Morphological and taxonomic properties of the newly isolated Cotonvirus japonicus , a new lineage of the subfamily Megavirinae

Since 2003, various viruses from the subfamily Megavirinae in the family Mimiviridae have been isolated worldwide, including icosahedral mimiviruses and tailed tupanviruses. To date, the evolutionary relationship between tailed and non-tailed mimiviruses has not yet been elucidated. Here, we present the genomic and morphological features of a newly isolated giant virus, Cotonvirus japonicus (cotonvirus), belonging to the family Mimiviridae. It contains a linear double-stranded DNA molecule of 1.47 Mb, the largest among the reported viruses in the subfamily Megavirinae , excluding tupanviruses. Among its 1,306 predicted open reading frames, 1,149 (88.0%) were homologous to those of the family Mimiviridae . Several nucleocytoplasmic large DNA virus (NCLDV) core genes, aminoacyl-tRNA synthetase genes, and the host specificity of cotonvirus were highly similar to those of Mimiviridae lineages A, B, and C; however, lineage A was slightly closer to cotonvirus than to the others. Moreover, based on its genome size, the presence of two copies of 18S rRNA-like sequences, and period of infection cycle, cotonvirus is the most similar to the tupanviruses among the icosahedral mimiviruses. Interestingly, the cotonvirus utilizes Golgi apparatus-like vesicles for virion factory (VF) formation. Overall, we showed that cotonvirus is a novel lineage of the subfamily Megavirinae . Our findings support the diversity of icosahedral mimiviruses and provide mechanistic insights into the replication, VF formation, and evolution of the subfamily Mimivirinae . Importance We have isolated a new virus of an independent lineage belonging to the family Mimiviridae , subfamily Megavirinae , from the fresh water of a canal in Japan, named cotonvirus. In a proteomic tree, this new nucleocytoplasmic large DNA virus (NCLDV) is phylogenetically placed at the root of three lineages of the subfamily Megavirinae —lineages A (mimivirus), B (moumouvirus), and C (megavirus). Multiple genomic and phenotypic features of cotonvirus are similar to those of tupanviruses, rather than the A, B, or C lineages, and other genomic features and host specificity of cotonvirus are similar to those of the latter than the former. These results suggest that cotonvirus is a unique virus, with chimeric features of existing viruses of Megavirinae and uses Golgi apparatus-like vesicles of the host cells for virion factory (VF) formation. Thus, cotonvirus can provide novel insights into the evolution of mimiviruses and the underlying mechanisms of VF formation.

Quantitative Assessment of Nucleocytoplasmic Large DNA Virus and Host Interactions Predicted by Co-occurrence Analyses

ABSTRACT Nucleocytoplasmic large DNA viruses (NCLDVs) are highly diverse and abundant in marine environments. However, the knowledge of their hosts is limited because only a few NCLDVs have been isolated so far. Taking advantage of the recent large-scale marine metagenomics census, in silico host prediction approaches are expected to fill the gap and further expand our knowledge of virus-host relationships for unknown NCLDVs. In this study, we built co-occurrence networks of NCLDVs and eukaryotic taxa to predict virus-host interactions using Tara Oceans sequencing data. Using the positive likelihood ratio to assess the performance of host prediction for NCLDVs, we benchmarked several co-occurrence approaches and demonstrated an increase in the odds ratio of predicting true positive relationships 4-fold compared to random host predictions. To further refine host predictions from high-dimensional co-occurrence networks, we developed a phylogeny-informed filtering method, Taxon Interaction Mapper, and showed it further improved the prediction performance by 12-fold. Finally, we inferred virophage-NCLDV networks to corroborate that co-occurrence approaches are effective for predicting interacting partners of NCLDVs in marine environments. IMPORTANCE NCLDVs can infect a wide range of eukaryotes, although their life cycle is less dependent on hosts compared to other viruses. However, our understanding of NCLDV-host systems is highly limited because few of these viruses have been isolated so far. Co-occurrence information has been assumed to be useful to predict virus-host interactions. In this study, we quantitatively show the effectiveness of co-occurrence inference for NCLDV host prediction. We also improve the prediction performance with a phylogeny-guided method, which leads to a concise list of candidate host lineages for three NCLDV families. Our results underpin the usage of co-occurrence approaches for the metagenomic exploration of the ecology of this diverse group of viruses.

