The hypersaline soils of the Odiel Saltmarshes Natural Area in Southwest Spain harbor highly diverse microbial communities adapted to extreme conditions. However, their genomic diversity remains largely unexplored. In addition to high salinity, these soils are contaminated with heavy metals, creating a hostile environment of great interest for studying extremophilic microorganisms and their metabolic adaptations. This study aims to characterize the uncovered prokaryotic taxa as Candidatus species inhabiting the hypersaline soils of the Odiel Saltmarshes, based on their metagenomic assembled genomic sequences. The reconstructed genomes were assessed for quality based on completeness and contamination thresholds and subsequently taxonomically classified. Comparative genomic analysis of six high-quality MAGs revealed key metabolic traits related to survival under extreme salinity and heavy metal conditions. The findings provide new insights about microbial diversity of hypersaline environments and expand the catalog of known prokaryotic genomes. Detailed characterization of six novel Candidatus taxa highlights the unique adaptations of these microorganisms, enhancing our understanding of life in extreme habitats.

Subterranean estuaries (STEs) are key bioreactors regulating the quantity and chemical composition of groundwater-derived nitrogen (N) reaching coastal ecosystems. Yet, the microbial controls on N-cycling within these groundwater-seawater mixing zones remain poorly understood. We investigated the spatio-temporal variations in microbial communities and their N-cycling potential within an alluvial Mediterranean STE with high N concentration. We explored changes in microbial abundance, heterotrophic activity, taxonomic composition, and the abundance of N-cycling genes across groundwater samples collected at several depths and distances from the shoreline in winter and summer. Microbial abundance, activity, and diversity varied strongly across hydrochemical zones according to physicochemistry and aquifer depth but showed limited seasonality. Functional predictions suggested a complex, spatially structured suite of N pathways encoded by diverse taxa occupying different STE zones, and quantitative-PCR revealed niche partitioning between ammonia-oxidizing archaea, prevalent in fresh-groundwater, and bacterial denitrifiers enriched in deep-saline layers. Multiple linear model predictions showed a stronger fit for NO2 - and NH4 + concentrations when using microbial properties than when using environmental variables, highlighting their importance for understanding N cycling in STEs. Our results suggest that the functional potential of the STE microbiome is complex and spatially structured across hydrochemical zones, explaining spatial variations in STE N-cycling.

Photodegradation in lakes is a major sink for the toxin methylmercury (MeHg) in forest-wetland-lake ecosystems. Previous attempts to estimate annual rates of MeHg photodegradation in lakes have followed a "black-box approach", where the process has been related to incident sunlight rather than photon absorption. Here we use experimental data from three contrasting dark and clear boreal lakes to develop the first apparent quantum yield (AQY) model for spectral MeHg photodegradation rates in lakes. The model was proven universal by its ability to predict experimental data from 22 lakes representing five global regions, covering wide ranges in dissolved organic carbon (DOC), specific UV absorbance at 254 nm, and pH (1.8-39.5 mg C L-1, 1.7-5.7 L mg-1 C m-1 and 4.8-8.5, respectively). The AQY model manifests a dual role played by dissolved organic matter (DOM) as a sensitizer, by producing reactive transient species (RTS) upon photon absorbance, and as an inhibitor, by scavenging of RTS by antioxidants associated with aromatic structures in DOM. Using site-specific data on direct and diffuse solar irradiance, MeHg concentrations, and spectral light absorption properties in 1033 lakes, we estimate an annual MeHg photodegradation rate of 12.1 kg y-1 in the total volume of Swedish lakes. This value corresponds to 24% of the estimated 51 kg of MeHg that annually is transported with runoff from soil into the same lakes. By the AQY model, we calculate the first regional estimates of MeHg photodegradation in lakes of boreal and temperate Europe, temperate North America, subtropical North America, and tropical South America, providing a basis for the establishment of regional MeHg cycling models.

