values of AAI (72.1%), ANI (82.2%), and rpoB' gene similarity (90.7%) were proposed to differentiate genera within the family Haloferacaceae. According to these standards, a novel genus related to the genus Halobaculum was proposed to accommodate Halobaculum halophilum Gai3-2 T and Halobaculum salinum NJ-3-1 T. Five halophilic archaeal strains, DT31T, DT55T, DT92T, SYNS20T, and YSMS11T, isolated from a tidal flat and a marine solar saltern in China, were subjected to polyphasic classification. The phenotypic, phylogenetic, phylogenomic, and comparative genomic analyses revealed that strains DT31T (= CGMCC 1.18923 T = JCM 35417 T), DT55T (= CGMCC 1.19048 T = JCM 36147 T), DT92T (= CGMCC 1.19057 T = JCM 36148 T), SYNS20T (= CGMCC 1.62628 T = JCM 36154 T), and YSMS11T (= CGMCC 1.18927 T = JCM 34912 T) represent five novel species of the genus Halobaculum, for which the names, Halobaculum lipolyticum sp. nov., Halobaculum marinum sp. nov., Halobaculum litoreum sp. nov., Halobaculum halobium sp. nov., and Halobaculum limi sp. nov., are proposed.

Methylotrophic methanogenesis in the Archaeoglobi revealed by cultivation of Ca. Methanoglobus hypatiae from a Yellowstone hot spring.

Over the past decade, environmental metagenomics and polymerase chain reaction-based marker gene surveys have revealed that several lineages beyond just a few well-established groups within the Euryarchaeota superphylum harbor the genetic potential for methanogenesis. One of these groups are the Archaeoglobi, a class of thermophilic Euryarchaeota that have long been considered to live non-methanogenic lifestyles. Here, we enriched Candidatus Methanoglobus hypatiae, a methanogen affiliated with the family Archaeoglobaceae, from a hot spring in Yellowstone National Park. The enrichment is sediment-free, grows at 64-70°C and a pH of 7.8, and produces methane from mono-, di-, and tri-methylamine. Ca. M. hypatiae is represented by a 1.62 Mb metagenome-assembled genome with an estimated completeness of 100% and accounts for up to 67% of cells in the culture according to fluorescence in situ hybridization. Via genome-resolved metatranscriptomics and stable isotope tracing, we demonstrate that Ca. M. hypatiae expresses methylotrophic methanogenesis and energy-conserving pathways for reducing monomethylamine to methane. The detection of Archaeoglobi populations related to Ca. M. hypatiae in 36 geochemically diverse geothermal sites within Yellowstone National Park, as revealed through the examination of previously published gene amplicon datasets, implies a previously underestimated contribution to anaerobic carbon cycling in extreme ecosystems.

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.

Microscopic and metatranscriptomic analyses revealed unique cross-domain parasitism between phylum Candidatus Patescibacteria/candidate phyla radiation and methanogenic archaea in anaerobic ecosystems.

To verify whether members of the phylum Candidatus Patescibacteria parasitize archaea, we applied cultivation, microscopy, metatranscriptomic, and protein structure prediction analyses on the Patescibacteria-enriched cultures derived from a methanogenic bioreactor. Amendment of cultures with exogenous methanogenic archaea, acetate, amino acids, and nucleoside monophosphates increased the relative abundance of Ca. Patescibacteria. The predominant Ca. Patescibacteria were families Ca. Yanofskyibacteriaceae and Ca. Minisyncoccaceae, and the former showed positive linear relationships (r2 ≥ 0.70) Methanothrix in their relative abundances, suggesting related growth patterns. Methanothrix and Methanospirillum cells with attached Ca. Yanofskyibacteriaceae and Ca. Minisyncoccaceae, respectively, had significantly lower cellular activity than those of the methanogens without Ca. Patescibacteria, as extrapolated from fluorescence in situ hybridization-based fluorescence. We also observed that parasitized methanogens often had cell surface deformations. Some Methanothrix-like filamentous cells were dented where the submicron cells were attached. Ca. Yanofskyibacteriaceae and Ca. Minisyncoccaceae highly expressed extracellular enzymes, and based on structural predictions, some contained peptidoglycan-binding domains with potential involvement in host cell attachment. Collectively, we propose that the interactions of Ca. Yanofskyibacteriaceae and Ca. Minisyncoccaceae with methanogenic archaea are parasitisms.IMPORTANCECulture-independent DNA sequencing approaches have explored diverse yet-to-be-cultured microorganisms and have significantly expanded the tree of life in recent years. One major lineage of the domain Bacteria, Ca. Patescibacteria (also known as candidate phyla radiation), is widely distributed in natural and engineered ecosystems and has been thought to be dependent on host bacteria due to the lack of several biosynthetic pathways and small cell/genome size. Although bacteria-parasitizing or bacteria-preying Ca. Patescibacteria have been described, our recent studies revealed that some lineages can specifically interact with archaea. In this study, we provide strong evidence that the relationship is parasitic, shedding light on overlooked roles of Ca. Patescibacteria in anaerobic habitats.

