Abstract: The purpose of this study was to express the full-length, truncated and bivalent fusion proteins of group Ⅰ fowl adenovirus serotype 4 (FAdV-4) fiber2 and serotype 8b(FAdV-8b) fiber by prokaryotic expression system, and evaluate its immunogenicity. Based on good antigenic truncated fragments (FP and BP) of FAdV-4 fiber2 and FAdV-8b fiber, a bivalent fusion fragment was constructed by SOE-PCR. The truncated protein, full-length and fusion protein were individually expressed in E. coli and identified by SDS-PAGE and Western blot. The SPF chickens were immunized using purified recombinant proteins and challenged with FAdV-4 and FAdV-8b. Survival rate of each group was calculated, and serum antibody levels of immunized chickens were detected by indirect ELISA. Viral loads of cloacal swab and the mRNA expression levels of cytokines IL-4, IFN-γ and TNF-α in liver were detected by qPCR post-challenge and histopathological observation. The constructed recombinant plasmids were successfully soluble expression in E. coli. The survival rate of BP-FP4 group was 58.3%, and that of other groups was 100%. The antibody level of full-length group (FL group and BL group) was higher than that of its corresponding fragment group (FP group and BP group), but the difference was not significant (P>0.05). At 3 days and 7 days post-challenge, the viral load in C8b group was significantly higher than that of immunized group (P<0.05). At 7 days post-challenge, the viral load in BP-FP8b group was significantly higher than that in BP and BL groups (P<0.05), but there was no significant difference between BL and BP group (P>0.05). The mRNA expression of three cytokines (IL-4,IFN-γ and TNF-α) in immunized groups were significantly higher compared to the negative control group (P<0.05). The mRNA expression of IFN-γ and TNF-α in FL group were significantly lower than those in BP-FP4 group (P<0.05), and there was no significant difference between FL group and FP group (P>0.05). The mRNA expression of IL-4 and IFN-γ in BL group were significantly lower than those in BP-FP8b group (P<0.05). Histopathological observation revealed that the liver lesions in the challenge groups were the most severe, followed by the FP and BP groups, and no significant pathological changes were observed in the FL and BL groups. This study showed that the immunoprotection of the truncated protein and the full-length protein of FAdV-4 fiber2 and FAdV-8b was better than that of the fusion protein. Therefore, the truncated protein could be used to replace the full-length protein in the preparation of fowl adenovirus subunit vaccine.

Key words: fowl adenovirus, fiber, prokaryotic expression, subunit vaccine