畜牧兽医学报2023，Vol.54Issue(12)：5162-5170,9.DOI:10.11843/j.issn.0366-6964.2023.12.026
禽腺病毒4型fiber-2与8b型fiber截短融合蛋白的原核表达及其免疫原性分析
Prokaryotic Expression and Immunogenicity Analysis of Truncated Fusion Protein of FAdV-4 Fiber2 and FAdV-8b Fiber
摘要
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)fi-ber 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 frag-ment 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 chick-ens 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 histopatho-logical 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 cor-responding 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 cy-tokines(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-y in BL group were significantly lower than those in BP-FP8b group(P＜0.05).Histopathologi-cal observation revealed that the liver lesions in the challenge groups were the most severe,fol-lowed 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 prepara-tion of fowl adenovirus subunit vaccine.关键词
禽腺病毒/纤突蛋白/原核表达/亚单位疫苗Key words
fowl adenovirus/fiber/prokaryotic expression/subunit vaccine分类
农业科技引用本文复制引用
迟丽丽,王君娜,张宇名,刘健,陈志远,李树凡,尹燕博,徐守振..禽腺病毒4型fiber-2与8b型fiber截短融合蛋白的原核表达及其免疫原性分析[J].畜牧兽医学报,2023,54(12):5162-5170,9.基金项目
国家重点研发计划项目(2018YFD0501403) （2018YFD0501403）
山东省现代农业产业技术体系家禽产业创新团队建设(SDAIT-011-03) （SDAIT-011-03）