Incorporating saline microalgae biomass in anaerobic digester treating sewage sludge: Impact on performance and microbial populations.

The aim of this study was to acclimate anaerobic prokaryotes to saline microalgae biomass. Semi-continuous experiments were conducted using two 1.5 L mesophilic reactors for 10 weeks, (hydraulic retention time of 21 days). The first reactor was solely fed with sewage sludge (control), while the second received a mixture of sewage sludge and microalgal biomass (80/20 %w/w) cultivated at 70 g·L-1 salinity. The in-reactor salinity reached after the acclimation phase was 14 g·L-1. Biomethane production was comparable between the control and acclimated reactors (205 ± 29 NmLMethane·gVolatileSolids-1). Salinity tolerance assessment of methanogenic archaea revealed that salinity causing 50% inhibition of methane production increased from 10 to 27 g·L-1 after acclimation. Microbial diversity analyses revealed notable changes in methanogenic archaea populations during co-digestion of saline microalgae biomass, particularly methylotrophic (+27%) and acetotrophic (-26%) methanogens. This study has highlighted the possibility of treating efficiently saline microalgae in co-digestion with sewage sludge in future industrial biogas plants.

Extracellular vesicle formation in Euryarchaeota is driven by a small GTPase.

Since their discovery, extracellular vesicles (EVs) have changed our view on how organisms interact with their extracellular world. EVs are able to traffic a diverse array of molecules across different species and even domains, facilitating numerous functions. In this study, we investigate EV production in Euryarchaeota, using the model organism Haloferax volcanii. We uncover that EVs enclose RNA, with specific transcripts preferentially enriched, including those with regulatory potential, and conclude that EVs can act as an RNA communication system between haloarchaea. We demonstrate the key role of an EV-associated small GTPase for EV formation in H. volcanii that is also present across other diverse evolutionary branches of Archaea. We propose the name, ArvA, for the identified family of archaeal vesiculating GTPases. Additionally, we show that two genes in the same operon with arvA (arvB and arvC) are also involved in EV formation. Both, arvB and arvC, are closely associated with arvA in the majority of other archaea encoding ArvA. Our work demonstrates that small GTPases involved in membrane deformation and vesiculation, ubiquitous in Eukaryotes, are also present in Archaea and are widely distributed across diverse archaeal phyla.

Viral potential to modulate microbial methane metabolism varies by habitat.

Methane is a potent greenhouse gas contributing to global warming. Microorganisms largely drive the biogeochemical cycling of methane, yet little is known about viral contributions to methane metabolism (MM). We analyzed 982 publicly available metagenomes from host-associated and environmental habitats containing microbial MM genes, expanding the known MM auxiliary metabolic genes (AMGs) from three to 24, including seven genes exclusive to MM pathways. These AMGs are recovered on 911 viral contigs predicted to infect 14 prokaryotic phyla including Halobacteriota, Methanobacteriota, and Thermoproteota. Of those 24, most were encoded by viruses from rumen (16/24), with substantially fewer by viruses from environmental habitats (0-7/24). To search for additional MM AMGs from an environmental habitat, we generate metagenomes from methane-rich sediments in Vrana Lake, Croatia. Therein, we find diverse viral communities, with most viruses predicted to infect methanogens and methanotrophs and some encoding 13 AMGs that can modulate host metabolisms. However, none of these AMGs directly participate in MM pathways. Together these findings suggest that the extent to which viruses use AMGs to modulate host metabolic processes (e.g., MM) varies depending on the ecological properties of the habitat in which they dwell and is not always predictable by habitat biogeochemical properties.

Methane production and lignocellulosic degradation of wastes from rice, corn and sugarcane by natural anaerobic fungi-methanogens co-culture.

