Exploring environmental intra-species diversity through non-redundant pangenome assemblies.

At the genome level, microorganisms are highly adaptable both in terms of allele and gene composition. Such heritable traits emerge in response to different environmental niches and can have a profound influence on microbial community dynamics. As a consequence, any individual genome or population will contain merely a fraction of the total genetic diversity of any operationally defined "species", whose ecological potential can thus be only fully understood by studying all of their genomes and the genes therein. This concept, known as the pangenome, is valuable for studying microbial ecology and evolution, as it partitions genomes into core (present in all the genomes from a species, and responsible for housekeeping and species-level niche adaptation among others) and accessory regions (present only in some, and responsible for intra-species differentiation). Here we present SuperPang, an algorithm producing pangenome assemblies from a set of input genomes of varying quality, including metagenome-assembled genomes (MAGs). SuperPang runs in linear time and its results are complete, non-redundant, preserve gene ordering and contain both coding and non-coding regions. Our approach provides a modular view of the pangenome, identifying operons and genomic islands, and allowing to track their prevalence in different populations. We illustrate this by analysing intra-species diversity in Polynucleobacter, a bacterial genus ubiquitous in freshwater ecosystems, characterized by their streamlined genomes and their ecological versatility. We show how SuperPang facilitates the simultaneous analysis of allelic and gene content variation under different environmental pressures, allowing us to study the drivers of microbial diversification at unprecedented resolution.

Linking prokaryotic genome size variation to metabolic potential and environment.

While theories and models have appeared to explain genome size as a result of evolutionary processes, little work has shown that genome sizes carry ecological signatures. Our work delves into the ecological implications of microbial genome size variation in benthic and pelagic habitats across environmental gradients of the brackish Baltic Sea. While depth is significantly associated with genome size in benthic and pelagic brackish metagenomes, salinity is only correlated to genome size in benthic metagenomes. Overall, we confirm that prokaryotic genome sizes in Baltic sediments (3.47 Mbp) are significantly bigger than in the water column (2.96 Mbp). While benthic genomes have a higher number of functions than pelagic genomes, the smallest genomes coded for a higher number of module steps per Mbp for most of the functions irrespective of their environment. Some examples of this functions are amino acid metabolism and central carbohydrate metabolism. However, we observed that nitrogen metabolism was almost absent in pelagic genomes and was mostly present in benthic genomes. Finally, we also show that Bacteria inhabiting Baltic sediments and water column not only differ in taxonomy, but also in their metabolic potential, such as the Wood-Ljungdahl pathway or the presence of different hydrogenases. Our work shows how microbial genome size is linked to abiotic factors in the environment, metabolic potential and taxonomic identity of Bacteria and Archaea within aquatic ecosystems.

A consensus protocol for the recovery of mercury methylation genes from metagenomes.

Mercury (Hg) methylation genes (hgcAB) mediate the formation of the toxic methylmercury and have been identified from diverse environments, including freshwater and marine ecosystems, Arctic permafrost, forest and paddy soils, coal-ash amended sediments, chlor-alkali plants discharges and geothermal springs. Here we present the first attempt at a standardized protocol for the detection, identification and quantification of hgc genes from metagenomes. Our Hg-cycling microorganisms in aquatic and terrestrial ecosystems (Hg-MATE) database, a catalogue of hgc genes, provides the most accurate information to date on the taxonomic identity and functional/metabolic attributes of microorganisms responsible for Hg methylation in the environment. Furthermore, we introduce "marky-coco", a ready-to-use bioinformatic pipeline based on de novo single-metagenome assembly, for easy and accurate characterization of hgc genes from environmental samples. We compared the recovery of hgc genes from environmental metagenomes using the marky-coco pipeline with an approach based on coassembly of multiple metagenomes. Our data show similar efficiency in both approaches for most environments except those with high diversity (i.e., paddy soils) for which a coassembly approach was preferred. Finally, we discuss the definition of true hgc genes and methods to normalize hgc gene counts from metagenomes.

Microbial communities mediating net methylmercury formation along a trophic gradient in a peatland chronosequence.

