Application value of molecular pathology in diagnosis of cervical lymph node tuberculosis and determination of drug resistance

doi:10.3969/j.issn.1000-6621.2018.11.006

Abstract

Abstract:

Objective To evaluate the clinical application value of molecular pathological methods in the diagnosis of cervical lymph node tuberculosis and its drug resistance.Methods Ninety-seven paraffin-embedded cervical lymph node specimens with tuberculosis (the tuberculosis (TB) group) and 20 specimens with other diseases (the non-TB group) were collected between March 2010 and October 2013 from the Beijing Chest Hospital. Ziehl-Neelsen (Z-N) staining and fluorescent quantitative polymerase chain reaction (FQ-PCR) were both performed on each specimen. Diagnostic efficiency of both methods was analyzed, Rifampin (RFP) and isoniazid (INH) resis-tances were detected by probe melting curve PCR.Results The sensitivities of Z-N staining and FQ-PCR were 22.7% (22/97), 67.0% (65/97), respectively. FQ-PCR showed significantly higher sensitivity than that of Z-N staining (χ²=38.53, P<0.001). The specificities of these two techniques were both 100.0% (20/20). The positive predictive values of the Z-N staining and FQ-PCR were 100.0% (22/22) and 100.0% (65/65), respectively, the negative predictive values were 21.1% (20/95) and 38.5% (20/52), respectively, and the coincidence rates were 35.9% (42/117) and 72.6% (85/117). Among 41 FQ-PCR positive TB samples, effective RFP and INH genotypic drug sensitivity test results were obtained in 10 and 27 samples, respectively, including 1 RFP resistant case and 13 INH resistant cases. Conclusion Molecular pathological techniques showed high sensitivity and specificity in detecting Mycobacterium tuberculosis DNA in FFPE specimens of cervical lymph node tuberculosis. Furthermore, the new techniques made it possible for pathological diagnosis of drug-resistant tuberculosis, and it can provide valuable evidences for accurate diagnosis and appropriate treatment.

Key words: Tuberculosis, lymph node, Neck, Nucleic acid amplification techniques, DNA Probes, Diagnosis, Tuberculosis, multidrug-resistant

MU Jing,LIU Zi-chen,SONG Jing,LI Kun,CHE Nan-ying,LIU Hong-gang. Application value of molecular pathology in diagnosis of cervical lymph node tuberculosis and determination of drug resistance[J]. Chinese Journal of Antituberculosis, 2018, 40(11): 1170-1175. doi: 10.3969/j.issn.1000-6621.2018.11.006

