Isolation and characterisation of novel Methanocorpusculum species indicates the genus is ancestrally host-associated.

BACKGROUND: With an increasing interest in the manipulation of methane produced from livestock cultivation, the microbiome of Australian marsupials provides a unique ecological and evolutionary comparison with 'low-methane' emitters. Previously, marsupial species were shown to be enriched for novel lineages of Methanocorpusculum, as well as Methanobrevibacter, Methanosphaera, and Methanomassiliicoccales. Despite sporadic reports of Methanocorpusculum from stool samples of various animal species, there remains little information on the impacts of these methanogens on their hosts. RESULTS: Here, we characterise novel host-associated species of Methanocorpusculum, to explore unique host-specific genetic factors and their associated metabolic potential. We performed comparative analyses on 176 Methanocorpusculum genomes comprising 130 metagenome-assembled genomes (MAGs) recovered from 20 public animal metagenome datasets and 35 other publicly available Methanocorpusculum MAGs and isolate genomes of host-associated and environmental origin. Nine MAGs were also produced from faecal metagenomes of the common wombat (Vombatus ursinus) and mahogany glider (Petaurus gracilis), along with the cultivation of one axenic isolate from each respective animal; M. vombati (sp. nov.) and M. petauri (sp. nov.). CONCLUSIONS: Through our analyses, we substantially expand the available genetic information for this genus by describing the phenotypic and genetic characteristics of 23 host-associated species of Methanocorpusculum. These lineages display differential enrichment of genes associated with methanogenesis, amino acid biosynthesis, transport system proteins, phosphonate metabolism, and carbohydrate-active enzymes. These results provide insights into the differential genetic and functional adaptations of these novel host-associated species of Methanocorpusculum and suggest that this genus is ancestrally host-associated.

Reduced sulfide and methane in rising main sewer via calcium peroxide dosing: Insights from microbial physiological characteristics, metabolisms and community traits.

Although the biocidal effect of calcium peroxide (CaO2) has attracted increasing attention in wastewater and sludge management, its potential in the reduction of sulfide and methane from sewer is not tapped. This study aims to fill this gap through the long-term operated sewer reactors. Results showed one-time dose of 0.2% (w/v) CaO2 with 12-h exposure decreased the average sulfide and methane production by 80% during one week. The electron paramagnetic resonance and free radical quenching tests indicated free radicals from CaO2 decomposing posed a major contribution on sewer biofilms (•OH>•O2->alkali). Mechanistic analysis revealed extracellular polymeric matrix breakdown (e.g., protein secondary structure) and cell membrane damage were caused by the increased lipid peroxidation of cells and exacerbated intracellular reactive oxygen species under CaO2 stress. Moreover, the intracellular metabolic pathways, such as electrons provision and transfer, as well as pivotal enzymatic activities (e.g., APS reductase, sulfite reductase and coenzymes F420) were significantly impaired. RT-qPCR analysis unveiled the absolute abundances of dsrA and mcrA were decreased by 7.53-40.37% and 67.00-74.85%, respectively. Although this study broadens the application scope of CaO2 and provides in-depth understanding of advanced oxidation-based technology in sewer management, the pipe scale risk due to the release of calcium ions warrants further investigation.

Effect of propionate, monensin, and saccharomyces cerevisiae or their combination on production and rumen fermentation of holstein steers.

This study aimed to evaluate the effects of calcium propionate (PrCa), PrCa + monensin sodium (PrCa + Mon), and PrCa + Saccharomyces cerevisiae (PrCa + Sc) on the productive performance of Holstein steers. Twenty-four Holstein steers (270.0 ± 25.85 kg) were distributed individually into four treatments of six replicates. The treatments were control (no additives), PrCa (10 g/kg), PrCa + Mon (10 g/kg + 30 mg/kg), and PrCa + Sc (10 g/kg + 12.8 × 109 cfu). The steers were fed for 43 days, and afterwards, nutrient intake and digestibility as well as volatile fatty acids were determined, while the weight gained, feed efficiency, and CH4 production were calculated. Diet of PrCa + Sc had the highest (P < 0.0001) acid detergent fiber intake and propionate acid as well as the nutrient digestibility, with lowest (P < 0.0001) rumen acetic acid, methane, and protozoa concentration versus other diets. In conclusion, dietary inclusion of PrCa + Sc (10 g/kg + 12.8 × 109 cfu) improved nutrient digestibility, rumen fermentation, and reduced methane emission, thereby enhancing the possibility of ecofriendly ruminant farming.

