Importance of mobile genetic element immunity in numerically abundant Trichodesmium clades.

The colony-forming cyanobacteria Trichodesmium spp. are considered one of the most important nitrogen-fixing genera in the warm, low nutrient ocean. Despite this central biogeochemical role, many questions about their evolution, physiology, and trophic interactions remain unanswered. To address these questions, we describe Trichodesmium pangenomic potential via significantly improved genomic assemblies from two isolates and 15 new >50% complete Trichodesmium metagenome-assembled genomes from hand-picked, Trichodesmium colonies spanning the Atlantic Ocean. Phylogenomics identified ~four N2 fixing clades of Trichodesmium across the transect, with T. thiebautii dominating the colony-specific reads. Pangenomic analyses showed that all T. thiebautii MAGs are enriched in COG defense mechanisms and encode a vertically inherited Type III-B Clustered Regularly Interspaced Short Palindromic Repeats and associated protein-based immunity system (CRISPR-Cas). Surprisingly, this CRISPR-Cas system was absent in all T. erythraeum genomes, vertically inherited by T. thiebautii, and correlated with increased signatures of horizontal gene transfer. Additionally, the system was expressed in metaproteomic and transcriptomic datasets and CRISPR spacer sequences with 100% identical hits to field-assembled, putative phage genome fragments were identified. While the currently CO2-limited T. erythraeum is expected to be a 'winner' of anthropogenic climate change, their genomic dearth of known phage resistance mechanisms, compared to T. thiebautii, could put this outcome in question. Thus, the clear demarcation of T. thiebautii maintaining CRISPR-Cas systems, while T. erythraeum does not, identifies Trichodesmium as an ecologically important CRISPR-Cas model system, and highlights the need for more research on phage-Trichodesmium interactions.

Metagenome-Assembled Genome of a Novel Cyclobacteriaceae Epibiont of a Heterotrophic Diazotroph.

Here, we describe the metagenome-assembled genome (MAG) HetDA_MAG_SS2, in the family Cyclobacteriaceae. It was found in association with a HetDA cyanobiont isolated from a Station ALOHA Trichodesmium colony. Annotation suggests that HetDA_MAG_SS2 is a chemoorganoheterotroph with the potential for lithoheterotrophy, containing genes for aerobic respiration, mixed acid fermentation, dissimilatory nitrate reduction to ammonium, and sulfide oxidation.

Metagenome-Assembled Genome HetDA_MAG_MH13 of the Family Devosiaceae, from a Marine N2-fixing Cyanobacterial Enrichment Culture.

Here, we present a draft genome in the order Rhizobiales and family Devosiaceae. This draft genome comes from an enrichment of a heterocystous, cyanobacterial diazotroph (HetDA) that was originally living in association with Trichodesmium species. This Rhizobiales organism is proposed to be an anoxygenic phototroph capable of dissimilatory nitrate reduction to ammonia (DNRA).

Metagenome-Assembled Genome of an Alphaproteobacterium Isolated from an HetDA Enrichment from the North Pacific Subtropical Gyre.

Here, we present HetDA_MAG_SS10, a metagenome-assembled genome (MAG) from an enrichment of a heterocystous diazotroph originally living in association with Trichodesmium spp. obtained near Station ALOHA in the North Pacific Ocean. HetDA_MAG_SS10, an alphaproteobacterium in the order Micavibrionales, is proposed to be photoheterotrophic via rhodopsin and has the potential for dimethylsulfoniopropionate (DMSP) demethylation.

Genome Sequence of a Heterocystous Diazotroph Isolated from a Trichodesmium Consortium from the North Pacific Subtropical Gyre.

Diazotrophic cyanobacteria play a vital role in the nitrogen influx of the global marine ecosystem. In July 2010, colonies of Trichodesmium spp. were picked near Station ALOHA in the oligotrophic North Pacific Subtropical Gyre, and a novel heterocystous diazotroph (strain HetDA_MAG_MS3) belonging to the genus Rivularia was found living in close association; it was cultured and sequenced.

Metagenome-Assembled Genome of a Cyclobacteriaceae Bacterium, HetDA_MAG_MS6, Isolated from a Trichodesmium Consortium from Station ALOHA.

