Development of an ELISA for NNV-Specific Antibody Detection in Grouper Hatcheries in China

Grouper (Epinephelus sp.) is one of the most economically important aquaculture species in China and Southeast Asian countries. Nervous necrosis virus (NNV) infection causes significant mortality in grouper aquaculture. Detection of anti-NNV antibodies using the ELISA method has been reported as effective in preventing vertical transmission of NNV in sea bass and flounder. In this study, IgM antibodies were purified from the serum of groupers and subsequently used to immunize rabbits to obtain a secondary antibody. Additionally, the NNV capsid protein (CP) was cloned, over-expressed, and purified in Escherichia coli. Consequently, an ELISA method was developed to detect NNV-specific antibodies in groupers by utilizing purified recombinant NNV capsid protein (CP) and horseradish peroxidase (HRP)-labeled rabbit anti-grouper antibody. This ELISA system was applied in three asymptomatic grouper hatcheries in China, revealing significant variations in antibody levels. Our ELISA method can effectively detect NNV-specific antibodies and is simpler than other methods, making it suitable for subsequent large-scale applications in grouper.