Background: Pre-existing immunity to emerging influenza viruses informs pandemic risk assessment. We compared cross-reactive neuraminidase (NA) antibody levels against avian influenza A(H5N1) by age and birth (imprinting) cohorts defined by periods of human influenza A subtype (H1N1, H2N2 or H3N2) circulation over the past century.

Methods: We used anonymized, residual sera collected from ten age groups spanning one to >80 years during an August 2024 cross-sectional serosurvey conducted in British Columbia, Canada. We assessed NA inhibition antibody titres by enzyme-linked lectin assay against clade 2.3.4.4b H5N1, and 2009 H1N1pdm09 and 1968 H3N2 pandemic strains.

Results: Among 575 participants with median age 32 (IQR: 15-62) years, 404 (70%) had detectable anti-N1 titres >10 against H5N1, with 260 (45%), 182 (32%) and 98 (17%) having titres >40, >80 and >160, respectively. Anti-N1 titres against 2009 H1N1pdm09 and H5N1 were strongly correlated (r=0.86; 95%CI: 0.82-0.89), with both highest among young adults born 1997-2003 who were school-aged children during the 2009 H1N1 pandemic (427.9, 100.8), lowest among the youngest and least influenza-experienced pediatric cohorts born 2015-2023 (20.7, 6.8) and middle-aged adults born during the 1957-1967 H2N2 era (25.1, 10.7), thereafter increasing toward a similar secondary peak among the oldest cohorts born during the pre-1947 H1N1 era (387.3, 81.0).

Conclusions: A substantial proportion of the population has pre-existing anti-N1 against H5N1, with age-related variation reflecting H1N1 imprinting and exposure opportunities. Through heightened attack rates and shifts in immunological hierarchies, historic pandemics shape and expand the immune repertoire with implications for pre-immunity to emerging zoonotic threats.