副猪格拉瑟菌6个候选抗原对小鼠的免疫特性分析

doi:10.11843/j.issn.0366-6964.2023.05.028

摘要/Abstract

摘要： 旨在获得具有良好免疫效果的候选抗原,本研究原核表达了副猪格拉瑟菌6个具有潜在保护力的蛋白,对小鼠免疫后进行攻毒,以期筛选获得具有免疫保护效果较好的候选抗原,从而为开发具有交叉保护作用的副猪格拉瑟菌亚单位疫苗奠定基础。63只BALB/c小鼠随机平均分为9组:HPSNAG-1330、CyaY、HPSNAG-0978、HPSNAG-0140、AfuA以及Fe³⁺ABC-tbp 6个亚单位疫苗组、G. parasuis-5灭活苗组、攻毒对照组和空白组;亚单位疫苗组每种对应蛋白各免疫50 μg·只^-1,灭活苗组免疫4×10⁹CFU,一免与二免间隔14 d,二免14 d后攻毒,副猪格拉瑟菌Nakasaki菌株攻毒剂量为1.26×10⁹CFU。在二免14 d后检测特异性抗体水平、淋巴细胞增殖和IL-2、IL-6、IL-10、IFN-γ和TNF-α的表达,攻毒后观察小鼠的临床症状并观察肺、脾的组织病理变化。免疫组小鼠均能产生体液免疫反应;淋巴细胞增殖试验显示:除rAfuA外,其它重组蛋白均能刺激灭活苗组中小鼠淋巴细胞显著增殖(P＜0.05),而6个重组蛋白对相应的亚单位疫苗组和空白组小鼠淋巴细胞增殖均不显著;同时,CyaY组和HPSNAG-1330组小鼠机体的炎性反应和脾、肺的损伤程度相对温和,综合评价效果优于灭活苗组。本研究通过小鼠免疫保护试验筛选出CyaY和HPSNAG-1330两个具有免疫保护性的候选抗原,为后续开展多个抗原的联合免疫和猪体试验提供了素材。

关键词: 副猪格拉瑟菌, 亚单位疫苗, 免疫效果评价

Abstract: To obtain subunit antigens with good immune effect, in this study, six potentially protective subunit proteins of G. parasuis were expressed in prokaryotic cells. After immunizing mice with proteins, challenge experiments were performed to screen antigen candidates with better immune protection effects, thus laying the foundation for developing a subunit vaccine of G. parasuis with cross-protection effects. Sixty-three BALB/c mice were randomly divided into 9 groups, including HPSNAG-1330, CyaY, HPSNAG-0978, HPSNAG-0140, AfuA, Fe³⁺ABC-tbp, inactivated G. parasuis, challenge group and blank group. The mice were immunized the second time with an interval of 14 days. Among them, the 6 subunit vaccine groups were immunized with 50 μg of each protein per mouse, and the inactivated vaccine group was immunized with 4×10⁹ CFU per mouse. After 14 days of the secondary immunization, the mice were challenged with a dose of 1.26×10⁹ CFU. At the same time, the antibody levels, lymphocyte proliferation, IL-2, IL-6, IL-10, IFN-γ and TNF-α were detected. After the challenge, the clinical symptoms and survival rate of the mice were observed and recorded, and the pathological variations of the lung and spleen were also observed. Mice in all immunized groups can produce antibodies against the corresponding proteins. The lymphocyte proliferation experiment showed that recombinant proteins, except for the rAfuA, stimulated lymphocyte proliferation significantly in the inactivated vaccine group (P<0.05). At the same time, there were no significant changes in the lymphocyte proliferation of the mice in the subunit vaccine group and the blank group when stimulated with the corresponding immune proteins. The survival rate (60%) of the CyaY and HPSNAG-1330 groups was higher than that of the inactivated vaccine group (40%), and which of the other groups was lower than that of the inactivated vaccine group. The above results proved that CyaY and HPSNAG-1330 proteins can induce effective immune protection responses in mice. At the same time, the lesions of the spleens and lungs of the mice in CyaY and the HPSNAG-1330 are relatively mild. So, the protective effect of these two proteins is better than that of inactivated vaccine. In this study, two subunit antigens with immune-protective properties were screened by mouse immunization experiments, which provided materials for subsequently combined immunization of multiple antigens using pig model experiments.

Key words: Glaesserella parasuis, recombinant subunit vaccine, evaluation of the immune effect

中图分类号:

S858.285.1

伭婷, 杨凯一, 蔡金双, 耿琰, 李玉峰. 副猪格拉瑟菌6个候选抗原对小鼠的免疫特性分析[J]. 畜牧兽医学报, 2023, 54(5): 2073-2082.

XUAN Ting, YANG Kaiyi, CAI Jinshuang, GENG Yan, LI Yufeng. Analysis of the Immune Characteristics of the Six Recombinant Proteins of Glaesserella parasuis in Mice[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(5): 2073-2082.

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