Intranasal NS1-truncated live attenuated canine influenza vaccine confers superior protection compared to inactivated vaccine in beagles.

Canine influenza virus (CIV) H3N2 continues to circulate among companion animals, posing a zoonotic risk due to its potential for cross-species transmission. However, currently available inactivated vaccines offer limited mucosal immunity and suboptimal protection. Here, we developed a novel intranasal live attenuated CIV H3N2 vaccine carrying a truncated nonstructural protein 1 (NS1) gene and evaluated its safety, immunogenicity, and protective efficacy in beagle dogs. The NS1-truncated LAIV was well-tolerated and induced robust mucosal and systemic immune responses, including high titers of virus-specific secretory IgA. Following challenge with virulent CIV H3N2 at 120 days post-vaccination, LAIV-immunized dogs exhibited complete clinical protection and minimal viral shedding, whereas dogs receiving the inactivated vaccine showed moderate disease signs. These findings demonstrate that the NS1-truncated LAIV confers superior protection compared to conventional vaccines and represents a promising next-generation platform for canine influenza control within a One Health framework.