ABSTRACTIn fungi, viral infections frequently remain cryptic causing little or no phenotypic changes. It can indicate either a long history of coevolution or a strong immune system of the host. Some fungi are outstandingly ubiquitous and can be recovered from a great diversity of habitats. However, the role of viral infection in the emergence of environmental opportunistic species is not known. The genus of filamentous and mycoparasitic fungiTrichoderma(Hypocreales, Ascomycota) consists of more than 400 species, which mainly occur on dead wood, other fungi, or as endo- and epiphytes. However, some species are environmental opportunists because they are cosmopolitan, can establish in a diversity of habitats, and can also become pests on mushroom farms and infect immunocompromised humans. In this study, we investigated the library of 163Trichodermastrains isolated from grassland soils in Inner Mongolia, China, and found only four strains with signs of the mycoviral nucleic acids, including a strain ofT. barbatuminfected with a novel strain of thePolymycoviridaeand named and characterized here as Trichoderma barbatum polymycovirus 1 (TbPMV1). Phylogenetic analysis suggested that TbPMV1 was evolutionarily distinct from thePolymycoviridaeisolated either from Eurotialean fungi or from the orderMagnaportales. Although thePolymycoviridaeviruses were also known from HypocrealeanBeauveria bassiana, the phylogeny of TbPMV1 did not reflect the phylogeny of the host. Our analysis lays the groundwork for further in-depth characterization of TbPMV1 and the role of mycoviruses in the emergence of environmental opportunism inTrichoderma.IMPORTANCEAlthough viruses infect all organisms, our knowledge of some groups of eukaryotes remains limited. For instance, the diversity of viruses infecting fungi—mycoviruses—is largely unknown. However, the knowledge of viruses associated with industrially relevant and plant-beneficial fungi, such asTrichodermaspp. (Hypocreales, Ascomycota), may shed light on the stability of their phenotypes and the expression of beneficial traits. In this study, we screened the library of soilborneTrichodermastrains because these isolates may be developed into bioeffectors for plant protection and sustainable agriculture. Notably, the diversity of endophytic viruses in soilTrichodermawas outstandingly low. Only 2% of 163 strains contained traces of dsRNA viruses, including the new Trichoderma barbatum polymycovirus 1 (TbPMV1) characterized in this study. TbPMV1 is the first mycovirus found inTrichoderma. Our results indicate that the limited data prevent the in-depth study of the evolutionary relationship between soilborne fungi and is worth further investigation.