Characterisation of betanodavirus of finfishes from India and its purification using CsCl density gradient ultracentrifugation

Betanodavirus is a pathogen of many tropical and temperate marine and brackishwater fish species. It is an etiological agent of viral nervous necrosis (VNN) in fish larvae and juveniles, leading up to 100% mortality. One among the best and effective approaches to control viral diseases in aquaculture is vaccination. Purified viral particles are useful for the development of immunodiagnostics and vaccines. In this study, purification of betanodavirus was attempted through ultracentrifugation using CsCl density gradient. SSN-1 cells were infected with betanodavirus and after recording cytopathic effect (CPE), cells were harvested. Virus infection was confirmed by PCR assay using reported primers and sequencing. Transmission electron microscopy (TEM) was carried out for confirmation ofinfection. Virus from cell culture supernatants was purified using CsCl density gradient and ultracentrifugation. Purified virus was further confirmed by PCR assay, SDS-PAGE analysis and TEM. The purified particles were tested for infectivity by incubating with SSN-1 cell line for its biological activity. Keywords: Betanodavirus, CsCl gradient, Density gradient, SSN-1 cells, Ultracentrifugation TEM, VNN