Immune responses of orange-spotted grouper, Epinephelus coioides, against virus-like particles of betanodavirus produced in Escherichia coli.

Betanodaviruses are the causative agents of viral nervous necrosis (VNN), a serious disease of cultured marine fish worldwide. Virus-like particles (VLPs) are one of the good novel vaccine candidates to control this disease. Until now, betanodavirus vaccine studies mainly focused on the humoral immune response and mortality after virus challenge. However, little is known about the activation of genes responsible for cellular and innate immunity by vaccines. In the present study, VLPs of orange-spotted grouper nervous necrosis virus (OGNNV) were produced in prokaryotes and their ability to enter Asian sea bass cells was the same as native virus, suggesting that they possess a similar structure to OGNNV. VLPs immunogenicity was then determined by intramuscularly vaccinating Epinephelus coioides at different concentrations (1.5 or 15 μg g(-1) fish body weight, FBW) and immunizing frequencies (administration once, twice and thrice). A single vaccination with the dosage of 1.5 μg g(-1) FBW is enough to provoke high titer antibodies (average 3 fold higher than that of negative control) with strong neutralizing antibody titer as early as 1 week post immunization. Furthermore, quantitative PCR analysis revealed that eleven genes associated with humoral, cellular and innate immunities were up-regulated in the liver, spleen and head kidney at 12h post immunization, correlating with the early antibody response. In conclusion, we demonstrated that VLP vaccination induced humoral immune responses and activated genes associated with cellular and innate immunity against betanodavirus infection in orange-spotted grouper.