Depth-specific distribution of bacterial MAGs in permafrost active layer in Ny Ålesund, Svalbard (79°N).

Arctic soil microbial communities may shift with increasing temperatures and water availability from climate change. We examined temperature and volumetric liquid water content (VWC) in the upper 80 cm of permafrost-affected soil over 2 years (2018-2019) at the Bayelva monitoring station, Ny Ålesund, Svalbard. We show VWC increases with depth, whereas in situ temperature is more stable vertically, ranging from -5°C to 5 °C seasonally. Prokaryotic metagenome-assembled genomes (MAGs) were obtained at 2-4 cm vertical resolution collected while frozen in April 2018 and at 10 cm vertical resolution collected while thawed in September 2019. The most abundant MAGs were Acidobacteriota, Actinomycetota, and Chloroflexota. Actinomycetota and Chloroflexota increase with depth, while Acidobacteriota classes Thermoanaerobaculia Gp7-AA8, Blastocatellia UBA7656, and Vicinamibacteria Vicinamibacterales are found above 6 cm, below 6 cm, and below 20 cm, respectively. All MAGs have diverse carbon-degrading genes, and Actinomycetota and Chloroflexota have autotrophic genes. Genes encoding ß -glucosidase, N-acetyl-ß-D-glucosaminidase, and xylosidase increase with depth, indicating a greater potential for organic matter degradation with higher VWC. Acidobacteriota dominate the top 6 cm with their classes segregating by depth, whereas Actinomycetota and Chloroflexota dominate below ∼6 cm. This suggests that Acidobacteriota classes adapt to lower VWC at the surface, while Actinomycetota and Chloroflexota persist below 6 cm with higher VWC. This indicates that VWC may be as important as temperature in microbial climate change responses in Arctic mineral soils. Here we describe MAG-based Seqcode type species in the Acidobacteriota, Onstottus arcticum, Onstottus frigus, and Gilichinskyi gelida and in the Actinobacteriota, Mayfieldus profundus.

Chemolithoautotroph distributions across the subsurface of a convergent margin.

Subducting oceanic crusts release fluids rich in biologically relevant compounds into the overriding plate, fueling subsurface chemolithoautotrophic ecosystems. To understand the impact of subsurface geochemistry on microbial communities, we collected fluid and sediments from 14 natural springs across a ~200 km transect across the Costa Rican convergent margin and performed shotgun metagenomics. The resulting 404 metagenome-assembled genomes (MAGs) cluster into geologically distinct regions based on MAG abundance patterns: outer forearc-only (25% of total relative abundance), forearc/arc-only (38% of total relative abundance), and delocalized (37% of total relative abundance) clusters. In the outer forearc, Thermodesulfovibrionia, Candidatus Bipolaricaulia, and Firmicutes have hydrogenotrophic sulfate reduction and Wood-Ljungdahl (WL) carbon fixation pathways. In the forearc/arc, Anaerolineae, Ca. Bipolaricaulia, and Thermodesulfovibrionia have sulfur oxidation, nitrogen cycling, microaerophilic respiration, and WL, while Aquificae have aerobic sulfur oxidation and reverse tricarboxylic acid carbon fixation pathway. Transformation-based canonical correspondence analysis shows that MAG distribution corresponds to concentrations of aluminum, iron, nickel, dissolved inorganic carbon, and phosphate. While delocalized MAGs appear surface-derived, the subsurface chemolithoautotrophic, metabolic, and taxonomic landscape varies by the availability of minerals/metals and volcanically derived inorganic carbon. However, the WL pathway persists across all samples, suggesting that this versatile, energy-efficient carbon fixation pathway helps shape convergent margin subsurface ecosystems.

Novel taxa of Acidobacteriota implicated in seafloor sulfur cycling.

Acidobacteriota are widespread and often abundant in marine sediments, yet their metabolic and ecological properties are poorly understood. Here, we examined metabolisms and distributions of Acidobacteriota in marine sediments of Svalbard by functional predictions from metagenome-assembled genomes (MAGs), amplicon sequencing of 16S rRNA and dissimilatory sulfite reductase (dsrB) genes and transcripts, and gene expression analyses of tetrathionate-amended microcosms. Acidobacteriota were the second most abundant dsrB-harboring (averaging 13%) phylum after Desulfobacterota in Svalbard sediments, and represented 4% of dsrB transcripts on average. Meta-analysis of dsrAB datasets also showed Acidobacteriota dsrAB sequences are prominent in marine sediments worldwide, averaging 15% of all sequences analysed, and represent most of the previously unclassified dsrAB in marine sediments. We propose two new Acidobacteriota genera, Candidatus Sulfomarinibacter (class Thermoanaerobaculia, "subdivision 23") and Ca. Polarisedimenticola ("subdivision 22"), with distinct genetic properties that may explain their distributions in biogeochemically distinct sediments. Ca. Sulfomarinibacter encode flexible respiratory routes, with potential for oxygen, nitrous oxide, metal-oxide, tetrathionate, sulfur and sulfite/sulfate respiration, and possibly sulfur disproportionation. Potential nutrients and energy include cellulose, proteins, cyanophycin, hydrogen, and acetate. A Ca. Polarisedimenticola MAG encodes various enzymes to degrade proteins, and to reduce oxygen, nitrate, sulfur/polysulfide and metal-oxides. 16S rRNA gene and transcript profiling of Svalbard sediments showed Ca. Sulfomarinibacter members were relatively abundant and transcriptionally active in sulfidic fjord sediments, while Ca. Polarisedimenticola members were more relatively abundant in metal-rich fjord sediments. Overall, we reveal various physiological features of uncultured marine Acidobacteriota that indicate fundamental roles in seafloor biogeochemical cycling.

