Carbon Metabolism of Methylotrophic Methanogens and Asgard Archaea in Marine Sediments

Carbon is the central element of life, as it is involved in building up of biological constituents and energy metabolisms in the cell. Archaea, - the most recently recognized domain of life - hold a crucial phylogenetic position in the evolution of life, but for most archaeal phyla, little is known about their role and activity in carbon metabolism. Archaea inhabit a variety of environments such as soils, sediments, sea water, and the guts of animals. Specifically in marine sediments, Thaumarchaeota, Euryarchaeota, Bathyarchaeota, Woesearchaeota and Asgard archaea are commonly found in archaeal communities. Methanogens affiliated to Euryarchaeota are well-known players in carbon metabolism, i.e., acetoclastic, hydrogenotrophic, and methylotrophic methanogenesis. Based on pure culture studies and genomic evidence, significant amounts of the biomass of methylotrophic methanogens growing on methyl substrates is derived from inorganic carbon. However, the in situ activity of these methanogens in carbon assimilation is unclear as the large inorganic carbon pool in marine sediment potentially affects carbon utilization patterns. To address this hypothesis, we initially applied nucleic acid stable isotope probing (SIP) to detect methylotrophic methanogens in marine sediment incubations. SIP results showed that 13C-labeled dissolved inorganic carbon (DIC) is necessary to identify methylotrophic methanogens, as illustrated by the nucleic acid synthesis pathway in these methanogens that 70-80% of carbon stems from DIC rather than methanol. In parallel, lipid-SIP suggested that DIC contributed to more than 60% from incubations with sediment from the sulfate reduction zone (SRZ), i.e., 20% higher than expected from lipid synthesis pathway. We further unexpectedly found that up to 12% methane was formed from DIC in autoclaved slurry incubations inoculated with the marine methylotrophic methanogen Methanococcoides methylutens. Similarly, methane formation from CO2 during methylotrophic methanogenesis was also observed with SRZ sediment incubations. In the same sediment incubations a higher amount of inorganic carbon was incorporated into lipids than expected, indicating that more DIC was assimilated into biomass than expected. Thus, the CO2 conversion to methane and biomass may play an important role in marine sediments. In the most recently discovered super phylum of the Archaea, the Asgard archaea might hold the key to understand the evolutionary origin of eukaryotes. Unlike methanogens, however, the diversity, carbon metabolism and the activity of Asgard archaea in marine sediments are still unknown. In this study, five new groups of Asgard archaea namely Kariarchaeota, Balderarchaeota, Hodarchaeota, Lagarchaeota and Gerdarchaeota are reported. In experiments with 13C-DIC, potential electron donors and electron acceptors, subgroup of Asgard archaea i.e., Lokiarchaeota was detected in the heavy SIP fractions from the incubations amended with organic polymers or sulfur, suggesting their activities of carbon fixation, organic polymers (cellulose, lignin and humic acid) degradation and sulfur metabolism. Furthermore, metagenomes were sequenced from heavy fractions of DNA-SIP samples obtained in the aforementioned experiments and from DNA extracted from mangrove sediments in the southeast coast of China. These metagenomes indicate that Asgard archaea harbor pathways of inorganic carbon fixation and degradation of cellulose, protein, short-chain and medium-chain alkane as well as assimilatory sulfate reduction. Crucially, the methyl coenzyme M reductase genes found in Helarchaeota have extended the potential of short-chain hydrocarbon oxidation to the Asgard archaea in this study. Overall, these findings illustrate that Asgard archaea actively utilize organic and inorganic carbon at the same time in mixotrophic fashion, which might play critical roles in carbon cycling of marine sediments. In particular, the successful detection of methylotrophic methanogens and Asgard archaea in marine sediments by nucleic acid-SIP with 13C-DIC suggested a crucial role of inorganic carbon in carbon metabolisms of these archaea. Given that many archaea harbor the acetyl-CoA associated carbon fixation pathway, my findings indicate that inorganic carbon assimilation might be ubiquitous in archaea when supply or availability of organic carbon are not sufficient in marine sediments

Influence of sedimentary deposition on the microbial assembly process in Arctic Holocene marine sediments