Inhibitory mechanisms on dry anaerobic digestion: Ammonia, hydrogen and propionic acid relationship.

Inhibitory pathways in dry anaerobic digestion are still understudied and current knowledge on wet processes cannot be easily transferred. This study forced instability in pilot-scale digesters by operating at short retention times (40 and 33 days) in order to understand inhibition pathways over long term operation (145 days). The first sign of inhibition at elevated total ammonia concentrations (8 g/l) was a headspace hydrogen level over the thermodynamic limit for propionic degradation, causing propionic accumulation. The combined inhibitory effect of propionic and ammonia accumulation resulted in further increased hydrogen partial pressures and n-butyric accumulation. The relative abundance of Methanosarcina increased while that of Methanoculleus decreased as digestion deteriorated. It was hypothesized that high ammonia, total solids and organic loading rate inhibited syntrophic acetate oxidisers, increasing their doubling time and resulting in its wash out, which in turn inhibited hydrogenotrophic methanogenesis and shifted the predominant methanogenic pathway towards acetoclastic methanogenesis at free ammonia over 1.5 g/l. C/N increases to 25 and 29 reduced inhibitors accumulation but did not avoid inhibition or the washout of syntrophic acetate oxidising bacteria.

Spatio-temporal variations of activity of nitrate-driven anaerobic oxidation of methane and community structure of Candidatus Methanoperedens-like archaea in sediment of Wuxijiang river.

Nitrate-driven anaerobic oxidation of methane (AOM), catalyzing by Candidatus Methanoperedens-like archaea, is a new addition in the global CH4 cycle. This AOM process acts as a novel pathway for CH4 emission reduction in freshwater aquatic ecosystems; however, its quantitative importance and regulatory factors in riverine ecosystems are nearly unknown. Here, we examined the spatio-temporal changes of the communities of Methanoperedens-like archaea and nitrate-driven AOM activity in sediment of Wuxijiang River, a mountainous river in China. These archaeal community composition varied significantly among reaches (upper, middle, and lower reaches) and between seasons (winter and summer), but their mcrA gene diversity showed no significant spatial or temporal variations. The copy numbers of Methanoperedens-like archaeal mcrA genes were 1.32 × 105-2.47 × 107 copies g-1 (dry weight), and the activity of nitrate-driven AOM was 0.25-1.73 nmol CH4 g-1 (dry weight) d-1, which could potentially reduce 10.3% of CH4 emissions from rivers. Significant spatio-temporal variations of mcrA gene abundance and nitrate-driven AOM activity were found. Both the gene abundance and activity increased significantly from upper to lower reaches in both seasons, and were significantly higher in sediment collected in summer than in winter. In addition, the variations of Methanoperedens-like archaeal communities and nitrate-driven AOM activity were largely impacted by the sediment temperature, NH4+ and organic carbon contents. Taken together, both time and space scales need to be considered for better evaluating the quantitative importance of nitrate-driven AOM in reducing CH4 emissions from riverine ecosystems.

Bacterial aerobic methane cycling by the marine sponge-associated microbiome.