Deep groundwaters populated by diverse and active microbes are among the most energy and nutrient-limited ecosystems. Characteristics of this ecosystem (including nutrient and dispersal limitations, low cell densities, and an episodic growth strategy) interactively underpin the so far elusive eco-evolutionary dynamics of its microbiome. Here, we used genome-resolved modular metabolic analyses of disconnected deep groundwater sites in the Fennoscandian Shield to test how eco-evolutionary constraints in these deep groundwater ecosystems shape microbial genome architecture, metabolic versatility, and community assembly at different depths. The analysis revealed that lineages with larger genomes (≥ 2.6 Mb) maintained higher population sizes in the deepest and most oligotrophic groundwaters, whereas lineages with known metabolic dependencies, such as and DPANN, declined in relative abundance with depth. This pattern was interpreted as consistent with limited opportunities for sustained metabolic cross-feeding in these ecosystems. Moreover, while similar ecological niches based on cross-feeding interactions and potential primary production were available across different boreholes, distinct microbial lineages appeared to occupy these niches at each site. The findings provided new insights into the role of metabolic cross-feeding in genome evolution and community assembly of deep groundwater microbiomes. By extending the streamlining theory, this study underscores the critical influence of ecological interactions, particularly metabolic exchanges, in shaping microbial life under severe nutrient limitation, offering new insights into subsurface microbial communities.

Mercury (Hg) has been stored in permafrost peatlands for millennia. As permafrost thaw is predicted to increase with ongoing climate warming, Hg is at risk to be remobilized from those peatlands and hotspots for Hg methylation could potentially form. Monomethylmercury (MeHg) is a known neurotoxin and a health concern to northern communities if Hg is remobilized, transformed to MeHg and subsequently bioaccumulated in the food chain. It is uncertain how Hg cycles in thawing permafrost systems and how much of it could potentially be remobilized by thaw processes. In this study, we have investigated Hg dynamics in a permafrost peatland of northern Sweden, where a snow fence field experiment was set up in 2005 to simulate accelerated permafrost thaw through winter warming. We compared total mercury (THg) and MeHg concentrations in soil plots representing intact and thawed permafrost conditions, investigated seasonal variations and examined the coupling between microbial community composition and MeHg concentration. Similar stocks of both total THg and MeHg were observed in intact and thawed permafrost conditions, suggesting that 17 years of winter warming manipulation and accelerated permafrost thaw had not led to substantial Hg loss from the peat nor extensive MeHg production. The apparent stability of the Hg stocks contrasted with our hypothesis and with many previous studies. While there was no difference in microbial communities between treatments with or without accelerated thaw, putative methylators were more abundant in thaw plots in the fall. This indicates that permafrost thaw has increased the potential for Hg methylation, although these shifts have not yet been strong enough to measurably affect MeHg stocks. Our study emphasizes the complexity of Hg dynamics in thaw-affected permafrost landscapes and the need to consider thaw-related perturbations of the Hg cycle on various timescales.

The continental subsurface hosts energy-constrained groundwaters with a high diversity of ecologically elusive microorganisms adapted to the prevailing low-energy conditions. This study explored potential interactions among microbes using anaerobic enrichment incubations with three types of groundwater of contrasting hydrochemistry from the Äspö Hard Rock Laboratory, Sweden. Removing cells larger than 0.45 µm from the inoculum resulted in incubations enriched in populations characterized by very small genomes, including Patescibacteria, Nanobdellota, and Omnitrophota. These incubations had a higher diversity than non-fractionated incubations. However, cell numbers and community structure of the fractionated incubations did not change over an incubation period up to four months, despite high microbial diversity and experimental amendments with either simple (acetate) or more complex (cell lysate) carbon sources. In addition, network analysis on the groundwaters revealed multiple co-occurrences between populations affiliated with the Patescibacteria and the Desulfobacterota. Overall, these findings support that a considerable part of microbial diversity has a small cell size in these low energy groundwaters and strong co-occurrences among populations as an important survival strategy.

