Implementation of a Recombinase Polymerase Amplification-Driven Duplex Lateral Flow Diagnostic Kit for Concurrent Detecting Infectious Pancreatic Necrosis Virus and Viral Hemorrhagic Septicemia Virus: A Comparative Analysis of RPA and PCR Methods.

IF 2.2 3区 农林科学 Q2 FISHERIES
Cuneyt Tamer, Hanne Nur Kuruçay, Seda Gözel, Bahadır Müftoglu, Elif Nur Kökluoğlu, Hamza Kadi, Fatih Dogan, Serdar Yildirim, Semra Gumusova, Zafer Yazici, Manfred Weidmann, Harun Albayrak
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引用次数: 0

Abstract

Diagnosis time of infectious diseases in aquaculture is an important issue, and local and commercial enterprises generally do not prefer laboratory diagnosis of infectious diseases due to the long lead time and cost. This leads to the under-reporting of outbreaks in aquaculture farms and the recognition of infectious agents when they spread to many farm systems and cause significant economic loss. In this study, we aim to develop a fast and simple diagnostic test that can easily be applied under field conditions against Infectious pancreatic necrosis virus (IPNV) and Viral hemorrhagic septicemia virus (VHSV Ie), which are two crucial viral pathogens that cause significant problems in trout farming in Türkiye, both in hatcheries and farms. For this reason, we developed a lateral flow test strip for the duplex diagnosis of VHSV Ie and IPNV sp. based on Recombinase polymerase amplification (RPA), one of the molecular tests developed in recent years. We also compared Quantitative RT-PCR (q RT-PCR), Quantitative RT-RPA (rtRT-RPA) and lateral flow RT-RPA (lfRT-RPA) methods, which optimise both singleplex and duplex. As a result of this stage, the minimum detection limits of both rtRT-PCR, rtRT-RPA, and lfRT-RPA methods were determined. The lfRT-RPA test is almost as sensitive as rtRT-PCR and rtRT-RPA for the detection of duplex VHSV Ie, but for the detection of the IPNV sp., the lfRT-RPA test was found to be approximately 10-fold less sensitive than rtRT-PCR and rtRT-RPA. It is thought that the duplex lateral flow detection kit will allow timely diagnosis in the field for detecting infectious diseases of other aquaculture farms.

重组酶聚合酶扩增驱动双侧流诊断试剂盒同时检测感染性胰腺坏死病毒和病毒性出血性败血症病毒:RPA和PCR方法的比较分析
养殖业传染病的诊断时间是一个重要的问题,地方和商业企业普遍不喜欢实验室诊断传染病,因为提前期长,成本高。这导致对水产养殖场暴发的报告不足,以及当传染病传播到许多养殖场系统并造成重大经济损失时对其的认识不足。在这项研究中,我们的目标是开发一种快速、简单的诊断测试,可以很容易地在现场条件下应用于感染性胰腺坏死病毒(IPNV)和病毒性出血性败血症病毒(VHSV Ie),这两种重要的病毒病原体在 rkiye鳟鱼养殖中造成重大问题,无论是在孵化场还是养殖场。为此,我们基于近年来发展起来的分子检测技术之一重组酶聚合酶扩增技术(Recombinase polymerase amplification, RPA),开发了一种用于VHSV Ie和IPNV sp.双重诊断的横向流动试纸条。我们还比较了定量RT-PCR (q RT-PCR)、定量RT-RPA (rtRT-RPA)和横向流动RT-RPA (lfRT-RPA)方法,它们对单工和双工都进行了优化。在此阶段,确定了rtRT-PCR、rtRT-RPA和lfRT-RPA方法的最低检出限。lfRT-RPA检测双工VHSV Ie的灵敏度几乎与rtRT-PCR和rtRT-RPA相同,但对于IPNV sp的检测,lfRT-RPA检测的灵敏度比rtRT-PCR和rtRT-RPA低约10倍。认为该双侧流检测试剂盒可为其他养殖场的传染病检测提供现场及时诊断。
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来源期刊
Journal of fish diseases
Journal of fish diseases 农林科学-海洋与淡水生物学
CiteScore
4.60
自引率
12.00%
发文量
170
审稿时长
6 months
期刊介绍: Journal of Fish Diseases enjoys an international reputation as the medium for the exchange of information on original research into all aspects of disease in both wild and cultured fish and shellfish. Areas of interest regularly covered by the journal include: -host-pathogen relationships- studies of fish pathogens- pathophysiology- diagnostic methods- therapy- epidemiology- descriptions of new diseases
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