Biomass from agriculture, forestry, and urban wastes is a potential renewable organic resource for energy generation. Many investigations have demonstrated that anaerobic fungi and methanogens could be co-cultured to degrade lignocellulose for methane generation. Thus, this study aimed to evaluate the effect of natural anaerobic fungi-methanogens co-culture on the methane production and lignocellulosic degradation of wastes from rice, corn and sugarcane. Hu sheep rumen digesta was used to develop a natural anaerobic fungi-methanogen co-culture. The substrates were rice straw (RS), rich husk (RH), corn stover (CS), corn cobs (CC), and sugarcane baggage (SB). Production of total gas and methane, metabolization rate of reducing sugar, glucose, and xylose, digestibility of hemicellulose and cellulose, activity of carboxymethylcellulase and xylanase, and concentrations of total acid and acetate were highest (P < 0.05) in CC, moderate (P < 0.05) in RS and CS, and lowest (P < 0.05) in SB and RH. The pH, lactate and ethanol were lowest (P < 0.05) in CC, moderate (P < 0.05) in RS and CS, and lowest (P < 0.05) SB and RH. Formate was lowest (P < 0.05) in CC, RS and CS, moderate (P < 0.05) in SB, and lowest (P < 0.05) in RH. Therefore, this study indicated that the potential of methane production and lignocellulosic degradation by natural anaerobic fungi-methanogens co-culture were highest in CC, moderate in RS and CS, and lowest in SB and RH.

Effect of micro-aeration on syntrophic and methanogenic activity in anaerobic sludge.

Micro-aeration was shown to improve anaerobic digestion (AD) processes, although oxygen is known to inhibit obligate anaerobes, such as syntrophic communities of bacteria and methanogens. The effect of micro-aeration on the activity and microbial interaction in syntrophic communities, as well as on the potential establishment of synergetic relationships with facultative anaerobic bacteria (FAB) or aerobic bacteria (AB), was investigated. Anaerobic sludge was incubated with ethanol and increasing oxygen concentrations (0-5% in the headspace). Assays with acetate or H2/CO2 (direct substrates for methanogens) were also performed. When compared with the controls (0% O2), oxygen significantly decreased substrate consumption and initial methane production rate (MPR) from acetate or H2/CO2. At 0.5% O2, MPR from these substrates was inhibited 30-40%, and close to 100% at 5% O2. With ethanol, significant inhibition (>36%) was only observed for oxygen concentrations higher than 2.5%. Oxygen was consumed in the assays, pointing to the stimulation of AB/FAB by ethanol, which helped to protect the syntrophic consortia under micro-aerobic conditions. This highlights the importance of AB/FAB in maintaining functional and resilient syntrophic communities, which is relevant for real AD systems (in which vestigial O2 amounts are frequently present), as well as for AD systems using micro-aeration as a process strategy. KEY POINTS: •Micro-aeration impacts syntrophic communities of bacteria and methanogens. •Oxygen stimulates AB/FAB, maintaining functional and resilient consortia. •Micro-aeration studies are critical for systems using micro-aeration as a process strategy.

Members of the class Candidatus Ordosarchaeia imply an alternative evolutionary scenario from methanogens to haloarchaea.

The origin of methanogenesis can be traced to the common ancestor of non-DPANN archaea, whereas haloarchaea (or Halobacteria) are believed to have evolved from a methanogenic ancestor through multiple evolutionary events. However, due to the accelerated evolution and compositional bias of proteins adapting to hypersaline habitats, Halobacteria exhibit substantial evolutionary divergence from methanogens, and the identification of the closest methanogen (either Methanonatronarchaeia or other taxa) to Halobacteria remains a subject of debate. Here, we obtained five metagenome-assembled genomes with high completeness from soda-saline lakes on the Ordos Plateau in Inner Mongolia, China, and we proposed the name Candidatus Ordosarchaeia for this novel class. Phylogenetic analyses revealed that Ca. Ordosarchaeia is firmly positioned near the median position between the Methanonatronarchaeia and Halobacteria-Hikarchaeia lineages. Functional predictions supported the transitional status of Ca. Ordosarchaeia with the metabolic potential of nonmethanogenic and aerobic chemoheterotrophy, as did remnants of the gene sequences of methylamine/dimethylamine/trimethylamine metabolism and coenzyme M biosynthesis. Based on the similarity of the methyl-coenzyme M reductase genes mcrBGADC in Methanonatronarchaeia with the phylogenetically distant methanogens, an alternative evolutionary scenario is proposed, in which Methanonatronarchaeia, Ca. Ordosarchaeia, Ca. Hikarchaeia, and Halobacteria share a common ancestor that initially lost mcr genes. However, certain members of Methanonatronarchaeia subsequently acquired mcr genes through horizontal gene transfer from distantly related methanogens. This hypothesis is supported by amalgamated likelihood estimation, phylogenetic analysis, and gene arrangement patterns. Altogether, Ca. Ordosarchaeia genomes clarify the sisterhood of Methanonatronarchaeia with Halobacteria and provide new insights into the evolution from methanogens to haloarchaea.