BACKGROUND: Methanotrophy by the sponge-hosted microbiome has been mainly reported in the ecological context of deep-sea hydrocarbon seep niches where methane is either produced geothermically or via anaerobic methanogenic archaea inhabiting the sulfate-depleted sediments. However, methane-oxidizing bacteria from the candidate phylum Binatota have recently been described and shown to be present in oxic shallow-water marine sponges, where sources of methane remain undescribed. RESULTS: Here, using an integrative -omics approach, we provide evidence for sponge-hosted bacterial methane synthesis occurring in fully oxygenated shallow-water habitats. Specifically, we suggest methane generation occurs via at least two independent pathways involving methylamine and methylphosphonate transformations that, concomitantly to aerobic methane production, generate bioavailable nitrogen and phosphate, respectively. Methylphosphonate may be sourced from seawater continuously filtered by the sponge host. Methylamines may also be externally sourced or, alternatively, generated by a multi-step metabolic process where carnitine, derived from sponge cell debris, is transformed to methylamine by different sponge-hosted microbial lineages. Finally, methanotrophs specialized in pigment production, affiliated to the phylum Binatota, may provide a photoprotective function, closing a previously undescribed C1-metabolic loop that involves both the sponge host and specific members of the associated microbial community. CONCLUSION: Given the global distribution of this ancient animal lineage and their remarkable water filtration activity, sponge-hosted methane cycling may affect methane supersaturation in oxic coastal environments. Depending on the net balance between methane production and consumption, sponges may serve as marine sources or sinks of this potent greenhouse gas. Video Abstract.

Effects of Thymbra capitata essential oil on in vitro fermentation end-products and ruminal bacterial communities.

An in vitro trial was carried out to investigate the effects of natural Thymbra capitata essential oil (NEO) and its main compounds [including carvacrol, p-cymene, γ-terpinene given alone or in a synthetic combination (SEO)] on ruminal fermentation and the bacterial community using batch cultures inoculated with ruminal digesta and incubating two different basal diets [high-forage (F) and high-concentrate (C) diet]. After 24 h of incubation, primary fermentation end-products [gas, methane, volatile fatty acids (VFAs) and ammonia] and rumen microbial diversity were determined. NEO reduced the total VFA concentration (P < 0.05) only in the C diet. In contrast, SEO and carvacrol decreased the total VFA concentration (P < 0.05) only in the F diet. Methane production was not affected (P > 0.05) by any of the experimental treatments or diets evaluated. Microbial diversity analysis showed only a moderate effect of carvacrol and SEO on 13 genera, including, mainly, Atopobium and Blautia (involved in subacute ruminal acidosis) or Candidatus Saccharimonas (related to laminitis). In conclusion, T. capitata EO has a limited potential to attain nutritional or environmental benefits, but further research should be carried out to clarify its effects on animal health and microbial food safety.

Coupling methane and bioactive polysaccharide recovery from wasted activated sludge: A sustainable strategy for sludge treatment.

Bioactive polysaccharides (PSs) are valuable resources that can be extracted from waste activated sludge (WAS). The PS extraction process causes cell lysis that may enhance hydrolytic processes during anaerobic digestion (AD) and thus increase the methane production. Thus, coupling PSs and methane recovery from WAS could be an efficient and sustainable sludge treatment. In present study, we comprehensively evaluated this novel process from the efficiencies of different coupling strategies, properties of the extracted PSs, and environmental impacts. The results showed that when the PS extraction was before AD, it produced 76.03 ± 2.00 mL of methane per gram of volatile solids (VS) and afforded a PS yield of 6.3 ± 0.09% (w:w), with a PS sulfate content of 13.15% ± 0.06%. In contrast, when PS extraction was after AD, the methane production decreased to 58.14 ± 0.99 mL of methane per gram of VS and afforded a PS yield of 5.67% ± 0.18% (w:w) in VS, with a PS sulfate content of 2.60% ± 0.04%. When there were two PS extractions before and after AD, the methane production, PS yield and sulfate content were 76.03 ± 2.00 mL of methane per gram of VS, 11.54 ± 0.62% and 8.35 ± 0.12%, respectively. Then, the bioactivity of the extracted PSs was assessed by one anti-inflammation assay and three anti-oxidation assays, and statistical analysis revealed that these four bioactivities of PSs were influenced by their sulfate content, protein content and monosaccharide composition, especially the ratios of arabinose and rhamnose. Furthermore, the environmental impact analysis shows that S1 was the best in five environmental indicators compared with other three non-coupled processes. These findings suggest that the coupling PSs and methane recovery process should be further explored to determine its potential for large-scale sludge treatment.

