Exploring the mycovirome: novel and diverse mycoviruses inBotrytis cinerea

Botrytis cinereais a necrotrophic fungal pathogen that causes significant economic losses to many crops, including vegetables, fruits, and ornamental plants. The management ofB. cinereais difficult due to a rise in fungicide resistance. Harnessing mycoviruses that cause reduced virulence (hypovirulence) inB. cinereais a promising alternative. Over 100 mycoviruses have been identified inBotrytisspp. to date, including several hypovirulence-inducing mycoviruses. This research aimed to further explore, for the first time in Canada, the mycovirome ofB. cinereaand identify potential hypovirulence-inducing mycoviruses. Isolates ofB. cinereawere collected from fruits and vegetables in the province of Quebec. Fitness and pathogenicity criteria, including sclerotia production, colony morphotype, and lesion size were evaluated. A double-stranded RNA (dsRNA) extraction protocol tailored to the detection of mycoviruses was used to sequence dsRNA from 45 isolates with low fitness/pathogenicity, and an in-house bioinformatics workflow was used to profile the mycovirome. Mycoviruses were identified in 44/45 isolates. Most of these had positive single-stranded RNA or dsRNA genomes, and a small number had negative single-stranded RNA, single-stranded DNA, or reverse transcriptase RNA genomes. Following deep analysis of RNA-dependent RNA polymerase and replication initiation proteins, a total of 62 unique contigs were identified belonging to new strains of mycovirus species. Furthermore, four putative novel mycovirus species belonging toEndornaviridae,Botybirnaviridae,Peribunyaviridae, andBunyaviralestaxa were identified. Several mycovirus species positively and/or negatively co-occurred withB. cinereaisolates collected from strawberry or raspberry. This study revealed a high degree of diversity in the mycovirome ofB. cinerea.Species accumulation curve analysis indicated that, with the number of isolates characterized, we were unable to capture the full extent of expected diversity. Nevertheless, we identified potential hypovirulence-inducing mycoviruses, including Botrytis cinerea mitovirus 1, Botrytis cinerea hypovirus 1, and Botrytis porri botybirnavirus 1. Some of these novel mycoviruses belonged to taxa known to produce viral particles, which can be an interesting feature for their use as biocontrol agents (BCA).