Susceptibility of Goldfish to Cyprinid Herpesvirus 2 (CyHV-2) SH01 Isolated from Cultured Crucian Carp.

Cyprinid herpesvirus 2 (CyHV-2), a member of the Alloherpesviridae family belonging to the genus Cyprinivirus, is a fatal contagious aquatic pathogen that affects goldfish (Carassius auratus) and crucian carp (Carassius carassius). Although crucian carp and goldfish belong to the genus Carassius, it is unclear whether they are susceptible to the same CyHV-2 isolate. In addition, the origin of the crucian carp-derived CyHV-2 virus isolate remains unclear. CyHV-2 SH01 was isolated during herpesviral hematopoietic necrosis disease (HVHN) outbreaks in crucian carp at a local fish farm near Shanghai. CyHV-2 SH01 was confirmed by PCR and Western blot analysis of kidney, spleen, muscle, and blood tissue from the diseased crucian carp. Moreover, histopathological and ultra-pathological analyses revealed pathological changes characteristic of CyHV-2 SH01 infection in the tissues of the diseased crucian carp. In the present study, goldfish and crucian carp were challenged with CyHV-2 SH01 to elucidate viral virulence. We found that CyHV-2 SH01 could cause rapid and fatal disease progression in goldfish and crucian carp 24 h post-injection at 28 °C. Experimental infection of goldfish by injection indicated that the average virus titer in the kidney of the goldfish was 103.47 to 103.59 copies/mg. In addition, tissues exhibited the most prominent histopathological changes (cellular wrinkling and shrinkage, cytoplasmic vacuolation, fusion of the gill lamellae, and hepatic congestion) in CyHV-2 SH01-infected goldfish and crucian carp. Thus, crucian carp and goldfish showed a high sensitivity, with typical symptoms, to HVHN disease caused by CyHV-2 SH01.

Suppression effect of plant-derived berberine on cyprinid herpesvirus 2 proliferation and its pharmacokinetics in Crucian carp (Carassius auratus gibelio).

Cyprinid herpesvirus 2 (CyHV-2), which infects silver crucian carp including goldfish (Carassius auratus auratus) and Crucian carp (Carassius auratus gibelio) with high mortality, is an emerging viral pathogen worldwide. Previous studies showed that berberine (BBR), a bioactive plant-derived alkaloid, demonstrated potential antiviral actions against many different viruses. Here, we assessed the effect of berberine hydrochloride (BBH) on the replication of CyHV-2 in vitro and in vivo. Cytotoxicity assay indicated that 5-25 µg/mL BBH was non-toxic to the RyuF-2 cells. In viral inhibition assays, real time PCR was employed to titrate the genomic copy number of progeny virus, real time RT-PCR was applied to monitor the transcriptional levels of viral genes, and Western blot analysis was performed to detect the synthetic levels of viral proteins. The results demonstrated that BBH systematically impedes the viral gene transcription and suppressed the replication of CyHV-2 in RyuF-2 cells. In animal challenge test, BBH was confirmed to protect Crucian carps from CyHV-2 infection in a dose-dependent manner, which was supported by suppressed viral replication levels, reduced viral pathogenesis and higher survival rates. Furthermore, pharmacokinetics data of BBH in Crucian carp revealed its rapid absorption (Tmax of 1.5 h), suitable plasma half-life (t1/2z/h of 7-12 h depending on oral dosage), and dose-dependent drug exposure properties following oral administration (revealed by AUC0-t values). These findings shed light on repurposing BBH to treat CyHV-2 infections in silver crucian carp.