Peatlands are generally important sources of methylmercury (MeHg) to adjacent aquatic ecosystems, increasing the risk of human and wildlife exposure to this highly toxic compound. While microorganisms play important roles in mercury (Hg) geochemical cycles where they directly and indirectly affect MeHg formation in peatlands, potential linkages between net MeHg formation and microbial communities involving these microorganisms remain unclear. To address this gap, microbial community composition and specific marker gene transcripts were investigated along a trophic gradient in a geographically constrained peatland chronosequence. Our results showed a clear spatial pattern in microbial community composition along the gradient that was highly driven by peat soil properties and significantly associated with net MeHg formation as approximated by MeHg concentration and %MeHg of total Hg concentration. Known fermentative, syntrophic, methanogenic and iron-reducing metabolic guilds had the strong positive correlations to net MeHg formation, while methanotrophic and methylotrophic microorganisms were negatively correlated. Our results indicated that sulfate reducers did not have a key role in net MeHg formation. Microbial activity as interpreted from 16S rRNA sequences was significantly correlated with MeHg and %MeHg. Our findings shed new light on the role of microbial community in net MeHg formation of peatlands that undergo ontogenetic change.

Multiomics in the central Arctic Ocean for benchmarking biodiversity change.

Multiomics approaches need to be applied in the central Arctic Ocean to benchmark biodiversity change and to identify novel species and their genes. As part of MOSAiC, EcoOmics will therefore be essential for conservation and sustainable bioprospecting in one of the least explored ecosystems on Earth.

Expression Levels of hgcAB Genes and Mercury Availability Jointly Explain Methylmercury Formation in Stratified Brackish Waters.

Neurotoxic methylmercury (MeHg) is formed by microbial methylation of inorganic divalent Hg (HgII) and constitutes severe environmental and human health risks. The methylation is enabled by hgcA and hgcB genes, but it is not known if the associated molecular-level processes are rate-limiting or enable accurate prediction of MeHg formation in nature. In this study, we investigated the relationships between hgc genes and MeHg across redox-stratified water columns in the brackish Baltic Sea. We showed, for the first time, that hgc transcript abundance and the concentration of dissolved HgII-sulfide species were strong predictors of both the HgII methylation rate and MeHg concentration, implying their roles as principal joint drivers of MeHg formation in these systems. Additionally, we characterized the metabolic capacities of hgc+ microorganisms by reconstructing their genomes from metagenomes (i.e., hgc+ MAGs), which highlighted the versatility of putative HgII methylators in the water column of the Baltic Sea. In establishing relationships between hgc transcripts and the HgII methylation rate, we advance the fundamental understanding of mechanistic principles governing MeHg formation in nature and enable refined predictions of MeHg levels in coastal seas in response to the accelerating spread of oxygen-deficient zones.

mOTUpan: a robust Bayesian approach to leverage metagenome-assembled genomes for core-genome estimation.

Recent advances in sequencing and bioinformatics have expanded the tree of life by providing genomes for uncultured environmentally relevant clades, either through metagenome-assembled genomes or through single-cell genomes. While this expanded diversity can provide novel insights into microbial population structure, most tools available for core-genome estimation are sensitive to genome completeness. Consequently, a major portion of the huge phylogenetic diversity uncovered by environmental genomic approaches remains excluded from such analyses. We present mOTUpan, a novel iterative Bayesian method for computing the core genome for sets of genomes of highly diverse completeness range. The likelihood for each gene cluster to belong to core or accessory genome is estimated by computing the probability of its presence/absence pattern in the target genome set. The core-genome prediction is computationally efficient and can be scaled up to thousands of genomes. It has shown comparable estimates to state-of-the-art tools Roary and PPanGGOLiN for high-quality genomes and is capable of using genomes at lower completeness thresholds. mOTUpan wraps a bootstrapping procedure to estimate the quality of a specific core-genome prediction, as the accuracy of each run will depend on the specific completeness distribution and the number of genomes in the dataset under scrutiny. mOTUpan is implemented in the mOTUlizer software package, and available at github.com/moritzbuck/mOTUlizer, under GPL 3.0 license.

Candidatus Methylumidiphilus Drives Peaks in Methanotrophic Relative Abundance in Stratified Lakes and Ponds Across Northern Landscapes.

