北美H3N8亚型犬流感病毒双重RT-PCR检测方法的建立  

Duplex RT-PCR assay for detection of North American H3N8 subtype canine influenza virus

作　　者：宋晶伟 王晨曦[1] 张谞霄 孙洪磊[1] 彭金山[2] 蒲娟[1] SONG Jing-wei WANG Chen-xi ZHANG Xu-xiao SUN Hong-lei PENG Jin-shan PU Juan(College of Veterinary Medicine, China Agricultural University, Beijing 100193, China BeijingBureau of Agriculture, Beijing 100029, China)

机构地区：[1]中国农业大学动物医学院,北京海淀100193 [2]北京市农业局,北京西城100029

出　　处：《中国兽医杂志》2017年第8期26-28,31,共4页Chinese Journal of Veterinary Medicine

基　　金："十二五"国家科技支撑项目(2013BAD12B01)

摘　　要：H3N8亚型犬流感病毒在北美地区广泛流行,并引发犬的呼吸道疾病,但目前中国尚未有犬感染H3N8亚型犬流感病毒的报道。随着近几年宠物贸易的繁盛,犬流感流行区域可能逐渐扩大,增加北美H3N8亚型犬流感病毒传入中国的可能。因此,需要建立一种针对北美H3N8亚型犬流感病毒的快速检测方法。本研究中,我们建立了针对北美H3N8亚型犬流感病毒的HA和NA基因片段的双重RT-PCR检测方法。该方法可特异性地检出北美H3N8亚型犬流感病毒HA和NA基因,而未检出H3N2亚型犬流感病毒及其他亚型流感病毒、犬瘟热病毒、传染性肝炎病毒、犬细小病病毒、副流感病毒;检测方法灵敏度可达0.1 pg/μL。利用该方法对临床样本进行了检测,证实该检测方法可靠。该检测方法的建立为监测北美H3N8亚型犬流感病毒提供了良好的技术支撑,可用于外来犬流感病毒的检测。H3N8 subtype canine influenza virus has spread through North America and resulted in respiratory diseases. However, at present the case of dogs infected with the virus has not been reported in China. As the pet trade has flourished in recent years,it also increases the possibility of transmitting the virus into our country. Therefore, it is necessary to establish a specific and sensi-tive method for the detection of the H3N8 subtype canine influenza virus. In this study, a duplex RT-PCR（d RT-PCR） was estab-lished. Two primer sets targeting the hemagglutinin（HA） gene and the neuraminidase（NA） gene of North American H3N8 subtypecanine influenza virus were designed, respectively were designed. The amplification by the duplex RT-PCR assay was positive forNorth American H3N8 subtype canine influenza virus but negative for H3N2 subtype canine influenza virus, other subtypes（H1N1and H9N2） of influenza viruses, canine distemper virus, infectious canine hepatitis virus, canine parvovirus and parainfluenza virus.The sensitivity was 0. 1 pg/μL. The detection of clinical samples was carried out, and proved that this method was feasible. This de-tection method will provide effective technical support for monitoring the North American H3N8 subtype canine influenza virus andcan be used in the surveillance of exotic diseases.

关 键 词：H3N8 犬流感病毒 双重RT-PCR 

分 类 号：S852.655[农业科学—基础兽医学]