Woeseiales transcriptional response to shallow burial in Arctic fjord surface sediment.

Distinct lineages of Gammaproteobacteria clade Woeseiales are globally distributed in marine sediments, based on metagenomic and 16S rRNA gene analysis. Yet little is known about why they are dominant or their ecological role in Arctic fjord sediments, where glacial retreat is rapidly imposing change. This study combined 16S rRNA gene analysis, metagenome-assembled genomes (MAGs), and genome-resolved metatranscriptomics uncovered the in situ abundance and transcriptional activity of Woeseiales with burial in four shallow sediment sites of Kongsfjorden and Van Keulenfjorden of Svalbard (79°N). We present five novel Woeseiales MAGs and show transcriptional evidence for metabolic plasticity during burial, including sulfur oxidation with reverse dissimilatory sulfite reductase (dsrAB) down to 4 cm depth and nitrite reduction down to 6 cm depth. A single stress protein, spore protein SP21 (hspA), had a tenfold higher mRNA abundance than any other transcript, and was a hundredfold higher on average than other transcripts. At three out of the four sites, SP21 transcript abundance increased with depth, while total mRNA abundance and richness decreased, indicating a shift in investment from metabolism and other cellular processes to build-up of spore protein SP21. The SP21 gene in MAGs was often flanked by genes involved in membrane-associated stress response. The ability of Woeseiales to shift from sulfur oxidation to nitrite reduction with burial into marine sediments with decreasing access to overlying oxic bottom waters, as well as enter into a dormant state dominated by SP21, may account for its ubiquity and high abundance in marine sediments worldwide, including those of the rapidly shifting Arctic.

Evidence for a Growth Zone for Deep-Subsurface Microbial Clades in Near-Surface Anoxic Sediments.

Global marine sediments harbor a large and highly diverse microbial biosphere, but the mechanism by which this biosphere is established during sediment burial is largely unknown. During burial in marine sediments, concentrations of easily metabolized organic compounds and total microbial cell abundance decrease. However, it is unknown whether some microbial clades increase with depth. We show total population increases in 38 microbial families over 3 cm of sediment depth in the upper 7.5 cm of White Oak River (WOR) estuary sediments. Clades that increased with depth were more often associated with one or more of the following: anaerobes, uncultured, or common in deep marine sediments relative to those that decreased. Maximum doubling times (in situ steady-state growth rates could be faster to balance cell decay) were estimated as 2 to 25 years by combining sedimentation rate with either quantitative PCR (qPCR) or the product of the fraction read abundance of 16S rRNA genes and total cell counts (FRAxC). Doubling times were within an order of magnitude of each other in two adjacent cores, as well as in two laboratory enrichments of Cape Lookout Bight (CLB), NC, sediments (average difference of 28% ± 19%). qPCR and FRAxC in sediment cores and laboratory enrichments produced similar doubling times for key deep subsurface uncultured clades Bathyarchaeota (8.7 ± 1.9 years) and Thermoprofundales/MBG-D (4.1 ± 0.7 years). We conclude that common deep subsurface microbial clades experience a narrow zone of growth in shallow sediments, offering an opportunity for selection of long-term subsistence traits after resuspension events.IMPORTANCE Many studies show that the uncultured microbes that dominate global marine sediments do not actually increase in population size as they are buried in marine sediments; rather, they exist in a sort of prolonged torpor for thousands of years. This is because, although studies have shown biomass turnover in these clades, no evidence has ever been found that deeper sediments have larger populations for specific clades than shallower layers. We discovered that they actually do increase population sizes during burial, but only in the upper few centimeters. This suggests that marine sediments may be a vast repository of mostly nongrowing microbes with a thin and relatively rapid area of cell abundance increase in the upper 10 cm, offering a chance for subsurface organisms to undergo natural selection.

Mineralized microbialites as archives of environmental evolution, Laguna Negra, Catamarca Province, Argentina.