Proteomic profiling of robust acetoclastic methanogen in chrysene-altered anaerobic digestion: Global dissection of enzymes.

Methanogen is a pivotal player in pollution treatment and energy recovery, and emerging pollutants (EPs) frequently occur in methanogen-applied biotechnology such as anaerobic digestion (AD). However, the direct effect and underlying mechanism of EPs on crucial methanogen involved in its application still remain unclear. The positive effect of chrysene (CH) on semi-continuous AD of sludge and the robust methanogen was dissected in this study. The methane yield in the digester with CH (100 mg/kg dry sludge) was 62.1 mL/g VS substrate, much higher than that in the control (46.1 mL/g VS substrate). Both methane production from acetoclastic methanogenesis (AM) and the AM proportion in the methanogenic pathway were improved in CH-shaped AD. Acetoclastic consortia, especially Methanosarcina and functional profiles of AM were enriched by CH in favor of the corresponding methanogenesis. Further, based on pure cultivation exposed to CH, the methanogenic performance, biomass, survivability and activity of typical Methanosarcina (M. barkeri) were boosted. Notably, iTRAQ proteomics revealed that the manufacturing (transcription and translation), expression and biocatalytic activity of acetoclastic metalloenzymes, particularly tetrahydromethanopterin S-methyltransferase and methyl-coenzyme M reductase with cobalt/nickel-cofactor (F430 and cobalamin), and acetyl-CoA decarbonylase/synthase with cobalt/nickel-active site, of M. barkeri were upregulated significantly with fold changes in the range of 1.21-3.20 due to the CH presence. This study shed light on EPs-affecting industrially crucial methanogen at the molecular biology level during AD and had implications in the technical relevance of methanogens.

Effect of grassland cutting frequency, species mixture, wilting and fermentation pattern of grass silages on in vitro methane yield.

Mitigating enteric methane (CH4) emissions is crucial as ruminants account for 5% of global greenhouse gas emissions. We hypothesised that less frequent harvesting, use of crops with lower WSC concentration, ensiling at low crop dry matter (DM) and extensive lactic acid fermentation would reduce in vitro CH4 production. Timothy (T), timothy + red clover mixture (T + RC) or perennial ryegrass (RG), cut either two or three times per season, was wilted to 22.5% or 37.5% DM and ensiled with or without formic acid-based additive. Silages were analysed for chemical composition and fermentation products. In vitro CH4 production was measured using an automated gas in vitro system. Methane production was, on average, 2.8 mL/g OM lower in the two-cut system than in the three-cut system (P < 0.001), and 1.9 mL/g OM lower in T than in RG (P < 0.001). Silage DM did not affect CH4 production (P = 0.235), but formic acid increased CH4 production by 1.2 mL/g OM compared to the untreated silage (P = 0.003). In conclusion, less frequent harvesting and extensive silage fermentation reduce in vitro CH4 production, while RG in comparison to T resulted in higher production of CH4.

The effects of feeding dried browse leaves on rumen ammonia levels, methanogens and protozoa amplification of sheep in the Coastal Savannah of Ghana.

