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Relative abundances of all microbial PiCRUST-inferred functional pathways for all samples, based on 16S rRNA amplicon sequencing data from a mire-wide survey (2015) and co-analyzed autochamber site samples (2014-2015). The 16S rRNA amplicon sequencing data is available under NCBI BioProject PRJNA1236848. The sample metadata and SRA accessions are available at https://doi.org/10.5281/zenodo.15047156. FUNDING: National Aeronautics and Space Administration, Interdisciplinary Science program: From Archaea to the Atmosphere (award # NNX17AK10G). National Science Foundation, Biology Integration Institutes Program: EMERGE Biology Integration Institute (award # 2022070). United States Department of Energy Office of Biological and Environmental Research, Genomic Science Program: The IsoGenie Project (grant #s DE-SC0004632, DE-SC0010580, and DE-SC0016440). Sequencing was performed using startup funding from the University of Arizona to Virginia Rich. We thank the Swedish Polar Research Secretariat and SITES for the support of the work done at the Abisko Scientific Research Station. SITES is supported by the Swedish Research Council's grant 4.3-2021-00164. 

Stordalen Mire microbial ASV table (mire-wide_ASV_table.tsv) and taxonomy (mire-wide_taxonomy.tsv), based on 16S rRNA amplicon sequencing data from a mire-wide survey (2015) and co-analyzed autochamber site samples (2014-2015). The 16S rRNA amplicon sequencing data is available under NCBI BioProject PRJNA1236848. The sample metadata and SRA accessions are available at https://doi.org/10.5281/zenodo.15047156. FUNDING: National Aeronautics and Space Administration, Interdisciplinary Science program: From Archaea to the Atmosphere (award # NNX17AK10G). National Science Foundation, Biology Integration Institutes Program: EMERGE Biology Integration Institute (award # 2022070). United States Department of Energy Office of Biological and Environmental Research, Genomic Science Program: The IsoGenie Project (grant #s DE-SC0004632, DE-SC0010580, and DE-SC0016440). Sequencing was performed using startup funding from the University of Arizona to Virginia Rich. We thank the Swedish Polar Research Secretariat and SITES for the support of the work done at the Abisko Scientific Research Station. SITES is supported by the Swedish Research Council's grant 4.3-2021-00164. 

Stordalen Mire sample metadata from a mire-wide survey (2015) and co-analyzed autochamber site samples (2014-2015). These samples were analyzed by 16S rRNA amplicon sequencing, and the 16S data is available under NCBI BioProject PRJNA1236848. Column descriptions for this metadata file: The first 4 columns (sample_name, SRA library_ID, SRA accession, BioSample) include sample & library names and accessions in NCBI. The sample_name column also matches the SampleID__ attribute in the EMERGE Database (EMERGE-DB; https://emerge-db.asc.ohio-state.edu/). The next 7 columns (SampleID, Habitat, Depth, Description, Source, Site, Origin) are the metadata used for the 16S data analysis (results available at https://doi.org/10.5281/zenodo.15047596 and https://doi.org/10.5281/zenodo.15047715). The final 9 columns (Latitude, Longitude, Date, Full Site Name, Core #, DepthMin (cm), DepthMax (cm), DepthAvg (cm), pH_porewater) provide other metadata, including latitude/longitude, sampling dates, full site and core names, depths, and porewater pH, standardized to match the nomenclature in the EMERGE-DB. FUNDING: National Aeronautics and Space Administration, Interdisciplinary Science program: From Archaea to the Atmosphere (award # NNX17AK10G). National Science Foundation, Biology Integration Institutes Program: EMERGE Biology Integration Institute (award # 2022070). United States Department of Energy Office of Biological and Environmental Research, Genomic Science Program: The IsoGenie Project (grant #s DE-SC0004632, DE-SC0010580, and DE-SC0016440). Sequencing was performed using startup funding from the University of Arizona to Virginia Rich. We thank the Swedish Polar Research Secretariat and SITES for the support of the work done at the Abisko Scientific Research Station. SITES is supported by the Swedish Research Council's grant 4.3-2021-00164. 

