Characterization of red-spotted grouper nervous necrosis virus isolated from ovarian fluids of asymptomatic wild Asian seabass, Lates calcarifer

Abstract

Viral nervous necrosis (VNN) caused by nervous necrosis virus is a serious disease affecting a wide range of marine, brackishwater and freshwater fishes. The disease causes acute mortalities in larval and early juvenile stages while adult fishes are asymptomatic and remain as carriers of the virus. In the present study, nervous necrosis virus was isolated from ovarian fluids of asymptomatic wild-collected Asian seabass, Lates calcarifer in SSN-1 cell line. The complete genome of the virus was amplified by PCR from plaque purified virus and sequenced. Bioinformatic analysis of the sequence revealed that the isolate belonged to red-spotted grouper nervous necrosis virus. The virus purified from infected SSN-1 cell culture supernatants by cesium chloride gradient ultracentrifugation resulted in two bands in the gradient. Both the bands on SDS-PAGE analysis and western blot revealed a single 40 kDa capsid protein. The virus replication was optimum at 26–28 °C in vitro and the virus can be completely inactivated using 3 mM binary ethylenimine in 32 h at 25 °C. The isolated virus was infectious and caused 100% mortality in Asian seabass larvae. The inactivated virus when injected to Asian seabass fingerlings produced neutralizing antibody titre ranging from 1:320 to 1: 1280. The isolate is a potential candidate for production of an inactivated vaccine against VNN.