Tewari Deepanker, Zellers Corey, Acland Helen, Pedersen Janice C
Microbiology Services, Pennsylvania Veterinary Laboratory, Harrisburg, PA 17110, USA.
J Clin Virol. 2007 Oct;40(2):142-5. doi: 10.1016/j.jcv.2007.07.011. Epub 2007 Sep 17.
Highly pathogenic H5N1 avian influenza (AI) poses a grave risk to human health. An important aspect of influenza control is rapid diagnosis.
This study describes the efficiency of AI-RNA extraction utilizing silica-based magnetic beads with robotics and its detection with an influenza A matrix gene real-time RT-PCR from tracheal swabs, and compares it to virus isolation and manual spin column extractions.
Analytical sensitivity was assessed by performing dilution analysis and detection of H2N2 AI viral RNA. Diagnostic sensitivity and specificity was assessed by analyzing tracheal swabs collected from H7N2 infected and uninfected chickens.
Both manual and robotic extractions detected AI virus at 1log(10)EID(50)/ml. Diagnostic sensitivity and specificity of matrix gene detection with the automated extraction method for chicken tracheal swab specimens was similar to that of virus isolation and the manual extraction method. There were only three discordant results among 212 tested specimens.
The main advantages of automated robotic viral nucleic acid extraction are high throughput processing; hands-free operation; and reduction in human and technical error. This study demonstrates successful detection of influenza A virus with magnetic beads utilizing the Qiagen MagAttract cell kit on a BioRobot M48 platform.
高致病性H5N1禽流感对人类健康构成严重威胁。流感防控的一个重要方面是快速诊断。
本研究描述了利用基于硅胶的磁珠结合机器人技术进行禽流感病毒RNA提取的效率，以及用甲型流感病毒基质基因实时逆转录聚合酶链反应（RT-PCR）从气管拭子中进行检测的情况，并将其与病毒分离和手动旋转柱提取法进行比较。
通过进行稀释分析和检测H2N2禽流感病毒RNA来评估分析灵敏度。通过分析从感染和未感染H7N2的鸡采集的气管拭子来评估诊断灵敏度和特异性。
手动和机器人提取法均能检测到1log(10)EID(50)/ml的禽流感病毒。鸡气管拭子标本自动提取法检测基质基因的诊断灵敏度和特异性与病毒分离法和手动提取法相似。在212份检测标本中仅有3个不一致的结果。
自动化机器人病毒核酸提取的主要优点是高通量处理、无需人工操作以及减少人为和技术误差。本研究证明了在BioRobot M48平台上利用Qiagen MagAttract细胞试剂盒通过磁珠成功检测甲型流感病毒。
