Molecular characterization of a novel mycovirus in the cultivated mushroom, Lentinula edodes

A cDNA library was constructed from a dsRNA purified from the fruiting body of L. edodes. The virus was tentatively named L. edodes mycovirus HKB (LeV). Based on the deduced RNA-dependent RNA polymerase (RdRp) sequence, phylogenetic analysis of LeV was conducted. Because no virion particles associated with the dsRNA were observed by electron microscopic observation, atomic force microscopy (AFM) observation was chosen for achieving molecular imaging of the virus.The 11,282-bp genome of LeV was obtained. The genome encoded two open reading frames (ORFs). ORF1 coded for a hypothetical protein and ORF2 for a putative RdRp, respectively. In addition, a region coding for a NUDIX domain was present in ORF1. There was a 62-bp intergenic region between ORF1 and RdRp. Similarity with coat protein of mycoviruses was not found within the whole sequence. Based on phylogenetic analysis of the putative RdRp sequence, LeV grouped into a clade with dsRNA found in the basidiomycetes Phlebiopsis gigantea and Helicobasidium mompa. The clade was placed apart from the Totiviridae and Chrysoviridae families. As suggested from the genome sequence, AFM revealed that the structure of LeV was linear unencapsidated dsRNA.The results suggest that LeV represents a novel family of mycoviruses, found thus far only among the basidiomycetes.In the 1970s, viruses that infect the cultivated mushroom Lentinula edodes, or shiitake, were extensively studied in Japan [1-3], and three morphologically distinct viruses were detected by electron microscopy [1,3]. However, unlike La France disease of the white button mushroom [4,5], mycoviruses have not been associated with shiitake diseases because these mycoviruses have commonly been found in healthy fruiting bodies [1,3]. In the USA, dsRNAs have also been observed in shiitake strains, but these appeared to be latent [6]. In the 1970s, shiitake cultivation was performed by inoculating mycelium spawn on oak logs; however, this labor-intensive method was grad