When polyethylene terephthalate microplastics meet Perfluorooctane sulfonate in thermophilic biogas upgrading system: Their effect on methanogenesis.

Microplastics (MPs) and Perfluorooctane sulfonate (PFOS) are two hard-biodegradable pollutants widely existing in the waste streams treated by anaerobic digestion. However, their synergistic effect on methanogenic metabolism is still unknown. This study investigated the impact of polyethylene terephthalate (PET) MPs alone and co-existing with PFOS on CO2 conversion to CH4 in a thermophilic biogas upgrading system. The results showed that either PET MPs addition alone or coexisting with PFOS improved the ultimate CH4 percentage and increased CO2 utilization rate. When Fe0 was added into the reactors with PET to enhance the interspecies electron transfer, a potential defluorination was observed with a defluorination rate of 15.88 ± 1.53%. Exposure of the reactor to PFOS of 300 µg/L could change the methanogenic pathway, resulting in a newly emerged Methanomassiliicoccus with dominance of 16%. Furthermore, under the exposure of PFOS, the number of predicted genes regulating enzymes in methanogenic steps from CO2 increased. These results suggest that the co-existence of PET MPs and PFOS will not inhibit the activity of hydrotrophic methanogenes, and a portion of PFOS may be biodegraded during the methanogenesis under Fe0 regulation.

Biochar mitigates the adverse effects of antimony on methanogenic activity: role as methane production-enhancer.

Antimony, extensively used in energy applications, poses toxicity and contamination concerns, especially in anaerobic environments where its impact on microbial activity is poorly understood. Emerging remedies, like biochar, show promise in soil and water treatment. This study investigates biochar's influence on methanogenic activity under Sb(V) and Sb(III) stress using anaerobic sludge as inoculum and lactate as the carbon source. Sb(III) and Sb(V) were introduced at varied concentrations (5-80 mg/L), with or without biochar, monitoring changes in biogas production, pH, Sb, and lactate levels over time. Experiments with Sb(V) also involved calculating mass balance and electron distribution. Results showcased the following significant enhancements: biochar notably improved COD removal and biogas production in Sb(III) spiked conditions, up to 5-fold and 2-fold increases, respectively. Sb(III) removal reached up to 99% with biochar, while in high Sb(V) concentrations, biochar reduced the adverse effect on biogas production by 96%. Adsorption capacities favored biomass (60.96 mg Sb(III)/gVSS, and 22.4 mg Sb(V)/gVSS) over biochar (3.33 mg Sb(III)/g, and 1.61 mg Sb(V)/g) for both Sb species. This study underscores biochar's potential to mitigate metalloid impact on methanogenic activity while aiding Sb removal from liquid phase, suggesting promising implications for remediation and methane production enhancement strategies.

Anaerobic co-digestion of corn straw, sewage sludge and fresh leachate: Focusing on synergistic/antagonistic effects and microbial mechanisms.

Effects of sewage sludge (SS) and fresh leachate (FL) addition on corn straw (CS) digestion and underlying mechanisms were investigated. Co-digestion of CS, SS and FL significantly increased cumulative methane production by 7.2-61.1%. Further analysis revealed that co-digestion acted mainly on slowly degradable substrates and exerted dual effects on methane production potential, which was closely related to the volatile solids (VS) content. Antagonistic effects of co-digestion resulted from the dominance of norank_c_Bathyarchaeia, a mixotrophic methanogen that may generate methane inefficiently and consume existing methane. The synergistic enhancement of methane production (0.7-12.7%) was achieved in co-digestion with 33.5-45.5% of total VS added as SS and FL. Co-digestion with more balanced nutrients and higher buffering capacity enriched Actinobacteriota, Firmicutes, and Synergistota, thereby facilitating the substrate degradation. Furthermore, the predominant acetoclastic methanogens, increased hydrogenotrophic methanogens, and decreased methylotrophic methanogens in the digester combined to prompt the synergy.

Distinct microbial hydrogen and reductant disposal pathways explain interbreed variations in ruminant methane yield.