Figures/Tables 3

References 22

[1]	World Health Organization . Global tuberculosis report 2018. Geneva: World Health Organization, 2018.
[2]	Golden MP, Vikram HR . Extrapulmonary tuberculosis: an overview. Am Fam Physician, 2005,72(9):1761-1768. doi: 10.1016/j.pop.2005.09.005 URL pmid: 16300038
[3]	唐神结, 高文 . 临床结核病学. 北京: 人民卫生出版社, 2011.
[4]	胡春梅, 郭晶, 严虹 , 等. 荧光定量PCR探针熔解曲线法在结核分枝杆菌耐药基因检测中的应用. 中国防痨杂志, 2016,38(1):38-41. doi: 10.3969/j.issn.1000-6621.2016.01.009 URL
[5]	Che NY, Huang SJ, Ma Y , et al. Comparison of histological, microbiological, and molecular methods in diagnosis of patients with TBLN having different anti-TB treatment background. Biomed Environ Sci, 2017,30(6):418-425.
[6]	Chandrasekhar B, Prayaga AK . Utility of concentration method by modified bleach technique for the demonstration of acid-fast bacilli in the diagnosis of tuberculous lymphadenopathy. J Cytol, 2012,29(3):165-168. doi: 10.4103/0970-9371.101160 URL pmid: 3480763
[7]	Wu RI, Mark EJ, Hunt JL . Staining for acid-fast bacilli in surgical pathology: practice patterns and variations. Hum Pathol, 2012,43(11):1845-1851. doi: 10.1016/j.humpath.2012.01.006 URL pmid: 22542129
[8]	刘燕翔, 胥萍, 吴妹英 , 等. 144例颈部肉芽肿性淋巴结炎患者病灶标本的病理与病原学分析. 结核病与肺部健康杂志, 2017,6(1):36-40. doi: 10.3969/j.issn.2095-3755.2017.01.010 URL
[9]	车南颖, 曲杨, 张晨 , 等. 结核分枝杆菌Ag85B蛋白表达特点及其病理学诊断价值. 中华病理学杂志, 2014,43(9):600-603. doi: 10.3760/cma.j.issn.0529-5807.2014.09.006 URL
[10]	Purohit MR, Mustafa T, Wiker HG , et al. Immunohistochemical diagnosis of abdominal and lymph node tuberculosis by detecting Mycobacterium tuberculosis complex specific antigen MPT64. Diagn Pathol, 2007,2:36. doi: 10.1186/1746-1596-2-36 URL pmid: 17894882
[11]	Hillemann D, Richter E, Rüsch-Gerdes S . Use of the BACTEC Mycobacteria Growth Indicator Tube 960 automated system for recovery of Mycobacteria from 9,558 extrapulmonary specimens, including urine samples. J Clin Microbiol, 2006,44(11):4014-4017. doi: 10.1128/JCM.00829-06 URL pmid: 17005737
[12]	Brodie D, Schluger NW . The diagnosis of tuberculosis. Clin Chest Med, 2005,26(2):247-271. doi: 10.1016/j.ccm.2005.02.012 URL
[13]	Lee YJ, Kim S, Kang Y , et al. Does polymerase chain reaction of tissue specimens aid in the diagnosis of tuberculosis? J Pathol Transl Med, 2016,50(6):451-458. doi: 10.4132/jptm.2016.08.04 URL pmid: 5122730
[14]	Raslan WF, Rabaan A, Al-Tawfiq JA . The predictive value of Gen-Probe’s amplified Mycobacterium tuberculosis direct test compared with culturing in paraffin-embedded lymph node tissue exhibiting granulomatous inflammation and negative acid fast stain. J Infect Public Health, 2014,7(4):251-256. doi: 10.1016/j.jiph.2013.11.002 URL pmid: 24602771
[15]	Fu YC, Liao IC, Chen HM , et al. Detection of Mycobacterium tuberculosis complex in paraffin-embedded tissues by the new automated abbott realtime mtb assay. Ann Clin Lab Sci, 2016,46(4):412-417.
[16]	罗春英, 王建东, 王璇 , 等. 荧光定量聚合酶链反应检测石蜡包埋组织结核杆菌的应用价值. 中华病理学杂志, 2012,41(8):562-563. doi: 10.3760/cma.j.issn.0529-5807.2012.08.013 URL
[17]	穆晶, 赵丹, 刘子臣 , 等. 荧光定量PCR技术在骨关节结核石蜡包埋标本检测中的应用价值. 中国防痨杂志, 2016,38(4):277-281. doi: 10.3969/j.issn.1000-6621.2016.04.009 URL
[18]	张海青, 车南颖 . 我国结核病病理学诊断和研究的现状与展望. 中国防痨杂志, 2014,36(9):793-797. doi: 10.3969/j.issn.1000-6621.2014.09.009 URL
[19]	中华医学会结核病学分会,结核病病理学诊断专家共识编写组. 中国结核病病理学诊断专家共识. 中华结核和呼吸杂志, 2017,40(6):419-425. doi: 10.3760/cma.j.issn.1001-0939.2017.06.005 URL
[20]	Polepole P, Kabwe M, Kasonde M , et al. Performance of the Xpert MTB/RIF assay in the diagnosis of tuberculosis in formalin-fixed, paraffin-embedded tissues. Int J Mycobacteriol, 2017,6(1):87-93. doi: 10.4103/2212-5531.201892 URL pmid: 28317811
[21]	García-Basteiro AL, Ismail MR, Carrilho C , et al. The role of Xpert MTB/RIF in diagnosing pulmonary tuberculosis in post-mortem tissues. Sci Rep, 2016,6:20703. doi: 10.1038/srep20703 URL pmid: 4748254
[22]	Zhao Y, Xu S, Wang L , et al. National survey of drug-resis-tant tuberculosis in China. N Engl J Med, 2012,366(23):2161-2170. doi: 10.1056/NEJMoa1108789 URL pmid: 22670902

检测方法	结核组(例)		非结核组(例)		敏感度 (%)	特异度 (%)	阳性预测值 (%)	阴性预测值 (%)	符合率 (%)
检测方法	阳性	阴性	阳性	阴性	敏感度 (%)	特异度 (%)	阳性预测值 (%)	阴性预测值 (%)	符合率 (%)
抗酸染色	22	75	0	20	22.7	100.0	100.0	21.1	35.9
FQ-PCR	65	32	0	20	67.0	100.0	100.0	38.5	72.6

例号	性别	年龄 (岁)	抗酸染色	探针熔解曲线法		MTB培养 (培养方法)
例号	性别	年龄 (岁)	抗酸染色	利福平	异烟肼	MTB培养 (培养方法)
1	女	32	阴性	敏感	耐药	-^b
2	女	42	阴性	-^a	耐药	未生长(MGIT 960)
3	男	31	阳性	-^a	耐药	未生长(MGIT 960)
4	女	34	阴性	-^a	耐药	-^b
5	男	40	阳性	-^a	耐药	未生长(MGIT 960)
6	女	26	阴性	敏感	耐药	未生长(罗氏培养)
7	男	20	阴性	-^a	耐药	-^b
8	女	23	阴性	-^a	耐药	未生长(MGIT 960)
9	女	44	阳性	-^a	耐药	-^b
10	女	20	阴性	-^a	耐药	未生长(罗氏培养)
11	女	58	阳性	敏感	耐药	-^b
12	女	17	阴性	-^a	耐药	未生长(罗氏培养)
13	女	24	阴性	-^a	耐药	未生长(罗氏培养)
14	女	26	阴性	耐药	-^a	-^b