对巴基斯坦腹泻犬直肠拭子样本中扩增的部分 VP-2 基因进行分子分析证实了犬细小病毒遗传变异株 CPV-2a 的流行，并检测到猫泛白细胞减少症病毒 (FPV) 的序列。
Molecular analysis of partial VP-2 gene amplified from rectal swab samples of diarrheic dogs in Pakistan confirms the circulation of canine parvovirus genetic variant CPV-2a and detects sequences of feline panleukopenia virus (FPV).
机构信息
Virology Lab, Center of Agricultural Biochemistry and Biotechnology (CABB), University of Agriculture, PO Box 38040, Jail road, Faisalabad, 38000, Pakistan.
Department of Clinical Medicine and Surgery, University of Agriculture, Faisalabad, Pakistan.
出版信息
Virol J. 2018 Mar 15;15(1):45. doi: 10.1186/s12985-018-0958-y.
BACKGROUND
The infection in dogs due to canine parvovirus (CPV), is a highly contagious one with high mortality rate. The present study was undertaken for a detailed genetic analysis of partial VP2 gene i.e., 630 bp isolated from rectal swab samples of infected domestic and stray dogs from all areas of district Faisalabad. Monitoring of viruses is important, as continuous prevalence of viral infection might be associated with emergence of new virulent strains.
METHODS
In the present study, 40 rectal swab samples were collected from diarrheic dogs from different areas of district Faisalabad, Pakistan, in 2014-15 and screened for the presence of CPV by immunochromatography. Most of these dogs were stray dogs showing symptoms of diarrhea. Viral DNA was isolated and partial VP2 gene was amplified using gene specific primer pair Hfor/Hrev through PCR. Amplified fragments were cloned in pTZ57R/T (Fermentas) and completely sequenced. Sequences were analyzed and assembled by the Lasergene DNA analysis package (v8; DNAStar Inc., Madison, WI, USA).
RESULTS
The results with immunochromatography showed that 33/40 (82%) of dogs were positive for CPV. We were able to amplify a fragment of 630 bp from 25 samples. In 25 samples the sequences of CPV-2a were detected showing the amino acid substitution Ser297Ala and presence of amino acid (426-Asn) in partial VP2 protein. Interestingly the BLAST analysis showed the of feline panleukopenia virus (FPV) sequences in 3 samples which were already positive for new CPV-2a, with 99% sequence homology to other FPV sequences present in GenBank.
CONCLUSIONS
Phylogenetic analysis showed clustering of partial CPV-VP-2 gene with viruses from China, India, Japan and Uruguay identifying a new variant, whereas the 3 FPV sequences showed immediate ancestral relationship with viruses from Portugal, South Africa and USA. Interesting observation was that CPV are clustering away from the commercial vaccine strains. In this work we provide a better understanding of CPV prevailing in Pakistan at molecular level. The detection of FPV could be a case of real co-infection or a case of dual presence, due to ingestion of contaminated food.
背景
犬细小病毒（CPV）引起的犬感染具有高度传染性和高死亡率。本研究对从巴基斯坦费萨拉巴德地区所有地区的感染家养和流浪犬的直肠拭子样本中分离出的部分 VP2 基因（630bp）进行了详细的遗传分析。病毒监测非常重要，因为病毒的持续流行可能与新的毒力株的出现有关。
方法
在本研究中，于 2014-15 年从巴基斯坦费萨拉巴德不同地区的腹泻犬中采集了 40 份直肠拭子样本，通过免疫层析法检测 CPV 的存在。这些犬大多为流浪犬，表现出腹泻症状。使用基因特异性引物对 Hfor/Hrev 通过 PCR 从病毒 DNA 中扩增部分 VP2 基因。将扩增片段克隆到 pTZ57R/T（Fermentas）中，并进行完全测序。通过 Lasergene DNA 分析包（v8；DNAStar Inc.，Madison，WI，USA）对序列进行分析和组装。
结果
免疫层析检测结果显示，33/40（82%）犬 CPV 阳性。我们能够从 25 个样本中扩增出 630bp 的片段。在 25 个样本中检测到 CPV-2a 的序列，显示部分 VP2 蛋白中丝氨酸 297 突变为丙氨酸和存在氨基酸（426-Asn）。有趣的是，3 个样本的 BLAST 分析显示猫泛白细胞减少症病毒（FPV）序列，这些样本已经对新型 CPV-2a 呈阳性，与 GenBank 中其他 FPV 序列的同源性为 99%。
结论
系统进化分析显示，部分 CPV-VP-2 基因与来自中国、印度、日本和乌拉圭的病毒聚类，确定了一个新的变异株，而 3 个 FPV 序列与来自葡萄牙、南非和美国的病毒有直接的祖先关系。有趣的是，CPV 与商业疫苗株聚类分离。在这项工作中，我们从分子水平上更好地了解了巴基斯坦流行的 CPV。FPV 的检测可能是真正的合并感染，也可能是由于摄入了受污染的食物而出现双重存在。
Zotero 插件