荧光PCR探针熔解曲线技术检测涂阳患者痰标本中结核分枝杆菌耐药性的价值

目的评估荧光PCR探针熔解曲线(MeltPro)技术检测涂阳患者痰标本中MTB耐药性的效能。方法收集2016年9月至2018年8月西安市胸科医院500例痰涂片阳性患者的痰标本,分别进行痰涂片镜检、GeneXpert MTB/RIF检测、BACTEC MGIT 960(简称“MGIT 960”)液体培养、表型药物敏感性试验(简称“药敏试验”),并应用MeltPro法检测痰标本中MTB菌株对利福平、异烟肼、二线注射类和氟喹诺酮类等抗结核药物的耐药性,并以表型药敏试验为标准,评价MeltPro法的检测效能。结果以表型药敏试验结果为标准,MeltPro法检测MTB对利福平耐药性的敏感度和特异度分别为98.7%(76/77)和94.2%(343/364),检测对异烟肼耐药性的敏感度和特异度分别为82.3%(102/124)和96.2%(304/316),检测对阿米卡星耐药性的敏感度和特异度分别为8/9和99.5%(432/434),检测对卷曲霉素耐药性的敏感度和特异度分别为6/7和99.1%(432/436),检测对左氧氟沙星耐药性的敏感度和特异度分别为90.6%(48/53)和99.2%(386/389),检测对莫西沙星耐药性的敏感度和特异度分别为88.1%(37/42)和96.5%(386/400)。以表型药敏试验结果为标准,GeneXpertMTB/RIF检测MTB对利福平耐药性的敏感度和特异度分别为94.8%(73/77)和94.0%(342/364)。GeneXpert-MTB/RIF和MeltPro法与表型药敏试验结果一致性均较好(Kappa值分别为0.81和0.84),但MeltPro法检测效能与表型药敏试验检测结果的一致性更高。结论MeltPro法检测涂阳患者痰标本中MTB菌株对利福平、异烟肼,以及二线注射类和氟喹诺酮类抗结核药物耐药性有较好的效能。

The value of fluorescent PCR probe melting curve technique in detecting anti-tuberculosis drug resistance of Mycobacterium tuberculosis using sputum samples from smear-positive patients

Objective To evaluate the detection performance of the fluorescent PCR probe melting curve (MeltPro) technique in detecting anti-tuberculosis drug resistance using smear-positive sputum specimens. Methods Sputum specimens from 500 sputum-positive tuberculosis patients in Xi’an Chest Hospital from September 2016 to August 2018 were collected. Sputum smear microscopy, GeneXpert MTB /RIF, BECTEC MGIT 960 liquid culture, phenotypic drug susceptibility test and MeltPro technique were applied to detect the drug resistance to rifampicin, isoniazid, second-line injection drugs and fluoroquinolones. Phenotypic drug susceptibility test was used as the reference standard to assess the detection performance. Results The sensitivity and specificity of MeltPro technique for rifampicin resistance detection were 98.7% (76/77) and 94.2% (343/364), for isoniazid were 82.3% (102/124) and 96.2% (304/316), for amikacin were 8/9 and 99.5% (432/434), for capreomycin were 6/7 and 99.1% (432/436), for levofloxacin were 90.6% (48/53) and 99.2% (386/389), and for moxifloxacin were 88.1% (37/42) and 96.5% (386/400), respectively. The sensitivity and specificity of GeneXpert MTB/RIF for detection of M.tuberculosis rifampicin resistance were 94.8% (73/77) and 94.0% (342/364), respectively. The results of both GeneXpert-MTB/RIF and MeltPro methods were well consistent with the phenotypic susceptibility test (Kappa values were 0.81 and 0.84, respectively), but the MeltPro method was more consistent. Conclusion MeltPro technique shows a good capability in detecting the drug resistance of anti-tuberculosis drugs of MTB including rifampicin, isoniazid, second-line injection drugs and fluoroquinolone using smear-positive sputum specimen.