Boreal lakes and ponds produce two-thirds of the total natural methane emissions above the latitude of 50° North. These lake emissions are regulated by methanotrophs which can oxidize up to 99% of the methane produced in the sediments and the water column. Despite their importance, the diversity and distribution of the methanotrophs in lakes are still poorly understood. Here, we used shotgun metagenomic data to explore the diversity and distribution of methanotrophs in 40 oxygen-stratified water bodies in boreal and subarctic areas in Europe and North America. In our data, gammaproteobacterial methanotrophs (order Methylococcales) generally dominated the methanotrophic communities throughout the water columns. A recently discovered lineage of Methylococcales, Candidatus Methylumidiphilus, was present in all the studied water bodies and dominated the methanotrophic community in lakes with a high relative abundance of methanotrophs. Alphaproteobacterial methanotrophs were the second most abundant group of methanotrophs. In the top layer of the lakes, characterized by low CH4 concentration, their abundance could surpass that of the gammaproteobacterial methanotrophs. These results support the theory that the alphaproteobacterial methanotrophs have a high affinity for CH4 and can be considered stress-tolerant strategists. In contrast, the gammaproteobacterial methanotrophs are competitive strategists. In addition, relative abundances of anaerobic methanotrophs, Candidatus Methanoperedenaceae and Candidatus Methylomirabilis, were strongly correlated, suggesting possible co-metabolism. Our data also suggest that these anaerobic methanotrophs could be active even in the oxic layers. In non-metric multidimensional scaling, alpha- and gammaproteobacterial methanotrophs formed separate clusters based on their abundances in the samples, except for the gammaproteobacterial Candidatus Methylumidiphilus, which was separated from these two clusters. This may reflect similarities in the niche and environmental requirements of the different genera within alpha- and gammaproteobacterial methanotrophs. Our study confirms the importance of O2 and CH4 in shaping the methanotrophic communities and suggests that one variable cannot explain the diversity and distribution of the methanotrophs across lakes. Instead, we suggest that the diversity and distribution of freshwater methanotrophs are regulated by lake-specific factors.

Energy efficiency and biological interactions define the core microbiome of deep oligotrophic groundwater.

While oligotrophic deep groundwaters host active microbes attuned to the low-end of the bioenergetics spectrum, the ecological constraints on microbial niches in these ecosystems and their consequences for microbiome convergence are unknown. Here, we provide a genome-resolved, integrated omics analysis comparing archaeal and bacterial communities in disconnected fracture fluids of the Fennoscandian Shield in Europe. Leveraging a dataset that combines metagenomes, single cell genomes, and metatranscriptomes, we show that groundwaters flowing in similar lithologies offer fixed niches that are occupied by a common core microbiome. Functional expression analysis highlights that these deep groundwater ecosystems foster diverse, yet cooperative communities adapted to this setting. We suggest that these communities stimulate cooperation by expression of functions related to ecological traits, such as aggregate or biofilm formation, while alleviating the burden on microorganisms producing compounds or functions that provide a collective benefit by facilitating reciprocal promiscuous metabolic partnerships with other members of the community. We hypothesize that an episodic lifestyle enabled by reversible bacteriostatic functions ensures the subsistence of the oligotrophic deep groundwater microbiome.

Freshwater Chlorobia Exhibit Metabolic Specialization among Cosmopolitan and Endemic Populations.

