Detection of foot-and-mouth disease virus: comparative diagnostic sensitivity of two independent real-time reverse transcription-polymerase chain reaction assays
Detection of foot-and-mouth disease virus: comparative diagnostic sensitivity of two independent real-time reverse transcription-polymerase chain reaction assays
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DOI：
10.1177/104063870601800114
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发表时间：
2006-01-01
影响因子：
1.5
通讯作者：
Beckham, TR
中科院分区：
文献类型：
--
作者：
King, DP;Ferris, NP;Beckham, TR
快速准确的诊断是有效控制口蹄疫的关键。现已认识到，逆转录聚合酶链式反应(RT-PCR)检测在临床标本中口蹄疫病毒(FMDV)的常规检测中具有十分重要的作用。本研究的目的是比较针对5‘非翻译区(5’UTR)和RNA聚合酶(3D)的两种独立的实时RT-PCR(RRT-PCR)方法检测临床标本中口蹄疫病毒的能力。从疑似口蹄疫病例的上皮组织悬液中提取的347份RNA样本中，88.1%(347份)的检测结果与两种方法的结果一致。两种试验之间的比较突出了19株口蹄疫病毒分离株(13株为5‘非编码区，6株为3D试验)，它们在I试验中没有产生信号，但在另一种试验中产生了阳性信号。选定菌株的基因组目标序列突出了引物或探针区的核苷酸替换，从而为RRT-PCR检测中产生的阴性结果提供了充分的解释。这些数据说明了持续监测流行的口蹄疫病毒野毒株的重要性，以确保RRT-PCR方法的设计符合目的，并表明使用多个诊断靶点可以进一步提高分子方法检测口蹄疫病毒的灵敏度。
Rapid and accurate diagnosis is central to the effective control of foot-and-mouth disease (FMD). It is now recognized that reverse-transcription polymerase chain reaction (RT-PCR) assays call play all important role in the routine detection of FMD virus (FMDV) in clinical samples. The aim of this study was to compare the ability of 2 independent real-time RT-PCR (rRT-PCR) assays targeting the 5' untranslated region (5'UTR) and RNA polymerase (3D) to detect FMDV in clinical samples. There was concordance between the results generated by the 2 assays for 88.1% (347 of 394) of RNA samples extracted from Suspensions of epithelial tissue obtained from Suspect FMD cases. The comparison between the 2 tests highlighted 19 FMDV isolates (13 for the 5'UTR and 6 for the 3D assay), which failed to produce a signal in I assay but gave a positive signal in the other. The sequence of the genomic targets of selected isolates highlighted nucleotide substitutions in the primer or probe regions, thereby providing ail explanation for negative results generated in the rRT-PCR assays. These data illustrate the importance of the continuous monitoring of circulating FMDV field strains to ensure the design of the rRT-PCR assay remains fit for purpose and suggest that the use of multiple diagnostic targets could further enhance the sensitivity of molecular methods for the detection of FMDV.