分枝杆菌属基因限制性内切酶谱分析

Restriction enzyme pattern analysis of Mycobacteria genosome

目的探讨基因限制性内切酶谱分析对分枝杆菌属检出的敏感性、特异性和准确性。并达到快速准确检出结核分枝杆菌和非结核分枝杆菌属的目的。方法首先以分枝杆菌属公共分子量６５０００抗原蛋白的基因，应用聚合酶链反应（ＰＣＲ）扩增编码其基因，此基因为各分枝杆菌所共有。扩增产物再分别经ＢｓｔＥⅡ和ＨａｅⅢ限制性内切酶酶切后的图型模式（ＰＲＡ），鉴定临床标本的结核分枝杆菌及非结核性分枝杆菌。结果ＰＲＡ法检测的灵敏度高。检出了２３株分枝杆菌标准菌株，得出特异的基因图谱。结核分枝杆菌复合体的酶切模式有其独特之处。并检测临床标本３２３份，其中痰标本２０６份，胸水４２份，尿液３５份。内切酶确诊结核分枝杆菌的阳性率分别为６７．９％，４８．０％，５．７％。结论对分枝杆菌采用限制性内切酶谱分析，检出结核和非结核分枝杆菌快速准确，并可提高阳性率，降低假阴性。

Objective\ To investigate the sensitivity, accuracy and specificity of the gene restrictive enzyme pattern analysis inorder to identify mycobacterium complex quickly and accurately. Methods The 65kD antigen protein gene which was shared by mycobacterium was amplified and encoded by polymerase chain reaction(PCR) method. Then the PCR products were cleavaged by restriction enzyme BstⅡ and Hae Ⅲ. By this way,the clinical samples of M. tuberculosis and M. non tuberculosis were identified. [WT5”HZ]Results The special gene maps were obtained by comparing with 23 reference strains of mycobacterium with PCR restriction enzyme pattern analysis (PRA). The species of 322 clinical isolates have been determined. The positivity rate in sputum, pleural fluid and urine was 67.9%, 48.0%, 5.7%, respectively. Conclusions This results indicate that the method for differential diagnosis between M. tuberculosis and others with detection PRA is quick and accurate.