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Methylmercury is a potent bioaccumulating neurotoxin that is produced by specific microorganisms that methylate inorganic mercury. Methylmercury production in diverse anaerobic bacteria and archaea was recently linked to the hgcAB genes. However, the full phylogenetic and metabolic diversity of mercury-methylating microorganisms has not been fully unraveled due to the limited number of cultured experimentally verified methylators and the limitations of primer-based molecular methods. Here, we describe the phylogenetic diversity and metabolic flexibility of putative mercury-methylating microorganisms by hgcAB identification in publicly available isolate genomes and metagenome-assembled genomes (MAGs) as well as novel freshwater MAGs. We demonstrate that putative mercury methylators are much more phylogenetically diverse than previously known and that hgcAB distribution among genomes is most likely due to several independent horizontal gene transfer events. The microorganisms we identified possess diverse metabolic capabilities spanning carbon fixation, sulfate reduction, nitrogen fixation, and metal resistance pathways. We identified 111 putative mercury methylators in a set of previously published permafrost metatranscriptomes and demonstrated that different methylating taxa may contribute to hgcA expression at different depths. Overall, we provide a framework for illuminating the microbial basis of mercury methylation using genome-resolved metagenomics and metatranscriptomics to identify putative methylators based upon hgcAB presence and describe their putative functions in the environment. IMPORTANCE Accurately assessing the production of bioaccumulative neurotoxic methylmercury by characterizing the phylogenetic diversity, metabolic functions, and activity of methylators in the environment is crucial for understanding constraints on the mercury cycle. Much of our understanding of methylmercury production is based on cultured anaerobic microorganisms within the Deltaproteobacteria, Firmicutes, and Euryarchaeota. Advances in next-generation sequencing technologies have enabled large-scale cultivation-independent surveys of diverse and poorly characterized microorganisms from numerous ecosystems. We used genome-resolved metagenomics and metatranscriptomics to highlight the vast phylogenetic and metabolic diversity of putative mercury methylators and their depth-discrete activities in thawing permafrost. This work underscores the importance of using genome-resolved metagenomics to survey specific putative methylating populations of a given mercury-impacted ecosystem.

Mercury (Hg) methylation is a microbially mediated process that converts inorganic Hg into bioaccumulative, neurotoxic methylmercury (MeHg). The metabolic activity of methylating organisms is highly dependent on biogeochemical conditions, which subsequently influences MeHg production. However, our understanding of the ecophysiology of methylators in natural ecosystems is still limited. Here, we identified potential locations of MeHg production in the anoxic, sulfidic hypolimnion of a freshwater lake. At these sites, we used shotgun metagenomics to characterize microorganisms with the Hg-methylation gene hgcA. Putative methylators were dominated by hgcA sequences divergent from those in well-studied, confirmed methylators. Using genome-resolved metagenomics, we identified organisms with hgcA (hgcA+) within the Bacteroidetes and the recently described Kiritimatiellaeota phyla. We identified hgcA+ genomes derived from sulfate-reducing bacteria, but these accounted for only 22% of hgcA+ genome coverage. The most abundant hgcA+ genomes were from fermenters, accounting for over half of the hgcA gene coverage. Many of these organisms also mediate hydrolysis of polysaccharides, likely from cyanobacterial blooms. This work highlights the distribution of the Hg-methylation genes across microbial metabolic guilds and indicate that primary degradation of polysaccharides and fermentation may play an important but unrecognized role in MeHg production in the anoxic hypolimnion of freshwater lakes.

“ Candidatus Accumulibacter” is the most studied PAO, with a primary role in biological nutrient removal. However, the species-level taxonomy of this lineage is convoluted due to the use of different phylogenetic markers or genome sequencing approaches. Here, we redefined the phylogeny of these organisms, proposing a comprehensive approach which could be used to address the classification of other diverse and uncultivated lineages.

This leadership development approach revolves around a set of competencies of highly effective senior leaders in higher education, developed in consultation with university presidents and vice presidents, former American Council on Education (ACE) Fellows, and others. The Class of 1999-2000 ACE Fellows, aspiring senior leaders in higher education, used these outcomes to assess their knowledge and skills, to plan learning activities to address identified gaps, and to document their learning during the Fellowship. Reflecting, developing, and documenting their leadership using the outcomes promotes continuing leadership development of Fellows upon return to their campuses and throughout their careers.