Tanniferous browse leaves are reported to inhibit methanogens and protozoa activity in the rumen, thus contributing to a reduction of methane emission. This study evaluated the influence of feeding dried browse leaves to sheep on rumen ammonia concentration, the base pair at which protozoa and methanogens were amplified and double stranded DNA concentration (dsDNA) from rumen fluid and faeces. The eight treatments were urea treated rice straw, Albizzia lebbek (AL), Moringa oleifera (MO), Millettia thonningii (MT), AL + MO, AL + MT, AL + MO + MT and MO + MT. After feeding 32 ram lambs for 3 months, one ram lamb on each of the eight experimental diets was randomly selected and slaughtered to obtain rumen fluid. Genomic DNAs were extracted from methanogen and protozoa strains obtained from rumen liquor and from faecal matter of sheep. Rumen ammonia was determined using spectrophotometer. Methanogens and protozoa from rumen fluid and faeces were amplified at 1100 base pair, 200-1100 base pair, 320-1100 base pair and 200-750 base pair respectively. Rumen ammonia concentration, dsDNA from rumen fluid and faeces ranged from 14.51 to 23.01 mg/dl, 65 to 900 µg/ml and 100 to 950 µg/g respectively. The rumen ammonia concentration met the requirements for efficient growth of microbes. The presence of methanogens and protozoa in the rumen fluid and in the faeces indicated that dried browse diets were able to inhibit the growth of both protozoa and methanogens in the rumen by eliminating them, and thus, were excreted in the faeces. Hence, feeding of dried browse leaves can contribute to lower methane emission.

Effect of canola oil supplementation level on total tract digestion, ruminal fermentation, and methane emissions of cows grazing Urochloa sp. supplemented with a fixed amount of concentrate.

Four rumen-cannulated cows (Bos taurus × Bos indicus, 657 ± 92 kg body weight, BW) in a rotational grazing (Urochloa sp.) system were assigned to different canola oil (CO) inclusion levels, 0.0, 0.40, 0.80, and 1.2 g/kg according to shrunk body weight (SBW, BW adjusted for gastrointestinal filling) in a 4 × 4 Latin Square design to evaluate CO on the CH4 emissions and dietary energy intake. CH4 emissions were estimated using an infrared analyzer methodology (Sniffer method). Grass intake and fecal production were estimated using Cr2O3 as an external marker. CO supplementation increased (linear effect, P ≤ 0.05) total dry matter and gross energy intake with a linear increase (P = 0.09) in neutral detergent fiber (NDF) intake. While digestible energy (Mcal/kg) linearly increased with increasing CO supplementation level (linear effect, P < 0.05), total tract digestion of organic matter, NDF, and CP was comparable (P > 0.05) between levels. Maximal CO supplementation (1.2 g/kg SBW) significantly decreased total ruminal protozoa population, acetate:propionate ratio, and enteric methane production (g/kg DMI) by 9, 5.3, and 17.5%, respectively. This study showed that, for cows grazing tropical forages, CO can be supplemented up to 1.2 g/kg SBW (5.8% of the total diet) without negatively affecting intake and nutrient digestion while reducing ruminal fermentation efficiency and enteric methane emission (≤ 17.5%).

Association of the chemical composition and nutritional value of forage resources in Colombia with methane emissions by enteric fermentation.

In the livestock sector, strategies are available to mitigate gas emissions, such as methane, one of the alternatives that have shown potential correspondence to changes in the composition of the diet. The main aim of this study was to analyze the influence of methane emissions with data on enteric fermentation obtained from the Electronic Data Gathering, Analysis, and Retrieval (EDGAR) database and based on forecasts of methane emissions by enteric fermentation with an autoregressive integrated moving average (ARIMA) model and the application of statistical tests to identify the association between methane emissions from enteric fermentation and the variables of the chemical composition and nutritional value of forage resources in Colombia. The results reported positive correlations between methane emissions and the variables ash content, ethereal extract, neutral detergent fiber (NDF), and acid detergent fiber (ADF) and negative correlations between methane emissions and the variables percentage of unstructured carbohydrates, total digestible nutrients (TDN), digestibility of dry matter, metabolizable energy (MERuminants), net maintenance energy (NEm), net energy gain (NEg), and net lactation energy (NEI). The variables with the most significant influence on the reduction of methane emissions by enteric fermentation are the percentage of unstructured carbohydrates and the percentage of starch. In conclusion, the analysis of variance and the correlations between the chemical composition and the nutritive value of forage resources in Colombia help to understand the influence of diet variables on methane emissions of a particular family and with it in the application of strategies of mitigation.

Biogas utilization without desulfurization pretreatment in a bioelectrochemical system.

