Molecular investigations of ericoid and orchid mycorrhizal fungi reveal new mycoviral taxa

Fungal viruses, or mycoviruses, are widely distributed in all major groups of fungi. Mycovirus infection may have diverse effects on the fungal host phenotype, including the decrease of growth, sporulation, virulence and secondary metabolite production, as well as the improvement of abiotic stress tolerance or pathogenicity. In the case of fungal endophytes, mycoviruses have been found to positively influence also the host plant phenotype, for example conferring heat tolerance. Indeed, a better understanding of these mycovirus-fungus-plant tripartite interactions and of their ecological roles will enhance the potential use of mycoviruses in improving plant health. Virology has so far concentrated on viruses infecting important plant pathogens, while little is known on the virus communities associated with saprotrophic or mutualistic fungi, despite their central role in the maintenance of soil ecosystem services and in plant stress tolerance. Among mutualistic fungi, ectomycorrhizal, arbuscular and orchid mycorrhizal (ORM) fungi have been previously shown to harbor mycoviruses, while ericoid mycorrhizal (ERM) fungi have never been investigated so far. We have investigated a collection of ORM and ERM fungi, isolated from different host plants in different geographic sites and grown in axenic cultures, by high-throughput sequencing of their transcriptomes (RNAseq) to include the genomic sequences of RNA-viruses. The analysis of fungal metaviromes focused on the identification, by similarity searches, of RNA dependent RNA polymerase (RdRP), the only universal gene among RNA viruses also used for phylogenomic reconstruction. Although RdRPs are characterized by extensive sequence divergence, they contain three conserved motifs that are required for polymerase activity, forming the so called "palm domain". Two distinct narnavirus-like sequences and one ourmia-like virus have been found respectively in 3 and 2 Oidiodendron maius strains out of 37 ERM strains. We suggest that the two contigs related to narnavirus represent a bisegmented virus, that splits the RdRP palm domain in two distinct proteins, encoded by each of the two viral genomic segments, therefore representing a genomic organization never observed before. We found that 9 out of 12 tested ORM isolates harbored mycoviruses, including a new mitovirus, a new barnavirus and two new endornaviruses, related to those already found but different enough to be considered new species. For the first time a minus stranded RNA virus was detected in ORM fungi, belonging to the Bunyavirales order, and having an unprecedented genomic organization for this orders, being tri-cistronic and carrying the putative RdRP and putative nucleocapsid in ambisense orientation on the same genomic RNA. In addition, a number of ORM isolates harbored a group of ambisense bicistronic viruses that may represent new lineages, as they could not be reliably assigned to any described evolutionary trajectories. This work represents a first insight into the virome of ERM fungi and adds new data for the virome of ORM fungi. A functional characterization of these viral taxa will provide new information on their possible effects on the fungal host phenotype, and on their role in the multitrophic interaction with the host plant.