A collection of Trichoderma isolates from natural environments in Sardinia, a biodiversity hotspot, reveals a complex virome that includes negative-stranded mycoviruses with unprecedented genome organizations

The Trichoderma genus includes soil-inhabiting fungi that provide important ecological services in their interaction with plants and other fungi. They are exploited for biocontrol. A collection of Trichoderma isolates from the Sardinia island (a biodiversity hotspot) had been previously characterized. Here we started a characterization of the viral components associated to 113 selected Trichoderma isolates, representatives of the collection. We carried out NGS sequencing of ribosome depleted total RNA following a bioinformatic pipeline that detects virus RNA-dependent RNA polymerases (RdRP) and other conserved virus protein sequences. This pipeline detected 17 viral RdRPs. Two of them correspond to viruses already detected in other regions of the world. The remaining 15 represent isolates of new virus species: surprisingly, eight of them are from new negative stranded RNA viruses, which for the first time are reported in the genus Trichoderma. Among them is a cogu-like virus, very closely related to plant-infecting viruses. Regarding the positive strand viruses, it is noticeable the presence of an ormycovirus belonging to a recently characterized group of bi-segmented ssRNA genome viruses with still uncertain phylogenetic assignment. Finally, for the first time we report a bipartite mononegavirales-infecting fungi: the proteins encoded by the second genomic RNA were used to re-evaluate a number of viruses in the Penicillimonavirus and Plasmopamonavirus genera, here shown to be bipartite and to encode a conserved polypeptide having structural conservation with the nucleocapsid (NC) domain of members of the Rabhdoviridae. IMPORTANCE Trichoderma is a genus of fungi of great biotechnological impact in multiple industrial fields. The possibility to investigate a diverse collection of Trichoderma isolates allowed us to characterize both double-stranded and single-stranded virus genomes belonging to three of the major phyla that constitute the RNA viral kingdom, thus further increasing the taxa of viruses infecting this genus. To our knowledge here we report for the first time negative-stranded RNA viruses infecting Trichoderma spp. and through in silico structural analysis a new conserved domain of nucleocapsids common among some mymonavirids. Obtaining such a library of mycoviruses could be the basis for further development of targeted virus-induced gene silencing or gene editing (VIGS/VIGE) tools; in addition, the many biotechnological applications of this fungus, will require to assess the qualitative (commercial) stability of strains, linked to positive or negative effects caused by mycovirus infections.