Viral ecogenomics across the Porifera

Background Viruses directly affect the most important biological processes in the ocean via their regulation of prokaryotic and eukaryotic populations. Marine sponges form stable symbiotic partnerships with a wide diversity of microorganisms and this high symbiont complexity makes them an ideal model for studying viral ecology. Here, we used morphological and molecular approaches to illuminate the diversity and function of viruses inhabiting nine sponge species from the Great Barrier Reef and seven from the Red Sea. Results Viromic sequencing revealed host-specific and site-specific patterns in the viral assemblages, with all sponge species dominated by the bacteriophage order Caudovirales but also containing variable representation from the nucleocytoplasmic large DNA virus families Mimiviridae, Marseilleviridae, Phycodnaviridae, Ascoviridae, Iridoviridae, Asfarviridae and Poxviridae. Whilst core viral functions related to replication, infection and structure were largely consistent across the sponge viromes, functional profiles varied significantly between species and sites largely due to differential representation of putative auxiliary metabolic genes (AMGs) and accessory genes, including those associated with herbicide resistance, heavy metal resistance and nylon degradation. Furthermore, putative AMGs varied with the composition and abundance of the sponge-associated microbiome. For instance, genes associated with antimicrobial activity were enriched in low microbial abundance sponges, genes associated with nitrogen metabolism were enriched in high microbial abundance sponges and genes related to cellulose biosynthesis were enriched in species that host photosynthetic symbionts. Conclusions Our results highlight the diverse functional roles that viruses can play in marine sponges and are consistent with our current understanding of sponge ecology. Differential representation of putative viral AMGs and accessory genes across sponge species illustrate the diverse suite of beneficial roles viruses can play in the functional ecology of these complex reef holobionts.

A55 Molecular systematics of sturgeon nucleocytoplasmic large DNA viruses

Namao virus (NV) is a sturgeon nucleocytoplasmic large DNA virus (sNCLDV) that can cause a lethal disease of the integumentary system in lake sturgeon Acipenser fulvescens. As a group, the sNCLDV have not been assigned to any currently recognized taxonomic family of viruses. In this study, a dataset of NV DNA sequences was generated and assembled as two non-overlapping contigs of 306 and 448 base pairs (bp) and then used to conduct a comprehensive systematics analysis using Bayesian phylogenetic inference for NV, other sNCLDV, and representative members of six families of the NCLDV superfamily. The phylogeny of NV was reconstructed using protein homologues encoded by nine nucleocytoplasmic virus orthologous genes (NCVOGs): NCVOG0022—mcp, NCVOG0038—DNA polymerase B elongation subunit, NCVOG0076—VV A18-type helicase, NCVOG0249—VV A32-type ATPase, NCVOG0262—AL2 VLTF3-like transcription factor, NCVOG0271—RNA polymerase II subunit II, NCVOG0274—RNA polymerase II subunit I, NCVOG0276—ribonucleotide reductase small subunit, and NCVOG1117—mRNA capping enzyme. The accuracy of our phylogenetic method was evaluated using a combination of Bayesian statistical analysis and congruence analysis. Stable tree topologies were obtained with datasets differing in target molecule(s), sequence length, and taxa. Congruent topologies were obtained in phylogenies constructed using individual protein datasets and when four proteins were used in a concatenated approach. The major capsid protein phylogeny indicated that ten representative sNCLDV form a monophyletic group comprised of four lineages within a polyphyletic Mimi-Phycodnaviridae group of taxa. Overall, the analyses revealed that Namao virus is a member of the Mimiviridae family with strong and consistent support for a clade containing NV and CroV as sister taxa.

Analyses of the Kroon Virus Major Capsid Gene and Its Transcript Highlight a Distinct Pattern of Gene Evolution and Splicing among Mimiviruses

ABSTRACT The inclusion of Mimiviridae members in the putative monophyletic nucleocytoplasmic large DNA virus (NCLDV) group is based on genomic and phylogenomic patterns. This shows that, along with other viral families, they share a set of genes known as core or “hallmark genes,” including the gene for the major capsid protein (MCP). Although previous studies have suggested that the maturation of mimivirus MCP transcripts is dependent on splicing, there is little information about the processing of this transcript in other mimivirus isolates. Here we report the characterization of a new mimivirus isolate, called Kroon virus (KV) mimivirus. Analysis of the structure, synteny, and phylogenetic relationships of the MCP genes in many mimivirus isolates revealed a remarkable variation at position and types of intronic and exonic regions, even for mimiviruses belonging to the same lineage. In addition, sequencing of KV and Acanthamoeba polyphaga mimivirus (APMV) MCP transcripts has shown that inside the family, even related giant viruses may present different ways to process the MCP mRNA. These results contribute to the understanding of the genetic organization and evolution of the MCP gene in mimiviruses. IMPORTANCE Mimivirus isolates have been obtained by prospecting studies since 2003. Based on genomic and phylogenomic studies of conserved genes, these viruses have been clustered together with members of six other viral families. Although the major capsid protein (MCP) gene is an important member of the so-called “hallmark genes,” there is little information about the processing and structure of this gene in many mimivirus isolates. In this work, we have analyzed the structure, synteny, and phylogenetic relationships of the MCP genes in many mimivirus isolates; these genes showed remarkable variation at position and types of intronic and exonic regions, even for mimiviruses belonging to the same lineage. These results contribute to the understanding of the genetic organization and evolution of the MCP gene in mimiviruses.