Photosynthetic bacteria from the class Chlorobia (formerly phylum Chlorobi) sustain carbon fixation in anoxic water columns. They harvest light at extremely low intensities and use various inorganic electron donors to fix carbon dioxide into biomass. Until now, most information on the functional ecology and local adaptations of Chlorobia members came from isolates and merely 26 sequenced genomes that may not adequately represent natural populations. To address these limitations, we analyzed global metagenomes to profile planktonic Chlorobia cells from the oxyclines of 42 freshwater bodies, spanning subarctic to tropical regions and encompassing all four seasons. We assembled and compiled over 500 genomes, including metagenome-assembled genomes (MAGs), single-amplified genomes (SAGs), and reference genomes from cultures, clustering them into 71 metagenomic operational taxonomic units (mOTUs or "species"). Of the 71 mOTUs, 57 were classified within the genus Chlorobium, and these mOTUs represented up to ∼60% of the microbial communities in the sampled anoxic waters. Several Chlorobium-associated mOTUs were globally distributed, whereas others were endemic to individual lakes. Although most clades encoded the ability to oxidize hydrogen, many lacked genes for the oxidation of specific sulfur and iron substrates. Surprisingly, one globally distributed Scandinavian clade encoded the ability to oxidize hydrogen, sulfur, and iron, suggesting that metabolic versatility facilitated such widespread colonization. Overall, these findings provide new insight into the biogeography of the Chlorobia and the metabolic traits that facilitate niche specialization within lake ecosystems.IMPORTANCE The reconstruction of genomes from metagenomes has helped explore the ecology and evolution of environmental microbiota. We applied this approach to 274 metagenomes collected from diverse freshwater habitats that spanned oxic and anoxic zones, sampling seasons, and latitudes. We demonstrate widespread and abundant distributions of planktonic Chlorobia-associated bacteria in hypolimnetic waters of stratified freshwater ecosystems and show they vary in their capacities to use different electron donors. Having photoautotrophic potential, these Chlorobia members could serve as carbon sources that support metalimnetic and hypolimnetic food webs.

Comprehensive dataset of shotgun metagenomes from oxygen stratified freshwater lakes and ponds.

Stratified lakes and ponds featuring steep oxygen gradients are significant net sources of greenhouse gases and hotspots in the carbon cycle. Despite their significant biogeochemical roles, the microbial communities, especially in the oxygen depleted compartments, are poorly known. Here, we present a comprehensive dataset including 267 shotgun metagenomes from 41 stratified lakes and ponds mainly located in the boreal and subarctic regions, but also including one tropical reservoir and one temperate lake. For most lakes and ponds, the data includes a vertical sample set spanning from the oxic surface to the anoxic bottom layer. The majority of the samples were collected during the open water period, but also a total of 29 samples were collected from under the ice. In addition to the metagenomic sequences, the dataset includes environmental variables for the samples, such as oxygen, nutrient and organic carbon concentrations. The dataset is ideal for further exploring the microbial taxonomic and functional diversity in freshwater environments and potential climate change impacts on the functioning of these ecosystems.

A Genomic Perspective Across Earth's Microbiomes Reveals That Genome Size in Archaea and Bacteria Is Linked to Ecosystem Type and Trophic Strategy.

Our view of genome size in Archaea and Bacteria has remained skewed as the data has been dominated by genomes of microorganisms that have been cultivated under laboratory settings. However, the continuous effort to catalog Earth's microbiomes, specifically propelled by recent extensive work on uncultivated microorganisms, provides an opportunity to revise our perspective on genome size distribution. We present a meta-analysis that includes 26,101 representative genomes from 3 published genomic databases; metagenomic assembled genomes (MAGs) from GEMs and stratfreshDB, and isolates from GTDB. Aquatic and host-associated microbial genomes present on average the smallest estimated genome sizes (3.1 and 3.0 Mbp, respectively). These are followed by terrestrial microbial genomes (average 3.7 Mbp), and genomes from isolated microorganisms (average 4.3 Mbp). On the one hand, aquatic and host-associated ecosystems present smaller genomes sizes in genera of phyla with genome sizes above 3 Mbp. On the other hand, estimated genome size in phyla with genomes under 3 Mbp showed no difference between ecosystems. Moreover, we observed that when using 95% average nucleotide identity (ANI) as an estimator for genetic units, only 3% of MAGs cluster together with genomes from isolated microorganisms. Although there are potential methodological limitations when assembling and binning MAGs, we found that in genome clusters containing both environmental MAGs and isolate genomes, MAGs were estimated only an average 3.7% smaller than isolate genomes. Even when assembly and binning methods introduce biases, estimated genome size of MAGs and isolates are very similar. Finally, to better understand the ecological drivers of genome size, we discuss on the known and the overlooked factors that influence genome size in different ecosystems, phylogenetic groups, and trophic strategies.

Vertical stratification patterns of methanotrophs and their genetic controllers in water columns of oxygen-stratified boreal lakes.

