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中国防痨杂志 ›› 2018, Vol. 40 ›› Issue (12): 1291-1295.doi: 10.3969/j.issn.1000-6621.2018.12.010

• 论著 • 上一篇    下一篇

术后标本行分子生物学检测对结核病患者诊疗的价值

温子禄,王琳,王军,陈辉,李洪伟,朱益军,蒋韶宁,张舒林,宋言峥   

  1. 201508 上海市(复旦大学附属)公共卫生临床中心科学研究部(温子禄、张舒林),胸外科(王琳、王军、陈辉、李洪伟、朱益军、蒋韶宁、宋言峥);
  • 收稿日期:2018-10-26 出版日期:2018-12-10 发布日期:2018-12-10
  • 基金资助:
    “十三五”国家科技重大专项(2017ZX10201301-003-002);上海市公共卫生临床中心院级科研课题(KY-GW-2018-19)

The value of molecular biological detection of postoperative specimens in the diagnosis and treatment of tuberculosis patients

WEN Zi-lu,WANG Lin,WANG Jun,CHEN Hui,LI Hong-wei,ZHU Yi-jun,JIANG Shao-ning,ZHANG Shu-lin,SONG Yan-zheng.   

  1. Department of Scientific Research, Shanghai Public Health Clinical Center, Institutes of Biomedical Sciences, Fudan University, Shanghai 201508, China
  • Received:2018-10-26 Online:2018-12-10 Published:2018-12-10

摘要:

目的 初步探讨分子生物学检测技术在结核病患者术后快速诊断中的价值。方法 回顾性分析2015年1月至2018年2月期间在上海市公共卫生临床中心胸外科接受手术治疗、临床资料齐全的170例结核病患者。170例患者中,男100例(58.82%)、女70例(41.18%); 1~18岁年龄组18例(10.59%)、19~65岁年龄组142例(83.53%)、66~82岁年龄组10例(5.88%);肺结核39例(22.94%)、肺外结核131例(77.06%)。取术后标本采用PCR膜条杂交及二代测序技术的分子生物学方法进行菌种鉴定和耐药相关基因检测。所有患者术前进行晨痰及穿刺活检,或者术中取样进行抗酸染色涂片检测。在进行痰涂片的同时,进行分枝杆菌改良罗氏固体培养、BACTEC MGIT 960快速培养及药物敏感性试验(简称“药敏试验”)。结果 (1)痰液抗酸染色阳性率为9.41%(16/170),其中肺结核患者的阳性率为23.08%(9/39), 而穿刺活检或者术中取样(脓液或分泌物等)的抗酸染色阳性率为51.76%(88/170)。(2)分枝杆菌培养阳性率为27.65%(47/170),其中耐药结核病检出率为14.12%(24/170)。(3)术后标本分子生物学菌种鉴定阳性率为51.76%(88/170);PCR膜条杂交显示,7例为NTM,4例培养联合MPB64抗原检测结果显示为NTM,但未鉴定到菌种;3例培养结果为阴性,无法进行MPB64抗原检测。7例标本进行了分子生物学菌种鉴定,1例为瘰疬分枝杆菌、戈登分枝杆菌复合感染,3例为胞内分枝杆菌, 1例为海分枝杆菌, 1例为鸟分枝杆菌,1例为脓肿分枝杆菌。(4)除去7例NTM感染患者,剩余的163例进行了分子生物学耐药基因检测,对一线抗结核药物耐药基因的检出率为54.60%(89/163), 对二线抗结核药物耐药基因的检出率为49.69%(81/163);与培养后药敏试验结果相比,药敏试验阳性的标本中有29.79%(14/47)在分子生物学耐药基因检测时的结果为阴性,而分子生物学耐药基因检测与药敏试验检测结果阳性的一致性为90.00%(27/30);分子生物学耐药基因检测与BACTEC MGIT 960快速培养药敏试验联合判断对耐药结核病的检出率为63.19%(103/163)。结论 分子生物学检测技术在术后标本的快速诊断中具有重要的价值,可区分结核分枝杆菌、非结核分枝杆菌及是否耐药,为结核病患者术后进一步诊治提供重要参考。

关键词: 结核, 外科手术, 分子诊断技术, 实验室技术和方法, 对比研究, 结果评价(卫生保健)

Abstract:

Objective To preliminarily explore the value of molecular biological detection technology in the rapid diagnosis of postoperative tuberculosis patients.Methods We conducted a retrospective analysis of 170 patients with tuberculosis who underwent surgical treatment and had complete clinical data in the Shanghai Public Health Clinical Center from January 2015 to February 2018. Of the 170 patients, 100 (58.82%) were male and 70 (41.18%) were female; there were 18 cases (10.59%) in the 1 to 18 age group, 142 cases (83.53%) in the 19 to 65 age group, and 10 cases (5.88%) in the 66 to 82 age group. A total of 39 patients (22.94%) were pulmonary tuberculosis, and 131 patients (77.06%) had extrapulmonary tuberculosis. PCR membrane hybridization and second-generation sequencing techniques were applied for strain identification and drug resistance-related gene detection in postoperative specimens. All patients underwent acid-fast staining in sputum, needle biopsy before surgery, or intraoperative sampling, as well as Roche solid culture, BACTEC-MGIT 960 rapid culture, and drug susceptibility test in sputum.Results (1) The positive rate of acid-fast staining of sputum was 9.41% (16/170), and the positive rate of acid-fast staining of pulmonary tuberculosis sputum was 23.08% (9/39), while the positive rate of acid-fast staining was 51.76% (88/170) in the biopsy or intraoperative sampling. (2) The positive rate of mycobacterial culture was 27.65% (47/170); the positive rate of phenotypic susceptibility test was 14.12% (24/170). (3) The positive rate of postoperative molecular biology species identification was 51.76% (88/170). The result of PCR membrane Hybridization showed that 7 cases were identified as non-tuberculous mycobacteria; 4 cases were also identified as non-tuberculous mycobacteria by culture combined with MPB64 antigen detection, but 3 cases were not identified as non-tuberculous mycobacteria by culture method and could not be detected by MPB64 antigen detection. The seven specimens identified for strain identification by molecular biological detection were one case with co-infection of Mycobacterium phlei and Mycobacterium gordon, three cases of intracellular mycobacteria, one case of Mycobacterium marinum, one case of Mycobacterium avium, and one with Mycobacterium abscessus. (4) The drug sensitivity of the remaining 163 cases were detected by molecular biological method. The detection rate of first-line anti-tuberculosis drug resistance genes was 54.60% (89/163), while the rate was 49.69% (81/163) in the second-line anti-tuberculosis drugs. Compared with the results of drug sensitivity test after culture, 29.79% (14/47) of the samples with positive drug susceptibility test were negative in the detection of resistance gene by molecular biological technology. The consistency of molecular biological resistance gene detection and drug susceptibility test was 90.00% (27/30). The molecular biological and BACTEC-MGIT 960 rapid culture drug sensitivity test composite detection rate was 63.19% (103/163).Conclusion Molecular biological detection technology is of great value in the rapid diagnosis of postoperative specimens. It can distinguish between Mycobacterium tuberculosis and non-tuberculous mycobacteria and can identify whether the strain is drug resistant, which provides an important reference for further diagnosis and treatment of tuberculosis patients.

Key words: Tuberculosis, Surgical procedures, operative, Molecular diagnostic techniques, Laboratory techniques and procedures, Comparative study, Outcome assessment (health care)