捻转血矛线虫LAMP-LFD检测方法的建立  

Establishment of a LAMP-LFD Detection Method for Haemonchus contortus

作　　者：段钰 汤洪宁 项继忠 吴飞 马光旭 杜爱芳[1] 杨怡[1] DUAN Yu;TANG Hong-ning;XIANG Ji-zhong;WU Fei;MA Guang-xu;DU Ai-fang;YANG Yi(College of Animal Science,Zhejiang University,Zhejiang Key Laboratory of Preventive Veterinary Medicine,Hangzhou,Zhejiang,310058,China;Huzhou Miemieyang Animal Husbandry Co.,Ltd,Huzhou,Zhejiang,313023,China)

机构地区：[1]浙江大学动物科学学院,浙江省动物预防医学重点实验室,浙江杭州310058 [2]湖州咩咩羊牧业有限公司,浙江湖州313023

出　　处：《动物医学进展》2024年第3期72-77,共6页Progress In Veterinary Medicine

基　　金：浙江省基础公益研究计划项目(LGN20C180005,LQ23C180006);浙江省自然科学基金重点项目(LZ22C180003)。

摘　　要：为了建立一种用于检测捻转血矛线虫的特异、灵敏、快速的检测方法,用环介导等温扩增技术(LAMP)结合横向流动试纸条(LFD),针对捻转血矛线虫mt-COⅠ基因的保守序列设计特异性引物和探针,对反应温度、反应时间、Mg^(2+)浓度、dNTP浓度进行优化,建立了捻转血矛线虫环介导等温扩增技术与横向流动试纸条检测技术(LAMP-LFD)相结合的核酸检测方法。结果显示,建立的LAMP-LFD检测方法最低检测限为100 fg,对奥斯特线虫、夏伯特线虫、毛首线虫DNA检测均为阴性;50份临床样品检测结果与粪便虫卵检测法结果的符合率为96%。该方法操作方便、结果判定快速简单,有望成为流行区域和基层检测捻转血矛线虫的新方法。In order to establish a specific,sensitive and rapid detection method for Haemonchus contortus,the loop-mediated isothermal amplification(LAMP)combined with the lateral flow dipstick(LFD)was used in this experiment.A set of specific primers and probes were designed for the conserved sequence of the mt-COⅠgene of H.contortus.The LAMP-LFD method for detecting H.contortus was established by optimizing reaction temperature,reaction time,Mg^(2+)concentration and dNTP concentration.The results showed that the minimum detection limit of the established LAMP-LFD method was 100 fg.The test results were negative for,Ostertagia circumcincta,Chabertia ovina,Trichocephalus.The coincidence rate of the results from 50 clinical samples between LAMP-LFD and fecal egg counting was 96%.With its convenience to operate and rapidity to determine the results,LAMP-LFD is expected to be a new method for detecting H.contortus in epidemic areas and grass roots.

关 键 词：捻转血矛线虫 环介导等温扩增 横向流动试纸条 mt-COⅠ 

分 类 号：S852.731[农业科学—基础兽医学]