Utilizing biogas as a fuel for heating and power generation usually requires desulfurization pretreatment. In this study, the biogas utilization without desulfurization pretreatment in a bioelectrochemical system (BES) was explored. The results showed that the biogas-fueled BES was successfully started up within 36 d and the presence of hydrogen sulfide promoted both methane consumption and electricity generation. The optimal performance (i.e., a methane consumption of 0.523 ± 0.004 mmol/d, a peak voltage of 577 ± 1 mV, a coulomb production of 37.86 ± 0.43C/d, a coulombic efficiency of 9.37 ± 0.06 % and the maximum power density of 2.070 W/m3) was obtained under bicarbonate buffer solution and 40 °C conditions. The addition of 1 mg/L sulfide and 5 mg/L L-cysteine facilitated methane consumption and electricity generation. In the anode biofilm, the dominant bacteria were Sulfurivermis, unclassified_o__Ignavibacteriales and Lentimicrobium, while Methanobacterium, Methanosarcina and Methanothrix were the predominant archaea. Besides, the metagenomics profiles reveal that anaerobic methane oxidation and electricity generation were closely related to sulfur metabolism. These findings provide a novel approach for utilizing biogas without desulfurization pretreatment.

Internal driving mechanism of microbial community and metabolic pathway for psychrophilic anaerobic digestion by microbial electrolysis cell.

The system that microbial electrolysis cell coupled anaerobic digestion (termed MEC-AD) with metal organic framework-modified cathode was operated under different voltage levels (0-1.2 V) at 20 °C. The maximum methane yield increased to 0.23 ± 0.01 LCH4 g-1COD at 0.9 V, with 28% improvement compared to 0 V (0.18 ± 0.01 LCH4 g-1COD). Moreover, total volatile fatty acid and propionate accumulation decreased by 32% and 15% at 0.9 V, indicating the system has potential to alleviate acidity suppression. Acidogens and electroactive microorganisms was clearly enriched with increasing applied voltage. Specifically, the abundance of Smithella increased, which could degrade propionate to acetate. Methanosaeta was dominant, accounting for ca. 40.1%∼55.1% of the archaea community at 0.3-1.2 V. Furthermore, the system reinforced psychrophilic methanogenesis by activating important enzymes involved in related metabolism pathways. Overall, this study provides perspective on the future practical application for the regulation of psychrophilic AD in electrochemically integrated bioreactors.

Development of a dynamic model for prediction of energy in milk protein, lactose, fat, and enteric methane emissions in goats based on energy balance and indirect calorimetry studies.

Feed costs are overwhelmingly the largest expense for dairy producers. Thus, improving milk production efficiency (milk fat and protein are the main incomes for farmers) is of great economic importance in the dairy industry. The main objective of this study was to develop a dynamic energy partitioning model to describe and quantify how dietary energy from carbohydrate, protein, and fat is transferred to milk (protein, lactose, and fat) in dairy goats. In addition, due to increasing worldwide concerns regarding livestock contribution to global warming, methane (CH4) emission was quantified. For modeling purposes, 158 individual goat observations were used and randomly split into 2/3 for model development and 1/3 for internal evaluation. For external evaluation, 20 different energy balance studies from the literature (77 observations) were evaluated. The Root Mean Square Prediction Error (RMSPE) was 13.2% for loss of energy in CH4, 16.8% for energy in fat, 19.4% for energy in protein, and 22.3 energy in lactose. Mean bias was around zero for all variables and the slope bias was zero for milk energy in lactose, close to 1% for milk fat (1.01%), and around 3% and 10% for protein and CH4, respectively. Random bias was greater than 85% for energy in CH4 and milk energy components indicating non-systematic errors and that the equation in the model fitted the data properly. Analyses of residuals appeared to be randomly distributed around zero. Slopes of regression lines for residuals vs. predicted were positive for milk fat energy, zero for lactose, and negative for milk energy in protein and CH4. This model suggested for use with mixed diets and by-products to obtain balanced macronutrient supply, methane emissions, and milk performance during mid lactation could be an interesting tool to help farmers simulate scenarios that increase milk fat and protein, evaluate CH4 emissions, without the costs of running animal trials.

