用DNA探针检测肺炎衣原体

DETECTION OF CHLAMYDIA PNEUMONIAE BY DNA PROBE BLOT

目的建立一种敏感而特异的肺炎衣原体分子生物学检测方法。方法用ＰＣＲ扩增约４６０ｂｐ的肺炎衣原体种特异性基因片段并标记成探针，建立ＤＮＡ探针杂交检测肺炎衣原体的方法。结果：探针只与肺炎衣原体ＡＲ－３９、ＶＲ１３１０株ＤＮＡ呈阳性杂交斑点，与肺炎衣原体ＡＲ－３９株全ＤＮＡＰｓｔＩ酶切片段在２．５ｋｂ处有一阳性杂交带，与其它两种衣原体、Ｑ热立克次体、嗜肺军团菌、大肠杆菌、绿脓杆菌、金黄色葡萄球菌ＤＮＡ斑点膜无阳性杂交信号。从１５２例有呼吸道疾患的住院病人鼻咽拭子和肺泡灌洗液中检出阳性１８例，阳性率１２％。结论本文建立的ＤＮＡ探针检测肺炎衣原体方法具有较高的敏感性和特异性，可用于批量临床标本的检测。

Aim To establish a sensitive and specific molecular biological method for detecting Chlamydia pneumoniae of respiratory tract infection patients Methods A DNA blot assay was developed by coating DNA of Chlamydia pneumoniae and/or digested from clinical samples on nitrocellulose(NC)membrane,blotting with a DNA probe labeled with DIG Results\ There were no positive blotting dot in Chlamydia trachomatis,Chlamydia psittaci,Coxiella burnetii,Legionella pneumophilia,Esherichia coli,Pseudomonas aeruginosa and Staphylococcus aureus except Chlamydia pneumoniae The sensitivity could be improved lpg The positive rate was 12%(18/153)in detection of nasopharyngeal swabs and bronchoalveolar lavage samples collected from 152 pneumonic hospitalized patients Conclusions\ This method was not only sensitive,rapid and specific but also could be applied to detect batch samples