Expert consensus on detection of Mycobacterium tuberculosis drug resistance

doi:10.3969/j.issn.1000-6621.2019.02.003

Abstract

Abstract:

Drug-resistant tuberculosis (DR-TB) especially multidrug-resistant tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB) are one of the most difficult problems to be solved in clinical practice. Anti-tuberculous drug susceptibility tests (DST) provide experimental basis for the diagnosis of DR-TB. This expert consensus briefly introduces the common M.tuberculosis (MTB) DST methods (including phenotypic DST and molecular DST) used in clinic, proposes the strategies of MTB drug resistance detection and correct interpretation of the results, points out the problems of DST detection technology and clinical application, and prospects the development of DST in the future.

Key words: Mycobacterium tuberculosis, Microbial sensitivity tests, Tuberculosis,multidrug-resistant, Laboratory techniques and procedures

Editorial Board of Chinese Journal of Antituberculosis,Basic and Clinical Groups of Tuberculosis Control Branch of China International Exchange and Promotive Association for Medical and Health Care. Expert consensus on detection of Mycobacterium tuberculosis drug resistance[J]. Chinese Journal of Antituberculosis, 2019, 41(2): 129-137. doi: 10.3969/j.issn.1000-6621.2019.02.003

Figures/Tables 2

References 82

[1]	中国防痨协会基础专业委员会. 结核病诊断实验室检验规程. 北京: 中国教育文化出版社, 2006.
[2]	Piubello A, Aït-Khaled N, Caminero JA , et al. Field guide for the management of drug-resistant tuberculosis. Paris: International Union Against Tuberculosis and Lung Disease, 2018.
[3]	郭苏珊, 林健雄, 郭金镇 , 等. BACTEC MGIT 960用于结核分枝杆菌直接药敏试验的研究. 中国病原生物学杂志, 2014,9(10):946-948,952.
[4]	钱雪琴, 曹宇硕, 曹竹君 , 等. BACTEC MGIT 960系统培养分枝杆菌时污染菌的分析. 中国卫生检验杂志, 2014,24(17):2498-2501, 2504.
[5]	李静, 王智存, 白广红 , 等. MicroDST ^TM微孔板检测法对抗结核一、二线药物敏感性试验的临床价值 . 中华肺部疾病杂志(电子版), 2018,11(5):583-587.
[6]	Abdel-Rahman SM, Erfan D, Abdel-Latif W , et al. Evaluation of Sensititre ^® MYCOTB panel for the susceptibility testing of Mycobacterium tuberculosis to first and second lines anti-tuberculosis drugs . Clin Med Diag, 2016,6(1):13-19.
[7]	Bang D . The management of tuberculosis: epidemiology, resistance and monitoring. Dan Med Bull, 2010,57(11):B4213.
[8]	中华人民共和国国家卫生和计划生育委员会.WS 288—2017 肺结核诊断. 2017 -11-09.
[9]	European Centre for Disease Prevention and Control. ERLN-TB expert opinion on the use of therapid molecular assays for the diagnosis of tuberculosis and detection of drug-resistance. Stockholm: European Centre for Disease Prevention and Control, 2013.
[10]	Rinder H, Mieskes KT, Löscher T . Heteroresistance in Mycobacterium tuberculosis. Int J Tuberc Lung Dis, 2001,5(4):339-345.
[11]	Alonso M, Palacios JJ, Herranz M , et al. Isolation of Mycobacterium tuberculosis strains with a silent mutation in rpoB leading to potential misassignment of resistance category. J Clin Microbiol, 2011,49(7):2688-2690. doi: 10.1128/JCM.00659-11 URL
[12]	Kawulur Hanna SI, Ngili Y . Study in silico and characterization of mutations in rpoB and katG genes in patients of tuberculosis in Jayapura, Papua province, Indonesia. Der Pharmacia Lettre, 2016,8(14):85-91.
