Jiang X, Wang J, Graham D Y, Estes M K
Division of Molecular Virology, Baylor College of Medicine, Houston, Texas 77030.
J Clin Microbiol. 1992 Oct;30(10):2529-34. doi: 10.1128/jcm.30.10.2529-2534.1992.
A method of reverse transcription (RT) and polymerase chain reaction (PCR) for the detection of Norwalk virus in human stools was developed. A cationic detergent, cetyltrimethylammonium bromide, was found to effectively remove from stool extracts factors that inhibit the RT-PCR assay. The specificities of the tests were shown by hybridization of the amplified DNA with Norwalk virus-specific cDNA probes and a consistent correlation between virus detection in stools and infection of volunteers. RT-PCR detected virus in stool samples diluted 10(-4) and was about 100 times more sensitive than dot blot hybridization. In serial stool samples collected before and at different times after inoculation of 10 volunteers with Norwalk virus, 37 of 55 were positive by RT-PCR, but only 27 were positive by dot blot hybridization (chi 2 = 22.96; P less than 0.001). Further application of this method should allow detection of Norwalk virus in food or environmental samples such as shellfish and shellfish waters.
已开发出一种用于检测人类粪便中诺沃克病毒的逆转录（RT）和聚合酶链反应（PCR）方法。发现一种阳离子去污剂十六烷基三甲基溴化铵能有效去除粪便提取物中抑制RT-PCR检测的因子。通过将扩增的DNA与诺沃克病毒特异性cDNA探针杂交以及粪便中病毒检测与志愿者感染之间的一致相关性，证明了该检测方法的特异性。RT-PCR能检测到稀释至10^(-4) 的粪便样本中的病毒，其灵敏度比斑点印迹杂交高约100倍。在10名志愿者接种诺沃克病毒之前及之后不同时间收集的系列粪便样本中，55份样本中有37份通过RT-PCR呈阳性，但通过斑点印迹杂交仅27份呈阳性（χ² = 22.96；P < 0.001）。该方法的进一步应用应能检测食品或环境样本（如贝类及贝类养殖水域）中的诺沃克病毒。
