A novel dsRNA virus was identified from the arboreal ant Camponotus yamaokai. The complete nucleotide sequence analysis of the virus revealed that the virus consisted of 5704 bp with two ORFs. ORF1 (3084 nt) encoded a putative capsid protein. ORF2 (1977 nt) encoded a viral RNA-dependent RNA polymerase (RdRp). ORF2 could be translated as a fusion with the ORF1 product by a - 1 frameshift in the overlapping ORF1. Phylogenetic analyses based on the RdRp revealed that the virus from C. yamaokai was most likely a novel totivirus, but it was not closely related to the previously known totiviruses in arthropods. Transmission electron microscopy revealed isometric virus particles of ~30 nm diameter in the cytoplasm, which was consistent with the characteristics of the family Totiviridae. The virus was detected by reverse transcription-PCR in all caste members and developmental stages of ants, including eggs, larvae, pupae, adult workers, alates (male and female) and queens. To our knowledge, this is the first report of a member of the family Totiviridae in a hymenopteran; the virus was designated Camponotus yamaokai virus.