Although shotgun metagenomic sequencing of microbiome samples enables partial reconstruction of strain-level community structure, obtaining high-quality microbial genome drafts without isolation and culture remains difficult. Here, we present an application of read clouds, short-read sequences tagged with long-range information, to microbiome samples. We present Athena, a de novo assembler that uses read clouds to improve metagenomic assemblies. We applied this approach to sequence stool samples from two healthy individuals and compared it with existing short-read and synthetic long-read metagenomic sequencing techniques. Read-cloud metagenomic sequencing and Athena assembly produced the most comprehensive individual genome drafts with high contiguity (>200-kb N50, fewer than ten contigs), even for bacteria with relatively low (20×) raw short-read-sequence coverage. We also sequenced a complex marine-sediment sample and generated 24 intermediate-quality genome drafts (>70% complete, <10% contaminated), nine of which were complete (>90% complete, <5% contaminated). Our approach allows for culture-free generation of high-quality microbial genome drafts by using a single shotgun experiment.
Marine sediments contain half of all marine microbial cells, and benthic archaea constitute an important part of those microorganisms. Studies on the activity of these archaea demonstrate these microorganisms are deeply entwined in carbon cycling within sediments and influence the availability of inorganic and organic carbon to the atmosphere and the deep subsurface. However, the metabolic capabilities of most benthic archaea are poorly characterized; therefore, their specific contributions to carbon cycling are unknown. Furthermore, the relationship between genetic and functional diversity of benthic archaeal lineages and the physicochemical and ecosystem controls on that diversity are unknown. This project aims to address these gaps in our knowledge, utilizing stable isotope probing to determine the incorporation of isotopically labelled substrates in conjunction with metagenomic and metatranscriptomic sequencing of benthic archaeal communities within sediments collected across a transect. This approach will identify the autotrophic, heterotrophic, and mixotrophic activity of known and uncharacterized benthic archaea, and assess differences in the magnitude of their activity. Furthermore, the proposed analyses will provide insight on the genetic diversity of expressed genes to help determine what genetic variations determine the archaeal community compositions in different environments and how these organisms relate to those found in the deep subsurface.