The present model using mixed diets with different by-products to obtain macronutrient balance, methane emission, and milk performance during mid lactation could be an interesting tool to help farmers, without the costs of running animal experiments. The dietary change from grain-based to partial replacement with agro-industrial-byproducts in mid-lactation dairy goats was accompanied by transformations in carbohydrate and fat energy transfer to support production. The output underscored that both oxidation of carbohydrate and fat interact to maintain milk energy output.

Biochar Facilitated Direct Interspecies Electron Transfer in Anaerobic Digestion to Alleviate Antibiotics Inhibition and Enhance Methanogenesis: A Review.

Efficient conversion of organic waste into low-carbon biofuels such as methane through anaerobic digestion (AD) is a promising technology to alleviate energy shortages. However, issues such as inefficient methane production and poor system stability remain for AD technology. Biochar-facilitated direct interspecies electron transfer (DIET) has recently been recognized as an important strategy to improve AD performance. Nonetheless, the underlying mechanisms of biochar-facilitated DIET are still largely unknown. For this reason, this review evaluated the role of biochar-facilitated DIET mechanism in enhancing AD performance. First, the evolution of DIET was introduced. Then, applications of biochar-facilitated DIET for alleviating antibiotic inhibition and enhancing methanogenesis were summarized. Next, the electrochemical mechanism of biochar-facilitated DIET including electrical conductivity, redox-active characteristics, and electron transfer system activity was discussed. It can be concluded that biochar increased the abundance of potential DIET microorganisms, facilitated microbial aggregation, and regulated DIET-associated gene expression as a microbial mechanism. Finally, we also discussed the challenges of biochar in practical application. This review elucidated the role of DIET facilitated by biochar in the AD system, which would advance our understanding of the DIET mechanism underpinning the interaction of biochar and anaerobic microorganisms. However, direct evidence for the occurrence of biochar-facilitated DIET still requires further investigation.

Molecular evidence for the production of labile, sulfur-bearing dissolved organic matter in the seep sediments of the South China Sea.

Cold seeps with methane-rich fluids leaking out of the seafloor usually support massive biomass of chemosynthetic organisms and associated fauna. A substantial amount of methane is converted to dissolved inorganic carbon by microbial metabolism, and this process also releases dissolved organic matter (DOM) into pore water. Here, pore water samples from "Haima cold seeps" sediments and the non-seep reference sediments in the northern South China Sea were analyzed for optical properties and molecular compositions of pore water DOM. Our results showed that the relative abundance of protein-like DOM, H/Cwa and molecular lability boundary percentage (MLBL%) in the seep sediments were significantly higher than those in the reference sediments, indicating that more labile DOM related to unsaturated aliphatic compounds is produced in the seep sediments. Spearman's correlation of the fluoresce and molecular data suggested that the humic-like components (C1 and C2) mainly constituted the refractory compounds (CRAM, highly unsaturated and aromatics compounds). In contrast, the protein-like component (C3) had high H/C ratios featuring high degree of DOM lability. The amount of S-containing formulas (CHOS and CHONS) was greatly elevated in the seep sediments, likely caused by abiotic and biotic sulfurization of DOM in the sulfidic environment. Although the abiotic sulfurization was proposed to have a stabilizing effect on organic matter, our results implied that the biotic sulfurization in the cold seep sediments would increase DOM lability. Overall, the labile DOM accumulated in the seep sediments is closely linked to methane oxidation, which not only support heterotrophic communities and but also likely have an impact on carbon and sulfur cycling in the sediments and the ocean.

Nitrogen and energy losses and methane emissions from beef cattle fed diets with gradual replacement of maize silage and concentrate with grass silage and corn-cob mix.

