Use of a portable real-time reverse transcriptase-polymerase chain reaction assay for rapid detection of foot-and-mouth disease virus
Use of a portable real-time reverse transcriptase-polymerase chain reaction assay for rapid detection of foot-and-mouth disease virus
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DOI：
10.2460/javma.2002.220.1636
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发表时间：
2002-06-01
期刊：
影响因子：
--
通讯作者：
Nelson, WM
中科院分区：
文献类型：
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作者：
Callahan, JD;Brown, F;Nelson, WM
目的-评估一种便携式实时逆转录聚合酶链式反应 (RT-PCR) 检测方法，旨在检测口蹄疫病毒 (FMDV) 的所有 7 种病毒血清型。设计实验室和动物研究。研究群体-在组织培养中生长的病毒和实验感染 FMDV 的动物。Procedure-1 牛、猪和羊感染了血清型 0 口蹄疫病毒。 24 小时后，动物被放置在单独的房间内，其中有 4 只未感染 FMDV 的相同物种的健康动物。定期采集口腔和鼻拭子标本、用 Probang 采集的口咽标本和血液样本，并观察动物的发烧情况和口蹄疫 (FMD) 的临床症状。对来自动物和组织培养物的样本进行了感染性病毒和病毒 RNA 的检测。结果 - 该检测检测到代表组织培养物中生长的所有 7 种 FMDV 血清型的病毒 RNA，但没有扩增一组选定的病毒，其中包括那些引起与 FMD 类似的水疱病的病毒；因此，该测定的特异性为 100%，具体取决于所选的组。该测定还达到或超过了实验感染动物样本的病毒培养物的敏感性。在许多情况下，该测定在临床疾病发作前 24 至 96 小时检测到口腔和鼻子中的病毒 RNA。 结论和临床相关性 - 测定试剂以玻璃化形式生产，允许在环境温度下储存和运输。该检测在便携式仪器上可在2小时或更短的时间内完成，从而为口蹄疫检测提供了一种快速、便携、灵敏、特异的方法。
Objective-To evaluate a portable real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assay designed to detect all 7 viral serotypes of foot-and-mouth disease virus (FMDV).Design-Laboratory and animal studies.Study Population-Viruses grown in tissue culture and animals experimentally infected with FMDV.Procedure-1 steer, pig, and sheep were infected with serotype 0 FMDV. Twenty-four hours later, animals were placed in separate rooms that contained 4 FMDV-free, healthy animals of the same species. Oral and nasal swab specimens, oropharyngeal specimens obtained with a probang, and blood samples were obtained at frequent intervals, and animals were observed for fever and clinical signs of foot-and-mouth disease (FMD). Samples from animals and tissue cultures were assayed for infectious virus and viral RNA.Results-The assay detected viral RNA representing all 7 FMDV serotypes grown in tissue culture but did not amplify a panel of selected viruses that included those that cause vesicular diseases similar to FMD; thus, the assay had a specificity of 100%, depending on the panel selected. The assay also met or exceeded sensitivity of viral culture on samples from experimentally infected animals. In many instances, the assay detected viral RNA in the mouth and nose 24 to 96 hours before the onset of clinical disease.Conclusions and Clinical Relevance-The assay reagents are produced in a vitrified form, which permits storage and transportation at ambient temperatures. The test can be performed in 2 hours or less on a portable instrument, thus providing a rapid, portable, sensitive, and specific method for detection of FMDV.