应用套式PCR检测嗜肺军团杆菌的实验研究

Detection of Legionella Pneumophila by Nested Polymerase Chain Reaction

应用套式聚合酶链反应（ＰＣＲ）检测１～６型嗜肺军团杆菌，两轮ＰＣＲ都分别出现了９９６ｂｐ和６５０ｂｐ的特异性扩增产物；而金黄色葡萄球菌等七种细菌均未出现物异性扩增。两轮ＰＣＲ分别可以检测１０ｐｇ和１０ｆｇ的模板ＤＮＡ。８份嗜肺军团杆菌含量分别为１～１０￣７ｃｆｕ／ｍｌ的痰感染模拟标本均出现了特异性扩增产物。提示该方法可用于临床，特别是检测含微量嗜肺军团杆菌的临床标本。

e amplified the samples of legionella pneumophila serogroups 1 to 6.The specific amplificationbands of 996bp in the first step and 650bp in the second step were observed respectively. Samples ofother 7 bacteria, including staphylococcus aureus, didn’t generate positive signals.The first and thesecond step PCR achieved the sensitivity as small as 10pg and 10fg of the target DNA respectively. Eight sputum samples which had been seeded with 1～10￣7 cfu/ml of legionella pneumophila were all pos-itive after nested PCR amplification. We demonstrated that the nested PCR method was superior in sen-sitivity and specificity for the detection of legionella pneumophila. The results suggested that thismethod could be applied in clinic, especially for detecting samples containing minute LP.