Abstract Licensed parenteral influenza vaccines induce systemic antibody responses and alleviate disease severity but do not efficiently prevent viral entry and transmission due to the lack of local mucosal immune responses. Here, we describe intranasal booster strategy with unadjuvanted recombinant hemagglutinin (HA) following initial mRNA-LNP vaccination, Prime and HA. This regimen establishes highly protective HA-specific mucosal immune memory responses in the respiratory tract. Intranasal HA boosters provided significantly reduced viral replication compared to parenteral mRNA-LNP boosters in both young and old mice. Correlation analysis revealed that slightly increased levels of nasal IgA are significantly associated with a reduced viral burden in the upper respiratory tract. Intranasal boosting with an antigenically distinct H1 HA conferred sterilizing immunity against heterologous H1N1 virus challenge. Additionally, a heterosubtypic intranasal H5 HA booster elicited cross-reactive mucosal humoral responses. Our work illustrates the potential of a nasal HA protein booster as a needle- and adjuvant-free strategy to prevent infection and disease from influenza A viruses. One Sentence Summary Adjuvant-free nasal booster induces protective immunity against influenza infection.

Intranasal hemagglutinin protein boosters induce robust mucosal immunity and cross-protection against influenza A viral challenge