Studies of epidemiology and Seroprevalence of bovine noroviruses in Germany
Studies of epidemiology and Seroprevalence of bovine noroviruses in Germany
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DOI：
10.1128/jcm.41.6.2300-2305.2003
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发表时间：
2003-06-01
影响因子：
9.4
通讯作者：
Clarke, IN
中科院分区：
文献类型：
--
作者：
Deng, Y;Batten, CA;Clarke, IN
耶拿病毒（JV）是一种引起犊牛腹泻的牛肠道杯状病毒。该病毒直径约30纳米，表面形态与人类诺瓦克病毒相似。最近描述了JV的基因组序列，并将该病毒确定为杯状病毒科诺如病毒属。本研究将开放阅读框2编码的JV衣壳基因克隆到杆状病毒转移载体pFastBac 1中，利用该载体转化大肠杆菌生成重组bacmid。用重组杆状病毒DNA转染昆虫细胞，可表达JV衣壳蛋白。重组JV衣壳蛋白自组装成与JV病毒粒子大小和外观相似的病毒样颗粒（VLPs）。将JV VLPs释放到细胞培养上清中，浓缩后经CsCl平衡梯度离心纯化。纯化的JV VLPs用于实验动物的超免疫。建立了抗原捕获酶联免疫吸附试验（ELISA），并对临床标本进行了初步鉴定，其中包含明确的人诺如病毒和牛腹泻样本，这些样本来自实验感染JV的小牛；该酶联免疫吸附试验仅对JV有特异性。采用ELISA对1999 - 2002年在德国图林根州、黑森州和巴伐利亚州的奶牛群中采集的381份腹泻样本进行了筛选；其中34份（8.9%）为JV感染阳性。使用抗JV ELISA筛选的824份血清或血浆样本进行的血清流行病学研究证实了JV的意外高流行率，结果显示图林根州99.1%的牛具有JV抗体。
Jena virus (JV) is a bovine enteric calicivirus that causes diarrhea in calves. The virus is approximately 30 nm in diameter and has a surface morphology similar to the human Norwalk virus. The genome sequence of JV was recently described, and the virus has been assigned to the genus Norovirus of the family Caliciviridae. In the present study, the JV capsid gene encoded by open reading frame 2 was cloned into the baculovirus transfer vector pFastBac 1, and this was used to transform Escherichia coli to generate a recombinant bacmid. Transfection of insect cells with the recombinant baculovirus DNA resulted in expression of the JV capsid protein. The recombinant JV capsid protein undergoes self-assembly into virus-like particles (VLPs) similar to JV virions in size and appearance. JV VLPs were released into the cell culture supernatant, concentrated, and then purified by CsCl equilibrium gradient centrifugation. Purified JV VLPs were used to hyperimmunize laboratory animals. An antigen capture enzyme-linked immunosorbent assay (ELISA) was developed and characterized initially with clinical specimens containing defined human noroviruses and bovine diarrheal samples from calves experimentally infected with JV; the ELISA was specific only for JV. The ELISA was used to screen 381 diarrheal samples collected from dairy herds in Thuringia, Hesse, and Bavaria, Germany, from 1999 to 2002; 34 of these samples (8.9%) were positive for JV infection. The unexpectedly high prevalence of JV was confirmed in a seroepidemiological study using 824 serum or plasma samples screened using an anti-JV ELISA, which showed that 99.1% of cattle from Thuringia have antibodies to JV.