Southern flounder, Paralichthys lethostigma, in the Pamlico River estuary in North Carolina experienced significant mortality from December 1981 through January 1982.  Fish were examined for bacterial and parasitic infection and for toxic residues from industrial and agricultural chemicals, but no significant findings resulted. Histopathological examination revealed extensive sloughing and necrosis of the mucosa of the pyloric caeca and intestine, and moderate to severe subacute inflammation of the submucosa of the pyloric caeca consisting almost entirely of lymphocytes. Internal organ homogenates were assayed for virus involvement, and an agent was isolated. The virus was recovered from dead flounder, but not from specimens captured live and appearing healthy. The virus was ether stable, partly inactivated by heating at 56°C for 1 hour, and partly neutralized by polyvalent-IPNV antiserum. Consistent with these observations, electron micrographs of negatively stained preparations showed an unenveloped, icosahedral virion about 55 mm in diameter. The virus isolated from southern flounder appears to be a new addition to the group of bisegmented, double-stranded RNA viruses isolated from fish, marine molluscs, birds, and insects.