[13]	Bhuju S, Fonseca Lde S, Marsico AG , et al. Mycobacterium tuberculosis isolates from Rio de Janeiro reveal unusually low correlation between pyrazinamide resistance and mutations in the pncA gene. Infect Genet Evol, 2013,19:1-6. doi: 10.1016/j.meegid.2013.06.008 URL
[14]	Ismail NA, Ismail MF, Noor SS , et al. Genotypic detection of rpoB and katG gene mutations associated with rifampicin and isoniazid resistance in Mycobacterium tuberculosis isolates: A local scenario (Kelantan). Malays J Med Sci, 2016,23(1):22-26.
[15]	Cheng S, Cui Z, Li Y , et al. Diagnostic accuracy of a molecular drug susceptibility testing method for the antituberculosis drug ethambutol: a systematic review and meta-analysis. J Clin Microbiol, 2014,52(8):2913-2924. doi: 10.1128/JCM.00560-14 URL
[16]	李静, 林日文, 张灿强 . Xpert MTB/RIF检测痰标本结核分枝杆菌与利福平耐受性的临床应用研究. 国际检验医学杂志, 2017,38(4):480-482.
[17]	Somoskovi A, Deggim V, Ciardo D , et al. Diagnostic implications of inconsistent results obtained with the Xpert MTB/RIF assay in detection of Mycobacterium tuberculosis isolates with an rpoB mutation associated with low-level rifampin resistance. J Clin Microbiol, 2013,51(9):3127-3129. doi: 10.1128/JCM.01377-13 URL
[18]	Pang Y, Dong H, Tan Y , et al. Rapid diagnosis of MDR and XDR tuberculosis with the MeltPro TB assay in China. Sci Rep, 2016,6:25330. doi: 10.1038/srep25330 URL
[19]	林明冠, 吴元东, 朱中元 , 等. 基因芯片技术在结核分枝杆菌耐药检测中的效果分析. 中国防痨杂志, 2018,40(1):58-62.
[20]	Idrees F, Irfan M, Jabeen K , et al. Diagnostic performance of genoType ^® MTBDRplus line probe assay in bronchoalveolar lavage for pulmonary tuberculosis diagnosis in sputum scarce and smear-negative patients . Int J Mycobacteriol, 2017,6(2):122-126. doi: 10.4103/ijmy.ijmy_42_17 URL
[21]	Theron G, Peter J, Richardson M , et al. GenoType ^® MTBDRsl assay for resistance to second-line anti-tuberculosis drugs.Cochrane Database Syst Rev, 2016, 9:CD010705.
[22]	Gao Y, Zhang Z, Deng J , et al. Multi-center evaluation of GenoType MTBDRsl line probe assay for rapid detection of pre-XDR and XDR Mycobacterium tuberculosis in China. J Infect, 2018,77(4):328-334. doi: 10.1016/j.jinf.2018.06.014 URL
[23]	中华人民共和国国家卫生健康委员会办公厅. 人间传染的病原微生物名录(2018年修订). 国卫办科教函[2018]848号. 北京:中华人民共和国国家卫生健康委员会办公厅, 2018.
[24]	World Health Organization . Guidelines for the programmatic management of drug-resistant tuberculosis. Geneva: World Health Organization, 2008.
[25]	World Health Organization. Policy statement: automated real-time nucleic acid amplification technology for rapid and simultaneousdetection of tuberculosis and rifampicin resistance: Xpert MTB/RIF system. Geneva: World Health Organization, 2011[2018-12-28]. .
