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. 2015 Mar 16:6:199.
doi: 10.3389/fmicb.2015.00199. eCollection 2015.

Genomic characteristics and environmental distributions of the uncultivated Far-T4 phages

Simon Roux  1 , François Enault  2 , Viviane Ravet  2 , Olivier Pereira  2 , Matthew B Sullivan  1
Affiliations

Genomic characteristics and environmental distributions of the uncultivated Far-T4 phages

Simon Roux et al. Front Microbiol. .

Abstract

Viral metagenomics (viromics) is a tremendous tool to reveal viral taxonomic and functional diversity across ecosystems ranging from the human gut to the world's oceans. As with microbes however, there appear vast swaths of "dark matter" yet to be documented for viruses, even among relatively well-studied viral types. Here, we use viromics to explore the "Far-T4 phages" sequence space, a neighbor clade from the well-studied T4-like phages that was first detected through PCR study in seawater and subsequently identified in freshwater lakes through 454-sequenced viromes. To advance the description of these viruses beyond this single marker gene, we explore Far-T4 genome fragments assembled from two deeply-sequenced freshwater viromes. Single gene phylogenetic trees confirm that the Far-T4 phages are divergent from the T4-like phages, genome fragments reveal largely collinear genome organizations, and both data led to the delineation of five Far-T4 clades. Three-dimensional models of major capsid proteins are consistent with a T4-like structure, and highlight a highly conserved core flanked by variable insertions. Finally, we contextualize these now better characterized Far-T4 phages by re-analyzing 196 previously published viromes. These suggest that Far-T4 are common in freshwater and seawater as only four of 82 aquatic viromes lacked Far-T4-like sequences. Variability in representation across the five newly identified clades suggests clade-specific niche differentiation may be occurring across the different biomes, though the underlying mechanism remains unidentified. While complete genome assembly from complex communities and the lack of host linkage information still bottleneck virus discovery through viromes, these findings exemplify the power of metagenomics approaches to assess the diversity, evolutionary history, and genomic characteristics of novel uncultivated phages.

Keywords: Caudovirales; T4 phages; capsid proteins; freshwater ecology; viral genomes.

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Figures

Figure 1
Figure 1
Phylogenetic tree computed from Gp23 (major capsid protein) multiple alignment. As this tree includes the PCR sequences used to define the Far-T4 group, the multiple alignment was trimmed to include only positions matching these amplicons. All nodes with bootstraps lower than 50 were collapsed. Far-T4 sequences are color-coded according to their original sample, and reference sequences are in black.
Figure 2
Figure 2
Comparison of Far-T4 contigs assembled from the Lake Pavin viromes. The two deeply sequenced viromes from Lake Pavin allowed for the assembly of large genome fragments, which are here compared in terms of gene similarity and order. Genes are color-coded according to their functional annotation, and major capsid genes (g23) are outlined in black. Significant sequence similarity between successive sequences are highlighted by gray squares, shaded according to percentage of amino-acid identity. Predicted proteins with no associated function are classified as “Uncharacterized phage proteins” when at least one similar sequence was detected in a known phage and “Hypothetical protein” otherwise. PurM, Phosphoribosylformylglycinami-dine cyclo-ligase; GTP Ch, GTP cyclohydrolase; DNA Pol, DNA polymerase; PRT, Phosphoribosyl transferase; Hsp, Heat shock protein.
Figure 3
Figure 3
Predicted structure of the Far-T4 major capsid protein (two views rotated by 90° through the y-axis). This model is based on the sequence from contig Pavin_2013_4m_8, and was predicted with I-Tasser based on the structure of the T4 phage vertex protein. Models are colored according to their conservation across T4-like phages, from least (red) to most (blue) conserved. Residues corresponding to insertions in the Far-T4 sequence not present in other T4 phages are highlighted with spheres.
Figure 4
Figure 4
Coverage of different Far-T4 clades in seawater and freshwater viral metagenomes. For each category of virome (freshwater and seawater), the coverage of each contig (i.e., number of bp similar to a contig normalized by the contig length) was computed from 2.5 millions of randomly selected reads from each virome. Coverage could not be assessed for Far-T4 clade 2 since no contigs are available for this clade.
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