Assembly-free single-molecule nanopore sequencing recovers complete virus genomes from natural microbial communities

Beaulaurier, John; Luo, Elaine; Eppley, John M.; Uyl, Paul Den; Dai, Xiaofeng; Turner, Daniel J.; Pendelton, Matthew; Juul, Sissel; Harrington, Eoghan; DeLong, Edward F.

doi:10.1101/619684

Cited by 9 publications

(7 citation statements)

References 59 publications

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“…4 ). It is likely that similar SAR11 prophages have gone undetected in short-read metagenomic surveys due to the inherent genetic complexity of SAR11 and marine phage 39 . PNP1-like prophages were identified in the subset of metagenomic surveys that sequenced complete virus genomes using fosmids or long-read sequencing.…”

Section: Methodsmentioning

confidence: 99%

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Lysogenic host–virus interactions in SAR11 marine bacteria

et al. 2020

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Host-virus interactions structure microbial communities, drive biogeochemical cycles, and enhance genetic diversity in nature 1 , 2 . Hypotheses proposed to explain the range of interactions that mediate these processes often invoke lysogeny 3 – 6 , a latent infection strategy used by temperate bacterial viruses to replicate in host cells until an induction event triggers the production and lytic release of free viruses. Most cultured bacteria harbor temperate viruses in their genomes (prophage) 7 . The absence of prophages in cultures of the dominant lineages of marine bacteria has contributed to an ongoing debate over the ecological significance of lysogeny and other viral life strategies in nature 6 , 8 – 15 . Here we report the discovery of prophages in cultured SAR11, the ocean’s most abundant clade of heterotrophic bacteria 16 , 17 . We show the concurrent production of cells and viruses, with enhanced virus production under carbon-limiting growth conditions. Evidence that related prophages are broadly distributed in the oceans suggests that similar interactions have contributed to the evolutionary success of SAR11 in nutrient limited systems.

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Section: Methodsmentioning

confidence: 99%

“…We identified closely related phage genomes in a metagenomic study of marine viruses from the Mediterranean Sea 38 and from the North Pacific 39 ( Fig. 4 ).…”

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confidence: 99%

Lysogenic host–virus interactions in SAR11 marine bacteria

et al. 2020

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“…790 from LAST to identify overlaps at 95% ANI across 100 bp-10 kbp within 200 bp of both ends; and iv. 1131 from NUCmer v3.1 [42] to identify direct terminal repeats 20bp-10kbp in length within 200 bp of both ends [43]. 1543 complete genomes were identified pooled amongst these four methods (Table S3).…”

Section: Genomic Completionmentioning

confidence: 99%

Double-stranded DNA virioplankton dynamics and reproductive strategies in the oligotrophic open ocean water column

et al. 2020

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Microbial communities are critical to ecosystem dynamics and biogeochemical cycling in the open oceans. Viruses are essential elements of these communities, influencing the productivity, diversity, and evolution of cellular hosts. To further explore the natural history and ecology of open-ocean viruses, we surveyed the spatiotemporal dynamics of double-stranded DNA (dsDNA) viruses in both virioplankton and bacterioplankton size fractions in the North Pacific Subtropical Gyre, one of the largest biomes on the planet. Assembly and clustering of viral genomes revealed a peak in virioplankton diversity at the base of the euphotic zone, where virus populations and host species richness both reached their maxima. Simultaneous characterization of both extracellular and intracellular viruses suggested depth-specific reproductive strategies. In particular, analyses indicated elevated lytic interactions in the mixed layer, more temporally variable temperate phage interactions at the base of the euphotic zone, and increased lysogeny in the mesopelagic ocean. Furthermore, the depth variability of auxiliary metabolic genes suggested habitat-specific strategies for viral influence on light-energy, nitrogen, and phosphorus acquisition during host infection. Most virus populations were temporally persistent over several years in this environment at the 95% nucleic acid identity level. In total, our analyses revealed variable distributional patterns and diverse reproductive and metabolic strategies of virus populations in the open-ocean water column.

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“…The long-read information bridges gaps in a short-read-only assembly that often occur due to intra- and interspecies repeats 6 . Complete viral genomes can be already identified from environmental samples without any assembly step via nanopore-based sequencing 7 . Combined with a reduction in cost per gigabase 8 and an increase in data output, the technologies for sequencing long reads quickly became suitable for metagenomic analysis 9–12 .…”

Section: Introductionmentioning

confidence: 99%

Metagenomics workflow for hybrid assembly, differential coverage binning, transcriptomics and pathway analysis (MUFFIN)

Damme

Hölzer

Viehweger

et al. 2020

Preprint

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Metagenomics has redefined many areas of microbiology. However, metagenome-assembled genomes (MAGs) are often fragmented, primarily when sequencing was performed with short reads. Recent long-read sequencing technologies promise to improve genome reconstruction. However, the integration of two different sequencing modalities makes downstream analyses complex. We, therefore, developed MUFFIN, a complete metagenomic workflow that uses short and long reads to produce high-quality bins and their annotations. The workflow is written by using Nextflow, a workflow orchestration software, to achieve high reproducibility and fast and straightforward use. This workflow also produces the taxonomic classification and KEGG pathways of the bins and can be further used by providing RNA-Seq data (optionally) for quantification and annotation. We tested the workflow using twenty biogas reactor samples and assessed the capacity of MUFFIN to process and output relevant files needed to analyze the microbial community and their function. MUFFIN produces functional pathway predictions and if provided de novo transcript annotations across the metagenomic sample and for each bin.Author SummaryRVD did the development and design of MUFFIN and wrote the first draft; BM and EBR did the critical reading and correction of the manuscript; MH did the critical reading of the manuscript and the general adjustments for the metagenomic workflow; AV did the critical reading of the manuscript and adjustments for the taxonomic classifications. CB supervised the project, did the workflow design, helped with the implementation, and revised the manuscript.

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Assembly-free single-molecule nanopore sequencing recovers complete virus genomes from natural microbial communities

Cited by 9 publications

References 59 publications

Lysogenic host–virus interactions in SAR11 marine bacteria

Lysogenic host–virus interactions in SAR11 marine bacteria

Double-stranded DNA virioplankton dynamics and reproductive strategies in the oligotrophic open ocean water column

Metagenomics workflow for hybrid assembly, differential coverage binning, transcriptomics and pathway analysis (MUFFIN)

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