[26]	Ioannidis P, Papaventsis D, Karabela S , et al. Cepheid GeneXpert MTB/RIF assay for Mycobacterium tuberculosis detection and rifampin resistance identification in patients with substantial clinical indications of tuberculosis and smear-negative microscopy results. J Clin Microbiol, 2011,49(8):3068-3070. doi: 10.1128/JCM.00718-11 URL
[27]	Boehme CC, Nicol MP, Nabeta P , et al. Feasibility, diagnostic accuracy, and effectiveness of decentralised use of the Xpert MTB/RIF test for diagnosis of tuberculosis and multidrug resistance: a multicentre implementation study. Lancet, 2011,377(9776):1495-1505. doi: 10.1016/S0140-6736(11)60438-8 URL
[28]	Armand S, Vanhuls P, Delcroix G , et al. Comparison of the Xpert MTB/RIF test with an IS6110-TaqMan real-time PCR assay for direct detection of Mycobacterium tuberculosis in respiratory and nonrespiratory specimens. J Clin Microbiol, 2011,49(5):1772-1776. doi: 10.1128/JCM.02157-10 URL
[29]	Sehgal IS, Dhooria S, Aggarwal AN , et al. Diagnostic performance of Xpert MTB/RIF in tuberculous pleural effusion: systematic review and meta-analysis. J Clin Microbiol, 2016,54(4):1133-1136. doi: 10.1128/JCM.03205-15 URL
[30]	World Health Organization. Policy statement: molecular line probe assays for rapid screening of patients at risk of multidrug-resistant tuberculosis (MDR-TB). Geneva: World Health Organization, 2008[2018-12-28]. Policy statement: molecular line probe assays for rapid screening of patients at risk of multidrug-resistant tuberculosis (MDR-TB). Geneva: World Health Organization, 2008[2018-12-28]. .
[31]	World Health Organization . Expert group report: molecular line probe assays for rapid screening of patients at risk of multidrug-resistant tuberculosis (MDR-TB). Geneva: World Health Organization, 2008.
[32]	World Health Organization . The use of molecular line probe assay for the detection of resistance to isoniazid and rifampicin: policy update. Geneva: World Health Organization, 2016.
[33]	王晶 . 不同检验方法对肺结核患者痰中结核分枝杆菌的检验价值分析. 临床检验杂志(电子版), 2018,7(1):43-44.
[34]	王超, 包训迪, 王庆 . 分子生物学方法在痰结核分枝杆菌检测中的应用价值分析. 今日健康, 2016,15(1):9.
[35]	郭新枝, 陈裕, 于永敏 , 等. 痰液BACTEC-MGIT 960快速结核菌培养对菌阴肺结核的临床诊断价值. 中国药物与临床, 2013,13(2):201-202.
[36]	World Health Organization . Policy guidance on drug susceptibility testing of second-line antituberculosis drugs. WHO/HTM/TB/2008.392. Geneva: World Health Organization, 2008.
[37]	高旭, 李静, 柳清云 , 等. 异质性耐药对结核分枝杆菌表型和基因型耐药检测结果的影响. 中华结核和呼吸杂志, 2014,37(4):260-265. doi: 10.3760/cma.j.issn.1001-0939.2014.04.007
[38]	李仁忠, 王黎霞 . 实验室新技术用于耐药结核病诊断流程的建议. 中华结核和呼吸杂志, 2016,39(7):568-569.
[39]	Williams DL, Spring L, Collins L , et al. Contribution of rpoB mutations to development of rifamycin cross-resistance in Mycobacterium tuberculosis. Antimicrob Agents Chemother, 1998,42(7):1853-1857. doi: 10.1128/AAC.42.7.1853 URL
[40]	Jamieson FB, Guthrie JL, Neemuchwala A , et al. Profiling of rpoB mutations and MICs for rifampin and rifabutin in Mycobacterium tuberculosis. J Clin Microbiol, 2014,52(6):2157-2162. doi: 10.1128/JCM.00691-14 URL pmid: 24740074
[41]	Nakata N, Kai M, Makino M . Mutation analysis of mycobacterial rpoB genes and rifampin resistance using recombinant Mycobacterium smegmatis. Antimicrob Agents Chemother, 2012,56(4):2008-2013. doi: 10.1128/AAC.05831-11 URL
[42]	Kurbatova EV, Cavanaugh JS, Shah NS , et al. Rifampicin-resistant Mycobacterium tuberculosis: susceptibility to isoniazid and other anti-tuberculosis drugs. Int J Tuberc Lung Dis, 2012,16(3):355-357. doi: 10.5588/ijtld.11.0542 URL
[43]	Mukinda FK , Theron D, van der Spuy GD, et al. Rise in ri-fampicin- monoresistant tuberculosis in Western Cape, South Africa. Int J Tuberc Lung Dis, 2012,16(2):196-202. doi: 10.5588/ijtld.11.0116 URL
[44]	胡晓红, 向启云, 韩宇 , 等. 基因芯片技术在宜昌地区结核分枝杆菌耐药情况监测中的应用. 中国病原生物学杂志, 2015,10(4):351-354.
