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We report the genome of Phormidium yuhuli AB48, which includes a circular chromosome and a circular plasmid (4,747,469 bp and 51,599 bp, respectively). This is currently the only closed reference genome of an isolate of the Phormidium genus, based on the Genome Taxonomy Database (GTDB), providing a potential model system for sustainable biotechnology innovation.

Anaerobic digestion of municipal mixed sludge produces methane that can be converted into renewable natural gas. To improve economics of this microbial mediated process, metabolic interactions catalyzing biomass conversion to energy need to be identified. Here, we present a two-year time series associating microbial metabolism and physicochemistry in a full-scale wastewater treatment plant. By creating a co-occurrence network with thousands of time-resolved microbial populations from over 100 samples spanning four operating configurations, known and novel microbial consortia with potential to drive methane production were identified. Interactions between these populations were further resolved in relation to specific process configurations by mapping metagenome assembled genomes and cognate gene expression data onto the network. Prominent interactions included transcriptionally active Methanolinea methanogens and syntrophic benzoate oxidizing Syntrophorhabdus, as well as a Methanoregulaceae population and putative syntrophic acetate oxidizing bacteria affiliated with Bateroidetes (Tenuifilaceae) expressing the glycine cleavage bypass of the Wood-Ljungdahl pathway.

[This corrects the article DOI: 10.3389/fbioe.2022.932695.].

Oxygen-deficient marine waters referred to as oxygen minimum zones (OMZs) or anoxic marine zones (AMZs) are common oceanographic features. They host both cosmopolitan and endemic microorganisms adapted to low oxygen conditions. Microbial metabolic interactions within OMZs and AMZs drive coupled biogeochemical cycles resulting in nitrogen loss and climate active trace gas production and consumption. Global warming is causing oxygen-deficient waters to expand and intensify. Therefore, studies focused on microbial communities inhabiting oxygen-deficient regions are necessary to both monitor and model the impacts of climate change on marine ecosystem functions and services. Here we present a compendium of 5,129 single-cell amplified genomes (SAGs) from marine environments encompassing representative OMZ and AMZ geochemical profiles. Of these, 3,570 SAGs have been sequenced to different levels of completion, providing a strain-resolved perspective on the genomic content and potential metabolic interactions within OMZ and AMZ microbiomes. Hierarchical clustering confirmed that samples from similar oxygen concentrations and geographic regions also had analogous taxonomic compositions, providing a coherent framework for comparative community analysis.

Here, we report metagenome-assembled genomes for "Candidatus Phormidium sp. strain AB48" and three cooccurring microorganisms from a biofilm-forming industrial photobioreactor environment, using the PacBio sequencing platform. Several mobile genetic elements, including a double-stranded DNA phage and plasmids, were also recovered, with the potential to mediate gene transfer within the biofilm community.

The anaerobic oxidation of methane coupled to sulfate reduction is a microbially mediated process requiring a syntrophic partnership between anaerobic methanotrophic (ANME) archaea and sulfate-reducing bacteria (SRB). Based on genome taxonomy, ANME lineages are polyphyletic within the phylum Halobacterota, none of which have been isolated in pure culture. Here, we reconstruct 28 ANME genomes from environmental metagenomes and flow sorted syntrophic consortia. Together with a reanalysis of previously published datasets, these genomes enable a comparative analysis of all marine ANME clades. We review the genomic features that separate ANME from their methanogenic relatives and identify what differentiates ANME clades. Large multiheme cytochromes and bioenergetic complexes predicted to be involved in novel electron bifurcation reactions are well distributed and conserved in the ANME archaea, while significant variations in the anabolic C1 pathways exists between clades. Our analysis raises the possibility that methylotrophic methanogenesis may have evolved from a methanotrophic ancestor.

Microbial sulfur metabolism contributes to biogeochemical cycling on global scales. Sulfur metabolizing microbes are infected by phages that can encode auxiliary metabolic genes (AMGs) to alter sulfur metabolism within host cells but remain poorly characterized. Here we identified 191 phages derived from twelve environments that encoded 227 AMGs for oxidation of sulfur and thiosulfate (dsrA, dsrC/tusE, soxC, soxD and soxYZ). Evidence for retention of AMGs during niche-differentiation of diverse phage populations provided evidence that auxiliary metabolism imparts measurable fitness benefits to phages with ramifications for ecosystem biogeochemistry. Gene abundance and expression profiles of AMGs suggested significant contributions by phages to sulfur and thiosulfate oxidation in freshwater lakes and oceans, and a sensitive response to changing sulfur concentrations in hydrothermal environments. Overall, our study provides fundamental insights on the distribution, diversity, and ecology of phage auxiliary metabolism associated with sulfur and reinforces the necessity of incorporating viral contributions into biogeochemical configurations.

