Identification of the viral determinant of hypovirulence and host range in Sclerotiniaceae of a genomovirus reconstructed from the plant metagenome

Uncharacterized viral genomes that encode circular replication-associated proteins of single-stranded DNA viruses have been discovered by metagenomics/metatranscriptomics approaches. Some of these novel viruses are classified under the newly formed Genomoviridae family. Here, we determine the host range of a novel genomovirus, SlaGemV-1, through the transfection of Sclerotinia sclerotiorum with infectious clones. Inoculating with the rescued virions, we further transfected Botrytis cinerea and Monilinia fruticola , two economically important members of family Sclerotiniaceae, and Fusarium oxysporum . SlaGemV-1 causes hypovirulence in S. sclerotiorum, B. cinerea , and M. fruticola . SlaGemV-1 also replicates in Spodoptera frugiperda insect cells, but not in Caenorhabditis elegans nor plants. By expressing viral-encoded genes separately through site-specific integration, the replication protein alone was sufficient to cause debilitation. Our study is the first to demonstrate the reconstruction of a metagenomically discovered genomovirus without known hosts with the potential of inducing hypovirulence, and the infectious clone allows for studying mechanisms of genomovirus-host interactions that are conserved across genera. Importance Little is known about the exact host range of widespread genomoviruses. The genome of soybean leaf-associated gemygorvirus-1 (SlaGemV-1) was originally assembled from a metagenomic/metatranscriptomic study without known hosts. Here, we rescued SlaGemV-1 and found that it could infect three important plant pathogenic fungi and Fall armyworm (S. frugiperda , Sf9) insect cells, but not a model nematode, C. elegans , or model plant species. Most importantly, SlaGemV-1 shows promise for inducing hypovirulence of the tested fungal species under family Sclerotiniaceae, including Sclerotinia sclerotiorum , Botrytis cinerea , Monilinia fruticola . The viral determinant of hypovirulence was further identified as replication initiation protein. As a proof of concept, we demonstrate that viromes discovered in plant metagenome can be a valuable genetic resource when novel viruses are rescued and characterized for their host range.