Here, we describe the metagenome-assembled genome (MAG) HetDA_MAG_MS6. HetDA_MAG_MS6 was obtained from an enrichment of the heterocystous diazotroph HetDA, which was isolated near Station ALOHA. The MAG was placed in the Cyclobacteriaceae family and is predicted to be a chemoorganoheterotroph with the potential for ammonia uptake, phosphonate transport, and sulfolipid biosynthesis.

Genome Sequence of the Euphotic Hawaiian Gammaproteobacterium HetDA_MAG_MS8, in the Order Nevskiales, Family Oceanococcaceae, Genus Oceanococcus.

We present a metagenome-assembled genome (MAG), HetDA_MAG_MS8, that was determined to be unique via relative evolutionary divergence (RED) scores and average nucleotide identity (ANI) values. HetDA_MAG_MS8 is in the order Nevskiales, genus Oceanococcus, and was assembled from a heterocytous cyanobiont enrichment from the Hawaii Ocean Time Series. HetDA_MAG_MS8 is predicted to be a facultative, aerobic, anoxygenic photolithoheterotroph that has the potential for sulfide oxidation and dimethylsulfoniopropionate (DMSP) synthesis.

Microbial Populations Are Shaped by Dispersal and Recombination in a Low Biomass Subseafloor Habitat.

The subseafloor is a vast habitat that supports microorganisms that have a global scale impact on geochemical cycles. Many of the endemic microbial communities inhabiting the subseafloor consist of small populations under growth-limited conditions. For small populations, stochastic evolutionary events can have large impacts on intraspecific population dynamics and allele frequencies. These conditions are fundamentally different from those experienced by most microorganisms in surface environments, and it is unknown how small population sizes and growth-limiting conditions influence evolution and population structure in the subsurface. Using a 2-year, high-resolution environmental time series, we examine the dynamics of microbial populations from cold, oxic crustal fluids collected from the subseafloor site North Pond, located near the mid-Atlantic ridge. Our results reveal rapid shifts in overall abundance, allele frequency, and strain abundance across the time points observed, with evidence for homologous recombination between coexisting lineages. We show that the subseafloor aquifer is a dynamic habitat that hosts microbial metapopulations that disperse frequently through the crustal fluids, enabling gene flow and recombination between microbial populations. The dynamism and stochasticity of microbial population dynamics in North Pond suggest that these forces are important drivers in the evolution of microbial populations in the vast subseafloor habitat. IMPORTANCE The cold, oxic subseafloor is an understudied habitat that is difficult to access, yet important to global biogeochemical cycles and starkly different compared to microbial habitats on the surface of the Earth. Our understanding of microbial evolution and population dynamics is largely molded by studies of microbes living in surface habitats that can host 10 to 1,000 times more microbial biomass than is frequently observed in the subsurface. This study provides an opportunity to observe population dynamics within a low biomass, growth-limited environment and reveals that microbial populations in the subseafloor are influenced by changes in selection pressure and gene sweeps. In addition, recombination between strains that have dispersed from elsewhere within the aquifer has an important impact on the evolution of microbial populations. Much of the microbial life on the planet exists under growth-limited conditions, and the subseafloor provides a natural laboratory to explore how life evolves in such environments.

Marine Dadabacteria exhibit genome streamlining and phototrophy-driven niche partitioning.

The remineralization of organic material via heterotrophy in the marine environment is performed by a diverse and varied group of microorganisms that can specialize in the type of organic material degraded and the niche they occupy. The marine Dadabacteria are cosmopolitan in the marine environment and belong to a candidate phylum for which there has not been a comprehensive assessment of the available genomic data to date. Here in, we assess the functional potential of the marine pelagic Dadabacteria in comparison to members of the phylum that originate from terrestrial, hydrothermal, and subsurface environments. Our analysis reveals that the marine pelagic Dadabacteria have streamlined genomes, corresponding to smaller genome sizes and lower nitrogen content of their DNA and predicted proteome, relative to their phylogenetic counterparts. Collectively, the Dadabacteria have the potential to degrade microbial dissolved organic matter, specifically peptidoglycan and phospholipids. The marine Dadabacteria belong to two clades with apparent distinct ecological niches in global metagenomic data: a clade with the potential for photoheterotrophy through the use of proteorhodopsin, present predominantly in surface waters up to 100 m depth; and a clade lacking the potential for photoheterotrophy that is more abundant in the deep photic zone.

MetaSanity: an integrated microbial genome evaluation and annotation pipeline.