[45]	刘银萍, 王杰, 张俊仙 , 等. 结核分枝杆菌rpoB基因突变与三种利福霉素类抗结核药物耐受相关性研究. 中国病原生物学杂志, 2016,11(11):982-985.
[46]	Wengenack NL, Uhl JR, St Amand AL , et al. Recombinant Mycobacterium tuberculosis KatG(S315T) is a competent catalase-peroxidase with reduced activity toward isoniazid. J Infect Dis, 1997,176(3):722-727. doi: 10.1086/jid.1997.176.issue-3 URL
[47]	刘银萍, 王杰, 张俊仙 , 等. 对异烟肼与丙硫异烟胺耐药的结核分枝杆菌临床分离株检测及相关基因突变的研究. 中国防痨杂志, 2016,38(9):718-721.
[48]	Wu XQ, Lu Y, Zhang JX , et al. Detection of the mutations in katG 315 and inhA-15 of Mycobacterium tuberculosis strains isolated from Chinese patients. Chin Med J (Engl), 2006,119(3):230-233. doi: 10.1097/00029330-200602010-00011 URL
[49]	Machado D, Perdigão J, Ramos J , et al. High-level resistance to isoniazid and ethionamide in multidrug-resistant Mycobacterium tuberculosis of the Lisboa family is associated with inhA double mutations. J Antimicrob Chemother, 2013,68(8):1728-1732. doi: 10.1093/jac/dkt090 URL
[50]	Vilchèze C, Jacobs WR Jr . Resistance to isoniazid and ethionamide in Mycobacterium tuberculosis: Genes, mutations, and causalities. Microbiol Spectr, 2014, 2(4):MGM2-0014- 2013.
[51]	Plinke C, Cox HS, Zarkua N , et al. embCAB sequence variation among ethambutol-resistant Mycobacterium tuberculosis isolates without embB306 mutation. J Antimicrob Chemother, 2010,65(7):1359-1367. doi: 10.1093/jac/dkq120 URL
[52]	Wu XQ, Liang JQ, Zhang JX , et al. Detection and evaluation of the mutations of embB gene in ethambutol-susceptible and resistant Mycobacterium tuberculosis isolates from China. Chin Med J (Engl), 2005,118(20):1739-1741.
[53]	Mokrousov I, Otten T, Vyshnevskiy B , et al. Detection of embB306 mutations in ethambutol-susceptible clinical isolates of Mycobacterium tuberculosis from Northwestern Russia: implications for genotypic resistance testing. J Clin Microbiol, 2002,40(10):3810-3813. doi: 10.1128/JCM.40.10.3810-3813.2002 URL
[54]	Bastos ML, Hussain H, Weyer K , et al. Treatment outcomes of patients with multidrug- and extensive drug-resistant tuberculosis according to drug susceptibility testing to first- and second-line drugs: an individual patient data meta-analysis. Clin Infect Dis, 2014,59(10):1364-1374. doi: 10.1093/cid/ciu619 URL
[55]	Plinke C, Walter K, Aly S , et al. Mycobacterium tuberculosis embB codon 306 mutations confer moderately increased resis-tance to ethambutol in vitro and in vivo. Antimicrob Agents Chemother, 2011,55(6):2891-2896. doi: 10.1128/AAC.00007-10 URL
[56]	Whitfield MG, Soeters HM, Warren RM , et al. A global perspective on pyrazinamide resistance: systematic review and Meta-analysis. PLoS One, 2015,10(7):e0133869. doi: 10.1371/journal.pone.0133869 URL
[57]	Ramirez-Busby SM, Valafar F . Systematic review of mutations in pyrazinamidase associated with pyrazinamide resistance in Mycobacterium tuberculosis clinical isolates. Antimicrob Agents Chemother, 2015,59(9):5267-5277. doi: 10.1128/AAC.00204-15 URL
[58]	Chang KC, Yew WW, Zhang Y . Pyrazinamide susceptibility testing in Mycobacterium tuberculosis: a systematic review with meta-analyses. Antimicrob Agents Chemother, 2011,55(10):4499-4505. doi: 10.1128/AAC.00630-11 URL
[59]	Miotto P, Cabibbe AM, Feuerriegel S , et al. Mycobacterium tuberculosis pyrazinamide resistance determinants: a multicenter study. MBio, 2014,5(5):e01819-14.
