Hasoksuz Mustafa, Alekseev Konstantin, Vlasova Anastasia, Zhang Xinsheng, Spiro David, Halpin Rebecca, Wang Shiliang, Ghedin Elodie, Saif Linda J
Food Animal Health Research Program, Department of Veterinary Preventive Medicine, Ohio Agricultural Research and Development Center, The Ohio State University, 1680 Madison Avenue, Wooster, OH 44691, USA.
J Virol. 2007 May;81(10):4981-90. doi: 10.1128/JVI.02361-06. Epub 2007 Mar 7.
Coronaviruses (CoVs) possess large RNA genomes and exist as quasispecies, which increases the possibility of adaptive mutations and interspecies transmission. Recently, CoVs were recognized as important pathogens in captive wild ruminants. This is the first report of the isolation and detailed genetic, biologic, and antigenic characterization of a bovine-like CoV from a giraffe (Giraffa camelopardalis) in a wild-animal park in the United States. CoV particles were detected by immune electron microscopy in fecal samples from three giraffes with mild-to-severe diarrhea. From one of the three giraffe samples, a CoV (GiCoV-OH3) was isolated and successfully adapted to serial passage in human rectal tumor 18 cell cultures. Hemagglutination assays, receptor-destroying enzyme activity, hemagglutination inhibition, and fluorescence focus neutralization tests revealed close biological and antigenic relationships between the GiCoV-OH3 isolate and selected respiratory and enteric bovine CoV (BCoV) strains. When orally inoculated into a BCoV-seronegative gnotobiotic calf, GiCoV-OH3 caused severe diarrhea and virus shedding within 2 to 3 days. Sequence comparisons and phylogenetic analyses were performed to assess its genetic relatedness to other CoVs. Molecular characterization confirmed that the new isolate belongs to group 2a of the mammalian CoVs and revealed closer genetic relatedness between GiCoV-OH3 and the enteric BCoVs BCoV-ENT and BCoV-DB2, whereas BCoV-Mebus was more distantly related. Detailed sequence analysis of the GiCoV-OH3 spike gene demonstrated the presence of a deletion in the variable region of the S1 subunit (from amino acid 543 to amino acid 547), which is a region associated with pathogenicity and tissue tropism for other CoVs. The point mutations identified in the structural proteins (by comparing GiCoV-OH3, BCoV-ENT, BCoV-DB2, and BCoV-Mebus) were most conserved among GiCoV-OH3, BCoV-ENT, and BCoV-DB2, whereas most of the point mutations in the nonstructural proteins were unique to GiCoV-OH3. Our results confirm the existence of a bovine-like CoV transmissible to cattle from wild ruminants, namely, giraffes, but with certain genetic properties different from those of BCoVs.
冠状病毒（CoVs）拥有庞大的RNA基因组，以准种形式存在，这增加了适应性突变和跨物种传播的可能性。最近，CoVs被认为是圈养野生反刍动物中的重要病原体。这是关于从美国一个野生动物公园的长颈鹿（Giraffa camelopardalis）中分离出一种类牛CoV并进行详细的基因、生物学和抗原特性分析的首次报告。通过免疫电子显微镜在三只患有轻度至重度腹泻的长颈鹿的粪便样本中检测到了CoV颗粒。从三只长颈鹿样本中的一个样本中，分离出一种CoV（GiCoV-OH3），并成功适应在人直肠肿瘤18细胞培养物中连续传代。血凝试验、受体破坏酶活性、血凝抑制和荧光灶中和试验显示，GiCoV-OH3分离株与选定的呼吸道和肠道牛CoV（BCoV）毒株之间存在密切的生物学和抗原关系。当口服接种到一只BCoV血清阴性的无菌小牛中时，GiCoV-OH3在2至3天内引起严重腹泻和病毒排出。进行了序列比较和系统发育分析，以评估其与其他CoVs的遗传相关性。分子特征证实，新分离株属于哺乳动物CoVs的2a组，并显示GiCoV-OH3与肠道BCoVs BCoV-ENT和BCoV-DB2之间的遗传关系更密切，而BCoV-Mebus的关系更远。对GiCoV-OH3刺突基因的详细序列分析表明，S1亚基可变区（从氨基酸543到氨基酸547）存在缺失，这是一个与其他CoVs的致病性和组织嗜性相关的区域。在结构蛋白中鉴定出的点突变（通过比较GiCoV-OH3、BCoV-ENT、BCoV-DB2和BCoV-Mebus）在GiCoV-OH3、BCoV-ENT和BCoV-DB2中最为保守，而非结构蛋白中的大多数点突变是GiCoV-OH3独有的。我们的结果证实了存在一种可从野生反刍动物（即长颈鹿）传播给牛的类牛CoV，但具有与BCoVs不同的某些遗传特性。