The vertical structuring of methanotrophic communities and its genetic controllers remain understudied in the water columns of oxygen-stratified lakes. Therefore, we used 16S rRNA gene sequencing to study the vertical stratification patterns of methanotrophs in two boreal lakes, Lake Kuivajärvi and Lake Lovojärvi. Furthermore, metagenomic analyses were performed to assess the genomic characteristics of methanotrophs in Lovojärvi and the previously studied Lake Alinen Mustajärvi. The methanotroph communities were vertically structured along the oxygen gradient. Alphaproteobacterial methanotrophs preferred oxic water layers, while Methylococcales methanotrophs, consisting of putative novel genera and species, thrived, especially at and below the oxic-anoxic interface and showed distinct depth variation patterns, which were not completely predictable by their taxonomic classification. Instead, genomic differences among Methylococcales methanotrophs explained their variable vertical depth patterns. Genes in clusters of orthologous groups (COG) categories L (replication, recombination and repair) and S (function unknown) were relatively high in metagenome-assembled genomes representing Methylococcales clearly thriving below the oxic-anoxic interface, suggesting genetic adaptations for increased stress tolerance enabling living in the hypoxic/anoxic conditions. By contrast, genes in COG category N (cell motility) were relatively high in metagenome-assembled genomes of Methylococcales thriving at the oxic-anoxic interface, which suggests genetic adaptations for increased motility at the vertically fluctuating oxic-anoxic interface.

Deltaproteobacteria and Spirochaetes-Like Bacteria Are Abundant Putative Mercury Methylators in Oxygen-Deficient Water and Marine Particles in the Baltic Sea.

Methylmercury (MeHg), a neurotoxic compound biomagnifying in aquatic food webs, can be a threat to human health via fish consumption. However, the composition and distribution of the microbial communities mediating the methylation of mercury (Hg) to MeHg in marine systems remain largely unknown. In order to fill this knowledge gap, we used the Baltic Sea Reference Metagenome (BARM) dataset to study the abundance and distribution of the genes involved in Hg methylation (the hgcAB gene cluster). We determined the relative abundance of the hgcAB genes and their taxonomic identity in 81 brackish metagenomes that cover spatial, seasonal and redox variability in the Baltic Sea water column. The hgcAB genes were predominantly detected in anoxic water, but some hgcAB genes were also detected in hypoxic and normoxic waters. Phylogenetic analysis identified putative Hg methylators within Deltaproteobacteria, in oxygen-deficient water layers, but also Spirochaetes-like and Kiritimatiellaeota-like bacteria. Higher relative quantities of hgcAB genes were found in metagenomes from marine particles compared to free-living communities in anoxic water, suggesting that such particles are hotspot habitats for Hg methylators in oxygen-depleted seawater. Altogether, our work unveils the diversity of the microorganisms with the potential to mediate MeHg production in the Baltic Sea and pinpoint the important ecological niches for these microorganisms within the marine water column.

Streamlined and Abundant Bacterioplankton Thrive in Functional Cohorts.

While fastidious microbes can be abundant and ubiquitous in their natural communities, many fail to grow axenically in laboratories due to auxotrophies or other dependencies. To overcome auxotrophies, these microbes rely on their surrounding cohort. A cohort may consist of kin (ecotypes) or more distantly related organisms (community) with the cooperation being reciprocal or nonreciprocal and expensive (Black Queen hypothesis) or costless (by-product). These metabolic partnerships (whether at single species population or community level) enable dominance by and coexistence of these lineages in nature. Here we examine the relevance of these cooperation models to explain the abundance and ubiquity of the dominant fastidious bacterioplankton of a dimictic mesotrophic freshwater lake. Using both culture-dependent (dilution mixed cultures) and culture-independent (small subunit [SSU] rRNA gene time series and environmental metagenomics) methods, we independently identified the primary cohorts of actinobacterial genera "Candidatus Planktophila" (acI-A) and "Candidatus Nanopelagicus" (acI-B) and the proteobacterial genus "Candidatus Fonsibacter" (LD12). While "Ca Planktophila" and "Ca. Fonsibacter" had no correlation in their natural habitat, they have the potential to be complementary in laboratory settings. We also investigated the bifunctional catalase-peroxidase enzyme KatG (a common good which "Ca Planktophila" is dependent upon) and its most likely providers in the lake. Further, we found that while ecotype and community cooperation combined may explain "Ca Planktophila" population abundance, the success of "Ca. Nanopelagicus" and "Ca. Fonsibacter" is better explained as a community by-product. Ecotype differentiation of "Ca. Fonsibacter" as a means of escaping predation was supported but not for overcoming auxotrophies.IMPORTANCE This study examines evolutionary and ecological relationships of three of the most ubiquitous and abundant freshwater bacterial genera: "Ca Planktophila" (acI-A), "Ca. Nanopelagicus" (acI-B), and "Ca. Fonsibacter" (LD12). Due to high abundance, these genera might have a significant influence on nutrient cycling in freshwaters worldwide, and this study adds a layer of understanding to how seemingly competing clades of bacteria can coexist by having different cooperation strategies. Our synthesis ties together network and ecological theory with empirical evidence and lays out a framework for how the functioning of populations within complex microbial communities can be studied.

