Enhanced protection through genotype-matched bivalent H9N2-Newcastle disease virus vaccination: Comparative efficacy against contemporary field strains in specific-pathogen-free chickens.

Newcastle Disease Virus (NDV) and Low Pathogenic Avian Influenza (LPAI H9N2) infections cause substantial economic losses in poultry production globally. Despite widespread vaccination, outbreaks persist due to antigenic mismatches between vaccine and circulating field strains, highlighting the need for improved vaccines with better genetic and antigenic matching. This study evaluated the efficacy of a novel bivalent inactivated vaccine (Aviline FLU H9 ND-K) containing genetically matched H9N2 (Saudi Arabian strain, G1 lineage B) and NDV (LaSota genotype II and genotype VII.1.1) strains against contemporary field isolates. The novel vaccine was compared to a commercial vaccine containing heterologous H9N2 (UAE origin, G1 lineage C) and NDV (Ulster strain, genotype II) antigens in a controlled challenge study. Seventy specific-pathogen-free (SPF) chickens were allocated to seven groups (n = 10/group) and vaccinated subcutaneously at 4 weeks of age. Four weeks post-vaccination, birds were challenged with either LPAI A/chicken/Egypt/SP3/2022 (H9N2) or NDV-B7-RLQP-CH-EG-12 (genotype VII). Clinical protection, antibody responses (hemagglutination inhibition), viral shedding (qRT-PCR), and cytokine expression interleukin-6 (IL-6), interferon-gamma (IFN-γ) were comprehensively assessed. Both vaccines conferred 100 % protection against clinical disease and mortality. However, the Aviline FLU H9 ND-K vaccine induced significantly higher antibody titers against both H9N2 (p < 0.001) and NDV (p < 0.05 at 7- and 14-days post-challenge). Viral shedding was substantially reduced in the genotype-matched vaccine groups, with H9N2 shedding eliminated by 7 post-challenge (dpc) with marked reduction by 5 dpc compared to 7 dpc with the commercial vaccine (p < 0.01). Similarly, NDV shedding ceased by 7 dpc in the Aviline FLU H9 ND-K group but persisted in the commercial vaccine group (p < 0.001). Pro-inflammatory cytokine expression was more effectively modulated in birds receiving the genotype-matched vaccine, with significantly lower IL-6 levels (p < 0.01) and higher IFN-γ expression (p < 0.05) compared to birds receiving the commercial vaccine. These findings demonstrate that the genotype-matched vaccine provides superior immunogenicity, enhanced viral clearance, and improved immunomodulatory effects, emphasizing the critical importance of antigenic matching in vaccine development for effective control of evolving avian pathogens. The potential impact on viral evolution and transmission dynamics within vaccinated populations warrants further investigation.