比较聚合酶链反应（PCR）和噬菌斑法检测和计数污染海水中的噬菌体。
Comparison of PCR and plaque assay for detection and enumeration of coliphage in polluted marine waters.
出版信息
Appl Environ Microbiol. 1997 Nov;63(11):4564-6. doi: 10.1128/aem.63.11.4564-4566.1997.
A total of 68 marine samples from various sites impacted by sewage and storm waters were analyzed by both the plaque assay and a reverse transcriptase (RT) PCR technique for F(sup+)-specific coliphage. The coliphage levels detected by the plaque assay averaged 1.90 x 10(sup4) PFU/100.0 ml. Using a most probable number (MPN) PCR approach, the levels averaged 2.40 x 10(sup6) MPN-PCR units/100.0 ml. Two samples were positive by RT-PCR but negative by plaque assay, and 12 samples were positive by plaque assay but negative by RT-PCR (levels lower than 11.00 PFU/100.0 ml). The host system used for the plaque assay may detect somatic coliphage in addition to the F(sup+)-specific coliphage. When it is used as an indicator of pollution, contamination may be missed with more restrictive systems. The difference in results may be due to the sensitivity, specificity, or inhibition of RT-PCR in marine samples. This study provides information on quantifying PCR results by an MPN method and insights into interpretation of PCR data for detection of viruses in marine environments.
共分析了来自受污水和雨水影响的不同地点的 68 个海洋样本，分别采用噬菌斑法和逆转录（RT）PCR 技术检测 F（sup+）-特异性噬菌体。噬菌斑法检测到的噬菌体水平平均为 1.90 x 10（sup4）PFU/100.0 ml。使用最可能数（MPN）PCR 方法，水平平均为 2.40 x 10（sup6）MPN-PCR 单位/100.0 ml。有两个样本通过 RT-PCR 呈阳性但噬菌斑法呈阴性，有 12 个样本噬菌斑法呈阳性但 RT-PCR 呈阴性（水平低于 11.00 PFU/100.0 ml）。噬菌斑法所用的宿主系统可能除了 F（sup+）-特异性噬菌体外，还能检测到体细胞噬菌体。当它被用作污染指标时，更严格的系统可能会错过污染。结果的差异可能是由于 RT-PCR 在海洋样本中的灵敏度、特异性或抑制作用。本研究提供了通过 MPN 方法量化 PCR 结果的信息，并深入了解了海洋环境中病毒检测的 PCR 数据解释。
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