SUMMARY: As the importance of microbiome research continues to become more prevalent and essential to understanding a wide variety of ecosystems (e.g. marine, built, host associated, etc.), there is a need for researchers to be able to perform highly reproducible and quality analysis of microbial genomes. MetaSanity incorporates analyses from 11 existing and widely used genome evaluation and annotation suites into a single, distributable workflow, thereby decreasing the workload of microbiologists by allowing for a flexible, expansive data analysis pipeline. MetaSanity has been designed to provide separate, reproducible workflows that (i) can determine the overall quality of a microbial genome, while providing a putative phylogenetic assignment, and (ii) can assign structural and functional gene annotations with varying degrees of specificity to suit the needs of the researcher. The software suite combines the results from several tools to provide broad insights into overall metabolic function. Importantly, this software provides built-in optimization for 'big data' analysis by storing all relevant outputs in an SQL database, allowing users to query all the results for the elements that will most impact their research. AVAILABILITY AND IMPLEMENTATION: MetaSanity is provided under the GNU General Public License v.3.0 and is available for download at https://github.com/cjneely10/MetaSanity. This application is distributed as a Docker image. MetaSanity is implemented in Python3/Cython and C++. Instructions for its installation and use are available within the GitHub wiki page at https://github.com/cjneely10/MetaSanity/wiki, and additional instructions are available at https://cjneely10.github.io/year-archive/. MetaSanity is optimized for users with limited programing experience. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

The reconstruction of 2,631 draft metagenome-assembled genomes from the global oceans.

Microorganisms play a crucial role in mediating global biogeochemical cycles in the marine environment. By reconstructing the genomes of environmental organisms through metagenomics, researchers are able to study the metabolic potential of Bacteria and Archaea that are resistant to isolation in the laboratory. Utilizing the large metagenomic dataset generated from 234 samples collected during the Tara Oceans circumnavigation expedition, we were able to assemble 102 billion paired-end reads into 562 million contigs, which in turn were co-assembled and consolidated in to 7.2 million contigs ≥2 kb in length. Approximately 1 million of these contigs were binned to reconstruct draft genomes. In total, 2,631 draft genomes with an estimated completion of ≥50% were generated (1,491 draft genomes >70% complete; 603 genomes >90% complete). A majority of the draft genomes were manually assigned phylogeny based on sets of concatenated phylogenetic marker genes and/or 16S rRNA gene sequences. The draft genomes are now publically available for the research community at-large.

290 metagenome-assembled genomes from the Mediterranean Sea: a resource for marine microbiology.

The Tara Oceans Expedition has provided large, publicly-accessible microbial metagenomic datasets from a circumnavigation of the globe. Utilizing several size fractions from the samples originating in the Mediterranean Sea, we have used current assembly and binning techniques to reconstruct 290 putative draft metagenome-assembled bacterial and archaeal genomes, with an estimated completion of ≥50%, and an additional 2,786 bins, with estimated completion of 0-50%. We have submitted our results, including initial taxonomic and phylogenetic assignments, for the putative draft genomes to open-access repositories for the scientific community to use in ongoing research.

BinSanity: unsupervised clustering of environmental microbial assemblies using coverage and affinity propagation.

Metagenomics has become an integral part of defining microbial diversity in various environments. Many ecosystems have characteristically low biomass and few cultured representatives. Linking potential metabolisms to phylogeny in environmental microorganisms is important for interpreting microbial community functions and the impacts these communities have on geochemical cycles. However, with metagenomic studies there is the computational hurdle of 'binning' contigs into phylogenetically related units or putative genomes. Binning methods have been implemented with varying approaches such as k-means clustering, Gaussian mixture models, hierarchical clustering, neural networks, and two-way clustering; however, many of these suffer from biases against low coverage/abundance organisms and closely related taxa/strains. We are introducing a new binning method, BinSanity, that utilizes the clustering algorithm affinity propagation (AP), to cluster assemblies using coverage with compositional based refinement (tetranucleotide frequency and percent GC content) to optimize bins containing multiple source organisms. This separation of composition and coverage based clustering reduces bias for closely related taxa. BinSanity was developed and tested on artificial metagenomes varying in size and complexity. Results indicate that BinSanity has a higher precision, recall, and Adjusted Rand Index compared to five commonly implemented methods. When tested on a previously published environmental metagenome, BinSanity generated high completion and low redundancy bins corresponding with the published metagenome-assembled genomes.