High peatland methane emissions following permafrost thaw: enhanced acetoclastic methanogenesis during early successional stages

Permafrost thaw in northern peatlands often leads to increased methane (CH4) emissions, but gaps remain in our understanding of the underlying controls responsible for increased emissions and the duration for which they persist. We assessed how shifting ecological conditions affect microbial communities, and the magnitude and stable isotopic signature (δ13C) of CH4 emissions along a thermokarst bog transect in boreal western Canada. Thermokarst bogs develop following permafrost thaw when dry, elevated peat plateaus collapse and become saturated and dominated by Sphagnum mosses. We differentiated between a young and a mature thermokarst bog stage (~30 and years ~200 since thaw, respectively). The young bog located along the thermokarst edge, was wetter, warmer and dominated by hydrophilic vegetation compared to the mature bog. Using 16S rRNA gene high throughput sequencing, we show that microbial communities were distinct near the surface and converged with depth, but lesser differences remained down to the lowest depth (160 cm). Microbial community analysis and δ13C data from CH4 surface emissions and dissolved gas depth profiles show that hydrogenotrophic methanogenesis was the dominant pathway at both sites. However, the young bog was found to have isotopically heavier δ13C-CH4 in both dissolved gases profiles and surface CH4 emissions, suggesting that acetoclastic methanogenesis was relatively more enhanced throughout the young bog peat profile. Furthermore, young bog CH4 emissions were three times greater than the mature bog. Our study suggests that interactions between ecological conditions and methanogenic communities enhance CH4 emissions in young thermokarst bogs, but these favorable conditions only persist for the initial decades after permafrost thaw.

Prediction of Genes That Function in Methanogenesis and CO2 Pathways in Extremophiles

Gaet’ale (GAL) and Mud’ara (MUP) are two hypersaline ponds located in the Danakil Depression recharged by underground water from the surrounding highlands. These two ponds have different pH, salinity, and show variation in the concentration of many ionic components. Metagenomic analysis concludes that GAL is dominated by bacteria as in the case of the other hypersaline and acidic ponds in the Danakil Depression. However, Archaea dominated the ponds of MUP. In the current study, the application of SEED and KEGG helped to map the ordered steps of specific enzyme catalyzed reaction in converting CO2 into cell products. We predict that highly efficient and light-independent carbon fixation involving phosphoenolpyruvate carboxylase takes place in MUP. On the contrary, genes encoding enzymes involved in hydrogenotrophic and acetoclastic methanogenesis appeared solely in ponds of GAL, implying the biological source of the hazardous methane gas in that environment. Based on the investigation of the sources of the genes of interest, it is clear that cooperative interactions between members of the two communities and syntrophic metabolism is the main strategy adapted to utilize inorganic carbon as a carbon source in both MUP and GAL. This insight can be used to design biotechnological applications of microbial communities in production of methane biogas or to minimize CO2 emissions.

Chasing the metabolism of novel syntrophic acetate-oxidizing bacteria in thermophilic methanogenic chemostats

Background: Acetate is the major intermediate of anaerobic digestion of organic waste to CH4. In anaerobic methanogenic systems, acetate degradation is carried out by either acetoclastic methanogenesis or a syntrophic degradation by a syntrophy of acetate oxidizers and hydrogenotrophic methanogens. Due to challenges in isolation of syntrophic acetate-oxidizing bacteria (SAOB), the diversity and metabolism of SAOB, as well as the mechanisms of their interactions with methanogenic partners remain poorly understood. Results: In this study, we successfully enriched previously unknown SAOB by operating continuous thermophilic anaerobic chemostats fed with acetate, propionate, butyrate, or isovalerate as the sole carbon and energy source. They represent novel clades belonging to Clostridia, Thermoanaerobacteraceae, Anaerolineae, and Gemmatimonadetes. In these SAOB, acetate is degraded through reverse Wood-Ljungdahl pathway or an alternative pathway mediated by the glycine cleavage system, while the SAOB possessing the latter pathway dominated the bacterial community. Moreover, H2 is the major product of the acetate degradation by these SAOB, which is mediated by [FeFe]-type electron-confurcating hydrogenases, formate dehydrogenases, and NADPH reoxidation complexes. We also identified the methanogen partner of these SAOB in acetate-fed chemostat, Methanosarcina thermophila, which highly expressed genes for CO2-reducing methanogenesis and hydrogenases to supportively consuming H2 at transcriptional level. Finally, our bioinformatical analyses further suggested that these previously unknown syntrophic lineages were prevalent and might play critical roles in thermophilic methanogenic reactors. Conclusion: This study expands our understanding on the phylogenetic diversity and in situ biological functions of uncultured syntrophic acetate degraders, and presents novel insights on how they interact with their methanogens partner. These knowledges strengthen our awareness on the important role of SAO in thermophilic methanogenesis and may be applied to manage microbial community to improve the performance and efficiency of anaerobic digestion. Keywords: Thermophilic anaerobic digestion, Microbial community, Syntrophic acetate oxidation, Glycine cleavage, Energy conservation