Changes in the sequence of an organism's genome, i.e., mutations, are the raw material of evolution. The frequency and location of mutations can be constrained by specific molecular mechanisms, such as diversity-generating retroelements (DGRs). DGRs have been characterized from cultivated bacteria and bacteriophages, and perform error-prone reverse transcription leading to mutations being introduced in specific target genes. DGR loci were also identified in several metagenomes, but the ecological roles and evolutionary drivers of these DGRs remain poorly understood. Here, we analyze a dataset of >30,000 DGRs from public metagenomes, establish six major lineages of DGRs including three primarily encoded by phages and seemingly used to diversify host attachment proteins, and demonstrate that DGRs are broadly active and responsible for >10% of all amino acid changes in some organisms. Overall, these results highlight the constraints under which DGRs evolve, and elucidate several distinct roles these elements play in natural communities.

Viruses play an important role in the ecology and biogeochemistry of marine ecosystems. Beyond mortality and gene transfer, viruses can reprogram microbial metabolism during infection by expressing auxiliary metabolic genes (AMGs) involved in photosynthesis, central carbon metabolism, and nutrient cycling. While previous studies have focused on AMG diversity in the sunlit and dark ocean, less is known about the role of viruses in shaping metabolic networks along redox gradients associated with marine oxygen minimum zones (OMZs). Here, we analyzed relatively quantitative viral metagenomic datasets that profiled the oxygen gradient across Eastern Tropical South Pacific (ETSP) OMZ waters, assessing whether OMZ viruses might impact nitrogen (N) cycling via AMGs. Identified viral genomes encoded six N-cycle AMGs associated with denitrification, nitrification, assimilatory nitrate reduction, and nitrite transport. The majority of these AMGs (80%) were identified in T4-like Myoviridae phages, predicted to infect Cyanobacteria and Proteobacteria, or in unclassified archaeal viruses predicted to infect Thaumarchaeota. Four AMGs were exclusive to anoxic waters and had distributions that paralleled homologous microbial genes. Together, these findings suggest viruses modulate N-cycling processes within the ETSP OMZ and may contribute to nitrogen loss throughout the global oceans thus providing a baseline for their inclusion in the ecosystem and geochemical models.

Monitoring population dynamics in co-culture is necessary in engineering microbial consortia involved in distributed metabolic processes or biosensing applications. However, it remains difficult to measure strain-specific growth dynamics in high-throughput formats. This is especially vexing in plate-based functional screens leveraging whole-cell biosensors to detect specific metabolic signals. Here, we develop an experimental high-throughput co-culture system to measure and model the relationship between fluorescence and cell abundance, combining chassis-independent recombinase-assisted genome engineering (CRAGE) and whole-cell biosensing with a PemrR-green fluorescent protein (GFP) monoaromatic reporter used in plate-based functional screening. CRAGE was used to construct Escherichia coli EPI300 strains constitutively expressing red fluorescent protein (RFP) and the relationship between RFP expression and optical density (OD600) was determined throughout the EPI300 growth cycle. A linear equation describing the increase of normalized RFP fluorescence during deceleration phase was derived and used to predict biosensor strain dynamics in co-culture. Measured and predicted values were compared using flow cytometric detection methods. Induction of the biosensor lead to increased GFP fluorescence normalized to biosensor cell abundance, as expected, but a significant decrease in relative abundance of the biosensor strain in co-culture and a decrease in bulk GFP fluorescence. Taken together, these results highlight sensitivity of population dynamics to variations in metabolic activity in co-culture and the potential effect of these dynamics on the performance of functional screens in plate-based formats. The engineered strains and model used to evaluate these dynamics provide a framework for optimizing growth of synthetic co-cultures used in screening, testing and pathway engineering applications.

