The PB1-F2 protein of the influenza A viruses (IAVs) can act as a virulence factor in mice. Its contribution to the virulence of IAV in swine, however, remains largely unexplored. In this study, we chose two genetically related H3N2 triple-reassortant IAVs to assess the impact of PB1-F2 in virus replication and virulence in pigs. Using reverse genetics, we disrupted the PB1-F2 ORF of A/swine/Wisconsin/14094/99 (H3N2) (Sw/99) and A/turkey/Ohio/313053/04 (H3N2) (Ty/04). Removing the PB1-F2 ORF led to increased expression of PB1-N40 in a strain-dependent manner. Ablation of the PB1-F2 ORF (or incorporation of the N66S mutation in the PB1-F2 ORF, Sw/99 N66S) affected the replication in porcine alveolar macrophages ... More
The PB1-F2 protein of the influenza A viruses (IAVs) can act as a virulence factor in mice. Its contribution to the virulence of IAV in swine, however, remains largely unexplored. In this study, we chose two genetically related H3N2 triple-reassortant IAVs to assess the impact of PB1-F2 in virus replication and virulence in pigs. Using reverse genetics, we disrupted the PB1-F2 ORF of A/swine/Wisconsin/14094/99 (H3N2) (Sw/99) and A/turkey/Ohio/313053/04 (H3N2) (Ty/04). Removing the PB1-F2 ORF led to increased expression of PB1-N40 in a strain-dependent manner. Ablation of the PB1-F2 ORF (or incorporation of the N66S mutation in the PB1-F2 ORF, Sw/99 N66S) affected the replication in porcine alveolar macrophages of only the Sw/99 KO (PB1-F2 knockout) and Sw/99 N66S variants. The Ty/04 KO strain showed decreased virus replication in swine respiratory explants, whereas no such effect was observed in Sw/99 KO, compared with the wild-type (WT) counterparts. In pigs, PB1-F2 did not affect virus shedding or viral load in the lungs for any of these strains. Upon necropsy, PB1-F2 had no effect on the lung pathology caused by Sw/99 variants. Interestingly, the Ty/04 KO-infected pigs showed significantly increased lung pathology at 3 days post-infection compared with pigs infected with the Ty/04 WT strain. In addition, the pulmonary levels of interleukin (IL)-6, IL-8 and gamma interferon were regulated differentially by the expression of PB1-F2. Taken together, these results indicate that PB1-F2 modulates virus replication, virulence and innate immune responses in pigs in a strain-dependent fashion.
