[From boots on the ground to nucleotides in the sequencer: a century of advances in the study of the plant virus ecology]. / De la paire de bottes à la paire de bases : de l'intérêt d'étudier l'écologie des viromes de plantes.

Plant virus ecology began to be explored at the end of the 19th century. Since then, major advances have revealed complex virus-host-vector interactions in a variety of environments. These advances have been accelerated by development of new technologies for virus detection and characterization, the latest of which being high-throughput sequencing (HTS). HTS technologies have proved to be effective for non-targeted characterization of all or nearly all viruses present in a sample without requiring prior information about virus identity, as would be needed for virus-targeted tests. Phytoviromic studies have thus made important advances, including characterization of the complex interactions between phytovirus dynamics and the structure of multi-species host communities, and documentation of the effects of anthropogenic ecosystem simplification on plant virus emergence and diversity. However, such studies must overcome challenges at every stage, from plant sampling to bioinformatics analysis. This review summarizes major advances in plant virus ecology, in association with technological developments, and presents key considerations for use of HTS in the study of the ecology of phytovirus communities.

Phytovirome Analysis of Wild Plant Populations: Comparison of Double-Stranded RNA and Virion-Associated Nucleic Acid Metagenomic Approaches.

Metagenomic studies have indicated that the diversity of plant viruses was until recently far underestimated. As important components of ecosystems, there is a need to explore the diversity and richness of the viruses associated with plant populations and to understand the drivers shaping their diversity in space and time. Two viral sequence enrichment approaches, double-stranded RNA (dsRNA) and virion-associated nucleic acids (VANA), have been used and compared here for the description of the virome of complex plant pools representative of the most prevalent plant species in unmanaged and cultivated ecosystems. A novel bioinformatics strategy was used to assess viral richness not only at the family level but also by determining operational taxonomic units (OTU) following the clustering of conserved viral domains. A large viral diversity dominated by novel dsRNA viruses was detected in all sites, while a large between-site variability limited the ability to draw a clear conclusion on the impact of cultivation. A trend for a higher diversity of dsRNA viruses was nevertheless detected in unmanaged sites (118 versus 77 unique OTUs). The dsRNA-based approach consistently revealed a broader and more comprehensive diversity for RNA viruses than the VANA approach, whatever the assessment criterion. In addition, dissimilarity analyses indicated both approaches to be largely reproducible but not necessarily convergent. These findings illustrate features of phytoviromes in various ecosystems and a novel strategy for precise virus richness estimation. These results allow us to reason methodological choices in phytovirome studies and likely in other virome studies where RNA viruses are the focal taxa.IMPORTANCE There are today significant knowledge gaps on phytovirus populations and on the drivers impacting them but also on the comparative performance-methodological approaches for their study. We used and compared two viral sequence enrichment approaches, double-stranded RNAs (dsRNA) and virion-associated nucleic acids (VANA), for phytovirome description in complex pools representative of the most prevalent plant species in unmanaged and cultivated ecosystems. Viral richness was assessed by determining operational taxonomic units (OTU) following the clustering of conserved viral domains. There is some limited evidence of an impact of cultivation on viral populations. These results provide data allowing us to reason the methodological choices in virome studies. For researchers primarily interested in RNA viruses, the dsRNA approach is recommended because it consistently provided a more comprehensive description of the analyzed phytoviromes, but it understandably underrepresented DNA viruses and bacteriophages.

Harnessing plant viruses in the metagenomics era: from the development of infectious clones to applications.

Recent metagenomic studies which focused on virus characterization in the entire plant environment have revealed a remarkable viral diversity in plants. The exponential discovery of viruses also requires the concomitant implementation of high-throughput methods to perform their functional characterization. Despite several limitations, the development of viral infectious clones remains a method of choice to understand virus biology, their role in the phytobiome, and plant resilience. Here, we review the latest approaches for efficient characterization of plant viruses and technical advances built on high-throughput sequencing and synthetic biology to streamline assembly of viral infectious clones. We then discuss the applications of plant viral vectors in fundamental and applied plant research as well as their technical and regulatory limitations, and we propose strategies for their safer field applications.