Microeukaryotes are abundant, diverse, and morphologically and functionally complex. They represent one of the largest groups of primary producers and are involved in major ecosystem functions such as nutrient transformations and maintenance of food webs. Although ~74,000 species of microeukaryotes are estimated to exist, very few are available in culture and only ~3000 are represented by reference genomes and transcriptomes. This limited representation significantly hinders the study of microeukaryotes in their natural environments. Single-cell transcriptomics with sequencing of full-length transcripts has potential to bypass this limitation. In this study, we employ Smart-seq3xpress for taxonomic identification and functional characterization of non-model mixotrophic freshwater microeukaryotes in a culture-free and reference-free manner, while also enabling inferences to be made for their prokaryotic associations. Computational analyses revealed 22 distinct microeukaryote taxa upon sequencing of 1520 randomly sampled cells, with assemblies of 12 good quality de novo partial transcriptomes, followed by functional annotation using sequence and structure homology. Gene expression analysis of the transcriptome for the most abundant microeukaryote in the sample, Rhodomonas, revealed a transcriptomic dip a few hours after experimental shading, followed by a gradual recovery in the next few days. Compared to samples in illuminated conditions, there was widespread downregulation of photosynthesis-, lysosome-, and carbon metabolism-related pathways in the shade, but the prevailing associations with prokaryotic lineages was unaffected by the light conditions. The different microeukaryotes, all present in the same environmental sample, featured distinct prokaryotic associates that were consistent across illumination conditions. This reference-free and culture-free study of 22 different microeukaryotic taxa sets the stage for high-throughput single cell transcriptomics to study microeukaryotic diversity in complex natural ecosystems, for their population-level taxonomic identity, sub-population-level transcriptomic profiles and metabolic states, and individual-level prokaryotic associations.

The effect of spatial factors reflecting dispersal potential between sites versus local environmental conditions on freshwater planktonic communities remains poorly understood. We assessed differences in the relative importance of local and seasonal environmental conditions versus spatial factors in explaining differences in community composition (i.e., beta-diversity patterns) in microbial plankton of 39 mid-latitude Chilean lakes spanning representative ecological gradients in altitude, mixing depth and water chemistry. The assemblages were taxonomically profiled by paired-end high throughput sequencing of the V3-V4 region of the 16S and the V4 region of the 18S rRNA genes. Variation partitioning analyses revealed that the explanatory power of environmental and seasonal factors versus spatial variables and their mutual overlap varied considerably among taxa and functional groups. More than 12 % of the variation in community structure was uniquely explained by environmental factors in the phytoplankton groups Dinophyta, Ochrophyta and Cyanobacteria, as well as in oligotrophic ultramicrobacteria, such as small rhodopsin containing Actinobacteria and LD12 Alphaproteobacteria. In phago-heterotrophic and saprotrophic groups, including heterotrophic micro-eukaryotes, and Bacteroidetes, environmental factors explained a smaller or even insignificant portion of the differences in the community structure. Our findings suggest that in Chilean lake microplankton, complex traits related to ecological and trophic strategy appear to affect the relative effect of local environmental properties on their community composition and hence the strength of species sorting along limnological gradients.

With climate change-induced sea ice decline in the Arctic Ocean, nitrogen is expected to become an increasingly important determinant of primary productivity. Nitrogen fixation is the conversion of molecular nitrogen to bioavailable ammonium by microorganisms called diazotrophs. Here, we report nitrogen fixation rates, diazotroph composition, and expression under different stages of declining sea ice in the Central Arctic Ocean (multiyear ice, five stations) and the Eurasian Arctic (marginal ice zone, seven stations). Nitrogen fixation in the Central Arctic Ocean was positively correlated with primary production, ranging from 0.4 ± 0.1 to 2.5 ± 0.87 nmol N L-1 d-1. Along two transects across the marginal ice zone, nitrogen fixation varied between days and ice regime from below detection up to 5.3 ± 3.65 nmol N L-1 d-1 associated with an ice-edge phytoplankton bloom. We show nitrogen fixation in sea ice-covered waters of the Arctic Ocean and provide insight into present and active non-cyanobacterial diazotrophs in the region.