Complementary Substrates-Brewery Wastewater and Piggery Effluent—Assessment and Microbial Community Profiling in a Hybrid Anaerobic Reactor

A hybrid anaerobic reactor was operated under the complementary effluents concept to reduce the unbalanced/inhibitory capacity of the provided piggery effluent. Brewery wastewater was chosen to complement piggery effluent (60:40% v/v, respectively). The HRT reduction from 6.7 to 3.0 days allowed the testing of an organic load increase from 4.5 to 10.0 g COD/L·d, which resulted in the almost doubling of biogas production. Biogas volumes (1.2 and 2.1 L/L·d, respectively) associated with its quality (>77% CH4) revealed that the hybrid anaerobic reactor responded positively to the operational changes and that piggery effluent can be advantageously digested using the brewery wastewater as the complementary effluent. The unit bottom and the packing bed were the main functional sections recognized in the hybrid. At the beginning of anaerobic digestion, bacterial populations belonged mostly to Bacteroidales (33%) and Clostridiales (35%). The process stability and the biogas quality at 3-d HRT were related to a change in the structure composition, since Flavobacteriales (18%), Bacillales (7%), Pseudomonadales (11%) and members of the Alcaligenaceae family (5%) also integrated the microbial communities. An evident change had also occurred in archaeal populations at this phase. Methanosaeta became the dominant genus (95%), confirming that acetoclastic methanogenesis was the main way for methane production.

Lignin intermediates lead to phenyl acid formation and microbial community shifts in meso- and thermophilic batch reactors

Background Lignin intermediates resulting from lignocellulose degradation have been suspected to hinder anaerobic mineralisation of organic materials to biogas. Phenyl acids like phenylacetate (PAA) are early detectable intermediates during anaerobic digestion (AD) of aromatic compounds. Studying the phenyl acid formation dynamics and concomitant microbial community shifts can help to understand the microbial interdependencies during AD of aromatic compounds and may be beneficial to counteract disturbances. Results The length of the aliphatic side chain and chemical structure of the benzene side group(s) had an influence on the methanogenic system. PAA, phenylpropionate (PPA), and phenylbutyrate (PBA) accumulations showed that the respective lignin intermediate was degraded but that there were metabolic restrictions as the phenyl acids were not effectively processed. Metagenomic analyses confirmed that mesophilic genera like Fastidiosipila or Syntrophomonas and thermophilic genera like Lactobacillus, Bacillus, Geobacillus, and Tissierella are associated with phenyl acid formation. Acetoclastic methanogenesis was prevalent in mesophilic samples at low and medium overload conditions, whereas Methanoculleus spp. dominated at high overload conditions when methane production was restricted. In medium carbon load reactors under thermophilic conditions, syntrophic acetate oxidation (SAO)-induced hydrogenotrophic methanogenesis was the most important process despite the fact that acetoclastic methanogenesis would thermodynamically be more favourable. As acetoclastic methanogens were restricted at medium and high overload conditions, syntrophic acetate oxidising bacteria and their hydrogenotrophic partners could step in for acetate consumption. Conclusions PAA, PPA, and PBA were early indicators for upcoming process failures. Acetoclastic methanogens were one of the first microorganisms to be impaired by aromatic compounds, and shifts to syntrophic acetate oxidation coupled to hydrogenotrophic methanogenesis occurred in thermophilic reactors. Previously assumed associations of specific meso- and thermophilic genera with anaerobic phenyl acid formation could be confirmed.

Hydrogenotrophic Methanogenesis Under Alkaline Conditions

A cement-based geological disposal facility (GDF) is one potential option for the disposal of intermediate level radioactive wastes. The presence of both organic and metallic materials within a GDF provides the opportunity for both acetoclastic and hydrogenotrophic methanogenesis. However, for these processes to proceed, they need to adapt to the alkaline environment generated by the cementitious materials employed in backfilling and construction. Within the present study, a range of alkaline and neutral pH sediments were investigated to determine the upper pH limit and the preferred route of methane generation. In all cases, the acetoclastic route did not proceed above pH 9.0, and the hydrogenotrophic route dominated methane generation under alkaline conditions. In some alkaline sediments, acetate metabolism was coupled to hydrogenotrophic methanogenesis via syntrophic acetate oxidation, which was confirmed through inhibition studies employing fluoromethane. The absence of acetoclastic methanogenesis at alkaline pH values (>pH 9.0) is attributed to the dominance of the acetate anion over the uncharged, undissociated acid. Under these conditions, acetoclastic methanogens require an active transport system to access their substrate. The data indicate that hydrogenotrophic methanogenesis is the dominant methanogenic pathway under alkaline conditions (>pH 9.0).