反向斑点杂交法快速检测嗜肺军团菌

目的　探讨早期、快速检测嗜肺军团菌的方法。方法　根据嗜肺军团菌特异的巨噬细胞感染增强子mip基因设计引物 ,通过聚合酶链反应合成一段特异的 2 6 0bp长链DNA探针 ,采用反向斑点杂交法检测生物素标记细菌DNA ,并应用于痰标本的检测。结果　所合成的 2 6 0bpDNA探针具有高度特异性 ,只和嗜肺军团菌杂交 ,与其他细菌、真菌、病毒无交叉反应 ,该探针最低可检测出 1ng的细菌DNA。杂交法和培养法分别检测 10 0份痰标本 ,两者的阳性率分别为 8%和 2 %。结论　该方法快速、特异 ,对嗜肺军团菌的早期诊断具有较高的应用价值。

Rapid detection of Legionella pneumophila by reverse Dot- blot hybridization

Objective To explore a rapid method for the detection of legionella pneumophila. Methods A pair of primers were designed according to the mip gene of legionella pneumophila. A specific 260 bp DNA probe was synthesized by polymerase chain reaction. Bacterial DNA labeled with biotin and sputum specimens were detected by reverse Dot-blot hybridization. Results The 260 bp DNA probe synthesized was highly specific. The probe only hybridized with legionella pneumophila without cross reaction with other bacteria, fungi or viruses. The probe can detect as low as 1 ng of bacterium DNA. One hundred sputum specimens were identified with the hybridization and culture method, with a positive rate of 8% and 2% respectively. Conclusion This method is of high value in rapid and specific identification of legionella pneumophila.