2014-2015年黑龙江三个地区犬冠状病毒分子流行病学调查及病毒的初步分离
发布时间：2018-09-14 10:54
【摘要】：犬冠状病毒病是由犬冠状病毒(Canine coronavirus,CCoV)引起的一种急性、高度接触性胃肠道传染病。在德国1971年首次报道后,该病毒在亚洲、欧洲、南美洲、北美洲等多个国家和地区相继被报道并呈现全球流行趋势,给养犬业带来极大的危害。近年来,随着CCoV高致病性变异毒株的发现及病毒致病性增强在欧洲国家陆续报道,该病再一次引起了广泛的关注。在过去五年,我国关于CCoV的流行情况及遗传进化性等报道较少。鉴于此,本试验通过对2014年5月~2015年4月,在黑龙江三个地区采集的腹泻犬粪便样品,进行CCoV的检测、遗传进化性分析、病毒的初步分离。明确黑龙江三个地区CCoV的基本流行情况,为CCoV的防控以及后续相关研究提供理论基础。为了明确黑龙江三个地区CCoV的流行现状及遗传进化性,本试验采集201份腹泻犬粪便拭子样品,采用RT-PCR方法针对CCoVM和S基因片段进行扩增,对扩增序列进行测序及分子进化性分析。M基因RT-PCR结果显示,201份样品中57份样品为CCoV阳性(28.36%,57/201),其中CCoV-Ⅰ型和CCoV-Ⅱ型分别占15.79%(9/57)和84.21%(48/57)。序列分析显示,57个CCoV毒株的M基因序列核苷酸同源性为88.4%~100%,推导氨基酸序列同源性为93%~100%;57个CCoV毒株与中国参考株HF3比较,核苷酸同源性为88.7%~96.2%,推导氨基酸序列同源性为92.2%~97.4%;CCoV-Ⅰ型和CCoV-Ⅱ型毒株与中国参考毒株和其他国家参考毒株相比较呈现遗传多样性。57个CCoV阳性样品与CPV-2、CaKV和CBo V混合感染率分别为31.58%(18/57)、33.33%(19/57)、5.26%(3/57)。犬感染CCoV的年龄、免疫情况、地区与季节性都有较大的差异。57个CCoV阳性样品中S基因扩增出28份,其中26个CCoV-Ⅱa亚型毒株,1个CCoV-Ⅱb亚型毒株和1个CCoV-Ⅰ型毒株。S基因序列分析显示,26个CCoV-Ⅱa亚型毒株之间的核苷酸同源性为86.3%~100%,推导氨基酸序列同源性为87.6%~100%;26个CCoV-Ⅱa亚型毒株与中国参考株V1之间核苷酸同源性为89.6%~92.2%,推导氨基酸序列同源性为91.1%~97.5%。26个CCoV-Ⅱa亚型毒株与中国参考株和欧洲、北美、南美洲、亚洲参考株相比较呈现出不同的进化史。本试验选取单纯感染的23株CCoV阳性样品,利用MDCK进行病毒分离,其中9株样品盲传至第4代出现明显CPE,通过RT-PCR鉴定为阳性,继续盲传第7代眼观CPE消失,RT-PCR鉴定为阴性;将9株样品重新接种到MDCK细胞,盲传至第5代眼观4株出现明显CPE,通过巢式RT-PCR鉴定为阳性,继续盲传,第6代无眼观CPE存在,巢式RT-PCR鉴定为阴性。本研究揭示,黑龙江三个地区腹泻犬中同时存在CCoV-Ⅰ型、CCoV-Ⅱa亚型和CCoV-Ⅱb亚型,且CCoV-Ⅱ型毒株具有较高的感染率及遗传多样性。在黑龙江三个地区CCoV与CPV-2、CaKV有着很高的混合感染率。CCoV在MDCK细胞上能出现CPE,但是病毒增殖极其不稳定。
[Abstract]:Canine coronavirus (canine coronavirus) is an acute, highly contagious gastrointestinal infection caused by canine coronavirus (Canine coronavirus,CCoV). After the first report in Germany in 1971, the virus has been reported in Asia, Europe, South America, North America and many other countries and regions. In recent years, with the discovery of highly pathogenic variant strains of CCoV and the increasing of viral pathogenicity in European countries, the disease has once again attracted wide attention. In the past five years, there have been few reports on the prevalence and genetic evolution of CCoV in China. In view of this, from May 2014 to April 2015, the fecal samples of diarrhea dogs collected in three areas of Heilongjiang Province were tested for CCoV, genetic evolution analysis and preliminary isolation of the virus. In order to provide theoretical basis for the prevention and control of CCoV and related research, the basic epidemic situation of CCoV in the three regions of Heilongjiang Province is clarified. In order to identify the prevalence and genetic evolution of CCoV in three regions of Heilongjiang Province, 201 fecal swabs from diarrhea dogs were collected and amplified by RT-PCR for CCoVM and S gene fragments. The results of sequencing and molecular evolution analysis showed that 57 of the 201 samples were CCoV positive (28.36 / 57), and CCoV- 鈪，
本文编号：2242500
[Abstract]:Canine coronavirus (canine coronavirus) is an acute, highly contagious gastrointestinal infection caused by canine coronavirus (Canine coronavirus,CCoV). After the first report in Germany in 1971, the virus has been reported in Asia, Europe, South America, North America and many other countries and regions. In recent years, with the discovery of highly pathogenic variant strains of CCoV and the increasing of viral pathogenicity in European countries, the disease has once again attracted wide attention. In the past five years, there have been few reports on the prevalence and genetic evolution of CCoV in China. In view of this, from May 2014 to April 2015, the fecal samples of diarrhea dogs collected in three areas of Heilongjiang Province were tested for CCoV, genetic evolution analysis and preliminary isolation of the virus. In order to provide theoretical basis for the prevention and control of CCoV and related research, the basic epidemic situation of CCoV in the three regions of Heilongjiang Province is clarified. In order to identify the prevalence and genetic evolution of CCoV in three regions of Heilongjiang Province, 201 fecal swabs from diarrhea dogs were collected and amplified by RT-PCR for CCoVM and S gene fragments. The results of sequencing and molecular evolution analysis showed that 57 of the 201 samples were CCoV positive (28.36 / 57), and CCoV- 鈪，
本文编号：2242500
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