Diets based on large proportions of grassland-based feed are uncommon in forage-based intensive beef production, thus contradicting governmental or commercial strategies to promote the use of grassland-based feed in ruminant production systems. Compared with typical maize silage/concentrate diets, grassland-based diets are associated with impaired nitrogen (N) and energy utilisation because of the comparably lower energy and higher CP content of these feeds. However, quantitative studies concerning the effects of increased dietary proportions of grassland-derived feeds on N and energy losses and utilisation and on methane emissions are missing and the compensation potential of using a limited proportion of an energy-rich forage is unknown. Therefore, we tested five diets with varying types and proportions of forage and concentrate. Three diets consisted of grass silage, maize silage, and concentrate in ratios of, g/kg DM, 100:600:300 (G100; control), 300:500:200 (G300), and 500:300:200 (G500), respectively. Two diets were composed of grass silage, corn-cob mix (CCM), and concentrate in ratios of, g/kg DM, 500:300:200 (G500CCM), and 750:150:100 (G750CCM), respectively. A high-protein concentrate (270 g CP/kg DM) was fed to G100, whereas a low-protein concentrate (140 g CP/kg DM) was used in the remaining diets. Diets were fed throughout the entire fattening period to groups of six Limousin-crossbred bulls each. When weighing 246 ± 18 kg, each animal underwent a 7-day total daily faeces and urine collection, which was followed by measuring methane emissions in respiration chambers for 48 h. Total DM intake was similar across all diets, whereas the N intake varied (P < 0.05). Urinary N loss (g/day) was the highest for G750CCM (28.2) and G100 (26.6) and lowest for G500CCM (15.2) and G300 (16.9) (P < 0.001). Energy utilisation was comparable among all groups. Metabolisable energy intake decreased numerically only with increasing proportions of grass silage in the diet. Substituting maize silage with CCM counteracted the loss in metabolisable energy intake. Absolute methane emissions were not different across the groups, but methane emission intensity (mg/g body protein retention) varied (P < 0.05), being numerically lower for G100 (349) and G500CCM (401) compared with the other groups (488 on average). In conclusion, the results show that the grass silage proportion in beef cattle diets can be substantially increased when strategically combined with energy-dense forages, such as CCM. This also limits the need for concentrate and additional protein sources; in addition, the associated urinary N emissions, which are potentially noxious to the environment, are avoided.

Taxonomic and enzymatic basis of the cellulolytic microbial consortium KKU-MC1 and its application in enhancing biomethane production.

Lignocellulosic biomass is a promising substrate for biogas production. However, its recalcitrant structure limits conversion efficiency. This study aims to design a microbial consortium (MC) capable of producing the cellulolytic enzyme and exploring the taxonomic and genetic aspects of lignocellulose degradation. A diverse range of lignocellulolytic bacteria and degrading enzymes from various habitats were enriched for a known KKU-MC1. The KKU-MC1 was found to be abundant in Bacteroidetes (51%), Proteobacteria (29%), Firmicutes (10%), and other phyla (8% unknown, 0.4% unclassified, 0.6% archaea, and the remaining 1% other bacteria with low predominance). Carbohydrate-active enzyme (CAZyme) annotation revealed that the genera Bacteroides, Ruminiclostridium, Enterococcus, and Parabacteroides encoded a diverse set of cellulose and hemicellulose degradation enzymes. Furthermore, the gene families associated with lignin deconstruction were more abundant in the Pseudomonas genera. Subsequently, the effects of MC on methane production from various biomasses were studied in two ways: bioaugmentation and pre-hydrolysis. Methane yield (MY) of pre-hydrolysis cassava bagasse (CB), Napier grass (NG), and sugarcane bagasse (SB) with KKU-MC1 for 5 days improved by 38-56% compared to non-prehydrolysis substrates, while MY of prehydrolysed filter cake (FC) for 15 days improved by 56% compared to raw FC. The MY of CB, NG, and SB (at 4% initial volatile solid concentration (IVC)) with KKU-MC1 augmentation improved by 29-42% compared to the non-augmentation treatment. FC (1% IVC) had 17% higher MY than the non-augmentation treatment. These findings demonstrated that KKU-MC1 released the cellulolytic enzyme capable of decomposing various lignocellulosic biomasses, resulting in increased biogas production.