Constructed wetlands are widely used to reduce nutrient loading to downstream waters, but they can also emit methane, a potent greenhouse gas. This trade-off between water quality benefits and climate impacts is driven by microbial processes that remain poorly understood in winter. We examined microbial community composition and methane-cycling potential in surface water samples from constructed wetlands in two agricultural regions of Sweden during the winter season, focusing on the effects of emergent vegetation and environmental conditions. Western wetlands, characterized by higher total nitrogen and dissolved oxygen, exhibited significantly greater microbial diversity and more complex co-occurrence networks than eastern wetlands. At the phylum level, Actinobacteriota and Firmicutes were more abundant in the west, while Bacteroidota dominated the east. The effects of emergent vegetation were region-specific: in the west, vegetated zones supported higher diversity and enrichment of plant-associated taxa. Several taxa affiliated with methanotrophs showed higher relative abundance in vegetated zones of the western wetlands, suggesting vegetation may enhance methane oxidation potential in surface waters, even though methane concentrations were similar. Overall, winter microbial networks remained structured, emphasizing the need for integrated microbial and biogeochemical studies to guide wetland design features, such as vegetation and nutrient regimes, that support both methane mitigation and nutrient retention in cold-climate agricultural landscapes.

Seagrass meadow ecosystems offer several valuable ecosystem services in coastal regions around the world. Recent studies have suggested that one such important service is reduction of pathogenic bacteria, specifically Vibrio spp., in adjacent waters. The specific mechanisms of pathogen reduction remain unclear, although increased sedimentation has been suggested as one likely process for pathogens to be quenched from the water column. Whether Vibrio spp. persist in the sediment or in other compartments of the seagrass meadow is currently unknown, but it has been shown that marine surface biofilms can function as reservoirs of pathogenic vibrios. This general feature may also apply to seagrass and sediment surfaces. In this study, we investigated the relative abundance and community ecology of Vibrio spp. bacteria in Baltic Sea seagrass meadows using both culturing and culture-independent methods. While we did not detect a significant reduction of Vibrio spp. in the water column above unvegetated sites as compared to seagrass meadows, we observed high relative abundances of Vibrio spp. on seagrass roots. This supports previous observations that marine surfaces are selectively colonized by Vibrio spp., implying that these habitats are important for the persistence and possibly release of Vibrio spp. into the water column. Our results emphasize the need to understand the interactions of pathogenic bacteria with coastal habitats, including interactions with host organisms such as seagrasses that provide biofilm microenvironments, in order to understand how diseases associated with these organisms develop.

In the Boreal region, extreme seasonal variations in day-night length expose communities to dynamic light and temperature fluctuations. Freshwater bacterioplankton, representing key ecosystem components, faces climate-driven shifts; yet the fixed day-length patterns determined by latitude underscore the importance of studying light's role in predicting ecosystem responses. We investigated bacterial community composition in a brown peat bog and a clear oligotrophic lake across seasons with contrasting light regimes: the summer solstice (> 20 h of daylight) and the autumn equinox (equal day-night length). Using amplicon sequencing of 16S rRNA transcripts, alongside measurements of physicochemical parameters, organic matter characterisation and dissolved carbon dioxide and methane gas measurements, we found no diel cycling in the lake during either period or in the peat bog near the summer solstice. However, the structure of bacterial peat bog communities exhibited cyclic changes over diel cycles at the autumn equinox. Twelve amplicon sequence variants, including both phototrophic and heterotrophic taxa, increased in abundance at all measured morning sampling times. These findings provide valuable insights into the diel patterns of boreal lentic habitats and their bacterioplankton communities, highlighting the absence of diel fluctuations in some systems and seasons, while revealing cyclic dynamics in others, driven by conditionally rare taxa.

The extensive use of Aqueous Film Forming Foam (AFFF) has led to substantial contamination by per- and polyfluoroalkyl substances (PFAS) of soils and groundwater at many firefighting training sites in Sweden and worldwide. PFAS is a group of extremely persistent anthropogenic substances that pose risk for adverse effect even at low levels. There is consequently a need to understand the potential for natural degradation of these compounds and the controlling environmental factors. Understanding the microbial capacity to degrade and transform PFAS is also crucial for comprehending their transport in soil and groundwater and for the development of a potential bioremediation technique. Despite commonly referred to as “forever chemicals”, there is emerging evidence of PFAS being microbially degraded in laboratory settings. The aim of this study is to investigate the microbial degradation capacity of the natural bacteria at two firefighting training sites (FFTS) contaminated with PFAS. Utilizing a sonic drill, soil samples were collected from both above and below the water table from FFTS near Örnsköldsvik and Sundsvall Timrå airports in Sweden. Enrichment cultures were initiated by mixing these soil samples with four different growth media—two for aerobic and two for anaerobic incubations. The incubation conditions, aerobic or anaerobic, were determined dependent on if the sample was taken above or below the groundwater level. All incubations were spiked with perfluorooctanesulfonic acid (PFOS), perfluorooctanoic acid (PFOA), 6:2 fluorotelomer sulfonic acid (6:2FTSA) and perfluorooctane sulfonaminde (FOSA) to reach a concentration of 9ppm. Samples from the incubations were taken at monthly intervals to screen for fluoride production, as an indicator for PFAS degradation, using ion chromatography. Using this approach, we aim to uncover the capability of the natural microbial community at these sites to degrade PFAS. In the second phase of this study, this will be followed by careful analysis of degradation products with the aim to identify degradation pathways.

