Distribution of marine birnavirus in cultured marine fish species from Kagawa Prefecture, Japan T Isshiki 1 , T Nagano 1 , K Kanehira 2 and S Suzuki 2 1 Kagawa Prefectural Fisheries Experiment Station, Kagawa, Japan 2 Centre for Marine Environmental Studies, Ehime University, Ehime, Japan Abstract To determine the distribution of marine birnavirus (MABV) in cultured populations of different marine fish species, 1291 pooled tissue samples from 2672 fish belonging to 22 species and one hybrid were collected from Kagawa Prefecture, Japan, during 1999–2001. Using cell-culture MABV was isolated from three species: yellowtail, Seriola quinqueradiata Temminck & Schlegel (positive number/sample number, 10/419), amberjack, S. dumerili (Risso) (4/72), and Japanese flounder, Paralichthys olivaceus (Temminck & Schlegel) (41/481). Using PCR on MABV-negative samples, the MABV genome was detected in the same three species [yellowtail (9/409), amberjack (4/68) and Japanese flounder (93/440)] and two additional species, spotted hali- but, Verasper variegatus (Temminck & Schlegel) (5/11), and goldstriped amberjack, S. lalandi Valenciennes (1/5). These MABV-positive species can be taxonomically divided into two groups: the genus Seriola and flatfish. In Japanese flounder, MABV was detected during all seasons, and the infection rate was correlated with water temperature. Aquaculture sites with MABV-positive fish were evenly distributed over the surveyed area, suggesting that MABV is widely distributed at aquaculture sites in Kagawa Prefecture. The nucleotide sequence at the variable region, the VP2/NS junction, revealed that the 39th base mutation occurs host- specifically for flatfish. Flatfish are suspected to be the main reservoir of MABV and might be respon- sible for establishing the infection cycle in aqua- culture environments. Keywords: birnavirus, distribution, natural hosts, flounder, yellowtail, mutation. Introduction Marine birnavirus (MABV) is a member of the family Birnaviridae, genus Aquabirnavirus, which comprises species that have a two-segmented dou- ble-stranded RNA genome within an icosahedral non-enveloped single-shelled capsid of about 60 nm in diameter. Aquabirnaviruses occur in a variety of aquatic animals in freshwater, brackish and seawater environments (Leong, Brown, Dobos, Kibenge, Ludert, Muller, Mundt & Nicholson 2000). Infectious pancreatic necrosis virus (IPNV) is a well-known aquabirnavirus associated with an acute and contagious disease in salmonids (Wolf 1988). MABV includes yellowtail ascites virus (YTAV), which is the causative agent of yellowtail ascites (Sorimachi & Hara 1985), and other aquabirnaviruses isolated from various marine fish and shellfish (Kusuda, Nishi, Hosono & Suzuki 1993; Suzuki & Nojima 1999). MABV is very close to IPNV, however, MABV strains can be distin- guished from IPNV strains by the genogrouping based on the nucleotide sequence in the VP2/NS region, which lies between the major capsid protein VP2 and viral protease NS in the large segment A (Hosono, Suzuki & Kusuda 1996). MABV can also be distinguished based on the RNA polymerase gene (Zhang & Suzuki 2003). Marine birnavirus has been isolated only from epizootics in yellowtail, Seriola quinqueradiata Temminck & Schlegel, where it causes ascites (Sorimachi & Hara 1985). However MABV is thought to cause disease in a variety of marine fish including Japanese flounder, Paralichthys olivaceus Journal of Fish Diseases 2004, 27, 89–98 Correspondence Dr Tadashi Isshiki, Faculty of Bioresources, Mie University, 1515 Kamihama, Tsu, Mie 514-8507, Japan (e-mail: isshiki@bio.mie-u.ac.jp) 89 Ó 2004 Blackwell Publishing Ltd