Microbial communities in their natural habitats consist of closely related populations that may exhibit phenotypic differences and inhabit distinct niches. However, connecting genetic diversity to ecological properties remains a challenge in microbial ecology due to the lack of pure cultures across the microbial tree of life. ‘Candidatus Accumulibacter phosphatis’ is a polyphosphate-accumulating organism that contributes to the Enhanced Biological Phosphorus Removal (EBPR) biotechnological process for removing excess phosphorus from wastewater and preventing eutrophication from downstream receiving waters. Distinct Accumulibacter clades often co-exist in full-scale wastewater treatment plants and lab-scale enrichment bioreactors, and have been hypothesized to inhabit distinct ecological niches. However, since individual strains of the Accumulibacter lineage have not been isolated in pure culture to date, these predictions have been made solely on genome-based comparisons and enrichments with varying strain composition. Here, we used genome-resolved metagenomics and metatranscriptomics to explore the activity of co-existing Accumulibacter clades in an engineered bioreactor environment. We obtained four high-quality genomes of Accumulibacter strains that were present in the bioreactor ecosystem, one of which is a completely contiguous draft genome scaffolded with long reads. We identified core and accessory genes to investigate how gene expression patterns differ among the dominating strains. Using this approach, we were able to identify putative pathways and functions that may differ between Accumulibacter clades and provide key functional insights into this biotechnologically significant microbial lineage.IMPORTANCE‘Candidatus Accumulibacter phosphatis’ is a model polyphosphate accumulating organism that has been studied using genome-resolved metagenomics, metatranscriptomics, and metaproteomics to understand the EBPR process. Within the Accumulibacter lineage, several similar but diverging clades are defined by the polyphosphate kinase (ppk1) locus sequence identity. These clades are predicted to have key functional differences in acetate uptake rates, phage defense mechanisms, and nitrogen cycling capabilities. However, such hypotheses have largely been made based on gene-content comparisons of sequenced Accumulibacter genomes, some of which were obtained from different systems. Here, we performed time-series genome-resolved metatranscriptomics to explore gene expression patterns of co-existing Accumulibacter clades in the same bioreactor ecosystem. Our work provides an approach for elucidating ecologically relevant functions based on gene expression patterns between closely related microbial populations.

“ Candidatus Accumulibacter phosphatis” is a model polyphosphate-accumulating organism that has been studied using genome-resolved metagenomics, metatranscriptomics, and metaproteomics to understand the EBPR process. Within the Accumulibacter lineage, several similar but diverging clades are defined by the shared sequence identity of the polyphosphate kinase ( ppk1 ) locus.

24 Summary: 25Advances in high-throughput sequencing technologies and bioinformatic pipelines have 26 exponentially increased the amount of data that can be obtained from uncultivated microbial 27 lineages inhabiting diverse ecosystems. Various annotation tools and databases currently exist for 28 predicting the functional potential of sequenced genomes or microbial communities based upon 29 sequence identity. However, intuitive, reproducible, and user-friendly tools for further exploring 30 and visualizing functional guilds of microbial community metagenomic sequencing datasets 31 remains lacking. Here, we present metabolisHMM, a series of workflows for visualizing the 32 distribution of curated and user-provided Hidden Markov Models (HMMs) to understand 33 metabolic characteristics and evolutionary histories of microbial lineages. metabolisHMM 34 performs functional annotations with a set of curated or user-defined HMMs to 1) construct 35 ribosomal protein and single marker gene phylogenies, 2) summarize the presence/absence of 36 metabolic pathway markers, and 3) create heatmap visualizations of presence/absence summaries. 37 38 39 Availability and Implementation: metabolisHMM is freely available on Github at 40 https://github.com/elizabethmcd/metabolisHMM and on PyPi at 41 https://pypi.org/project/metabolisHMM/ under the GNU General Public License v3.0. 42 43 44 Contact: elizabethmcd93@gmail.com 45 46 47 48