Fungi play essential roles across ecosystems, yet their diversity in aquatic environments remains poorly understood compared to terrestrial systems. To address this gap, we analyzed metagenomes from 26 lakes in the boreal and subarctic zones, along with one tropical reservoir, to characterize fungal and fungal-like (Oomycota) community structure. We also examined environmental factors shaping these communities. Most variation in fungal composition was explained by lake identity, depth layer, and season, with total organic carbon as a significant explanatory variable. Despite geographic and time differences, dominant fungal phyla, orders, and genera were largely consistent across all lakes. However, genus-level variation indicated distinct community compositions likely influenced by differences in carbon substrate availability. Attempts to classify metagenomic reads down to the species level-illustrated here through the well-characterized oomycete genus Phytophthora-were constrained by the limited taxonomic resolution of current reference databases. While metagenomics offers powerful means to investigate entire microbial communities, our results underscore a persistent bottleneck: the insufficient representation of aquatic eukaryotic genomes in public databases.

Deoxygenation in aquatic ecosystems threatens biodiversity at all levels of functional and genetic diversity. Recent studies have shown the prevalence of microorganisms that transform mercury into neurotoxic methylmercury (mercury methylators-hgcA+ prokaryotes) in oxygen-deficient water columns. As climate warming expands coastal oxygen minimum zones, ongoing and near-future changes may ultimately lead to increased methylmercury formation. However, little is known about the presence of aquatic mercury methylators before the Industrial Revolution, marked by increased mercury emissions and deposition in the environment. Here we have detected hgcA genes in Black Sea sedimentary archives, with the highest abundance 9,000-5,500 years ago when anoxic conditions were documented in the water column. Historical sedimentary and modern water column data on mercury methylators provide valuable insights for projecting future methylmercury production in aquatic ecosystems impacted by ongoing deoxygenation. It also underscores the potential impacts of climate change on human exposure to methylmercury from mercury-contaminated seafood.

The effects of microplastic (MP) accumulation in freshwaters on organisms and ecosystem functions are poorly understood, as are the roles of MP particle properties in regulating these effects. In freshwater microcosms, we quantified variation in microbial communities and ecosystem functions and compared effects of MP concentration (0, 1000, 50000 particles/kgsediment), shape (sphere, fragment, fibre), and polymer (polyethylene, polyethylene terephthalate, polypropylene, polystyrene) with those of a model invertebrate consumer (Chironomus riparius). We detected multiple effects of specific MP properties, especially associated with MP fragments and fibres, and the polymer polypropylene. These effects included increases in microbial abundance, consumer biomass and ecosystem respiration, as well as decreases in microbial enzyme activity and water chlorophyll-a. MP presence was also associated with increased relative abundance of microbial taxa reported to degrade plastics. However, consumer presence mostly had stronger effects (effect sizes ranging from f 11 -313 %) than MP exposure (effect sizes ranging from f 1-89 %) on microbial communities and ecosystem functions. Furthermore, several MP effects were only detected when chironomid consumers were absent. Overall, our findings suggest that MP effects on microbes and ecosystem functions are often relatively small and variable, depending on particle properties and consumer presence. Nevertheless, the number of MP effects detected highlights the need for further investigations of interactions between MPs and other environmental drivers, to more thoroughly assess the risks of MP pollution for freshwater ecosystems.

