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Abstract: Avian influenza viruses (AIV) of the H9 subtype cause serious health problems in chickens, resulting in great economic losses to the poultry industry worldwide. The killed vaccine (KV) against H9 subtype AIV has been widely used in China since 1998 but has been linked with side effects in chickens and only partial protection. A few studies have demonstrated the immunostimulatory effects of the Hemagglutinating virus of Japan envelope (HVJ-E) in cancer therapy. In this study, the adjuvant efficacy and the protective effects of HVJ-E, in combination with H9N2 AI KV against AIV were evaluated. The maturation of murine dendritic cells treated by HVJ-E was verified by FACS in the current experiment, then the antibody hemagglutination inhibition (HI) titers and cytokines and the post-challenge virological profiles (oropharyngeal and cloacal virus shedding) were investigated to define the immune responses in chickens. Our findings indicate that HVJ-E could induce dendritic cell (DC) maturation in mice. Injection of HVJ-E in chickens resulted in raised levels of IFN-β and IFN-γ being present in sera suggesting a stimulatory effect in these animals. The antibody responses to AIV of chickens inoculated with HVJ-E adjuvanted killed H9-AIV were higher than those of chickens inoculated with oil adjvanted H9-HIV. Furthermore, although inoculation of either HVJ-E or oil adjuvanted AIV reduced virus shedding following challenge, compared to controls, HVJ-E adjuvanted AIV was more effective in reducing shedding than oil adjuvant. [Copyright &y& Elsevier]