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Cyanobacteria are highly abundant in the marine photic zone and primary drivers of the conversion of inorganic carbon into biomass. To date, all studied cyanobacterial lineages encode carbon fixation machinery relying upon form I Rubiscos within a CO₂-concentrating carboxysome. Here, we report that the uncultivated anoxic marine zone (AMZ) IB lineage of Prochlorococcus from pelagic oxygen-deficient zones (ODZs) harbors both form I and form II Rubiscos, the latter of which are typically noncarboxysomal and possess biochemical properties tuned toward low-oxygen environments. We demonstrate that these cyanobacterial form II enzymes are functional in vitro and were likely acquired from proteobacteria. Metagenomic analysis reveals that AMZ IB are essentially restricted to ODZs in the Eastern Pacific, suggesting that form II acquisition may confer an advantage under low-O₂conditions. AMZ IB populations express both forms of Rubisco in situ, with the highest form II expression at depths where oxygen and light are low, possibly as a mechanism to increase the efficiency of photoautotrophy under energy limitation. Our findings expand the diversity of carbon fixation configurations in the microbial world and may have implications for carbon sequestration in natural and engineered systems. 

Abstract Motivation Linking microbial community members to their ecological functions is a central goal of environmental microbiology. When assigned taxonomy, amplicon sequences of metabolic marker genes can suggest such links, thereby offering an overview of the phylogenetic structure underpinning particular ecosystem functions. However, inferring microbial taxonomy from metabolic marker gene sequences remains a challenge, particularly for the frequently sequenced nitrogen fixation marker gene, nitrogenase reductase (nifH). Horizontal gene transfer in recent nifH evolutionary history can confound taxonomic inferences drawn from the pairwise identity methods used in existing software. Other methods for inferring taxonomy are not standardized and require manual inspection that is difficult to scale. Results We present Phylogenetic Placement for Inferring Taxonomy (PPIT), an R package that infers microbial taxonomy from nifH amplicons using both phylogenetic and sequence identity approaches. After users place query sequences on a reference nifH gene tree provided by PPIT (n = 6317 full-length nifH sequences), PPIT searches the phylogenetic neighborhood of each query sequence and attempts to infer microbial taxonomy. An inference is drawn only if references in the phylogenetic neighborhood are: (1) taxonomically consistent and (2) share sufficient pairwise identity with the query, thereby avoiding erroneous inferences due to known horizontal gene transfer events. We find that PPIT returns a higher proportion of correct taxonomic inferences than BLAST-based approaches at the cost of fewer total inferences. We demonstrate PPIT on deep-sea sediment and find that Deltaproteobacteria are the most abundant potential diazotrophs. Using this dataset we show that emending PPIT inferences based on visual inspection of query sequence placement can achieve taxonomic inferences for nearly all sequences in a query set. We additionally discuss how users can apply PPIT to the analysis of other marker genes. Availability PPIT is freely available to non-commercial users at https://github.com/bkapili/ppit. Installation includes a vignette that demonstrates package use and reproduces the nifH amplicon analysis discussed here. The raw nifH amplicon sequence data have been deposited in the GenBank, EMBL, and DDBJ databases under BioProject number PRJEB37167. Supplementary information Supplementary data are available at Bioinformatics online. 

Abstract Diazotrophic microorganisms regulate marine productivity by alleviating nitrogen limitation. However, we know little about the identity and activity of diazotrophs in deep-sea sediments, a habitat covering nearly two-thirds of the planet. Here, we identify candidate diazotrophs from Pacific Ocean sediments collected at 2893 m water depth using 15N-DNA stable isotope probing and a novel pipeline for nifH sequence analysis. Together, these approaches detect an unexpectedly diverse assemblage of active diazotrophs, including members of the Acidobacteria, Firmicutes, Nitrospirae, Gammaproteobacteria, and Deltaproteobacteria. Deltaproteobacteria, predominately members of the Desulfobacterales and Desulfuromonadales, are the most abundant diazotrophs detected, and display the most microdiversity of associated nifH sequences. Some of the detected lineages, including those within the Acidobacteria, have not previously been shown to fix nitrogen. The diazotrophs appear catabolically diverse, with the potential for using oxygen, nitrogen, iron, sulfur, and carbon as terminal electron acceptors. Therefore, benthic diazotrophy may persist throughout a range of geochemical conditions and provide a stable source of fixed nitrogen over geologic timescales. Our results suggest that nitrogen-fixing communities in deep-sea sediments are phylogenetically and catabolically diverse, and open a new line of inquiry into the ecology and biogeochemical impacts of deep-sea microorganisms. 

Abstract Trace metals have been an important ingredient for life throughout Earth’s history. Here, we describe the genome-guided cultivation of a member of the elusive archaeal lineage Caldarchaeales (syn. Aigarchaeota ), Wolframiiraptor gerlachensis , and its growth dependence on tungsten. A metagenome-assembled genome (MAG) of W. gerlachensis encodes putative tungsten membrane transport systems, as well as pathways for anaerobic oxidation of sugars probably mediated by tungsten-dependent ferredoxin oxidoreductases that are expressed during growth. Catalyzed reporter deposition-fluorescence in-situ hybridization (CARD-FISH) and nanoscale secondary ion mass spectrometry (nanoSIMS) show that W. gerlachensis preferentially assimilates xylose. Phylogenetic analyses of 78 high-quality Wolframiiraptoraceae MAGs from terrestrial and marine hydrothermal systems suggest that tungsten-associated enzymes were present in the last common ancestor of extant Wolframiiraptoraceae . Our observations imply a crucial role for tungsten-dependent metabolism in the origin and evolution of this lineage, and hint at a relic metabolic dependence on this trace metal in early anaerobic thermophiles.