摘要
目的:探索实时荧光定量PCR(qRT-PCR)联合玻璃珠磨菌法对检测不同临床标本中结核分枝杆菌菌载量的优化作用。方法:回顾性搜集2008年1月1日至2016年12月31日首都医科大学附属北京胸科医院住院和门诊诊治的疑似结核病患者11570份不同临床标本的qRT-PCR检测结果并进行分析,主要包括痰液1487份、脓液273份、脑脊液1391份、气管(支气管)肺泡灌洗液41份和血液4650份等。再通过建立伪随机函数的方法从中选取82份临床诊断为肺结核的痰液标本,对其进行玻璃珠(直径0.3 mm)振荡磨菌前后qRT-PCR定量检测效果分析。结果:11570份不同临床标本的总检测阳性率为12.16%(1407/11570)。其中,气管(支气管)灌洗液的检出率最高[36.59%(15/41)],以门诊患者来源的阳性率最高[43.48%(10/23)];其次是脓液标本[35.53%(97/273)],以住院患者来源的阳性率最高[36.26%(95/262)];痰液标本位居第三[30.33%(451/1487)];其他标本阳性率介于10.43%(485/4650)~15.03%(209/1391)之间。对选取的82份痰标本行5 min的玻璃珠磨菌处理后,qRT-PCR检测的核酸浓度[44.45(9.05,71.62)×10^(7)拷贝/ml]较处理前[4.03(1.87,5.92)×10^(7)拷贝/ml]明显提高,差异有统计学意义(Z=2.120,P=0.016)。其中,有28份(34.15%)痰标本提高了菌株数量级,尤其是21份含菌量为10^(2)拷贝/ml的临床痰标本经振荡研磨后有14份(66.67%)提升到了10^(3)拷贝/ml菌量数量级,使阳性检出率由磨菌处理前的60.98%(50/82)提高到磨菌处理后的78.05%(64/82)。结论:qRT-PCR对灌洗液、脓液和痰液标本中结核分枝杆菌的检出率高,在其他标本中检出率低。采用玻璃珠振荡磨菌处理后可提高临床标本的阳性检出率。
Objective:To explore a method with improved efficiency for quantitatively detecting Mycobacterium tuberculosis(MTB)in different type of clinical samples by combining glass beads grinding with real-time quantitative PCR(qRT-PCR).Methods:A retrospective study was performed in Beijing Chest Hospital.Data of qRT-PCR examination performed between Jan.1st,2008 and Dec.31th 2016 were retrieved.The outcomes of 11570 clinical specimens collected from tuberculosis suspects of inpatients and outpatients were acquired,including 1487 sputa,273 purulent specimens,1391 cerebrospinal fluids(CSF),41 bronchoalveolar lavage fluids(BALF),and 4650 blood samples.Furthermore,82 specimens collected from pulmonary tuberculosis patients were chosen by pseudorandom function,the quantitative outcomes were compared between the procedures with or without grinding with glass beads(diameter 0.3 mm).Results:The overall positive rate of the different clinical specimen types was 12.16%(1407/11570).Among them,BALF acquired the highest positive rate(36.59%(15/41)),and the outpatients had higher positive rate(43.48%(10/23)).The second highest positive rate was among purulent specimens(35.53%(97/273)),of which the inpatients yielded higher positive rate(36.26%(95/262)).The positive rate of sputa ranked the third(30.33%(451/1487)).The positive rate of the other leftover samples ranged over 10.43%(485/4650)-15.03%(209/1391).In contrast to the method without grinding,the nucleic acid quantity of the specimen grinded by class beads increased evidently(44.45(9.05,71.62)×10^(7) copy/ml vs.4.03(1.87,5.92)×10^(7) copy/ml;Z=2.120,P=0.016).Additionally,28(34.15%)samples produced one order of magnitude increase outcomes,66.67%(14/21)samples from 10^(2) copy/ml to 10^(3) copy/ml by using glass bead grinding.Hence,the positive rate of the selected 82 specimens increased from 60.98%(50/82)to 78.05%(64/82).Conclusion:qRT-PCR had higher detection rates in BALF,pus and sputa than other sample types.Grinding specimen by glass bead could improve the positive detection
作者
戴广明
王芬
曹廷明
褚洪迁
黄海荣
孙照刚
DAI Guang-ming;WANG fen;CAO Ting-ming;CHU Hong-qian;HUANG Hai-rong;SUN Zhao-gang(Translational Medicine Center,Beijing Chest Hospital,Capital Medical University,Beijing 101149,China;Beijing Key Laboratory in Drug Resistant Tuberculosis Research,Beijing Tuberculosis and Thoracic Tumor Research Institute,Beijing 101149,China;National Tuberculosis Clinical Laboratory,Beijing Chest Hospital,Capital Medical University,Beijing 101149,China)
出处
《中国防痨杂志》
CAS
CSCD
2022年第5期450-454,共5页
Chinese Journal of Antituberculosis
基金
北京市教育委员会科研计划项目(KZ202110025034)
北京市科委首都特色基金重点项目(Z211100002921002)。
关键词
分枝杆菌
结核
分子诊断技术
实验室技术和方法
评价研究
Mycobacterium tuberculosis
Molecular diagnostic techniques
Laboratory techniques and procedures
Evaluation studies