Methanogenesis is an ancient metabolism of key ecological relevance, with direct impact on the evolution of Earth's climate. Recent results suggest that the diversity of methane metabolisms and their derivations have probably been vastly underestimated. Here, by probing thousands of publicly available metagenomes for homologues of methyl-coenzyme M reductase complex (MCR), we have obtained ten metagenome-assembled genomes (MAGs) belonging to potential methanogenic, anaerobic methanotrophic and short-chain alkane-oxidizing archaea. Five of these MAGs represent under-sampled (Verstraetearchaeota, Methanonatronarchaeia, ANME-1 and GoM-Arc1) or previously genomically undescribed (ANME-2c) archaeal lineages. The remaining five MAGs correspond to lineages that are only distantly related to previously known methanogens and span the entire archaeal phylogeny. Comprehensive comparative annotation substantially expands the metabolic diversity and energy conservation systems of MCR-bearing archaea. It also suggests the potential existence of a yet uncharacterized type of methanogenesis linked to short-chain alkane/fatty acid oxidation in a previously undescribed class of archaea ('Candidatus Methanoliparia'). We redefine a common core of marker genes specific to methanogenic, anaerobic methanotrophic and short-chain alkane-oxidizing archaea, and propose a possible scenario for the evolutionary and functional transitions that led to the emergence of such metabolic diversity.

Publication interests should not limit access to public data

In the version of this article initially published, the author A. Murat Eren was listed as A.M. Eren. The corresponding affiliation was given as the Knapp Center for Biomedical Discovery, rather than Department of Medicine, University of Chicago, Chicago, Illinois, USA, and Marine Biological Laboratory, Woods Hole, Massachusetts, USA. The errors have been corrected in the HTML and PDF versions of the article as of 29 November 2017. In the version of this article initially published, the following acknowledgment was omitted: A.L. was supported by the Russian Science Foundation (grant number 14-50-00069). The error has been corrected in the HTML and PDF versions of the article as of 7 December 2017.

Microbial communities drive biogeochemical cycles through networks of metabolite exchange that are structured along energetic gradients. As energy yields become limiting, these networks favor co-metabolic interactions to maximize energy disequilibria. Here we apply single-cell genomics, metagenomics, and metatranscriptomics to study bacterial populations of the abundant "microbial dark matter" phylum Marinimicrobia along defined energy gradients. We show that evolutionary diversification of major Marinimicrobia clades appears to be closely related to energy yields, with increased co-metabolic interactions in more deeply branching clades. Several of these clades appear to participate in the biogeochemical cycling of sulfur and nitrogen, filling previously unassigned niches in the ocean. Notably, two Marinimicrobia clades, occupying different energetic niches, express nitrous oxide reductase, potentially acting as a global sink for the greenhouse gas nitrous oxide.

Marine oxygen minimum zones (OMZs) are widespread regions of the ocean that are currently expanding due to global warming. While inhospitable to most metazoans, OMZs are hotspots for microbial mediated biogeochemical cycling of carbon, nitrogen and sulphur, contributing disproportionately to marine nitrogen loss and climate active trace gas production. Our current understanding of microbial community responses to OMZ expansion is limited by a lack of time-resolved data sets linking multi-omic sequence information (DNA, RNA, protein) to geochemical parameters and process rates. Here, we present six years of time-resolved multi-omic observations in Saanich Inlet, a seasonally anoxic fjord on the coast of Vancouver Island, British Columbia, Canada that undergoes recurring changes in water column oxygenation status. This compendium provides a unique multi-omic framework for studying microbial community responses to ocean deoxygenation along defined geochemical gradients in OMZ waters.

We present two standards developed by the Genomic Standards Consortium (GSC) for reporting bacterial and archaeal genome sequences. Both are extensions of the Minimum Information about Any (x) Sequence (MIxS). The standards are the Minimum Information about a Single Amplified Genome (MISAG) and the Minimum Information about a Metagenome-Assembled Genome (MIMAG), including, but not limited to, assembly quality, and estimates of genome completeness and contamination. These standards can be used in combination with other GSC checklists, including the Minimum Information about a Genome Sequence (MIGS), Minimum Information about a Metagenomic Sequence (MIMS), and Minimum Information about a Marker Gene Sequence (MIMARKS). Community-wide adoption of MISAG and MIMAG will facilitate more robust comparative genomic analyses of bacterial and archaeal diversity.

