The immune response of white sturgeon to envelope glycoproteins of white sturgeon herpesvirus type 2 (WSHV-2)

White sturgeon (Acipenser transmontanus) is an important fish species in the California aquaculture industry. The rearing of juvenile sturgeon at several farms in central and northern California has been plagued by annual losses due to viral infections. The white sturgeon herpesvirus type 2 (WSHV-2) is one of the viruses associated with these losses. Investigations into the severity of these losses at California sturgeon farms due to WSHV-2 led us to analyze the principal viral antigens responsible for the serological response of white sturgeon to WSHV-2. A total of 12 hybridoma cell lines secreting monoclonal antibodies (Mabs) against WSHV-2 were produced and characterized. Three Mabs designated A-8F, A-5H and B-2H, were chosen as affinity reagents and characterized initially with respect to various elution conditions by an enzyme-linked immunosorbent assay (ELISA). Of a number of elution buffers tested, 3 M MgCl₂ was the most efficient for recovering the 180 kD viral glycoprotein with Mab A-8F. A simple newly developed immunoaffinity column was used to purify the 180 kD glycoprotein. Five out of nine fish injected with the 180 kD viral glycoprotein responded by producing anti-180 kD antibodies as detected by ELISA and immunoblotting. Negligible neutralizing titers were found in those sera with ELISA titers. Primary and secondary immune responses were found in sturgeon injected with the purified 180 kD protein. Passive immunization with anti-180 KD serum produced in larger sturgeon did not protect juvenile sturgeon from experimentally-induced infections with WSHV-2.; However, they were found to be protected from experimental infections with WSHV-2 after passive transfer of serum with anti-WSHV-2 neutralizing activity from WSHV-2 hyperimmunized adults. An ELISA was developed and applied to detect the presence of anti-WSHV-2 antibodies in sera from WSHV-2 hyperimmunized fish, broodstock, farm-reared juveniles following a virus outbreak, and normal sturgeon. The prevalence of antibodies among these groups of fish were 100%, 100%, 78% and 9.78%, respectively. Immunoblots of sera from WSHV-2 hyperimmunized or broodstock white sturgeon exhibited different reactivity patterns with isolated WSHV-2 proteins that did not correlate with ELISA and PNT (plaque neutralization test) titers. However, the 180 kD and 120 kD proteins were frequently recognized under reducing conditions, while the 200 kD and 120 kD proteins were always recognized under non-reducing conditions. In the sera of broodstock sturgeon, the 120 kD protein was the most frequently recognized viral protein under reducing conditions, while the 200 kD and 120 kD proteins were always recognized under non-reducing conditions.