[60]	Scorpio A, Collins D, Whipple D , et al. Rapid differentiation of bovine and human tubercle bacilli based on a characteristic mutation in the bovine pyrazinamidase gene. J Clin Microbiol, 1997,35(1):106-110.
[61]	Sreevatsan S, Pan X, Zhang Y , et al. Mutations associated with pyrazinamide resistance in pncA of Mycobacterium tuberculosis complex organisms. Antimicrob Agents Chemother, 1997,41(3):636-640. doi: 10.1128/AAC.41.3.636 URL
[62]	Wu XQ, Lu Y, Zhang JX , et al. Detection of streptomycin resistance in Mycobacterium tuberculosis clinical isolates using four molecular methods in China. Yi Chuan Xue Bao, 2006,33(7):655-663.
[63]	He J, Zhu B, Yang Z , et al. Molecular analysis of the rpsL gene for rapid detection of streptomycin-resistant Mycobacterium tuberculosis: a meta-analysis. Scand J Infect Dis, 2014,46(8):585-592. doi: 10.3109/00365548.2014.918649 URL
[64]	Sugawara I, Zhang J, Li C . Cross-resistance of Mycobacterium tuberculosis isolates among streptomycin, kanamycin and amikacin. Indian J Exp Biol, 2009,47(6):520-522.
[65]	Pang Y, Dong H, Tan Y , et al. Rapid diagnosis of MDR and XDR tuberculosis with the MeltPro TB assay in China. Sci Rep, 2016,6:25330. doi: 10.1038/srep25330 URL
[66]	车洋, 杨天池, 于梅 , 等. 116例耐多药结核分枝杆菌临床分离株对4种注射用抗结核药物耐药及交叉耐药分析. 中国卫生检验杂志, 2016,26(12):1802-1804.
[67]	Tsukamura M, Mizuno S . Cross-resistance relationships among the aminoglycoside antibiotics in Mycobacterium tuberculosis. J Gen Microbiol, 1975,88(2):269-274. doi: 10.1099/00221287-88-2-269 URL
[68]	Du Q, Dai G, Long Q , et al. Mycobacterium tuberculosis rrs A1401G mutation correlates with high-level resistance to kanamycin, amikacin, and capreomycin in clinical isolates from mainland China. Diagn Microbiol Infect Dis, 2013,77(2):138-142. doi: 10.1016/j.diagmicrobio.2013.06.031 URL
[69]	Kambli P, Ajbani K, Nikam C , et al. Correlating rrs and eis promoter mutations in clinical isolates of Mycobacterium tuberculosis with phenotypic susceptibility levels to the second line injectables. Int J Mycobacteriol, 2016,5(1):1-6. doi: 10.1016/j.ijmyco.2015.09.001 URL
[70]	Avalos E, Catanzaro D, Catanzaro A , et al. Frequency and geographic distribution of gyrA and gyrB mutations associated with fluoroquinolone resistance in clinical Mycobacterium tuberculosis isolates: a systematic review. PLoS One, 2015,10(3):e0120470. doi: 10.1371/journal.pone.0120470 URL
[71]	Willby M, Sikes RD, Malik S , et al. Correlation between GyrA substitutions and ofloxacin, levofloxacin, and moxifloxacin cross-resistance in Mycobacterium tuberculosis. Antimicrob Agents Chemother, 2015,59(9):5427-5434. doi: 10.1128/AAC.00662-15 URL
[72]	张治国, 杜春英, 张倩 , 等. 我国结核分枝杆菌gyrA不同突变类型对氟喹诺酮类药物耐药水平的相关性研究. 中国防痨杂志, 2016,38(9):706-711.
