Graduate Thesis Or Dissertation
 

Isolation and characterization of a new reovirus from chum salmon (Oncorhynchus keta)

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  • This report describes the isolation of a new virus from adult chum salmon (Oncorhynchus keta) returning to the Tokushibetsu Hatchery, Hokkaido, Japan. The agent was isolated in the CHSE-214 cell line derived from chinook salmon (Oncorhynchus tshawytscha). The virus replicated in selected fish cell lines incubated between 10 and 20°C and produced a unique cytopathic effect characterized by the formation of circular plaque-like areas of cytoplasmic fusion. The nuclei of the cells were not affected. Physical and chemical studies have shown the agent, tentatively named chum salmon virus, to be a member of the family Reoviridae and distinct from any known virus of fish. Electron microscopy of negatively stained particles revealed a naked icosahedral virion 75 nm in diameter with a clearly defined double capsid. Treatment with α-chymotrypsin removed the outer capsid yielding 50-55 nm subviral particles with enhanced infectivity. Viral infectivity was not reduced by treatment with ether or chloroform. The virus was resistant to freeze-thaw and pH 3, but unstable at 56°C. It did not hemagglutinate human type 0 erythrocytes. No cross-neutralization was observed with antiserum against reovirus types 1, 2 or 3 or against infectious pancreatic necrosis virus. Viral replication was not inhibited by 5-fluoro-2'-deoxyuridine and infected cells stained with acridine orange showed typical reoviruslike cytoplasmic inclusions. Polyacrylamide gel electrophoresis of the viral genome revealed 10 segments of double-stranded RNA ranging in weight from 2.6 X 10⁶ to 0.32 X 10⁶ daltons with a total genome weight of 15.3 X 10⁶ daltons. Analysis of virion polypeptides on SDSpolyacrylamide gels indicated five major virion proteins with molecular weights of 145,000, 130,000, 75,000, 43,000 and 33,000 daltons were present in the intact virion. Two minor polypeptides of 80,000 and 30,000 daltons were also present. The density of the intact virion in CsC1 was 1.33 g/cc. The virus had limited pathogenicity for salmonid fish. Chum and chinook salmon infected with the virus exhibited a focal necrotizing hepatitis. No mortality could be attributed to these lesions which were active sites of viral synthesis. Rainbow trout (Salmo gairdneri) and kokanee salmon (Oncorhynchus nerka) showed no pathological changes in any organ system examined. The virus replicated in all four species tested.
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