Mercury methylating microbial communities of boreal forest soils.

The formation of the potent neurotoxic methylmercury (MeHg) is a microbially mediated process that has raised much concern because MeHg poses threats to wildlife and human health. Since boreal forest soils can be a source of MeHg in aquatic networks, it is crucial to understand the biogeochemical processes involved in the formation of this pollutant. High-throughput sequencing of 16S rRNA and the mercury methyltransferase, hgcA, combined with geochemical characterisation of soils, were used to determine the microbial populations contributing to MeHg formation in forest soils across Sweden. The hgcA sequences obtained were distributed among diverse clades, including Proteobacteria, Firmicutes, and Methanomicrobia, with Deltaproteobacteria, particularly Geobacteraceae, dominating the libraries across all soils examined. Our results also suggest that MeHg formation is also linked to the composition of non-mercury methylating bacterial communities, likely providing growth substrate (e.g. acetate) for the hgcA-carrying microorganisms responsible for the actual methylation process. While previous research focused on mercury methylating microbial communities of wetlands, this study provides some first insights into the diversity of mercury methylating microorganisms in boreal forest soils.

Methanogens and Iron-Reducing Bacteria: the Overlooked Members of Mercury-Methylating Microbial Communities in Boreal Lakes.

Methylmercury is a potent human neurotoxin which biomagnifies in aquatic food webs. Although anaerobic microorganisms containing the hgcA gene potentially mediate the formation of methylmercury in natural environments, the diversity of these mercury-methylating microbial communities remains largely unexplored. Previous studies have implicated sulfate-reducing bacteria as the main mercury methylators in aquatic ecosystems. In the present study, we characterized the diversity of mercury-methylating microbial communities of boreal lake sediments using high-throughput sequencing of 16S rRNA and hgcA genes. Our results show that in the lake sediments, Methanomicrobiales and Geobacteraceae also represent abundant members of the mercury-methylating communities. In fact, incubation experiments with a mercury isotopic tracer and molybdate revealed that only between 38% and 45% of mercury methylation was attributed to sulfate reduction. These results suggest that methanogens and iron-reducing bacteria may contribute to more than half of the mercury methylation in boreal lakes.IMPORTANCE Despite the global awareness that mercury, and methylmercury in particular, is a neurotoxin to which millions of people continue to be exposed, there are sizable gaps in the understanding of the processes and organisms involved in methylmercury formation in aquatic ecosystems. In the present study, we shed light on the diversity of the microorganisms responsible for methylmercury formation in boreal lake sediments. All the microorganisms identified are associated with the processing of organic matter in aquatic systems. Moreover, our results show that the well-known mercury-methylating sulfate-reducing bacteria constituted only a minor portion of the potential mercury methylators. In contrast, methanogens and iron-reducing bacteria were important contributors to methylmercury formation, highlighting their role in mercury cycling in the environment.

Novel Autotrophic Organisms Contribute Significantly to the Internal Carbon Cycling Potential of a Boreal Lake.