Under nitrogen deprivation (-N), cyanobacterium Synechocystis sp. PCC 6803 exhibits growth arrest, reduced protein content, and remarkably increased glycogen accumulation. However, producing glycogen under this condition requires a two-step process with cell transfer from normal to -N medium. Metabolic engineering and chemical treatment for rapid glycogen accumulation can bypass the need for two-step cultivation. For example, recent studies indicate that individually disrupting hydrogen (H2) or poly(3-hydroxybutyrate) (PHB) synthesis, or treatment with methyl viologen (MV), effectively increases glycogen accumulation in Synechocystis. Here we explore the effects of disrupted H2 or poly(3-hydroxybutyrate) synthesis, together with MV treatment to on enhanced glycogen accumulation in Synechocystis grown in normal medium. Wild-type cells without MV treatment exhibited low glycogen content of less than 6% w/w dry weight (DW). Compared with wild type, disrupting PHB synthesis combined with MV treatment did not increase glycogen content. Disrupted H₂ production without MV treatment yielded up to 11% w/w DW glycogen content. Interestingly, when combined, disrupted H2 production with MV treatment synergistically enhanced glycogen accumulation to 51% and 59% w/w DW within 3 and 7 days, respectively. Metabolomic analysis suggests that MV treatment mediated the conversion of proteins into glycogen. Metabolomic and transcriptional-expression analysis suggests that disrupted H2 synthesis under MV treatment positively influenced glycogen synthesis. Disrupted H₂ synthesis under MV treatment significantly increased NADPH levels. This increased NADPH content potentially contributed to the observed enhancements in antioxidant activity against MV-induced oxidants, O2 evolution, and metabolite substrates levels for glycogen synthesis in normal medium, ultimately leading to enhanced glycogen accumulation in Synechocystis. Key message: Combining disrupted hydrogen-gas synthesis and the treatment by photosynthesis electron-transport inhibitor significantly enhance glycogen production in cyanobacteria. [ABSTRACT FROM AUTHOR]

Over the past decade, high-throughput short-read 16S rRNA gene amplicon sequencing has eclipsed clone-dependent long-read Sanger sequencing for microbial community profiling. The transition to new technologies has provided more quantitative information at the expense of taxonomic resolution with implications for inferring metabolic traits in various ecosystems. We applied single-molecule real-time sequencing for microbial community profiling, generating full-length 16S rRNA gene sequences at high throughput, which we propose to name PhyloTags. We benchmarked and validated this approach using a defined microbial community. When further applied to samples from the water column of meromictic Sakinaw Lake, we show that while community structures at the phylum level are comparable between PhyloTags and Illumina V4 16S rRNA gene sequences (iTags), variance increases with community complexity at greater water depths. PhyloTags moreover allowed less ambiguous classification. Last, a platform-independent comparison of PhyloTags and in silico generated partial 16S rRNA gene sequences demonstrated significant differences in community structure and phylogenetic resolution across multiple taxonomic levels, including a severe underestimation in the abundance of specific microbial genera involved in nitrogen and methane cycling across the Lake's water column. Thus, PhyloTags provide a reliable adjunct or alternative to cost-effective iTags, enabling more accurate phylogenetic resolution of microbial communities and predictions on their metabolic potential.

The 'Atribacteria' is a candidate phylum in the Bacteria recently proposed to include members of the OP9 and JS1 lineages. OP9 and JS1 are globally distributed, and in some cases abundant, in anaerobic marine sediments, geothermal environments, anaerobic digesters and reactors and petroleum reservoirs. However, the monophyly of OP9 and JS1 has been questioned and their physiology and ecology remain largely enigmatic due to a lack of cultivated representatives. Here cultivation-independent genomic approaches were used to provide a first comprehensive view of the phylogeny, conserved genomic features and metabolic potential of members of this ubiquitous candidate phylum. Previously available and heretofore unpublished OP9 and JS1 single-cell genomic data sets were used as recruitment platforms for the reconstruction of atribacterial metagenome bins from a terephthalate-degrading reactor biofilm and from the monimolimnion of meromictic Sakinaw Lake. The single-cell genomes and metagenome bins together comprise six species- to genus-level groups that represent most major lineages within OP9 and JS1. Phylogenomic analyses of these combined data sets confirmed the monophyly of the 'Atribacteria' inclusive of OP9 and JS1. Additional conserved features within the 'Atribacteria' were identified, including a gene cluster encoding putative bacterial microcompartments that may be involved in aldehyde and sugar metabolism, energy conservation and carbon storage. Comparative analysis of the metabolic potential inferred from these data sets revealed that members of the 'Atribacteria' are likely to be heterotrophic anaerobes that lack respiratory capacity, with some lineages predicted to specialize in either primary fermentation of carbohydrates or secondary fermentation of organic acids, such as propionate.