[73]	Malik S, Willby M, Sikes D , et al. New insights into fluoroquinolone resistance in Mycobacterium tuberculosis: functional genetic analysis of gyrA and gyrB mutations. PLoS One, 2012,7(6):e39754. doi: 10.1371/journal.pone.0039754 URL
[74]	World Health Organization . The use of molecular line probe assays for the detection of resistance to second-line anti-tuberculosis drugs:Policy guidance. Geneva: World Health Organization, 2016.
[75]	Whitfield MG, Soeters HM, Warren RM , et al. A global perspective on pyrazinamide resistance: systematic review and Meta-analysis. PLoS One, 2015,10(7):e0133869. doi: 10.1371/journal.pone.0133869 URL
[76]	World Health Organization . Global tuberculosis report 2012. Geneva: World Health Organization, 2012.
[77]	Streicher EM, Maharaj K, York T , et al. Rapid sequencing of the Mycobacterium tuberculosis pncA gene for detection of pyra-zinamide susceptibility. J Clin Microbiol, 2014,52(11):4056-4057. doi: 10.1128/JCM.02438-14 URL
[78]	World Health Organization. Guidelines for the programmatic management of drug-resistant tuberculosis—2011 update. Geneva: World Health Organization, 2011[2018-12-28]. .
[79]	World Health Organization. Policy guidance on drug-susceptibility testing (DST) of second-line antituberculosis drugs. Geneva: World Health Organization, 2008[2018-12-28]. .
[80]	Kim SJ . Drug-susceptibility testing in tuberculosis: methods and reliability of results. Eur Respir J, 2005,25(3):564-569. doi: 10.1183/09031936.05.00111304 URL
[81]	Kim SJ, Espinal MA, Abe C , et al. Is second-line anti-tuberculosis drug susceptibility testing reliable? Int J Tuberc Lung Dis, 2004,8(9):1157-1158.
[82]	Kam KM, Sloutsky A, Yip CW , et al. Determination of critical concentrations of second-line anti-tuberculosis drugs with clinical and microbiological relevance. Int J Tuberc Lung Dis, 2010,14(3):282-288.

抗结核药物	检测的靶基因	编码的产物	靶区域,突变率	常检测的突变位点, 发生率及耐药水平
RFP、Rft和Rfb	rpoβ	RNA聚合酶β亚单位	507~533位密码子(81bp),90%~95%	531、526位密码子,30%~75%,高
INH	katG	过氧化氢酶-过氧化物酶	整个基因,60%~95%	315位密码子,70%~80%,低
	inhA	烯酰基运载蛋白还原酶	调节区,12%	-15,10%,低
EMB	embB	阿拉伯糖基转移酶	整个基因,36%~70.6%	306位密码子,47%~89%,高
Sm	rpsL	核糖体蛋白S12	43和88位密码子,40%~95%	K43R,40%~90%,高
	rrs	16S rRNA	整个基因碱基,29%	513位,10.3%,低
PZA	pncA	吡嗪酰胺酶	编码基因(约占97%)和启动子(约占3%),68%~97%	3~17、61~85和132~142位密码子,高
FQs	gyrA	DNA旋转酶A	67~106位密码子,75%~94%	D94N或H或G或Y或A、A90V、G88C、W91P和A83V;D94G,21%~32%,高; A90V,13%~20%,低
	gyrB	DNA旋转酶B	5%~15%	T511N、P592S;N538D、E540V 和R485C+T539N,低
Am、Km和Cm	rrs	16S rRNA	50%~84%	A1401G,60%~84%耐Km菌株、50%~80%耐Am或Cm菌株