Oxygen-stratified lakes are typical for the boreal zone and also a major source of greenhouse gas emissions in the region. Due to shallow light penetration, restricting the growth of phototrophic organisms, and large allochthonous organic carbon inputs from the catchment area, the lake metabolism is expected to be dominated by heterotrophic organisms. In this study, we test this assumption and show that the potential for autotrophic carbon fixation and internal carbon cycling is high throughout the water column. Further, we show that during the summer stratification carbon fixation can exceed respiration in a boreal lake even below the euphotic zone. Metagenome-assembled genomes and 16S profiling of a vertical transect of the lake revealed multiple organisms in an oxygen-depleted compartment belonging to novel or poorly characterized phyla. Many of these organisms were chemolithotrophic, potentially deriving their energy from reactions related to sulfur, iron, and nitrogen transformations. The community, as well as the functions, was stratified along the redox gradient. The autotrophic potential in the lake metagenome below the oxygenic zone was high, pointing toward a need for revising our concepts of internal carbon cycling in boreal lakes. Further, the importance of chemolithoautotrophy for the internal carbon cycling suggests that many predicted climate change-associated fluctuations in the physical properties of the lake, such as altered mixing patterns, likely have consequences for the whole-lake metabolism even beyond the impact to the phototrophic community.IMPORTANCE Autotrophic organisms at the base of the food web are the only life form capable of turning inorganic carbon into the organic form, facilitating the survival of all other organisms. In certain environments, the autotrophic production is limited by environmental conditions and the food web is supported by external carbon inputs. One such environment is stratified boreal lakes, which are one of the biggest natural sources of greenhouse gas emissions in the boreal region. Thus, carbon cycling in these habitats is of utmost importance for the future climate. Here, we demonstrate a high potential for internal carbon cycling via phototrophic and novel chemolithotrophic organisms in the anoxic, poorly illuminated layers of a boreal lake. Our results significantly increase our knowledge on the microbial communities and their metabolic potential in oxygen-depleted freshwaters and help to understand and predict how climate change-induced alterations could impact the lake carbon dynamics.

Model Communities Hint at Promiscuous Metabolic Linkages between Ubiquitous Free-Living Freshwater Bacteria.

Genome streamlining is frequently observed in free-living aquatic microorganisms and results in physiological dependencies between microorganisms. However, we know little about the specificity of these microbial associations. In order to examine the specificity and extent of these associations, we established mixed cultures from three different freshwater environments and analyzed the cooccurrence of organisms using a metagenomic time series. Free-living microorganisms with streamlined genomes lacking multiple biosynthetic pathways showed no clear recurring pattern in their interaction partners. Free-living freshwater bacteria form promiscuous cooperative associations. This notion contrasts with the well-documented high specificities of interaction partners in host-associated bacteria. Considering all data together, we suggest that highly abundant free-living bacterial lineages are functionally versatile in their interactions despite their distinct streamlining tendencies at the single-cell level. This metabolic versatility facilitates interactions with a variable set of community members.

Geobacteraceae are important members of mercury-methylating microbial communities of sediments impacted by waste water releases.

Microbial mercury (Hg) methylation in sediments can result in bioaccumulation of the neurotoxin methylmercury (MMHg) in aquatic food webs. Recently, the discovery of the gene hgcA, required for Hg methylation, revealed that the diversity of Hg methylators is much broader than previously thought. However, little is known about the identity of Hg-methylating microbial organisms and the environmental factors controlling their activity and distribution in lakes. Here, we combined high-throughput sequencing of 16S rRNA and hgcA genes with the chemical characterization of sediments impacted by a waste water treatment plant that releases significant amounts of organic matter and iron. Our results highlight that the ferruginous geochemical conditions prevailing at 1-2 cm depth are conducive to MMHg formation and that the Hg-methylating guild is composed of iron and sulfur-transforming bacteria, syntrophs, and methanogens. Deltaproteobacteria, notably Geobacteraceae, dominated the hgcA carrying communities, while sulfate reducers constituted only a minor component, despite being considered the main Hg methylators in many anoxic aquatic environments. Because iron is widely applied in waste water treatment, the importance of Geobacteraceae for Hg methylation and the complexity of Hg-methylating communities reported here are likely to occur worldwide in sediments impacted by waste water treatment plant discharges and in iron-rich sediments in general.