The sea-level rise during the Holocene (11–0 ky BP) and its resulting sedimentation and biogeochemical processes may control microbial life in Arctic sediments. To gain further insight into this interaction, we investigated a sediment core (up to 10.7 m below the seafloor) from the Chuckchi Shelf of the western Arctic Ocean using metabarcoding-based sequencing and qPCR to characterize archaeal and bacterial 16S rRNA gene composition and abundance, respectively. We found that Arctic Holocene sediments harbor local microbial communities, reflecting geochemical and paleoclimate separations. The composition of bacterial communities was more diverse than that of archaeal communities, and specifically distinct at the boundary layer of the sulfate–methane transition zone. Enriched cyanobacterial sequences in the Arctic middle Holocene (8–7 ky BP) methanogenic sediments remarkably suggest past cyanobacterial blooms. Bacterial communities were phylogenetically influenced by interactions between dispersal limitation and environmental selection governing community assembly under past oceanographic changes. The relative influence of stochastic and deterministic processes on the bacterial assemblage was primarily determined by dispersal limitation. We have summarized our findings in a conceptual model that revealed how changes in paleoclimate phases cause shifts in ecological succession and the assembly process. In this ecological model, dispersal limitation is an important driving force for progressive succession for bacterial community assembly processes on a geological timescale in the western Arctic Ocean. This enabled a better understanding of the ecological processes that drive the assembly of communities in Holocene sedimentary habitats affected by sea-level rise, such as in the shallow western Arctic shelves

Temperature Controls Crystalline Iron Oxide Utilization by Microbial Communities in Methanic Ferruginous Marine Sediment Incubations

Microorganisms can use crystalline iron minerals for iron reduction linked to organic matter degradation or as conduits for direct interspecies electron transfer (mDIET) to syntrophic partners, e.g., methanogens. The environmental conditions that lead either to reduction or conduit use are so far unknown. We investigated microbial community shifts and interactions with crystalline iron minerals (hematite and magnetite) in methanic ferruginous marine sediment incubations during organic matter (glucose) degradation at varying temperatures. Iron reduction rates increased with decreasing temperature from 30°C to 4°C. Both hematite and magnetite facilitated iron reduction at 4°C, demonstrating that microorganisms in the methanic zone of marine sediments can reduce crystalline iron oxides under psychrophilic conditions. Methanogenesis occurred, however, at higher rates with increasing temperature. At 30°C, both hematite and magnetite accelerated methanogenesis onset and maximum process rates. At lower temperatures (10°C and 4°C), hematite could still facilitate methanogenesis but magnetite served more as an electron acceptor for iron reduction than as a conduit. Different temperatures selected for different key microorganisms: at 30°C, members of genus Orenia, Halobacteroidaceae, at 10°C, Photobacterium and the order Clostridiales, and at 4°C Photobacterium and Psychromonas were enriched. Members of the order Desulfuromonadales harboring known dissimilatory iron reducers were also enriched at all temperatures. Our results show that crystalline iron oxides predominant in some natural environments can facilitate electron transfer between microbial communities at psychrophilic temperatures. Furthermore, temperature has a critical role in determining the pathway of crystalline iron oxide utilization in marine sediment shifting from conduction at 30°C to predominantly iron reduction at lower temperatures

Temperature controls crystalline iron oxide utilization by microbial communities in methanic ferruginous marine sediment incubations

Microorganisms can use crystalline iron minerals for iron reduction linked to organic matter degradation or as conduits for direct interspecies electron transfer (mDIET) to syntrophic partners, e.g., methanogens. The environmental conditions that lead either to reduction or conduit use are so far unknown. We investigated microbial community shifts and interactions with crystalline iron minerals (hematite and magnetite) in methanic ferruginous marine sediment incubations during organic matter (glucose) degradation at varying temperatures. Iron reduction rates increased with decreasing temperature from 30 degrees C to 4 degrees C. Both hematite and magnetite facilitated iron reduction at 4 degrees C, demonstrating that microorganisms in the methanic zone of marine sediments can reduce crystalline iron oxides under psychrophilic conditions. Methanogenesis occurred, however, at higher rates with increasing temperature. At 30 degrees C, both hematite and magnetite accelerated methanogenesis onset and maximum process rates. At lower temperatures (10 degrees C and 4 degrees C), hematite could still facilitate methanogenesis but magnetite served more as an electron acceptor for iron reduction than as a conduit. Different temperatures selected for different key microorganisms: at 30 degrees C, members of genus Orenia, Halobacteroidaceae, at 10 degrees C, Photobacterium and the order Clostridiales, and at 4 degrees C Photobacterium and Psychromonas were enriched. Members of the order Desulfuromonadales harboring known dissimilatory iron reducers were also enriched at all temperatures. Our results show that crystalline iron oxides predominant in some natural environments can facilitate electron transfer between microbial communities at psychrophilic temperatures. Furthermore, temperature has a critical role in determining the pathway of crystalline iron oxide utilization in marine sediment shifting from conduction at 30 degrees C to predominantly iron reduction at lower temperatures

