Co-infection of megalocytivirus and viral nervous necrosis virus in a very severe mass mortality of juvenile orange-spotted groupers (Epinephelus coioides)

Megalocytivirus is a large double-stranded DNA virus, and viral nervous necrosis virus (VNNV) is a piscine nodavirus comprising two single-stranded RNAs. Both of them are the major viral causative agents of grouper diseases worldwide. However, the co-infection of these two agents in the same mass mortality has not yet been reported. In mid-June 2011, a mass mortality of juvenile orange-spotted groupers occurred in a juvenile grouper rearing farm in Qionghai county, Hainan province, China. Approximately 270 000 juvenile groupers died in two weeks. The moribund fishes were sampled for viral agent detection by molecular investigation. Both megalocytivirus and VNNV were detected and designated as OGSIV-QH11 and GNNV-QH11, respectively. MCP/CP sequence analyses revealed that OGSIV-QH11 was an RSIV type of megalocytivirus and GNNV-QH11 was an RGNNV-type VNNV. Virus isolation was performed in the MFF-1 cell line, which is highly susceptible for megalocytivirus. OSGIV-QH11 can grow well in MFF-1 cells, as confirmed by transmission electron microscopy (TEM), indirect immunofluorescence assay and virus purification. GNNV-QH11 was detected in infected MFF-1 cells by RT-PCR but not by TEM. In summary, we first confirmed the co-infection of megalocytivirus and viral nervous necrosis virus in a mass mortality of juvenile orange-spotted groupers, and then isolated a megalocytivirus from the infected MFF-1 cells. Our study provides a better understanding of viral diseases in grouper aquaculture industry.

► A very severe mass mortality of juvenile groupers causing about 270,000 fish death was reported.
► Both megalocytivirus and VNNV were detected from this case of mass mortality of juvenile groupers.
► A new megalocytivirus (OSGIV-HN11) was isolated and partial characterized in MFF-1 cells.