Background: Avian influenza (AI) caused by AI virus subtype H9N2 is a prevalent viral disease with enormous economic losses in poultry farms through significant respiratory and gastrointestinal manifestations. The degree of protection obtained from a vaccine strongly depends on the level of antigenic similarity between challenge and vaccine virus. Aims: The study aimed at investigating the possible effects of continuous antigenic changes occurring in circulating Iranian viruses since 1998 on the commercial vaccines outcome by using vaccine seeds from earlier outbreaks for inhibiting viral replication in target organs of broilers challenged with the recent isolate. Methods: Ninety broilers at one day of age were randomly allocated into 5 groups and vaccinated with autogenous or commercial vaccines (A or B). Two remaining groups consisted of challenged without vaccination and intact birds. Quantitative real time-polymerase chain reaction (qRT-PCR) was performed on the trachea and faecal samples of challenged chickens with recent H9N2 virus to determine viral load. Moreover, humoral antibodies titers were evaluated by hemagglutination inhibition (HI) assay. Results: There was no significant difference in H9N2 viral load in the trachea among vaccinated groups on 5 days post challenge (DPC), but on 15 DPC, the autogenous vaccine significantly lowered viral load compared to commercial vaccines (P≤0.05). No significant differences in faecal swab’s viral load was observed between autogenous and commercial vaccine A, and both of them significantly inhibited viral load compared to unvaccinated group (P≤0.05). In addition, the autogenous vaccine elicited the highest HI titer. Conclusion: Inactivated vaccines that use isolates from previous outbreaks are no longer able to induce proper immunity against recent H9N2 viruses. It seems the time to change vaccine strains to more recent isolates that have better antigenic similarity with current circulating H9N2 viruses in the region has come.

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