Detection and sequence analysis of two novel co-infecting double-strand RNA mycoviruses in Ustilaginoidea virens

Four novel double-stranded RNA molecules, named dsRNA 1 (5124 bp), dsRNA 2(1711 bp), dsRNA 3 (1423 bp) and dsRNA 4 (855 bp), were detected in strain HNHS-1 of Ustilaginoidea virens, the causal agent of rice false smut disease. Sequence analysis showed that the dsRNA1 contains two overlapping open reading frames (ORF) potentially encoding proteins with modest levels of sequence similarity to the coat protein (CP) and putative RNA-dependent RNA polymerase (RdRp), respectively, of viruses of the family Totiviridae. The deduced gene product of the ORF encoded by dsRNA2 is homologous to putative RdRp of viruses in the family Partitiviridae; the ORF encoded by dsRNA3 shares some similarity to a hypothetical protein with unknown function. It is noteworthy that the dsRNA4 lacked integrated ORFs. Isomeric viral particles of about 40 nm in diameter were observed by transmission electron microscopy in a mycelium tissue preparation of strain HNHS-1-R1, a single-spore subculture of strain HNHS-1 containing only the dsRNA1 segment. Phylogenetic analysis and examination of the organization of the two putative RdRp sequences both indicated that there are at least two novel virus species present in strain HNHS-1. We named the two novel viruses Ustilaginoidea virens RNA virus 2 and Ustilaginoidea virens partitivirus 4, respectively.