This is a pre-release of this MAG set, which is now published in ENA under BioProject PRJNA386568.FILES: mags_emerge_20230110.tar.gz - Archive containing MAG files (.fna). metadata_MAGs_EMERGE.tsv - Table containing MIMAG(5.0)-formatted sample attributes, genome information, and other metadata for the MAGs. This table also includes JGI or NCBI genome accession #s for some additional MAGs that are not part of the .tar.gz archive.  NEW in Version 1.0.0: Added source metagenome accessions, including SRA runs (derived_from) and BioSamples (metaG_biosample), for all MAGs where this info was available. Added other metadata (including SampleID__, assembly methods, and sequencing technology) that was previously absent for the externally-cited MAGs.   FUNDING: This research is a contribution of the EMERGE Biology Integration Institute (https://emerge-bii.github.io/), funded by the National Science Foundation, Biology Integration Institutes Program, Award # 2022070. This study was also funded by the Genomic Science Program of the United States Department of Energy Office of Biological and Environmental Research, grant #s DE-SC0004632. DE-SC0010580. and DE-SC0016440. We thank the Swedish Polar Research Secretariat and SITES for the support of the work done at the Abisko Scientific Research Station. SITES is supported by the Swedish Research Council's grant 4.3-2021-00164. Data collected at the Joint Genome Institute was generated under the following awards: The majority of sequencing at JGI was supported by BER Support Science Proposal 503530 (DOI: 10.46936/10.25585/60001148), conducted by the U.S. Department of Energy Joint Genome Institute (https://ror.org/04xm1d337), a DOE Office of Science User Facility supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC02-05CH11231. Sequencing of SIP samples was performed under the Facilities Integrating Collaborations for User Science (FICUS) initiative (proposal 503547; award DOI: 10.46936/fics.proj.2017.49950/60006215) and used resources at the DOE Joint Genome Institute (https://ror.org/04xm1d337) and the Environmental Molecular Sciences Laboratory (https://ror.org/04rc0xn13), which are DOE Office of Science User Facilities. Both facilities are sponsored by the Office of Biological and Environmental Research and operated under Contract Nos. DE-AC02-05CH11231 (JGI) and DE-AC05-76RL01830 (EMSL). 

This release (MAGs v2) is a major new version of this metagenome-assembled genome (MAG) set. All previous releases on this page (which only differ in the metadata) are designated "MAGs v1." The current release (MAGs v2) uses CheckM2 v1.0.2 filtering (≥70% completeness, ≤10% contamination) to expand this dataset to include 36,419 MAGs, with the following subcategories: Cronin_v1:  Manually-curated subset of the "Field" category from MAGs v1. Cronin_v2:  MAGs from raw bin filtering on the same assemblies used to generate Cronin_v1. Woodcroft_v2:  MAGs from raw bin filtering on the same assemblies used to generate the MAGs reported in Woodcroft & Singleton et al. (2018). SIPS:  Updated genomes from samples originating from a stable isotope probing (SIP) incubation experiment by Moira Hough et al. ("SIP" in MAGs v1), re-analyzed due to read truncation and sample linkage issues in MAGs v1. JGI:  Expanded set of genomes from the Joint Genome Institute's metagenome annotation pipeline.   FILES: Emerge_MAGs_v2.tar.gz - Archive containing the MAG files (.fna). metadata_MAGs_v2_EMERGE.tsv - Table containing source sample names and accessions, GTDB taxonomy information, CheckM2 quality reports, NCBI GenomeBatch- and MIMAG(6.0)-formatted sample attributes and other metadata for the MAGs.    FUNDING: This research is a contribution of the EMERGE Biology Integration Institute (https://emerge-bii.github.io/), funded by the National Science Foundation, Biology Integration Institutes Program, Award # 2022070. This study was also funded by the Genomic Science Program of the United States Department of Energy Office of Biological and Environmental Research, grant #s DE-SC0004632. DE-SC0010580. and DE-SC0016440. We thank the Swedish Polar Research Secretariat and SITES for the support of the work done at the Abisko Scientific Research Station. SITES is supported by the Swedish Research Council's grant 4.3-2021-00164. Data collected at the Joint Genome Institute was generated under the following awards: The majority of sequencing at JGI was supported by BER Support Science Proposal 503530 (DOI: 10.46936/10.25585/60001148), conducted by the U.S. Department of Energy Joint Genome Institute (https://ror.org/04xm1d337), a DOE Office of Science User Facility supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC02-05CH11231. Sequencing of SIP samples was performed under the Facilities Integrating Collaborations for User Science (FICUS) initiative (proposal 503547; award DOI: 10.46936/fics.proj.2017.49950/60006215) and used resources at the DOE Joint Genome Institute (https://ror.org/04xm1d337) and the Environmental Molecular Sciences Laboratory (https://ror.org/04rc0xn13), which are DOE Office of Science User Facilities. Both facilities are sponsored by the Office of Biological and Environmental Research and operated under Contract Nos. DE-AC02-05CH11231 (JGI) and DE-AC05-76RL01830 (EMSL). 

