Using signature genes as tools to assess environmental viral ecology and diversity

Viruses (including bacteriophages) are the most abundant biological entities on the planet. As such, they are thought to have a major impact on all aspects of microbial community structure and function. Despite this critical role in ecosystem processes, the study of virus/phage diversity has lagged far behind parallel studies of the Bacterial and Eukaryotic kingdoms, largely due to the absence of any ‘universal phylogenetic marker’. Here we review the development and use of signature genes to investigate viral diversity, as a viable strategy for datasets of specific virus groups. Genes that have been used include those encoding structural proteins, portal protein, major capsid protein and tail sheath protein, auxiliary metabolism genes such as psbA, psbB and phoH, and several polymerase genes. These marker genes have been used in combination with PCR-based fingerprinting and/or sequencing strategies to investigate spatial, temporal and seasonal variation and diversity in a wide range of habitats.http://aem.asm.orghb201

A roadmap for genome-based phage taxonomy

Bacteriophage (phage) taxonomy has been in flux since its inception over four decades ago. Genome sequencing has put pressure on the classification system and recent years have seen significant changes to phage taxonomy. Here, we reflect on the state of phage taxonomy and provide a roadmap for the future, including the abolition of the order Caudovirales and the families Myoviridae, Podoviridae, and Siphoviridae. Furthermore, we specify guidelines for the demarcation of species, genus, subfamily and family-level ranks of tailed phage taxonomy

RNA-viromics reveals diverse communities of soil RNA viruses with the potential to affect grassland ecosystems across multiple trophic levels

The distribution and diversity of RNA viruses in soil ecosystems are largely unknown, despite their significant impact on public health, ecosystem functions, and food security. Here, we characterise soil RNA viral communities along an altitudinal productivity gradient of peat, managed grassland and coastal soils. We identified 3462 viral contigs in RNA viromes from purified virus-like-particles in five soil-types and assessed their spatial distribution, phylogenetic diversity and potential host ranges. Soil types exhibited minimal similarity in viral community composition, but with >10-fold more viral contigs shared between managed grassland soils when compared with peat or coastal soils. Phylogenetic analyses predicted soil RNA viral communities are formed from viruses of bacteria, plants, fungi, vertebrates and invertebrates, with only 12% of viral contigs belonging to the bacteria-infecting Leviviricetes class. 11% of viral contigs were found to be most closely related to members of the Ourmiavirus genus, suggesting that members of this clade of plant viruses may be far more widely distributed and diverse than previously thought. These results contrast with soil DNA viromes which are typically dominated by bacteriophages. RNA viral communities, therefore, have the potential to exert influence on inter-kingdom interactions across terrestrial biomes

Diversity and ecology of viruses in hyperarid desert soils

In recent years, remarkable progress has been made in the field of virus environmental ecology. In marine ecosystems for example, viruses are now thought to play pivotal roles in the biogeochemical cycling of nutrients and to be mediators of microbial evolution through horizontal gene transfer. In soils, the diversity and ecology of viruses is poorly understood, but evidence supports the view that these differ substantially from aquatic systems. Desert biomes cover ~33% of global land masses, yet the diversity and roles of viruses in these dominant ecosystems remain poorly understood. There is evidence that hot hyperarid desert soils are characterised by high levels of bacterial lysogens and low extracellular virus counts. In contrast, cold desert soils contain high extracellular virus titres. We suggest that the prevalence of microbial biofilms in hyperarid soils, combined with extreme thermal regimes, constitute strong selection pressures on both temperate and virulent viruses. Many desert soil virus sequences show low identity values to virus genomes in public databases, suggesting the existence of distinct and as yet uncharacterised soil phylogenetic lineages (e.g. cyanophages). We strongly advocate for amplification-free metavirome analyses while encouraging the classical isolation of phages from dominant and culturable microbial isolates in order to populate sequence databases. This review provides an overview of recent advances in the study of viruses in hyperarid soils, the factors that contribute to viral abundance and diversity in hot and cold deserts and suggests technical recommendations for future studies.The South African National Research Foundation and the University of Pretoria Genomics Research Institute. EMA is funded by the Claude Leon Foundation Postdoctoral Fellowship program.http://aem.asm.orghb2016Genetic

Phage annotation guide: Guidelines for assembly and high-quality annotation

All sequencing projects of bacteriophages (phages) should seek to report an accurate and comprehensive annotation of their genomes. This article defines 14 questions for those new to phage genomics that should be addressed before submitting a genome sequence to the International Nucleotide Sequence Database Collaboration or writing a publication

Viromic Analysis of Wastewater Input to a River Catchment Reveals a Diverse Assemblage of RNA Viruses

Enteric viruses cause gastrointestinal illness and are commonly transmitted through the fecal-oral route. When wastewater is released into river systems, these viruses can contaminate the environment. Our results show that we can use viromics to find the range of potentially pathogenic viruses that are present in the environment and identify prevalent genotypes. The ultimate goal is to trace the fate of these pathogenic viruses from origin to the point where they are a threat to human health, informing reference-based detection methods and water quality management.Detection of viruses in the environment is heavily dependent on PCR-based approaches that require reference sequences for primer design. While this strategy can accurately detect known viruses, it will not find novel genotypes or emerging and invasive viral species. In this study, we investigated the use of viromics, i.e., high-throughput sequencing of the biosphere’s viral fraction, to detect human-/animal-pathogenic RNA viruses in the Conwy river catchment area in Wales, United Kingdom. Using a combination of filtering and nuclease treatment, we extracted the viral fraction from wastewater and estuarine river water and sediment, followed by high-throughput RNA sequencing (RNA-Seq) analysis on the Illumina HiSeq platform, for the discovery of RNA virus genomes. We found a higher richness of RNA viruses in wastewater samples than in river water and sediment, and we assembled a complete norovirus genotype GI.2 genome from wastewater effluent, which was not contemporaneously detected by conventional reverse transcription-quantitative PCR (qRT-PCR). The simultaneous presence of diverse rotavirus signatures in wastewater indicated the potential for zoonotic infections in the area and suggested runoff from pig farms as a possible origin of these viruses. Our results show that viromics can be an important tool in the discovery of pathogenic viruses in the environment and can be used to inform and optimize reference-based detection methods provided appropriate and rigorous controls are included

High-level diversity of tailed phages, eukaryote-associated viruses, and virophage-like elements in the metaviromes of Antarctic soils

The metaviromes of two distinct Antarctic hyperarid desert soil communities have been characterized. Hypolithic communities, cyanobacterium-dominated assemblages situated on the ventral surfaces of quartz pebbles embedded in the desert pavement, showed higher virus diversity than surface soils, which correlated with previous bacterial community studies. Prokaryotic viruses (i.e., phages) represented the largest viral component (particularly Mycobacterium phages) in both habitats, with an identical hierarchical sequence abundance of families of tailed phages (Siphoviridae>Myoviridae>Podoviridae). No archaeal viruses were found. Unexpectedly, cyanophages were poorly represented in both metaviromes and were phylogenetically distant from currently characterized cyanophages. Putative phage genomes were assembled and showed a high level of unaffiliated genes, mostly from hypolithic viruses. Moreover, unusual gene arrangements in which eukaryotic and prokaryotic virus-derived genes were found within identical genome segments were observed. Phycodnaviridae and Mimiviridae viruses were the second-mostabundant taxa and more numerous within open soil. Novel virophage-like sequences (within the Sputnik clade) were identified. These findings highlight high-level virus diversity and novel species discovery potential within Antarctic hyperarid soils and may serve as a starting point for future studies targeting specific viral groups.IS