Shotgun metagenomics indicates novel family A DNA polymerases predominate within marine virioplankton

Abstract

Virioplankton have a significant role in marine ecosystems, yet we know little of the predominant biological characteristics of aquatic viruses that influence the flow of nutrients and energy through microbial communities. Family A DNA polymerases, critical to DNA replication and repair in prokaryotes, are found in many tailed bacteriophages. The essential role of DNA polymerase in viral replication makes it a useful target for connecting viral diversity with an important biological feature of viruses. Capturing the full diversity of this polymorphic gene by targeted approaches has been difficult; thus, full-length DNA polymerase genes were assembled out of virioplankton shotgun metagenomic sequence libraries (viromes). Within the viromes novel DNA polymerases were common and found in both double-stranded (ds) DNA and single-stranded (ss) DNA libraries. Finding DNA polymerase genes in ssDNA viral libraries was unexpected, as no such genes have been previously reported from ssDNA phage. Surprisingly, the most common virioplankton DNA polymerases were related to a siphovirus infecting an a-proteobacterial symbiont of a marine sponge and not the podoviral T7-like polymerases seen in many other studies. Amino acids predictive of catalytic efficiency and fidelity linked perfectly to the environmental clades, indicating that most DNA polymerase-carrying virioplankton utilize a lower efficiency, higher fidelity enzyme. Comparisons with previously reported, PCR-amplified DNA polymerase sequences indicated that the most common virioplankton metagenomic DNA polymerases formed a new group that included siphoviruses. These data indicate that slower-replicating, lytic or lysogenic phage populations rather than fast-replicating, highly lytic phages may predominate within the virioplankton.

Methods for the extraction, storage, amplification and sequencing of DNA from environmental samples

“…Instead, virus-focused metagenomics studies either concentrate virus-genomes via their virions or enrich for double-stranded RNA that is a broadly shared feature of virus replication. The virus-enriched nucleic acids are then sequenced by shotgun metagenomics methods (Bibby 2013) for the detection of viruses in environmental material, including marine water (Schmidt et al 2014), fresh water (Mohiuddin & Schellhorn 2015), soil (Ballaud et al 2016), crop plants (Aw et al 2016), native plants and weed plant species (Blouin et al 2016). We remain unable to recommend a single molecular approach to comprehensively target all of the diversity of viral material for metabarcoding studies in an unbiased manner, i.e.…”

Polar freshwater cyanophage S-EIV1 represents a new widespread evolutionary lineage of phages

“…PUP clade was identified by Breitbart et al (2004), Env clade 1-3 were identified by Labonté et al (2009). Env-4 was identified by Schmidt et al (2014). Env 5 is a new environmental clade that includes cyanophage S-EIV1.…”

“…Phylogenetic analysis of DNApolA with known phages, metagenomic sequences and amplicons (Breitbart et al, 2004;Labonté et al, 2009;Schmidt et al, 2014) reveals that S-EIV1 along with environmental sequences forms a previously unrecognized evolutionary group of DNApolA sequences (Figure 4). Further evidence for S-EIV1-like phages in aquatic systems was obtained from the recruitment of reads from environmental viral metagenomic data (Figures 6a and b).…”

Genetic diversity and temporal dynamics of phytoplankton viruses in East Lake, China

“…The scale bar represents the number of amino acid substitutions per residue. Huang et al, 2010;Schmidt et al, 2014). Phylogenetic analysis was conducted with 52 different polA amino acid sequences from this study and close relatives retrieved from GenBank ( Figure 3).…”