Pollution by brominated flame retardants (BFRs) and per- and polyfluoroalkyl substances (PFAS) has the potential to alter the composition and activity of natural bacterial communities. However, the variation of bacterial communities under the combined influence of both pollutants was not clear. In order to investigate the distinct impacts, two sampling campaigns were carried out in an estuary contaminated with both BFRs and PFAS. An intriguing finding was that the bacterial communities exhibited obvious different responses to PFAS and BFRs pollution. This implies that different types of organic pollutants exert diverse promoting or inhibitory effects on bacterial taxa, which may further result in distinct changes in the functions of microorganisms. Moreover, the results of the effects of individual PFAS and BFR substances on bacterial communities revealed that pollutants with different molecular structures have varying impacts on the microbial community. Certain bacterial taxa showed specific responses to BDE209 (Decabromodiphenyl ether), PBDEs (poly brominated diphenyl ethers), and PFO5DoA (perfluorinated 3,5,7,9, 11-pentaxododecanoic acid). This suggests that bacterial communities have a particular response to substances containing ether bonds or long perfluorocarbon chains. Additionally, a number of PFAS-associated bacterial taxa were consistent with the findings of our previous microcosm experiments. This confirms that the PFAS-related taxa monitored in this study truly reflect the response of the bacterial community to contamination. The present study offers novel perspectives on the relationships between emerging contaminants, traditional organic pollutants, and native bacterial communities, and is expected to facilitate the achievement of high-resolution bacteria-based organic pollution monitoring in coastal and estuary ecosystems.

Microbial methane generation (methanogenesis) is an important metabolic process in the terrestrial deep biosphere and is an analog to early Earth as it is proposed to be one of the most ancient metabolisms on Earth. Signs of methanogenesis in meteorite impact craters are of particular interest in this respect as these settings are proposed hot spots for deep microbial colonization of the upper crust. Yet, reports of active deep rock-hosted methanogenesis are scarce, particularly for methylotrophic methanogenesis, while reports from terrestrial meteorite impact craters are completely lacking. Here, we used indigenous communities in cultures enriched from 400-m deep fluids to confirm and characterize active methane production from several carbon donors, including indigenous oil, in a terrestrial impact crater at Siljan, Sweden. Metagenomic and metatranscriptomic data of the methane-producing cultures revealed a consortium dominated by Acetobacterium sp. KB-1 and Candidatus Methanogranum gryphiswaldense, mediating methanogenesis solely via the methyl-reduction pathway, and resulting in a δ13Cmethanol-methane isotope enrichment of up to 98.6‰. These results provide insights into methylotrophic methanogenesis in deep subsurface environments in general, and in particular in fractured meteorite impact structures.IMPORTANCEThis study revealed that microbes enriched from groundwater in a 380-m deep borehole within the Siljan meteorite impact crater in Sweden were capable of producing methane, a key greenhouse gas. This is especially significant because it is the first proof of active methanogens in an impact crater and showing a specific pathway of methane production-methylotrophic methanogenesis-is present in the deep terrestrial subsurface, an environment that is typically hard to study. These findings shed light on life in extreme conditions on Earth and show that meteorite craters can be biological hotspots, rich with ancient life processes.

Microbial community variation in estuaries had been a research hotspot in recent years, but the effects of hydrodynamic forces on bacterial community redistribution remain underexplored. This study presents fixed-point observations of bacterial community redistribution under hydrodynamic influence during both dry and wet seasons. Alpha diversity analysis indicated that the bacterial community in the dry season exhibited higher richness and diversity than in the wet season, likely due to weaker hydrodynamics and lower temperatures. Cooccurrence network analysis showed that particle-associated (PA) communities had lower network diameter, graph density, and average path length than free-living (FL) communities. The FL community was more affected by vertical mixing, while the PA community was more affected by bottom flow. It is speculated that variation in the FL community results from the mixing of seawater and river water bacteria, which are more affected by surface currents, whereas PA community variation is driven by sedimentation and the bottom flow transport of particles. Unlike FL taxa, which correlated with real-time flow, PA taxa showed a stronger correlation with the flow data 1 h in the past, likely due to the greater inertial resistance of particulate matter compared to bacteria. This study highlights the differential behavior of PA and FL communities under hydrodynamic forces, enhancing the understanding of factors influencing bacterial community variation in estuaries.