PDF(Pubmed)
Abstract：
Human adenoviruses (HAdVs) are common pathogens that are associated with a variety of diseases, including respiratory tract infections (RTIs). Without reliable, fast, and cost-effective detection methods for HAdVs, patients may be misdiagnosed and inappropriately treated. To address this problem, we have developed a multiplex loop-mediated isothermal amplification (LAMP) assay for the detection of the species Human adenovirus B (HAdV-B), Human adenovirus C (HAdV-C) and Human adenovirus E (HAdV-E) that cause RTIs. This multiplexing approach is based on the melting curve analysis of the amplicons with a specific melting temperature for each HAdV species. Without the need for typing of HAdVs, the LAMP results can be visually detected using colorimetric analysis. The assay reliably detects at least 375 copies of HAdV-B and -C and 750 copies of HAdV-E DNA per reaction in less than 35 min at 60 °C. The designed primers have no in silico cross-reactivity with other human respiratory pathogens. Validation on 331 nasal swab samples taken from patients with RTIs showed a 90-94% agreement rate with our in-house multiplex quantitative polymerase chain reaction (qPCR) method. Concordance between the quantitative and visual LAMP was 99%. The novel multiplexed LAMP could be an alternative to PCR for diagnostic purposes, saving personnel and equipment time, or could be used for point-of-care testing.
摘要：
人类腺病毒（HAdV）是与多种疾病相关的常见病原体，包括呼吸道感染（RTIs）。没有可靠的,快,以及HAdV的经济有效的检测方法，患者可能会被误诊和不当治疗。为了解决这个问题,我们开发了一种多重环介导等温扩增（LAMP）检测方法，用于检测人类腺病毒B（HAdV-B），引起RTI的人腺病毒C(HAdV-C)和人腺病毒E(HAdV-E)。该多重方法基于扩增子的解链曲线分析,对于每种HAdV种类具有特定的解链温度。不需要键入HAdV，可以使用比色分析直观地检测LAMP结果。该测定在60°C下在少于35分钟内每个反应可靠地检测至少375个拷贝的HAdV-B和-C以及750个拷贝的HAdV-EDNA。设计的引物与其他人类呼吸道病原体没有计算机交叉反应性。从RTI患者中采集的331个鼻拭子样本的验证显示，与我们的内部多重定量聚合酶链反应（qPCR）方法的符合率为90-94％。定量和视觉LAMP之间的一致性为99%。新型多重LAMP可以替代PCR用于诊断目的，节省人员和设备时间，或可用于即时测试。
