Abstract

Novel duck reovirus (NDRV) is a major pathogen that causes immunosuppression and secondary infections in ducklings, resulting in elevated mortality and severe economic losses to the global duck industry. However, there is currently no licensed vaccine available in China for the effective prevention and control of NDRV infections. To effectively control NDRV outbreaks, we developed a live-attenuated vaccine candidate against NDRV by serially passaging the NDRV E232 strain in specific-pathogen-free (SPF) chicken embryos and DF-1 cells. The resulting E232-P100 strain replicated efficiently in DF-1 cells, with viral titers reaching up to 10 7.0 TCID 50 /0.2 mL. No clinical symptoms or pathological lesions were observed in two-day-old Cherry Valley ducklings inoculated with E232-P100, and no virulence reversion was observed after five rounds of in vivo back-passage in ducklings. The minimum protective dose of the attenuated E232-P100 strain was 10 3.0 TCID 50 /0.2 mL. Evaluation of the onset of immune protection showed that complete protection was achieved by 5 days post-vaccination. Comparative genomic analysis revealed 18 amino acid substitutions between E232-P100 and the parental E232 strain, including three within the σC protein that are potentially associated with attenuation. Collectively, the E232-P100 strain represents a safe, stable, and highly protective live attenuated vaccine candidate for the prevention of NDRV infection, providing a promising foundation for the development of effective prevention strategies against NDRV epidemics in waterfowl populations.