Ruminants are essential for global food security, but these are major sources of the greenhouse gas methane. Methane yield is controlled by the cycling of molecular hydrogen (H2), which is produced during carbohydrate fermentation and is consumed by methanogenic, acetogenic, and respiratory microorganisms. However, we lack a holistic understanding of the mediators and pathways of H2 metabolism and how this varies between ruminants with different methane-emitting phenotypes. Here, we used metagenomic, metatranscriptomic, metabolomics, and biochemical approaches to compare H2 cycling and reductant disposal pathways between low-methane-emitting Holstein and high-methane-emitting Jersey dairy cattle. The Holstein rumen microbiota had a greater capacity for reductant disposal via electron transfer for amino acid synthesis and propionate production, catalyzed by enzymes such as glutamate synthase and lactate dehydrogenase, and expressed uptake [NiFe]-hydrogenases to use H2 to support sulfate and nitrate respiration, leading to enhanced coupling of H2 cycling with less expelled methane. The Jersey rumen microbiome had a greater proportion of reductant disposal via H2 production catalyzed by fermentative hydrogenases encoded by Clostridia, with H2 mainly taken up through methanogenesis via methanogenic [NiFe]-hydrogenases and acetogenesis via [FeFe]-hydrogenases, resulting in enhanced methane and acetate production. Such enhancement of electron incorporation for metabolite synthesis with reduced methanogenesis was further supported by two in vitro measurements of microbiome activities, metabolites, and public global microbiome data of low- and high-methane-emitting beef cattle and sheep. Overall, this study highlights the importance of promoting alternative H2 consumption and reductant disposal pathways for synthesizing host-beneficial metabolites and reducing methane production in ruminants.

Microorganism community structure: A characterisation of agrosystems from Madeira Archipelago.

Microbial diversity profoundly influences soil ecosystem functions, making it vital to monitor community dynamics to comprehend its structure. Our study focused on six agrosystems in Madeira Archipelago, analysing bacteria, archaea, fungi and AMF through classical microbiology and molecular techniques. Despite distinct edaphoclimatic conditions and management practices, bacterial structures exhibited similarities, with Alphaproteobacteria at 18%-20%, Bacilli at 11%-18% and Clostridia at 9%-14%. The predominance of copiothrophic groups suggested that soil nutrient content was the driver of these communities. Regarding archaea, the communities changed among sites, and it was evident that agrosystems provided niches for methanogens. The Crenarchaeota varied between 15% and 29%, followed by two classes of Euryarchaeota, Methanomicrobia (17%-25%) and Methanococci (4%-32%). Fungal communities showed consistent composition at the class level but had differing diversity indices due to management practices and soil texture. Sordaryomycetes (21%-28%) and Agaricomycetes (15%-23%) were predominant. Conversely, AMF communities appeared to be also influenced by the agrosystem, with Glomus representing over 50% of the community in all agrosystems. These insights into microbial groups' susceptibilities to environmental conditions are crucial for maintaining healthy soil and predicting climate change effects on agrosystems' productivity, resilience and sustainability. Additionally, our findings enable the development of more robust prediction models for agricultural practices.

Metabolism of novel potential syntrophic acetate-oxidizing bacteria in thermophilic methanogenic chemostats.

Acetate is a major intermediate in the anaerobic digestion of organic waste to produce CH4. In methanogenic systems, acetate degradation is carried out by either acetoclastic methanogenesis or syntrophic degradation by acetate oxidizers and hydrogenotrophic methanogens. Due to challenges in the isolation of syntrophic acetate-oxidizing bacteria (SAOB), the diversity and metabolism of SAOB and the mechanisms of their interactions with methanogenic partners are not fully characterized. In this study, the in situ activity and metabolic characteristics of potential SAOB and their interactions with methanogens were elucidated through metagenomics and metatranscriptomics. In addition to the reported SAOB classified in the genera Tepidanaerobacter, Desulfotomaculum, and Thermodesulfovibrio, we identified a number of potential SAOB that are affiliated with Clostridia, Thermoanaerobacteraceae, Anaerolineae, and Gemmatimonadetes. The potential SAOB possessing the glycine-mediated acetate oxidation pathway dominates SAOB communities. Moreover, formate appeared to be the main product of the acetate degradation by the most active potential SAOB. We identified the methanogen partner of these potential SAOB in the acetate-fed chemostat as Methanosarcina thermophila. The dominated potential SAOB in each chemostat had similar metabolic characteristics, even though they were in different fatty-acid-fed chemostats. These novel syntrophic lineages are prevalent and may play critical roles in thermophilic methanogenic reactors. This study expands our understanding of the phylogenetic diversity and in situ biological functions of uncultured syntrophic acetate degraders and presents novel insights into how they interact with methanogens.IMPORTANCECombining reactor operation with omics provides insights into novel uncultured syntrophic acetate degraders and how they perform in thermophilic anaerobic digesters. This improves our understanding of syntrophic acetate degradation and contributes to the background knowledge necessary to better control and optimize anaerobic digestion processes.