Subtype-specific neutralizing antibodies promote antigenic shift during influenza virus co-infection

Reassortment between different influenza strains occurs when they co-infect the same host cell. The emergence of a reassortant virus depends on both its intrinsic fitness and extrinsic factors, including preexisting humoral immunity. The generation of pandemic strains, such as H2N2 and H3N2, and zoonotic influenza A viruses, such as H5N6, H5N8, and H7N9, in birds is suggested to be the result of extensive selection by preexisting antibodies. To further explore the role of humoral immunity in reassortment, we generated two divergent fluorescent protein-expressing viruses and used strain-specific and cross-reactive monoclonal antibodies (mAbs) to assess the impact of cross-immunity on reassortment. Our results indicate that all mAbs altered the genotypic diversity and significantly reduced the release of progeny virions in co-infected cells both in vitro and in vivo. Moreover, antibody transfer studies in mice revealed protection from challenge with divergent pathogenicity profiles. Notably, selection driven by a strain-specific mAb depended on its neutralizing specificity, whereas the selection driven by broadly reactive mAbs was independent of neutralization specificity. Our findings demonstrate that preexisting neutralizing antibodies shape reassortment and that strain-specific neutralizing antibodies promote antigenic shift during co-infection, which is not the case for broadly cross-reactive antibodies that recognize influenza viruses from different subtypes.