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鲤疱疹病毒3型ORF148 DNA疫苗对建鲤鱼苗的免疫保护
作者:
作者单位:

1. 华南农业大学兽医学院, 广东 广州 510642;
2. 广东省农业科学院动物卫生研究所, 广东省畜禽疫病防治研究重点实验室, 农业农村部兽用药物与诊断技术广东科学观测实验站, 广东 广州 510640

作者简介:

吴静(1993-),女,硕士研究生,研究方向动物解剖与组织胚胎学.E-mail:20172027028@stu.scau.edu.cn

中图分类号:

S94

基金项目:

国家自然科学基金项目（31402347）；广东省科技计划项目（2016A020210029）；广东省海洋与渔业厅海港建设和渔业产业发展专项（A201701C04）；广州市科技计划项目（201707010216）；广东省自然科学基金项目（2018A030310193）.


Immunization of Cyprinus carpio var. jian with Cyprinid herpesvirus 3 ORF148 DNA vaccine against koi herpesvirus disease
Author:
  • WU Jing 1,2

    WU Jing

    College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China
    ;Key Laboratory of Livestock Disease Prevention of Guangdong ProvinceScientific Observation and Experiment Station of Veterinary Drugs and Diagnostic Techniques of Guangdong Province, Ministry of Agriculture and Rural AffairsInstitute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China
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  • LIU Zhenxing 2

    LIU Zhenxing

    Key Laboratory of Livestock Disease Prevention of Guangdong ProvinceScientific Observation and Experiment Station of Veterinary Drugs and Diagnostic Techniques of Guangdong Province, Ministry of Agriculture and Rural AffairsInstitute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China
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  • MA Yanping 2

    MA Yanping

    Key Laboratory of Livestock Disease Prevention of Guangdong ProvinceScientific Observation and Experiment Station of Veterinary Drugs and Diagnostic Techniques of Guangdong Province, Ministry of Agriculture and Rural AffairsInstitute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China
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  • LIANG Zhiling 2

    LIANG Zhiling

    Key Laboratory of Livestock Disease Prevention of Guangdong ProvinceScientific Observation and Experiment Station of Veterinary Drugs and Diagnostic Techniques of Guangdong Province, Ministry of Agriculture and Rural AffairsInstitute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China
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  • MA Jiangyao 2

    MA Jiangyao

    Key Laboratory of Livestock Disease Prevention of Guangdong ProvinceScientific Observation and Experiment Station of Veterinary Drugs and Diagnostic Techniques of Guangdong Province, Ministry of Agriculture and Rural AffairsInstitute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China
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  • HAO Le 2

    HAO Le

    Key Laboratory of Livestock Disease Prevention of Guangdong ProvinceScientific Observation and Experiment Station of Veterinary Drugs and Diagnostic Techniques of Guangdong Province, Ministry of Agriculture and Rural AffairsInstitute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China
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  • KE Hao 2

    KE Hao

    Key Laboratory of Livestock Disease Prevention of Guangdong ProvinceScientific Observation and Experiment Station of Veterinary Drugs and Diagnostic Techniques of Guangdong Province, Ministry of Agriculture and Rural AffairsInstitute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China
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  • FENG Guoqing 2

    FENG Guoqing

    Key Laboratory of Livestock Disease Prevention of Guangdong ProvinceScientific Observation and Experiment Station of Veterinary Drugs and Diagnostic Techniques of Guangdong Province, Ministry of Agriculture and Rural AffairsInstitute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China
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  • LI Yugu 1

    LI Yugu

    College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China

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Affiliation:

1. College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China;
2. Key Laboratory of Livestock Disease Prevention of Guangdong Province;Scientific Observation and Experiment Station of Veterinary Drugs and Diagnostic Techniques of Guangdong Province, Ministry of Agriculture and Rural Affairs;Institute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China

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    摘要:

