A multivalent mRNA-lipid nanoparticle vaccine containing eight hemagglutinin antigens elicited broad neutralizing antibody responses and protected against influenza A virus challenge in swine.

The diversity within H1 and H3 subtype influenza A viruses (IAV) in swine prevents effective vaccine control approaches with inactivated whole-virus vaccines. We addressed the challenge of controlling co-circulating hemagglutinin (HA) clades of swine IAV with the development of a multivalent mRNA-lipid nanoparticle (LNP) vaccine expressing 8 HA proteins to maximize genetic coverage. We applied a computational approach to select eight HA genes that represented 95% of the observed IAV detected in the United States between 2022 and 2025. Piglets were vaccinated and boosted intramuscularly with either individual HA mRNA-LNP or an 8-HA multivalent mRNA-LNP. Serum was collected to evaluate systemic antibody levels. Twenty-one days post-boost, pigs were challenged with a field relevant H1 1A.3.3.3-c3 IAV strain. The 8-HA multivalent mRNA-LNP vaccine induced neutralizing antibodies against all eight antigens and vaccinees were protected against lung lesions, with lesion scores similar to non-challenged animals. Homologous monovalent vaccination significantly reduced IAV detection in nasal secretions and in the lungs. Heterologous monovalent vaccination was not cross-protective but did not induce vaccine-associated enhanced respiratory disease. We provide evidence that monovalent and multivalent mRNA-LNP influenza vaccines elicited neutralizing antibody responses in pigs and protected against viral challenge. The versatility and capacity for rapidly updating the mRNA-LNP vaccine platform make it an appealing tool to improve animal health and minimize the circulation and diversity of IAV in swine. ImportanceInfluenza A virus is an important respiratory pathogen in swine, and zoonotic transmission of swine strains to humans remains a public health risk. Control strategies against IAV in swine herds rely heavily on biosecurity measures and vaccination. However, the antigenic diversity of IAV circulating in swine challenges current vaccination programs, and there is a need for broadly protective vaccines or platforms that can rapidly update components to reflect circulating diversity. mRNA-LNP vaccines have emerged as promising vaccine platforms, offering simultaneous delivery of multiple antigens, rapid development, scalable manufacturing, and potent immunogenicity. In this study, we assessed the immunogenicity and protective capacity of monovalent and multivalent mRNA-LNP vaccines encoding eight representative IAV HA antigens. To our knowledge, this is the first study to objectively select multiple representative endemic swine IAV strains by quantifying genetic diversity within the phylogeny and to apply this selection to rationally design and evaluate a multivalent HA mRNA-based influenza vaccine in the swine model.