Viral infections dynamically alter the composition and metabolic potential of marine microbial communities and the evolutionary trajectories of host populations with resulting feedback on biogeochemical cycles. It is quite possible that all microbial populations in the ocean are impacted by viral infections. Our knowledge of virus-host relationships, however, has been limited to a minute fraction of cultivated host groups. Here, we utilized single-cell sequencing to obtain genomic blueprints of viruses inside or attached to individual bacterial and archaeal cells captured in their native environment, circumventing the need for host and virus cultivation. A combination of comparative genomics, metagenomic fragment recruitment, sequence anomalies and irregularities in sequence coverage depth and genome recovery were utilized to detect viruses and to decipher modes of virus-host interactions. Members of all three tailed phage families were identified in 20 out of 58 phylogenetically and geographically diverse single amplified genomes (SAGs) of marine bacteria and archaea. At least four phage-host interactions had the characteristics of late lytic infections, all of which were found in metabolically active cells. One virus had genetic potential for lysogeny. Our findings include first known viruses of Thaumarchaeota, Marinimicrobia, Verrucomicrobia and Gammaproteobacteria clusters SAR86 and SAR92. Viruses were also found in SAGs of Alphaproteobacteria and Bacteroidetes. A high fragment recruitment of viral metagenomic reads confirmed that most of the SAG-associated viruses are abundant in the ocean. Our study demonstrates that single-cell genomics, in conjunction with sequence-based computational tools, enable in situ, cultivation-independent insights into host-virus interactions in complex microbial communities.

The "Latescibacteria" (formerly WS3), member of the Fibrobacteres-Chlorobi-Bacteroidetes (FCB) superphylum, represents a ubiquitous candidate phylum found in terrestrial, aquatic, and marine ecosystems. Recently, single-cell amplified genomes (SAGs) representing the "Latescibacteria" were obtained from the anoxic monimolimnion layers of Sakinaw Lake (British Columbia, Canada), and anoxic sediments of a coastal lagoon (Etoliko lagoon, Western Greece). Here, we present a detailed in-silico analysis of the four SAGs to gain some insights on their metabolic potential and apparent ecological roles. Metabolic reconstruction suggests an anaerobic fermentative mode of metabolism, as well as the capability to degrade multiple polysaccharides and glycoproteins that represent integral components of green (Charophyta and Chlorophyta) and brown (Phaeophycaea) algae cell walls (pectin, alginate, ulvan, fucan, hydroxyproline-rich glycoproteins), storage molecules (starch and trehalose), and extracellular polymeric substances (EPSs). The analyzed SAGs also encode dedicated transporters for the uptake of produced sugars and amino acids/oligopeptides, as well as an extensive machinery for the catabolism of all transported sugars, including the production of a bacterial microcompartment (BMC) to sequester propionaldehyde, a toxic intermediate produced during fucose and rhamnose metabolism. Finally, genes for the formation of gas vesicles, flagella, type IV pili, and oxidative stress response were found, features that could aid in cellular association with algal detritus. Collectively, these results indicate that the analyzed "Latescibacteria" mediate the turnover of multiple complex organic polymers of algal origin that reach deeper anoxic/microoxic habitats in lakes and lagoons. The implications of such process on our understanding of niche specialization in microbial communities mediating organic carbon turnover in stratified water bodies are discussed.

The ecological importance of viruses is now widely recognized, yet our limited knowledge of viral sequence space and virus-host interactions precludes accurate prediction of their roles and impacts. In this study, we mined publicly available bacterial and archaeal genomic data sets to identify 12,498 high-confidence viral genomes linked to their microbial hosts. These data augment public data sets 10-fold, provide first viral sequences for 13 new bacterial phyla including ecologically abundant phyla, and help taxonomically identify 7-38% of 'unknown' sequence space in viromes. Genome- and network-based classification was largely consistent with accepted viral taxonomy and suggested that (i) 264 new viral genera were identified (doubling known genera) and (ii) cross-taxon genomic recombination is limited. Further analyses provided empirical data on extrachromosomal prophages and coinfection prevalences, as well as evaluation of in silico virus-host linkage predictions. Together these findings illustrate the value of mining viral signal from microbial genomes.