聚合酶螺旋反应检测肺炎克雷伯菌与临床样本评价  

Suvery and Rapid Detection of Klebsiella Pneumoniae in Clinical Samples by Polymerase Spiral Reaction

作　　者：王泉 马瑞瑛 杨婷[1] 张亚丽 许苗 撒玉玲 陈清波 WANG Quan;MA Ruiying;YANG Ting;Zhang Yali;XU Miao;SA Yuling;CHENG Qingbo(Department of Laboratory Medicine,The Eighth Affiliated Hospital of Xinjiang Medical University,Xinjiang Urumqi 830049)

机构地区：[1]新疆医科大学第八附属医院检验科,新疆乌鲁木齐830049

出　　处：《生物化工》2021年第4期38-42,共5页Biological Chemical Engineering

摘　　要：目的:筛选聚合酶螺旋反应(Polymerase Spiral Reaction,PSR)检测呼吸道感染患者痰液中肺炎克雷伯菌的引物并进行初步应用。方法:基于肺炎克雷伯氏菌外膜蛋白基因设计PSR反应引物并进行筛选,对菌株特异性及最低检出限进行评估;于2019年9—12月从新疆医科大学第八附属医院连续收集270份疑似呼吸道感染患者的痰液样本,对痰标本进行细菌培养鉴定,PCR及实时荧光定量PCR检测。分别以痰培养及实时荧光定量PCR检测结果为参考,评价PSR检测痰液样本中肺炎克雷伯菌的检测效能。结果:实验结果表明该方法最低检出限可达到10^(2) CFU/mL,与其他18种常见致病菌无交叉反应。270例疑似呼吸道感染患者的临床样本痰培养法检出肺炎克雷伯菌阳性54例,PSR方法检测阳性68例。对比痰培养结果,PSR检测痰液样本中肺炎克雷伯菌的敏感度和特异度分别为96.30%(52/54),92.59%(200/216)。结合测序结果,其阳性预测值与阴性预测值分别为97.1%(68/70)及99.01%(200/202),与痰培养检测结果一致性较好(Kappa值为0.81)。此外,对比荧光定量PCR法结果,其敏感度、特异度分别为100%(62/62)和96.63%(201/208)。结论:PSR在痰液样本呈现了较高的检测性能,适合使用在临床上对肺炎克雷伯菌的快速检测。Objective:Selecting the primer and preliminary applicating of PSR to detect Klebsiella pneumoniae in sputum samples of patients with respiratory infection.Methods:The PSR primers targeting the specific genes KPN_04473 of K.pneumoniaewere designed,and the specificity,limit of detection were evaluated as well.A total of 270 sputum samples,collected from suspected respiratory infection patients by The Eighth Affiliated Hospital of Xin Jiang Medical University during September to December.2019,were identified by sputum culture,PSR and real-time PCR.The performance in detecting clinical samples of PSR was evaluated according to the results of aforementioned two methods,respectively.Results:The limit detection of PSR was found to be 10^(2) CFU/mL.Further experiments showed that the primer presented good specificity for detection of the targets,without cross reaction 18 other pathogens.In cases of 270 suspected respiratory infection samples,54 samples were positive for K.pneumoniae by sputum culture and 68 samples were positive by PSR method.Compared with the sputum culture,the sensitivity,specificity of PSR were 96.30%(52/54),92.59%(200/216).In combination with sequencing results,positive predictive value and negative predictive value were 97.1%(68/70)and 99.01%(200/202),respectively.The results were basically consistent with the sputum culture method(Kappa=0.81).Compared with real-time PCR,the sensitivity,specificity of PSR were 98.39%(61/62)and 96.63%(201/208).Conclusion:PSR presented high performance in detecting K.pneumoniae from sputum sample and was suitable for clinical use.

关 键 词：呼吸道感染 聚合酶螺旋反应 肺炎克雷伯菌 快速检测 

分 类 号：R446.5[医药卫生—诊断学]