    鲤疱疹病毒3型（Cyprinid herpesvirus 3，CyHV-3）又称为锦鲤疱疹病毒（Koi herpesvirus，KHV），是一种高传染性、致死性病毒。为开发新型CyHV-3 DNA疫苗，前期研究中将CyHV-3 ORF148基因插入pEGFP-N1构建重组质粒，在此基础上，本研究通过转染试验、间接免疫荧光试验证实pORF148-EGFP融合蛋白可以在CCB-J细胞系和建鲤体内表达；将重组质粒作为DNA疫苗，肌肉注射免疫建鲤鱼苗，ELISA检测表明免疫pEGFP-ORF148重组质粒可以显著提高建鲤血清特异性抗体水平；RT-qPCR检测显示，免疫pEGFP-ORF148重组质粒后，建鲤脾脏和头肾中的免疫相关基因如及基因表达量均显著提高。攻毒实验显示，CyHV-3攻毒21 d后，PBS组、pEGFP-N1组和pEGFP-ORF148组的建鲤存活率分别为30%、35%和85%，免疫pEGFP-ORF148可以显著提高建鲤的存活率（<0.01）。本研究旨在为CyHV-3 DNA疫苗的应用提供理论依据。

    Abstract:

    Cyprinid herpesvirus 3 (CyHV-3), also known as koi herpesvirus (KHV), is a highly contagious and lethal virus. At present, a kind of commercial attenuated vaccine has been used in Israel against this virus. However, this vaccine, as well as other reported gene-modified attenuated vaccines against CyHV-3 infection, cannot exclude the risk of reversion to virulence. In contrast to live vaccines, DNA vaccines are easy to produce and safe to apply. CyHV-3 has a genome of 295 kb, which is predicted to encode 164 different open reading frames (ORFs). Among them, ORF148 encodes a glycosylated type I membrane protein and bioinformatics analysis has shown that this protein has good immunogenicity. During our previous study, CyHV-3 ORF148 was inserted into pEGFP-N1, generating a pEGFP-ORF148 recombinant plasmid. In the present study, it was confirmed that a pORF148-EGFP fusion protein could be successfully expressed by transfection assay and immunofluorescence analysis, respectively. The carps, n=45), were intramuscularly injected with the pEGFP-ORF148 recombinant plasmid, pEGFP-N1 and PBS three times at two-week intervals. The results of ELISA showed that immunization with the pEGFP-ORF148 recombinant plasmid could significantly (<0.01) improve the tire of specific antibodies against CyHV-3. The RT-qPCR revealed that injection with the pEGFP-ORF148 recombinant plasmid caused 145.30-, 131-, 52.27-, 12.16-, 4.19-, 3.05-, 4.03-fold increases of , gene expression in the head kidney, respectively, when the PBS immunized carps served as a control group. In the spleen, the expression of from pEGFP-ORF148 recombinant plasmid injected carps were 22.44-, 4.63-, 6.14-, and 4.23 times higher than those of control group. Post-infection with CyHV-3, the survival rates of carps immunized with PBS, pEGFP-N1, and the pEGFP-ORF148 recombinant plasmid were 30%, 35% and 85%, respectively, suggesting that immunization with the pEGFP-ORF148 recombinant plasmid significantly protected the carps against the CyHV-3 infection (<0.01). In conclusion, DNA vaccination based on the pEGFP-ORF148 recombinant plasmid could elicit the activation of an immune response and production of a specific antibody to produce effective protection against CyHV-3 infection. Thus, this study highlights a potential DNA vaccine candidate against CyHV-3.

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吴静,刘振兴,马艳平,梁志凌,马江耀,郝乐,柯浩,冯国清,李玉谷.鲤疱疹病毒3型ORF148 DNA疫苗对建鲤鱼苗的免疫保护[J].中国水产科学,2020,27(4):454-462
WU Jing, LIU Zhenxing, MA Yanping, LIANG Zhiling, MA Jiangyao, HAO Le, KE Hao, FENG Guoqing, LI Yugu. Immunization of Cyprinus carpio var. jian with Cyprinid herpesvirus 3 ORF148 DNA vaccine against koi herpesvirus disease[J]. Journal of Fishery Sciences of China,2020,27(4):454-462

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