Unidad de Ictiopatología-Patologia Viral, Departamento de Microbiología y Parasitologia, Instituto de Acuicultura, Universidade de Santiago de Compostela, Santiago de Compostela, Spain.
Departamento de Microbiología, Facultad de Ciencias, Universidad de Malaga, Malaga, Spain.
J Appl Microbiol. 2018 Apr;124(4):977-989. doi: 10.1111/jam.13586. Epub 2017 Oct 22.
The reliability of polymerase chain reaction (PCR) techniques is an important issue in viral diagnosis, and it is even crucial when they must be applied for detection of viruses in asymptomatic carriers. The problems will arise when the aim is to study wild fish populations, where the viral loads and prevalence values are extremely low. We have evaluated several PCR procedures employed by two laboratories for monitoring fish captured in several oceanographic campaigns performed in the Gulf of Cádiz.
To evaluate the reliability of different diagnostics test used, we have re-analysed fish samples that had been previously subjected to diagnosis for a surveillance of viruses performed in 2010-2011 in wild fish populations. The following parameters were employed: the clinical sensitivity (Ss), the clinical specificity (Sp), the predictive positive value, the predictive negative value, and the positive and negative likelihood ratio (LR and LR ). For viral nervous necrosis virus, a RT-PCR procedure supplemented by nested PCR showed the highest values (100%) for all the parameters. For viral haemorrhagic septicaemia virus, the highest values were provided by RT-PCR supplemented by dot-blot hybridization. In the case of infectious pancreatic necrosis virus, none of the procedures yielded 100% for any parameter.
The results obtained for viral prevalence indicate: (i) that the conservation of the samples at -80°C did not affect to the capacity of detection of the virus in the tissues, and (ii) that the reproducibility of the diagnosis can be affected by factors including the staff experience and/or the materials employed. Finally, the use of a combination of procedures in advised to ensure the maximum reliability of the diagnosis when it is applied to asymptomatic fish populations.
This paper describes a strategy of combining diagnostic tests for the surveillance and monitoring of wild fish populations to reduce underestimation of the prevalence of viruses this type of populations.
聚合酶链反应（PCR）技术的可靠性是病毒诊断中的一个重要问题，当必须应用于无症状携带者的病毒检测时，这一点甚至至关重要。当目的是研究野生鱼类种群时，就会出现问题，因为这些病毒的载量和流行率极低。我们评估了两个实验室用于监测在加的斯湾进行的几次海洋学考察中捕获的鱼类所使用的几种 PCR 程序。
为了评估所使用的不同诊断测试的可靠性，我们重新分析了先前用于监测 2010-2011 年野生鱼类种群中病毒的鱼类样本。采用以下参数：临床灵敏度（Ss）、临床特异性（Sp）、预测阳性值、预测阴性值、阳性和阴性似然比（LR 和 LR）。对于神经坏死病毒，补充巢式 PCR 的 RT-PCR 程序显示出所有参数的最高值（100%）。对于病毒性出血性败血症病毒，RT-PCR 补充斑点杂交提供了最高值。对于传染性胰腺坏死病毒，没有任何程序的任何参数都达到 100%。
病毒流行率的结果表明：（i）-80°C 保存样品不会影响组织中病毒的检测能力；（ii）诊断的重现性可能受到包括人员经验和/或所用材料在内的因素的影响。最后，建议使用组合程序来确保将其应用于无症状鱼类种群时诊断的最大可靠性。
本文描述了一种结合诊断测试的策略，用于监测和监测野生鱼类种群，以减少对这类种群中病毒流行率的低估。
