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基于 qPCR 和 qLAMP 的甲壳动物虹彩病毒 1 (DIV1)快速检测方法的建立与比较。

Development and comparison of qPCR and qLAMP for rapid detection of the decapod iridescent virus 1 (DIV1).

机构信息

Institute of Food Safety and Nutrition, Jinan University, Guangzhou 510632, Guangdong, China.

School of Food Science and Engineering, South China University of Technology, Guangzhou 510641, Guangdong, China.

出版信息

J Invertebr Pathol. 2021 Jun;182:107567. doi: 10.1016/j.jip.2021.107567. Epub 2021 Mar 10.

Abstract

Decapod iridescent virus 1 (DIV1) is a new virus discovered in recent years that infects farmed shrimp. DIV1 is highly infectious and causes substantial economic loss to the aquaculture industry of China. To prevent and control the spread and outbreak of DIV1 in a timely manner, it is necessary to establish an efficient method for DIV1 diagnosis. In this study, quantitative real-time polymerase chain reaction (qPCR) and quantitative real-time loop-mediated isothermal amplification (qLAMP) detection methods were established based on the specific sequence of the viral ATPase gene. The results indicated that the minimum detection limits of qPCR and qLAMP were 1.9 × 10 copies/μL and 1.9 × 10 copies/μL, respectively; the designed primer had good specificity for DIV1 and did not react with 13 other viruses, including white spot syndrome virus (WSSV), Enterocytozoon hepatopenaei (EHP), acute hepatopancreatic necrosis disease (AHPND), infectious hypodermal and haematopoietic necrosis virus (IHHNV), etc. A total of 43 clinical samples suspected of DIV1 infection were diagnosed by qPCR and qLAMP. Our qPCR demonstrated results consistent with a qPCR assay published previously, and the diagnostic sensitivity (DSe) and diagnostic specificity (DSp) of qLAMP were 85.71% and 100%, respectively. This result indicates that qPCR and qLAMP have good accuracy in the detection of DIVI in clinical samples. As established in this study, qPCR and qLAMP combined with a comprehensive comparative analysis can provide effective new solutions for the detection of DIV1.

摘要

十足目虹彩病毒 1 型(DIV1)是近年来发现的一种感染养殖对虾的新型病毒。DIV1 具有高度传染性,给中国水产养殖业造成了巨大的经济损失。为了及时预防和控制 DIV1 的传播和爆发,有必要建立一种有效的 DIV1 诊断方法。本研究基于病毒 ATPase 基因的特异性序列,建立了定量实时聚合酶链反应(qPCR)和定量实时环介导等温扩增(qLAMP)检测方法。结果表明,qPCR 和 qLAMP 的最小检测限分别为 1.9×10 拷贝/μL 和 1.9×10 拷贝/μL;设计的引物对 DIV1 具有良好的特异性,与其他 13 种病毒(包括白斑综合征病毒(WSSV)、对虾肝肠胞虫(EHP)、急性肝胰腺坏死病(AHPND)、传染性皮下及造血组织坏死病毒(IHHNV)等)均无反应。用 qPCR 和 qLAMP 对 43 份疑似 DIV1 感染的临床样本进行了诊断。本研究建立的 qPCR 与之前发表的 qPCR 检测结果一致,qLAMP 的诊断灵敏度(DSe)和诊断特异性(DSp)分别为 85.71%和 100%。结果表明,qPCR 和 qLAMP 对临床样本中 DIVI 的检测具有良好的准确性。qPCR 和 qLAMP 结合综合比较分析,为 DIV1 的检测提供了有效的新解决方案。

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