# Phage‐Mediated Presentation of a Conserved HA2 Epitope From Influenza A Virus Elicits Significant IgY Antibody Responses in Broiler Chickens

## TL;DR

A phage displaying a conserved influenza virus peptide triggered strong antibody responses in chickens, suggesting potential for universal flu vaccines.

## Contribution

A phage-based delivery system expressing a conserved influenza HA2 epitope was developed and shown to elicit specific IgY antibody responses in broiler chickens.

## Key findings

- The HA2 1–9 peptide was successfully expressed on the surface of M13 phage and induced specific IgY antibody responses in chickens.
- Vaccination with the phage did not significantly reduce H9N2 viral shedding, indicating a need for regimen optimization.
- Phage display platforms show promise for antigen presentation and immune activation in vaccine development.

## Abstract

Peptides alone often exhibit limited immunogenicity, necessitating the development of effective antigen delivery systems to facilitate recognition and presentation, ultimately eliciting robust immune responses and activating T and B lymphocytes. Filamentous bacteriophages, such as M13, are recognized as efficient platforms for peptide expression and presentation via their capsid surfaces. The conserved amino acid sequence HA2 1–9 (GLFGAIAGF) from the haemagglutinin (HA) transmembrane protein of Influenza A virus (IAV) has been identified as a promising immunogen for eliciting broad‐spectrum immune responses against diverse IAV strains. In this study, the N‐terminal fragment of Protein VIII from M13 phage was genetically engineered to express the conserved HA2 1–9 sequence. High‐level expression of the HA2 peptide on the phage surface was confirmed via immunoblotting analysis. Birds were intramuscularly vaccinated with the recombinant M13 phage displaying the HA2 peptide and subsequently challenged intranasally with the H9N2 IAV subtype. The results demonstrated that the GLFGAIAGF peptide elicited specific immunoglobulin Y (IgY) antibody responses against the HA2 peptide in birds. However, vaccination did not lead to a significant reduction in the shedding of H9N2 virus in the trachea and cloaca. This study highlights the potential of phage display platforms for antigen expression and immune activation. While the conserved GLFGAIAGF epitope successfully induced specific IgY responses, the limited efficacy in reducing viral shedding underscores the need for further optimization of the vaccination regimen, as well as investigation of alternative delivery routes, such as intranasal or oral administration, to enhance protective efficacy.

The conserved region of HA2 peptide of Influenza A virus was expressed on the surface of Protein VIII from M13 phage. Immunization of broiler chickens by this hybrid phage showed high levels of antibody responses, highlighting its potential as a promising tool for universal vaccine development against Influenza A virus.