Environmental fluctuations are recorded in a variety of sedimentary archives of lacustrine depositional systems. Geochemical signals recovered from bottom sediments in closed-basin lakes are among the most sensitive paleoenvironmental indicators and are commonly used in reconstructing lake evolution. Microbialites (i.e., organosedimentary deposits accreted through microbial trapping and binding of detrital sediment or in situ mineral precipitation on organics [Palaios, 2, 1987, 241]), however, have been largely overlooked as paleoenvironmental repositories. Here, we investigate concentrically laminated mineralized microbialites from Laguna Negra, a high-altitude (4,100 m above sea level) hypersaline, closed-basin lake in northwestern Argentina, and explore the potential for recovery of environmental signals from these unique sedimentary archives. Spatial heterogeneity in hydrological regime helps define zones inside Laguna Negra, each with their own morphologically distinct microbialite type. Most notably, platey microbialites (in Zone 3A) are precipitated by evaporative concentration processes, while discoidal oncolites (in Zone 3C) are interpreted result from fluid mixing and biologically mediated nucleation. This spatial heterogeneity is reflected in petrographically distinct carbonate fabrics: micritic, botryoidal, and isopachous. Fabric type is interpreted to reflect a combination of physical and biological influences during mineralization, and paired C-isotope measurement of carbonate and organic matter supports ecological differences as a dominant control on C-isotopic evolution between zones. Laminae of Laguna Negra microbialites preserve a range of δ13 Ccarb from +5.75‰ to +18.25‰ and δ18 Ocarb from -2.04‰ to +9.28‰. Temporal trends of lower carbon and oxygen isotopic compositions suggest that the influence of CO2 degassing associated with evaporation has decreased over time. Combined, these results indicate that microbialite archives can provide data that aid in interpretation of both lake paleohydrology and paleoenvironmental change.

Methanogens in the Antarctic Dry Valley permafrost.

Polar permafrost is at the forefront of climate change, yet only a few studies have enriched the native methane-producing microbes that might provide positive feedbacks to climate change. Samples Ant1 and Ant2, collected in Antarctic Miers Valley from permafrost sediments, with and without biogenic methane, respectively, were evaluated for methanogenic activity and presence of methanogens. After a one-year incubation of both samples under anaerobic conditions, methane production was observed only at room temperature in microcosm Ant1 with CO2/H2 (20/80) as carbon and energy sources and was monitored during the subsequent 10 years. The concentration of methane in the headspace of microcosm Ant1 changed from 0.8% to a maximum of 45%. Archaeal 16S rRNA genes from microcosm Ant1 were related to psychrotolerant Methanosarcina lacustris. Repeated efforts at achieving a pure culture of this organism were unsuccessful. Metagenomic reads obtained for the methane-producing microcosm Ant1 were assembled and resulted in a 99.84% complete genome affiliated with the genus Methanosarcina. The metagenome assembled genome contained cold-adapted enzymes and pathways suggesting that the novel uncultured Methanosarcina sp. Ant1 is adapted to sub-freezing conditions in permafrost. This is the first methanogen genome reported from the 15 000 years old permafrost of the Antarctic Dry Valleys.

Interlaboratory quantification of Bacteria and Archaea in deeply buried sediments of the Baltic Sea (IODP Expedition 347).

Two common quantification methods for subseafloor microorganisms are catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH) and quantitative PCR (qPCR). Using these methods, we quantified Bacteria and Archaea in Baltic Sea basin sediments (IODP Exp. 347) down to 90 mbsf, testing the following hypotheses in an interlaboratory comparison: (1) proteinase K permeabilization of archaeal cell walls increases CARD-FISH accuracy and (2) qPCR varies by more than an order of magnitude between laboratories using similar protocols. CARD-FISH counts did not differ between permeabilization treatments, demonstrating that proteinase K did not increase accuracy of CARD-FISH counts. However, 91% of these counts were below the quantification limit of 1.3 × 107 cells cm-3. For qPCR, data varied between laboratories, but were largely within the same order of magnitude if the same primers were used, with 88% of samples being above the quantification limit. Copy number values were elevated by preparing a sediment slurry before DNA extraction: 3.88 × 106-2.34 × 109 16S rRNA gene copies cm-3 vs. 1.39 × 107-1.87 × 109 total cells cm-3. By qPCR, Bacteria were more abundant than Archaea, although they usually were within the same order of magnitude. Overall, qPCR is more sensitive than CARD-FISH, but both require optimization to consistently achieve both precision and accuracy.

Draft Genome Sequence of Antarctic Methanogen Enriched from Dry Valley Permafrost.

A genomic reconstruction belonging to the genus Methanosarcina was assembled from metagenomic data from a methane-producing enrichment of Antarctic permafrost. This is the first methanogen genome reported from permafrost of the Dry Valleys and can help shed light on future climate-affected methane dynamics.