Recovered microbial community structure is known to be influenced by sample storage conditions and nucleic acid extraction methods, and the impact varies by sample type. Peat soils store a large portion of soil carbon and their microbiomes mediate climate feedbacks. Here, we tested three storage conditions and five extraction protocols on peat soils from three physicochemically distinct habitats in Stordalen Mire, Sweden, revealing significant methodological impacts on microbial (here, meaning bacteria and archaea) community structure. Initial preservation method impacted alpha but not beta diversity, with in-field storage in LifeGuard buffer yielding roughly two-thirds the richness of in-field flash-freezing or transport from the field on ice (all samples were stored at −80 °C after return from the field). Nucleic acid extraction method impacted both alpha and beta diversity; one method (the PowerSoil Total RNA Isolation kit with DNA Elution Accessory kit) diverged from the others (PowerMax Soil DNA Isolation kit-High Humic Acid Protocol, and three variations of a modifiedPowerMax Soil DNA/RNA isolation kit), capturing more diverse microbial taxa, with divergent community structures. Although habitat and sample depth still consistently dominated community variation, method-based biases in microbiome recovery for these climatologically-relevant soils are significant, and underscore the importance of methodological consistency for accurate inter-study comparisons, long-term monitoring, and consistent ecological interpretations. 

Abstract Soil microorganisms are pivotal in the global carbon cycle, but the viruses that affect them and their impact on ecosystems are less understood. In this study, we explored the diversity, dynamics, and ecology of soil viruses through 379 metagenomes collected annually from 2010 to 2017. These samples spanned the seasonally thawed active layer of a permafrost thaw gradient, which included palsa, bog, and fen habitats. We identified 5051 virus operational taxonomic units (vOTUs), doubling the known viruses for this site. These vOTUs were largely ephemeral within habitats, suggesting a turnover at the vOTU level from year to year. While the diversity varied by thaw stage and depth‐related patterns were specific to each habitat, the virus communities did not significantly change over time. The abundance ratios of virus to host at the phylum level did not show consistent trends across the thaw gradient, depth, or time. To assess potential ecosystem impacts, we predicted hostsin silicoand found viruses linked to microbial lineages involved in the carbon cycle, such as methanotrophy and methanogenesis. This included the identification of viruses ofCandidatusMethanoflorens, a significant global methane contributor. We also detected a variety of potential auxiliary metabolic genes, including 24 carbon‐degrading glycoside hydrolases, six of which are uniquely terrestrial. In conclusion, these long‐term observations enhance our understanding of soil viruses in the context of climate‐relevant processes and provide opportunities to explore their role in terrestrial carbon cycling. 

ABSTRACT While wetlands are major sources of biogenic methane (CH₄), our understanding of resident microbial metabolism is incomplete, which compromises the prediction of CH₄emissions under ongoing climate change. Here, we employed genome-resolved multi-omics to expand our understanding of methanogenesis in the thawing permafrost peatland of Stordalen Mire in Arctic Sweden. In quadrupling the genomic representation of the site’s methanogens and examining their encoded metabolism, we revealed that nearly 20% of the metagenome-assembled genomes (MAGs) encoded the potential for methylotrophic methanogenesis. Further, 27% of the transcriptionally active methanogens expressed methylotrophic genes; forMethanosarcinalesandMethanobacterialesMAGs, these data indicated the use of methylated oxygen compounds (e.g., methanol), while forMethanomassiliicoccales, they primarily implicated methyl sulfides and methylamines. In addition to methanogenic methylotrophy, >1,700 bacterial MAGs across 19 phyla encoded anaerobic methylotrophic potential, with expression across 12 phyla. Metabolomic analyses revealed the presence of diverse methylated compounds in the Mire, including some known methylotrophic substrates. Active methylotrophy was observed across all stages of a permafrost thaw gradient in Stordalen, with the most frozen non-methanogenic palsa found to host bacterial methylotrophy and the partially thawed bog and fully thawed fen seen to house both methanogenic and bacterial methylotrophic activities. Methanogenesis across increasing permafrost thaw is thus revised from the sole dominance of hydrogenotrophic production and the appearance of acetoclastic at full thaw to consider the co-occurrence of methylotrophy throughout. Collectively, these findings indicate that methanogenic and bacterial methylotrophy may be an important and previously underappreciated component of carbon cycling and emissions in these rapidly changing wetland habitats. IMPORTANCEWetlands are the biggest natural source of atmospheric methane (CH₄) emissions, yet we have an incomplete understanding of the suite of microbial metabolism that results in CH₄formation. Specifically, methanogenesis from methylated compounds is excluded from all ecosystem models used to predict wetland contributions to the global CH₄budget. Though recent studies have shown methylotrophic methanogenesis to be active across wetlands, the broad climatic importance of the metabolism remains critically understudied. Further, some methylotrophic bacteria are known to produce methanogenic by-products like acetate, increasing the complexity of the microbial methylotrophic metabolic network. Prior studies of Stordalen Mire have suggested that methylotrophic methanogenesis is irrelevantin situand have not emphasized the bacterial capacity for metabolism, both of which we countered in this study. The importance of our findings lies in the significant advancement toward unraveling the broader impact of methylotrophs in wetland methanogenesis and, consequently, their contribution to the terrestrial global carbon cycle.