【摘要】 猪繁殖与呼吸综合征(porcine reproductive and respiratory syndrome, PRRS)在我国常年呈现流行趋势，国内防控措施主要以灭活疫苗和减毒活疫苗为主，但其仍具有一定的局限性。本研究旨在研制一种安全高效的PRRS疫苗。通过生物信息学预测糖蛋白5(glycoprotein 5, GP5)抗原表位集中区域，构建GP5抗原肽重组腺病毒，免疫小鼠(Mus musculus)后，评价其安全性、中和抗体效价、淋巴细胞干扰素-γ(interferon-gamma, IFN-γ)分泌水平、淋巴细胞增殖及细胞因子转录水平，分析其免疫效果。结果显示，经PCR和酶切鉴定，成功构建了重组腺病毒质粒pAdTrack-GP5aa130-170；对包装后的腺病毒进行Western blot鉴定，构建了能稳定表达目的蛋白的重组腺病毒。重组腺病毒免疫小鼠后，增重变化和精神状态良好。中和抗体效价、淋巴细胞IFN-γ释放水平及淋巴细胞增殖率显著高于对照组(P<0.05)，与商品苗相比差异不显著。淋巴细胞白细胞介素4(interleukin 4, IL-4)、IL-6、IL-10转录水平极显著高于对照组和商品苗组(P<0.01)，而肿瘤坏死因子-α(tumor necrosis factor-α, TNF-α)表达量无显著差异；血清中的IL-4、IFN-γ和TNF-α分泌量极显著高于对照组(P<0.01)。以上结果表明，本研究成功构建了表达GP5抗原表位区域的重组腺病毒，免疫小鼠后可诱导产生较高的中和抗体和细胞免疫反应，具有良好的免疫原性。研究结果为PRRS新型疫苗的研制提供了参考。更多还原

【Abstract】 Porcine reproductive and respiratory syndrome(PRRS) has shown a persistent epidemic trend in our country, with inactivated vaccines and attenuated live vaccines being the primary options domestically.However, these vaccines still have certain limitations. This study aims to develop a safe and efficient PRRS vaccine. By predicting the concentrated region of antigenic epitopes of glycoprotein 5(GP5) through bioinformatics, a GP5 antigenic peptide recombinant Adenovirus was constructed. After immunizing mice(Mus musculus), its safety, neutralizing antibody titer, lymphocyte interferon-gamma(IFN-γ) secretion level,lymphocyte proliferation, and cytokine transcription level were evaluated to analyze its immunogenic effect.The results showed that PCR and enzyme digestion successfully constructed the recombinant adenoviral plasmid pAdTrack-GP5aa130-170. Western blot identification of the packaged Adenovirus confirmed the construction of a recombinant Adenovirus capable of stably expressing the target protein. After immunization with recombinant Adenovirus, the mice showed good weight gain and mental state. The neutralizing antibody titer, lymphocyte IFN-γ release level, and lymphocyte proliferation rate were significantly higher than the control group(P<0.05), with no significant difference compared to the commercial vaccine. The transcription levels of lymphocyte interleukin 4(IL-4), IL-6, and IL-10 were extremely significantly higher than that in the control and commercial vaccine groups(P<0.01). At the same time, there was no significant difference in tumor necrosis factor-α(TNF-α) expression level. The secretion levels of IL-4, IFN-γ, and TNF-α in the serum were extremely significantly higher than the control group(P<0.01). These results indicated that this study successfully constructed a recombinant Adenovirus expressing the GP5 antigenic epitope region, which could induce a high level of neutralizing antibodies and cellular immune responses in mice after immunization, and had good immunogenicity. The findings provide a reference for developing a novel vaccine against PRRS.更多还原