A protein conserved in cypriniviruses is a major virulence factor of Cyprinid herpesvirus 3

Cyprinid herpesvirus 3 (CyHV-3) is the causative agent of a lethal disease in common and koi carp. Since its emergence, CyHV‑3 has caused severe economic losses worldwide creating a need for a safe and efficacious vaccine. In a previous study, we showed that a recombinant strain deleted for ORF56 and ORF57 exhibited a safety/efficacy profile compatible with its use as an attenuated recombinant vaccine.

In the present study, we investigated the relative contribution of the two genes to the attenuated phenotype observed. To reach this goal, a series of recombinants deleted either for ORF56 or ORF57 were produced. These recombinants were characterized in vitro for their correct molecular structure. In addition, immunofluorescence staining showed that the deletion of ORF56 did not abrogate the expression of ORF57, and vice versa. In vivo experiments demonstrated that the deletion of ORF57 explains most of the attenuation observed for the ORF56-57 deletion. Furthermore, we observed that ORF57 deletion induced in vitro a growth defect (reduction of both the production of infectious particles and the size of viral plaque). Orthologue of CyHV-3 ORF57 in Anguillid herpesvirus 1 (AngHV-1) has been shown to be a tegumental protein. Interestingly, using both qPCR and western blot based approaches, we demonstrated that the particles produced by the ORF57 deleted mutant are less infectious than those of the wild type virus.

In conclusion, this study demonstrates that ORF57 is a major virulence factor of CyHV-3. Importantly, as ORF57 is conserved in cypriniviruses, its orthologues could therefore represent a target for production of attenuated vaccine against several other major fish pathogens such as AngHV-1 and Cyprinid herpesvirus 2 (CyHV-2).