Crystalline iron oxides stimulate methanogenic benzoate degradation in marine sediment- derived enrichment cultures

Elevated dissolved iron concentrations in the methanic zone are typical geochemical signatures of rapidly accumulating marine sediments. These sediments are often characterized by co-burial of iron oxides with recalcitrant aromatic organic matter of terrigenous origin. Thus far, iron oxides are predicted to either impede organic matter degradation, aiding its preservation, or identified to enhance organic carbon oxidation via direct electron transfer. Here, we investigated the effect of various iron oxide phases with differing crystallinity (magnetite, hematite, and lepidocrocite) during microbial degradation of the aromatic model compound benzoate in methanic sediments. In slurry incubations with magnetite or hematite, concurrent iron reduction, and methanogenesis were stimulated during accelerated benzoate degradation with methanogenesis as the dominant electron sink. In contrast, with lepidocrocite, benzoate degradation, and methanogenesis were inhibited. These observations were reproducible in sediment-free enrichments, even after five successive transfers. Genes involved in the complete degradation of benzoate were identified in multiple metagenome assembled genomes. Four previously unknown benzoate degraders of the genera Thermincola (Peptococcaceae, Firmicutes), Dethiobacter (Syntrophomonadaceae, Firmicutes), Deltaproteobacteria bacteria SG8_13 (Desulfosarcinaceae, Deltaproteobacteria), and Melioribacter (Melioribacteraceae, Chlorobi) were identified from the marine sediment-derived enrichments. Scanning electron microscopy (SEM) and catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH) images showed the ability of microorganisms to colonize and concurrently reduce magnetite likely stimulated by the observed methanogenic benzoate degradation. These findings explain the possible contribution of organoclastic reduction of iron oxides to the elevated dissolved Fe2+ pool typically observed in methanic zones of rapidly accumulating coastal and continental margin sediments

Kohlenstoffmetabolismus von methylotrophen Methanogenen und Asgard-Archaeen in marinen Sedimenten

Carbon is the central element of life, as it is involved in building up of biological constituents and energy metabolisms in the cell. Archaea, - the most recently recognized domain of life - hold a crucial phylogenetic position in the evolution of life, but for most archaeal phyla, little is known about their role and activity in carbon metabolism. Archaea inhabit a variety of environments such as soils, sediments, sea water, and the guts of animals. Specifically in marine sediments, Thaumarchaeota, Euryarchaeota, Bathyarchaeota, Woesearchaeota and Asgard archaea are commonly found in archaeal communities. Methanogens affiliated to Euryarchaeota are well-known players in carbon metabolism, i.e., acetoclastic, hydrogenotrophic, and methylotrophic methanogenesis. Based on pure culture studies and genomic evidence, significant amounts of the biomass of methylotrophic methanogens growing on methyl substrates is derived from inorganic carbon. However, the in situ activity of these methanogens in carbon assimilation is unclear as the large inorganic carbon pool in marine sediment potentially affects carbon utilization patterns. To address this hypothesis, we initially applied nucleic acid stable isotope probing (SIP) to detect methylotrophic methanogens in marine sediment incubations. SIP results showed that 13C-labeled dissolved inorganic carbon (DIC) is necessary to identify methylotrophic methanogens, as illustrated by the nucleic acid synthesis pathway in these methanogens that 70-80% of carbon stems from DIC rather than methanol. In parallel, lipid-SIP suggested that DIC contributed to more than 60% from incubations with sediment from the sulfate reduction zone (SRZ), i.e., 20% higher than expected from lipid synthesis pathway. We further unexpectedly found that up to 12% methane was formed from DIC in autoclaved slurry incubations inoculated with the marine methylotrophic methanogen Methanococcoides methylutens. Similarly, methane formation from CO2 during methylotrophic methanogenesis was also observed with SRZ sediment incubations. In the same sediment incubations a higher amount of inorganic carbon was incorporated into lipids than expected, indicating that more DIC was assimilated into biomass than expected. Thus, the CO2 conversion to methane and biomass may play an important role in marine sediments. In the most recently discovered super phylum of the Archaea, the Asgard archaea might hold the key to understand the evolutionary origin of eukaryotes. Unlike methanogens, however, the diversity, carbon metabolism and the activity of Asgard archaea in marine sediments are still unknown. In this study, five new groups of Asgard archaea namely Kariarchaeota, Balderarchaeota, Hodarchaeota, Lagarchaeota and Gerdarchaeota are reported. In experiments with 13C-DIC, potential electron donors and electron acceptors, subgroup of Asgard archaea i.e., Lokiarchaeota was detected in the heavy SIP fractions from the incubations amended with organic polymers or sulfur, suggesting their activities of carbon fixation, organic polymers (cellulose, lignin and humic acid) degradation and sulfur metabolism. Furthermore, metagenomes were sequenced from heavy fractions of DNA-SIP samples obtained in the aforementioned experiments and from DNA extracted from mangrove sediments in the southeast coast of China. These metagenomes indicate that Asgard archaea harbor pathways of inorganic carbon fixation and degradation of cellulose, protein, short-chain and medium-chain alkane as well as assimilatory sulfate reduction. Crucially, the methyl coenzyme M reductase genes found in Helarchaeota have extended the potential of short-chain hydrocarbon oxidation to the Asgard archaea in this study. Overall, these findings illustrate that Asgard archaea actively utilize organic and inorganic carbon at the same time in mixotrophic fashion, which might play critical roles in carbon cycling of marine sediments. In particular, the successful detection of methylotrophic methanogens and Asgard archaea in marine sediments by nucleic acid-SIP with 13C-DIC suggested a crucial role of inorganic carbon in carbon metabolisms of these archaea. Given that many archaea harbor the acetyl-CoA associated carbon fixation pathway, my findings indicate that inorganic carbon assimilation might be ubiquitous in archaea when supply or availability of organic carbon are not sufficient in marine sediments

