Vaccine-induced antigenic drift of a human-origin H3N2 Influenza A virus in swine alters glycan binding and sialic acid avidity

Interspecies transmission of human influenza A viruses (FLUAV) to swine occurs frequently, yet the molecular factors driving adaptation remain poorly understood. Here we investigated how vaccine-induced immunity shapes the evolution of a human-origin H3N2 virus in pigs using an <i>in vivo</i> sustained transmission model. Pigs (seeders) were vaccinated with a commercial inactivated swine vaccine and then infected with an antigenically distinct FLUAV containing human-origin HA/NA. Contact pigs were introduced two days later. After 3 days, seeder pigs were removed, and new contacts introduced. This was repeated for a total of 4 contacts. Sequencing of nasal swab samples showed the emergence of mutations clustered near the HA receptor binding site, enabling immune escape and abolishing binding to N-glycolylneuraminic acid. Mutant viruses recognized α2,6-sialosides with 3 N-acetyllactosamine repeats, which are rare in swine lungs, while the parental virus bound structures with a minimum of 2 repeats. Adaptative HA mutations enhanced avidity for α2,6-linked sialic acid, likely compensating for the low abundance of extended glycans. Notably, residues outside the canonical HA binding pocket contribute to glycan binding, suggesting a trade-off between receptor breadth and avidity. These findings show that non-neutralizing immunity promotes viral adaptation by fine-tuning receptor engagement and immune evasion.