金胺O染色阳性痰涂片刮削物的GeneXpert MTB/RIF与实时荧光定量PCR检测研究  

Applying GeneXpert MTB/RIF assay and real-time fluorescence quantitative PCR into detection of Mycobacterium tuberculosis in sputum smear scraps stained with gold amine O

作　　者：王楠[1] 邓丽 牛群[1] 谢贝[1] 吴玲[1] 刘志辉[1] 孟繁荣[1] WANG Nan;DENG Li;NIU Qun;XIE Bei;WU Ling;LIU Zhi-hui;MENG Fan-rong(Institute of Pulmonary Diseases,Guangzhou Chest Hospital,Guangzhou,Guangdong 510095,China)

机构地区：[1]广州市胸科医院肺部疾病研究所,广东广州510095

出　　处：《热带医学杂志》2022年第10期1330-1333,共4页Journal of Tropical Medicine

基　　金：广东省自然科学基金(2018A030313550);广州市卫生健康科技项目(20201A011042);广州市高水平临床重点专科和培育专科建设项目(穗卫函[2019]1555号);广州市医学重点学科(结核病学)建设项目

摘　　要：目的探讨以金胺O染色阳性痰涂片刮削物为样本进行GeneXpert MTB/RIF与结核分枝杆菌(MTB)实时荧光定量PCR检测的可行性。方法刮取金胺O染色阳性1+、2+、3+和4+痰涂片50、30、20、20例,以GeneXpert MTB/RIF检测试剂盒检测刮削物结核菌利福平药物敏感性并计算结核菌检测阳性率。刮取金胺O染色阳性1+、2+、3+和4+痰涂片痰膜各50例于离心管中,实验组以1 mL 4%NaoH处理、对照组以1 mL无菌生理盐水处理后提取沉淀物基因组DNA,以结核分枝杆菌实时荧光定量PCR检测试剂盒检测其拷贝数。采用χ^(2)检验比较实验组和对照组结核菌阳性检出率差异。结果金胺O染色阳性1+、2+、3+和4+痰涂片刮削物GeneXpert MTB/RIF检测阳性率分别为58.00%、96.69%、100.00%、100.00%,其中1+、2+和4+痰涂片刮削物中检测到利福平耐药结核菌。金胺O染色阳性1+、2+、3+、4+痰涂片刮削物MTB实时荧光定量PCR检测阳性率,实验组分别为60.00%、38.00%、64.00%、90.00%,对照组分别为76.00%、80.00%、84.00%、94.00%;其中2+和3+样本阳性率实验组与对照组之间差异有统计学意义(χ^(2)=10.395、5.198,P均<0.05)。结论直接以金胺O染色阳性痰涂片刮削物为样本进行GeneXpert MTB/RIF检测和实时荧光定量PCR检测实际可行。Objective To explore the feasibility of applying GeneXpert MTB/RIF assay and real-time fluorescence quantitative PCR in detection of Mycobacterium tuberculosis(MTB)in sputum smear scraps stained with gold amine O.Methods Sputum smears stained by gold amine O with 1+(n=50),2+(n=30),3+(n=20),and 4+(n=20)positive results were scraped into tubes;the MTB rifampicin sensitivity were analysed through GeneXpert MTB/RIF test.Sputum smears stained by gold amine O with 1+,2+,3+,and 4+positive results,each with 50 slices,were scraped into tubes.Experimental group were treated with 1 mL 4%NaoH,while the control group with 1 mL sterile normal saline,and then genome DNA were extraced.The copy number of MTB genome DNA was detected by real-time fluorescence quantitative PCR kit,and the positive rate was calculated.Chi-square test was applied in exploring the difference of MTB positive rates between the experimental and the control groups.Results GeneXpert MTB/RIF test results indicated the MTB positive rate was 58.00%in 1+,96.69%in 2+,100.00%in 3+and 100.00%in 4+sputum smears.Rifampicin-resistant Mycobacterium tuberculosis was detected in 1+,2+and 4+sputum smears.Real-time fluorescence quantitative PCR results indicated the positive rates of MTB in 1+,2+,3+and 4+sputum smear were 60.00%,38.00%,64.00%and 90.00%in the experimental group,respectively,and 76.00%,80.00%,84.00%and 94.00%in the control group respectively.The differences of positive rates between experimental and control groups in sputum smears of 2+and 3+were significant(χ^(2)=10.395,5.198,all P<0.05).Conclusion It was feasible to detect MTB in sputum smears stained with positive gold amine O by GeneXpert MTB/RIF assay and real-time fluorescence quantitative PCR.

关 键 词：结核分枝杆菌 痰涂片 GeneXpert MTB/RIF 实时荧光定量PCR 

分 类 号：R52[医药卫生—内科学] R446.5[医药卫生—临床医学]