开发和评估一种环介导等温扩增检测方法，用于在现场条件下快速检测猪巨细胞病毒。
Development and evaluation of a loop-mediated isothermal amplification assay for the rapid detection of porcine cytomegalovirus under field conditions.
机构信息
Animal Science College of Guizhou University, Guiyang, Guizhou, China.
出版信息
Virol J. 2012 Dec 29;9:321. doi: 10.1186/1743-422X-9-321.
BACKGROUND
Porcine cytomegalovirus (PCMV) induces silent infection in adult pigs but more frequently causes fatal, generalized infection in newborn piglets. This study aimed to develop a new loop-mediated isothermal amplification (LAMP) method for the sensitive, rapid, and inexpensive detection of PCMV under field conditions.
METHODS
Tissue obtained from nine-week-old PCMV-free Landrace pigs or pig samples from postmortem examinations were analyzed. The samples were found to have clinical signs and lesions consistent with inclusion body rhinitis. Six specific primers were designed by targeting the PCMV DNA polymerase (DPOL) DNA. The LAMP reaction was optimized in a water bath. The sensitivity and specificity of LAMP and polymerase chain reaction (PCR) were compared.
RESULTS
PCMV DNA was amplified at 65°C, and the result could be detected as early as 30 min into the reaction. Positive reactions could be visualized by the naked eye as a color change brought on by the addition of SYBR Green. The sensitivity and specificity of LAMP were found to be similar to those of the PCR.
CONCLUSIONS
LAMP is a high-throughput technique for the detection of PCMV and has a high specificity, sensitivity and simplicity; these factors make it suitable for detection of PCMV under field conditions.
背景
猪巨细胞病毒（PCMV）在成年猪中引起潜伏感染，但在新生仔猪中更常引起致命的全身性感染。本研究旨在开发一种新的环介导等温扩增（LAMP）方法，用于在现场条件下灵敏、快速和廉价地检测 PCMV。
方法
分析了来自 9 周龄无 PCMV 的长白猪的组织或尸检后的猪样本。这些样本具有与包涵体鼻炎相一致的临床症状和病变。通过针对 PCMV DNA 聚合酶（DPOL）DNA 设计了六个特定引物。在水浴中优化了 LAMP 反应。比较了 LAMP 和聚合酶链反应（PCR）的灵敏度和特异性。
结果
在 65°C 下扩增 PCMV DNA，反应 30 分钟即可检测到结果。通过添加 SYBR Green 引起的颜色变化，可以通过肉眼观察到阳性反应。LAMP 的灵敏度和特异性与 PCR 相似。
结论
LAMP 是一种用于检测 PCMV 的高通量技术，具有高特异性、灵敏度和简单性；这些因素使其适合在现场条件下检测 PCMV。
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