新型鸭呼肠孤病毒在DF-1细胞系中的增殖特性

doi:10.3969/j.issn.1004-1524.2019.05.06

浙江农业学报 ›› 2019, Vol. 31 ›› Issue (5): 716-721.DOI: 10.3969/j.issn.1004-1524.2019.05.06

新型鸭呼肠孤病毒在DF-1细胞系中的增殖特性

云涛, 华炯钢, 叶伟成, 倪征, 陈柳, 张存^*

浙江省农业科学院畜牧兽医研究所, 浙江杭州 310021

收稿日期:2019-01-10 出版日期:2019-05-25 发布日期:2019-05-23
通讯作者: *张存,E-mail: Cunzhang2004@aliyun.com
作者简介:云涛(1978—),男,浙江杭州人,博士,副研究员,从事禽病毒病分子生物学研究。E-mail: yt-t@163.com
基金资助:
浙江省重大科技专项重点农业项目(2015C02012)

Proliferative characteristics of novel duck reovirus strain JDM10 in DF-1 cells

YUN Tao, HUA Jionggang, YE Weicheng, NI Zheng, CHEN Liu, ZHANG Cun^*

Institute of Animal Husbandry and Veterinary Sciences, Zhejiang Academy of Agriculture Sciences, Hangzhou 310021, China

Received:2019-01-10 Online:2019-05-25 Published:2019-05-23

摘要/Abstract

摘要： 为了研究新型鸭呼肠孤病毒(new-type duck reovirus,NDRV)在鸡胚成纤维细胞DF-1中的增殖特性,将NDRV JDM10毒株接种到DF-1细胞,连续传代后,通过观察病毒对细胞的致病变效应,测定半数组织感染量(TCID₅₀)、RT-PCR检测、间接免疫荧光(IFA)及Western-blot免疫学检测,探索NDRV对DF-1细胞的作用效果。结果表明,NDRV JDM10毒株在DF-1细胞中能有效增殖,产生明显的致病变效应;RT-PCR检测成功扩增出1条大小为1 001 bp的条带;病毒蛋白在细胞中获得了良好的表达,并与抗NDRV σC单克隆抗体发生特异性反应;NDRV JDM10毒株在感染DF-1细胞后会经历潜伏期、快速增长期、稳定期3个时期,并在72 h病毒效价达到峰值,TCID₅₀为1×10^-7.90·(0.1mL)^-1。本研究为进一步研究NDRV的致病机理和研制细胞疫苗奠定了基础。

关键词: 新型鸭呼肠孤病毒(NDRV), 鸡胚成纤维(DF-1)细胞, 增殖特性

Abstract: To explore the proliferative charactistics of novel duck reovirus (NDRV) in chicken embryo fibroblast cells (DF-1). NDRV JDM10 strain was inoculated into DF-1 cells, and after continuous passage. The pathological effect (CPE) of virus on cells, half of tissue infection (TCID₅₀), RT-PCR, indirect immunofluorescence (IFA) and western-blot immunology were studied. The results showed that NDRV JDM10 could effectivly proliferate in the DF-1 cells and produce typical CPE such as cell shedding, cell body enlargement and cell disruption. The σC gene (1 001 bp) was successfully amplified from the infected cells by RT-PCR. NDRV σC was expressed well in DF-1 cells and could specifically reacted with anti-NDRV σC monoclonal antibody. The one-step growth curve showed that NDRV JDM10 strains experienced three periods of incubation, rapid growth and stable phase after infection with DF-1 cells, and the virus titer reached peak at 72 h, and was TCID₅₀ of 1×10^-7.90·(0.1 mL)^-1. These results established foundation for further study of the pathogenesis of NDRV and the development of cell vaccines.

Key words: novel duck reovirus (NDRV), DF-1 cells, proliferative characteristics

中图分类号:

S852.65

云涛, 华炯钢, 叶伟成, 倪征, 陈柳, 张存. 新型鸭呼肠孤病毒在DF-1细胞系中的增殖特性[J]. 浙江农业学报, 2019, 31(5): 716-721.

YUN Tao, HUA Jionggang, YE Weicheng, NI Zheng, CHEN Liu, ZHANG Cun. Proliferative characteristics of novel duck reovirus strain JDM10 in DF-1 cells[J]. , 2019, 31(5): 716-721.

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新型鸭呼肠孤病毒在DF-1细胞系中的增殖特性

Proliferative characteristics of novel duck reovirus strain JDM10 in DF-1 cells

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