猪捷申病毒5型VP1蛋白的原核表达及间接ELISA检测方法的建立

doi:10.11843/j.issn.0366-6964.2025.02.036

畜牧兽医学报 ›› 2025, Vol. 56 ›› Issue (2): 883-889.doi: 10.11843/j.issn.0366-6964.2025.02.036

猪捷申病毒5型VP1蛋白的原核表达及间接ELISA检测方法的建立

邵永恒¹(), 倪民婷²^,⁴, 高梦玲², 汤娇²^,³, 张耕馨²^,⁴, 林圣宇²^,⁵, 刘光亮², 陈佳宁²^,*(), 王雯慧¹^,*()

1. 甘肃农业大学动物医学院, 兰州 730070
2. 中国农业科学院兰州兽医研究所动物疫病防控全国重点实验室, 兰州 730000
3. 山西农业大学动物医学院, 晋中 030801
4. 黑龙江八一农垦大学动物科技学院, 大庆 163319
5. 福建农林大学动物科学学院, 福州 350002

收稿日期:2024-03-13 出版日期:2025-02-23 发布日期:2025-02-26
通讯作者: 陈佳宁,王雯慧 E-mail:1203552240@qq.com;chenjianing@caas.cn;wwh777@126.com
作者简介:邵永恒(1996-)，男，甘肃天水人，硕士生，主要从事基础兽医学研究，E-mail：1203552240@qq.com
基金资助:
甘肃省自然科学基金(22JR5RA027)

Prokaryotic Expression of VP1 Protein to Porcine Teschovirus Type 5 and the Establishment of an Indirect ELISA Detection Method

SHAO Yongheng¹(), NI Minting²^,⁴, GAO Mengling², TANG Jiao²^,³, ZHANG Gengxin²^,⁴, LIN Shengyu²^,⁵, LIU Guangliang², CHEN Jianing²^,*(), WANG Wenhui¹^,*()

1. College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China
2. State Key Laboratory for Animal Disease Control and Prevention, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730000, China
3. College of Veterinary Medicine, Shanxi Agricultural University, Jinzhong 030801, China
4. College of Animal Science and Technology, Heilongjiang Bayi Agricultural University, Daqing 163319, China
5. College of Animal Science, Fujian Agriculture and Forestry University, Fuzhou 350002, China

Received:2024-03-13 Online:2025-02-23 Published:2025-02-26
Contact: CHEN Jianing, WANG Wenhui E-mail:1203552240@qq.com;chenjianing@caas.cn;wwh777@126.com

摘要/Abstract

摘要：

旨在建立针对猪捷申病毒(porcine teschovirus，PTV)5型的间接ELISA检测方法，为其防控提供物质储备。PTV是猪的一种急性、烈性病原，以高发病率和高致死率为主要特征。其临床表现包括脑炎、肺炎、繁殖障碍、心肌炎及腹泻等多种症状。但该病原亚型众多，不同亚型间临床表现差异较大，且缺乏高效、特异的血清学检测方法，阻碍了对PTV-5的防治。本研究针对危害较大的PTV-5构建了pET-30a-VP1重组质粒，通过原核表达获得了VP1蛋白，以纯化的VP1为包被抗原，经条件优化成功建立了针对PTV-5的间接ELISA方法。该方法与常见猪源病毒标准阳性血清无交叉反应，最低检测稀释度为1∶3 200，批内与批间变异系数均低于10%，具有良好的特异性、敏感性和重复性。利用该方法对东北地区279份临床样本进行检测，其阳性率为67.74%，与报道的流行情况相符。本研究建立的间接ELISA检测方法综合评价良好，为PTV-5的临床检测提供技术支持。

关键词: 猪捷申病毒, VP1蛋白, 原核表达, 间接ELISA

Abstract:

The aim of this study was to establish an indirect ELISA method for the detection of porcine teschovirus (PTV) type 5 and to provide a material reserve for its prevention and control. PTV is an acute virulent pathogen of pigs, which is characterized by high morbidity and mortality. The clinical manifestations include encephalitis, pneumonia, reproductive disorders, myocarditis, and diarrhea. PTV can be divided into various subtypes. The clinical symptoms vary from each other. Therefore, lacking of efficient and specific serological detection methods blocks the control and prevention of PTV-5. This study inserted the VP1 gene of PTV-5 into pET-30a plasmid and obtained the VP1 protein. After optimizing the conditions, an indirect ELISA method against the VP1 of PTV-5 was successfully established. The method has no cross-reactivity with the standard positive sera of common porcine viruses. The lowest detection dilution was 1∶3 200, and the coefficients of variation were less than 10% both intra-and inter-batch, showing good specificity, sensitivity, and reproducibility of the method. A total of 279 clinical samples collected from the northeast were detected by this method. The results showed that the positive rate was 67.74%, which was in consistent with previous report. The indirect ELISA assay established in this study has a good overall evaluation and provides technical support for the clinical detection of PTV-5.

Key words: porcine teschovirus, VP1, prokaryotic expression, indirect ELISA

中图分类号:

S855.3

邵永恒, 倪民婷, 高梦玲, 汤娇, 张耕馨, 林圣宇, 刘光亮, 陈佳宁, 王雯慧. 猪捷申病毒5型VP1蛋白的原核表达及间接ELISA检测方法的建立[J]. 畜牧兽医学报, 2025, 56(2): 883-889.

SHAO Yongheng, NI Minting, GAO Mengling, TANG Jiao, ZHANG Gengxin, LIN Shengyu, LIU Guangliang, CHEN Jianing, WANG Wenhui. Prokaryotic Expression of VP1 Protein to Porcine Teschovirus Type 5 and the Establishment of an Indirect ELISA Detection Method[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(2): 883-889.

图/表 4

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猪捷申病毒5型VP1蛋白的原核表达及间接ELISA检测方法的建立

Prokaryotic Expression of VP1 Protein to Porcine Teschovirus Type 5 and the Establishment of an Indirect ELISA Detection Method

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