Immunogen selection and prior immunity shape antibody breadth following immunisation with avian H5 hemagglutinin

Abstract

Avian influenza A viruses pose a persistent zoonotic threat to humans owing to their expanding host range and high case fatality rates. In particular, viruses from the 2.3.4.4b clade of the H5 subtype have now been detected in over 60 mammalian species, raising serious pandemic concerns. Understanding immune recognition of the H5 hemagglutinin (HA) is therefore critical for effective vaccine design and pandemic preparedness. To understand the breadth of cross-recognition induced by different H5 strains, we selected genetically diverse H5 human isolates from 2003-2023 and assessed neutralising antibody responses elicited by adjuvanted recombinant HA protein-based vaccines in C57BL/6 mice. Neutralisation activity of sera was determined against seven H5 HA variants using pseudotyped viruses and a PR8-reassortant virus in micro-neutralisation assays. Our results showed a wide variety of cross-strain neutralisation across H5 HA antigen variants. The conventional vaccine strain A/Indonesia/05/2005 displayed narrow activity against emerging clade 2.3.4.4b viruses, whereas ancestral variants exhibited cross-neutralisation profiles showing a diversity of breath but with limited potency. Polyvalent H5 HA formulations and nanoparticle-displayed H5 HA platforms substantially broadened cross-neutralisation against diverse H5 strains. To examine the impact of pre-existing immunity on H5 vaccine immunogenicity in mouse models, mice were primed with either seasonal influenza infection or quadrivalent influenza vaccine (QIV) prior to H5 HA immunisation. QIV pre-vaccination, but not prior influenza infection, enhanced subsequent neutralizing responses towards A/Fujian-Sanyuan/21099/2017 (clade 2.3.4.4b) H5. Collectively, our results demonstrate that immunogen selection and prior immunity shape antibody breadth following immunisation with avian A(H5) hemagglutinin. Importance Highly pathogenic avian influenza H5 viruses continue to spread across an unprecedented range of mammalian hosts, heightening the risk of a human pandemic. Current vaccine approaches for H5 rely on frequent recommendations of candidate vaccine viruses to match emerging H5 strains. Developing broadly protective H5 vaccines is thus a priority as part of pandemic preparedness. This study demonstrates a considerable variability in the potential of H5 vaccine antigens to induce neutralisation breadth, as well as the potential for multivalent vaccines and ferritin nanoparticle-based strategies to robustly augment immunity across antigenic variants. Furthermore, prior immunity established by seasonal QIV impacts the immunogenicity of subsequent 2.3.4.4b HA vaccination against H5 diversity. These findings provide critical insights to guide the rational design of broadly reactive H5 vaccines and inform pre-pandemic preparedness strategies.