鸭肿头出血症病毒在鸭胚原代成纤维细胞中增殖特性的研究

Proliferative characteristics of duck swollen head haemorrhagic disease virus in duck embryo primary fibroblasts

将鸭肿头出血症病毒（DSHDV）强毒株经尿囊腔接种10日龄鸭胚，传3代，取第3代尿囊液接种鸭胚成纤维细胞（DEF），传代培养至有细胞病变出现，并对接毒细胞进行TCID50动态检测和透射电镜观察。结果显示，第1代接毒细胞培养至120h出现变圆、脱落，并形成少量蚀斑，第2代培养至96～120h，DEF形成大量圆形或椭圆形的典型蚀斑，第3代培养至72～96h，DEF出现典型蚀斑；此后可稳定适应于DEF，典型细胞病变出现于72～96h。DSHDV在DEF上的TCID50值随传代次数的增加而增高，由第1代的10^2.72增加到第10代的10^6.82，最高毒价出现于96h，该点是收获病毒的最佳时间。经透射电镜观察，可见DSHDV粒子呈球形或椭圆形，大小为60～75nm，无囊膜，具有双层衣壳。结果证实，DSHDV强毒株已经适应了DEF，传代后可获得较高的病毒效价。

Duck swollen head haemorrhagic disease virus（DSHDV） was propagated by allantoic cavity inoculation of 10-day-old duck embryo for 3 passages. The allantoic fluid was inoculated onto duck embryo fibroblasts（DEFs） and then passaged until the cytopathic effect （CPE） appeared. Dynamic detection of TCID50 and observation by transmission electron microscopy of the infected cells were carried out. At 1st passage,the cells became round, fall off, and produced only a little plaque at hour 120. At 2nd passage, DSHDV produced the typical plaque during 96-120 h. At 3rd passage,characteristic plaques formed on cell monolayer during hour 72-96. Since then,the DSHDV was adapted to DEFs, and typical and consistent plaques were observed between hour 72-96. The TCID50 titer of DSHDV was getting higher from P1 10^2.75 to P10 10^6.82 Hour 96 at 3rd passage was the best time to harvest DSHDV because of the maximum virus titers. The virions were globular and ellipse, approximately 60-75 nm in size, without peplos and with double capsid under transmission electron microscope. These results confirmed that DSHDV was able to adapt to DEFs,and the virus titer was getting higher along with increase of passages.