Metatranscriptomics reveals different features of methanogenic archaea among global vegetated coastal ecosystems

Vegetated coastal ecosystems (VCEs; i.e., mangroves, saltmarshes, and seagrasses) represent important sources of natural methane emission. Despite recent advances in the understanding of novel taxa and pathways associated with methanogenesis in these ecosystems, the key methanogenic players and the contribution of different substrates to methane formation remain elusive. Here, we systematically investigate the community and activity of methanogens using publicly available metatranscriptomes at a global scale together with our in-house metatranscriptomic dataset. Taxonomic profiling reveals that 13 groups of methanogenic archaea were transcribed in the investigated VCEs, and they were predominated by Methanosarcinales. Among these VCEs, methanogens exhibited all the three known methanogenic pathways in some mangrove sediments, where methylotrophic methanogens Methanosarcinales/Methanomassiliicoccales grew on diverse methyl compounds and coexisted with hydrogenotrophic (mainly Methanomicrobiales) and acetoclastic (mainly Methanothrix) methanogens. Contrastingly, the predominant methanogenic pathway in saltmarshes and seagrasses was constrained to methylotrophic methanogenesis. These findings reveal different archaeal methanogens in VCEs and suggest the potentially distinct methanogenesis contributions in these VCEs to the global warming

Iron and sulfate reduction structure microbial communities in (sub-)Antarctic sediments

Permanently cold marine sediments are heavily influenced by increased input of iron as a result of accelerated glacial melt, weathering, and erosion. The impact of such environmental changes on microbial communities in coastal sediments is poorly understood. We investigated geochemical parameters that shape microbial community compositions in anoxic surface sediments of four geochemically differing sites (Annenkov Trough, Church Trough, Cumberland Bay, Drygalski Trough) around South Georgia, Southern Ocean. Sulfate reduction prevails in Church Trough and iron reduction at the other sites, correlating with differing local microbial communities. Within the order Desulfuromonadales, the family Sva1033, not previously recognized for being capable of dissimilatory iron reduction, was detected at rather high relative abundances (up to 5%) while other members of Desulfuromonadales were less abundant (<0.6%). We propose that Sva1033 is capable of performing dissimilatory iron reduction in sediment incubations based on RNA stable isotope probing. Sulfate reducers, who maintain a high relative abundance of up to 30% of bacterial 16S rRNA genes at the iron reduction sites, were also active during iron reduction in the incubations. Thus, concurrent sulfate reduction is possibly masked by cryptic sulfur cycling, i.e., reoxidation or precipitation of produced sulfide at a small or undetectable pool size. Our results show the importance of iron and sulfate reduction, indicated by ferrous iron and sulfide, as processes that shape microbial communities and provide evidence for one of Sva1033’s metabolic capabilities in permanently cold marine sediments