【摘要】 目的评价滚环扩增技术(rolling circle amplification,RCA)对痰标本中结核分枝杆菌利福平敏感性的直接快速检测效果。方法选取首都医科大学附属北京胸科医院住院的初治和复治涂阳肺结核患者24h痰标本32份,采用RCA法、直接测序法及传统药敏试验绝对浓度法测定痰标本中结核分枝杆菌对利福平的敏感性,并进行了比较。结果 32份痰标本中结核分枝杆菌,采用RCA法24株可检测到rpo B基因突变,其中23株单位点突变(516位2株,526位12株,531位9株),1株516位和526位联合突变,与直接测序结果均一致。32份痰标本的RCA方法检测与DNA测序结果和药物敏感性试验进行对比,28株结果均一致。4株结果不一致,其中2株RCA及测序无突变,药敏结果显示为耐药;2株RCA及测序显示有突变,药敏结果显示为敏感。结论RCA法用于痰标本中直接检测结核分枝杆菌对利福平的敏感性,同直接测序法结果一致,同传统药敏试验相比,RCA法能够大幅度节省时间,具有很好的临床应用和推广前景。更多还原

【Abstract】 Objective To evaluate the detection capability of direct and rapid of rolling circle amplifi cation(RCA) for rifampin(RIF) susceptibility of Mycobacterium tuberculosis. Methods By using the selected sputum specimens of 32 initial treatment and re-treatment sputum smear(SS+)pulmonary TB cases hospitalized in Beijing Chest Hospital, utilizing the RCA, direct DNA sequencing and conventional drug susceptibility testing absolute concentration method, detect the Mycobacterium tuberculosis sensitivity to RIF, furthermore compare the results from above three measurements. Results Of 32 Mycobacterium tuberculosis isolates, by using of RCA, 24 had mutations in the amplifi ed region of rpo B. 23 strains of mono point mutations, there were 2 occur at codon 516, 12 occur at codon 526 and 9 occur at codon 531, 1 strains 516 and 526-bit double locus mutation. The results are consistent with the results of sequencing. Compared to the results of three methods of 32 sputum specimens, 28 strains were consistent. There were 4 cases inconsistent, among them 2 strains of RCA and sequencing without mutations, drug susceptibility results showed resistance; 2 strains of RCA and sequencing showed mutations, drug susceptibility results showed that for sensitive. Conclusion In the sputum specimens RCA method used for the direct detection of Mycobacterium tuberculosis sensitivity to RIF has the same equal effects as sequencing. Compared to the conventional drug susceptibility testing, the RCA method can greatly save time, moreover it possess good clinical application and prospects.更多还原