Serological imprinting and vaccination history shape adult influenza antibody landscapes and H5N1 cross‑reactivity

Serological imprinting and vaccination history shape adult influenza antibody landscapes and H5N1 cross‑reactivity | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Serological imprinting and vaccination history shape adult influenza antibody landscapes and H5N1 cross‑reactivity Ralf Duerr, Kesi Wilson, Miilani Yonatan, Trishala Karmacharya, and 12 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8913230/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted You are reading this latest preprint version Abstract Influenza vaccines are central to mitigating the ~1 billion annual influenza cases, yet their effectiveness remains modest, and how immune history shapes protection against emerging strains is unclear. The rapid spread of highly pathogenic avian influenza H5N1 clade 2.3.4.4b in birds, poultry, and cattle has intensified concerns about pandemic risk, but how pre‑existing seasonal immunity and lifelong imprinting influence responses to this lineage is unknown. Here, we use a customized 26‑plex bead‑based immunoassay to longitudinally profile hemagglutinin (HA)‑ and neuraminidase (NA)‑specific antibodies in 30 adults in New York City over three consecutive influenza seasons (2021–2024). Seasonal vaccination induced broad, predominantly IgG1 responses that rose significantly one month post‑vaccination, waned between seasons, and left stable, antigen‑specific serological imprints. We observe low‑to‑intermediate but consistent antibody levels to contemporary H5N1 clade 2.3.4.4b HAs following routine seasonal vaccination, revealing partial H5 cross‑reactivity without known H5 exposure. For six antigens, baseline levels or vaccine‑induced boosts tracked with birth year in patterns that mirror historical subtype circulation, demonstrating subtype‑ and antigen‑specific imprinting, including for H5N1. These findings define how imprinting and vaccination history jointly structure adult influenza antibody landscapes and suggest that multiplex, personalized serology could inform tailored vaccination strategies and pre-pandemic H5N1 preparedness. Health sciences/Diseases/Infectious diseases/Influenza virus Biological sciences/Immunology/Adaptive immunity/Humoral immunity/Antibodies Biological sciences/Immunology/Vaccines Full Text Additional Declarations Yes there is potential Competing Interest. Ralf Duerr is a Scientific Advisory Board member for ProPharma. Mark J. Mulligan reports potential competing interests: clinical trials and laboratory research contract funding for vaccines and monoclonal antibodies for SARS-CoV-2 with Lilly, Pfizer, and Sanofi; personal fees for Scientific Advisory Board service from Merck, Meissa Vaccines, Inc., Sanofi, HilleVax, and Pfizer. All other authors declare no conflicts of interest. The mouse vaccination and challenge experiments were conducted in an Enhanced Pathogen Facility (EPF) and were approved by the Institutional Animal Care and Use Committee (IACUC) and institutional biosafety committees. All procedures were subject to institutional animal care and biosafety oversight; protocol details are available from the authors upon request Supplementary Files TableS2Infectioncohort.xlsx Table S2 TableS3Antigenandbeadinformation.xlsx Table S3 TableS1Vaccinationcohort.xlsx Table S1 TableS5Linearandlogisticregressiondata.xlsx Table S5 Suppl.InfoWilsonDuerretal.SerologicalImprinting20260213highres.pdf Supplementary Information Cite Share Download PDF Status: Under Review Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-8913230","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":599356836,"identity":"daf1a33e-804d-4f54-8845-8e6981bba49c","order_by":0,"name":"Ralf Duerr","email":"data:image/png;base64,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","orcid":"https://orcid.org/0000-0001-9569-9237","institution":"New York University - 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Ralf Duerr is a Scientific Advisory Board member for ProPharma. Mark J. Mulligan reports potential competing interests: clinical trials and laboratory research contract funding for vaccines and monoclonal antibodies for SARS-CoV-2 with Lilly, Pfizer, and Sanofi; personal fees for Scientific Advisory Board service from Merck, Meissa Vaccines, Inc., Sanofi, HilleVax, and Pfizer. All other authors declare no conflicts of interest.\u003c/p\u003e\n\u003cp\u003eThe mouse vaccination and challenge experiments were conducted in an Enhanced Pathogen Facility (EPF) and were approved by the Institutional Animal Care and Use Committee (IACUC) and institutional biosafety committees. All procedures were subject to institutional animal care and biosafety oversight; protocol details are available from the authors upon request\u003c/p\u003e","formattedTitle":"Serological imprinting and vaccination history shape adult influenza antibody landscapes and H5N1 cross‑reactivity","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"nature-portfolio","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"","title":"Nature Portfolio","twitterHandle":"","acdcEnabled":false,"dfaEnabled":false,"editorialSystem":"ejp","reportingPortfolio":"","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-8913230/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8913230/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"Influenza vaccines are central to mitigating the ~1 billion annual influenza cases, yet their effectiveness remains modest, and how immune history shapes protection against emerging strains is unclear. The rapid spread of highly pathogenic avian influenza H5N1 clade 2.3.4.4b in birds, poultry, and cattle has intensified concerns about pandemic risk, but how pre‑existing seasonal immunity and lifelong imprinting influence responses to this lineage is unknown. \r\nHere, we use a customized 26‑plex bead‑based immunoassay to longitudinally profile hemagglutinin (HA)‑ and neuraminidase (NA)‑specific antibodies in 30 adults in New York City over three consecutive influenza seasons (2021–2024). Seasonal vaccination induced broad, predominantly IgG1 responses that rose significantly one month post‑vaccination, waned between seasons, and left stable, antigen‑specific serological imprints. We observe low‑to‑intermediate but consistent antibody levels to contemporary H5N1 clade 2.3.4.4b HAs following routine seasonal vaccination, revealing partial H5 cross‑reactivity without known H5 exposure. For six antigens, baseline levels or vaccine‑induced boosts tracked with birth year in patterns that mirror historical subtype circulation, demonstrating subtype‑ and antigen‑specific imprinting, including for H5N1. 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