Rescue of a unique ovine parainfluenza virus type 3 strain via Red/ET assembly

Parainfluenza virus type 3 (PIV3) is an important pathogen that causes respiratory disease in humans and many animals, but ovine infection has been rarely reported. In this study, a strain of ovine parainfluenza virus type 3 (OPIV3) was isolated from a sheep farm in Tianjin, China, and named TJ2022. Phylogenetic analysis indicated a high homology between the TJ2022 strain and the OPIV3 TX01 strain. Compared with the TX01 strain, TJ2022 showed 17 and 13 unique amino acid mutations in the P and L proteins, respectively, and possessed a distinct gene end sequence, suggesting unique genetic evolution of TJ2022. Three-dimensional structural prediction analysis of the L protein indicated that characteristic mutations in the L protein may affect the virus's replication capacity. Host immune response analysis showed that the expression levels of IFN-α, IFN-β, and interferon stimulated genes (Mx1, IFI6, ISG15, and OAS1) were significantly elevated after TJ2022 infection. Furthermore, using the Red/ET homologous recombination system, we constructed an infectious clone of OPIV3 dependent on the T7 promoter (rTJ2022). The rTJ2022 exhibited genetic stability, growth kinetics, and host cell immune response similar to TJ2022. This study establishes the foundation for exploring OPIV3 infection and defense mechanisms, epidemiology, and the potential development of live vector vaccines.