Study on genes for cross-resistance to isoniazid and ethionamidein Mycobacterium tuberculosis
XIA Qiang1, LIU Haican2, ZHAO Xiuqin2, WAN Kanglin2, ZHAO Lili2
1. Hangzhou Di'an Medical Laboratory Center, Hangzhou, Zhejiang 310012, China; 2. Chinese Center for Disease Control and Prevention, Beijing 102206, China
Abstract：Objective To examine the association between the cross-resistance to ethionamide (Eto) and isoniazid (INH) and mutations of drug resistant genes in Mycobacterium tuberculosis (MTB), so as to provide the evidence for clinical diagnosis and treatment for multidrug-resistant (MDR) tuberculosis. Methods Totally 126 MTB clinical isolates were selected, including 88 MDR-MTB clinical isolates and 38 INH- and rifampicin (RFP)-sensitive isolates. The resistance to INH and Eto was tested in MTB clinical isolates using the drug susceptibility test, and the mutations in the spacer region of INH and Eto resistance-related katG, inhA, ethA, mshA, ndh, spacer region of oxyR-ahpC and inhA promoter were detected using PCR assay. The phenotypic resistance served as a gold standard, and the sensitivity, specificity and accuracy of gene mutation tests were calculated for detection of MTB clinical isolates cross-resistant to INH and Eto. Results Of the 126 MTB clinical isolates, there were 37 isolates cross-resistant to INH and Eto (29.37%), 51 isolates with resistance to INH and susceptibility to Eto (40.48%), 4 isolates with susceptibility to INH and resistance to Eto (3.17%) and 34 isolates with susceptibility to INH and Eto (26.98%). Among the 41 Eto-resistant MTB clinical isolates, there were 37 isolates with resistance to INH (90.24%). There were 64 MTB clinical isolates detected with katG mutations (50.79%), 4 isolates with mutation in the spacer region of oxyR-ahpC (3.17%), 2 isolates with inhA mutations (1.59%), and these isolates were all resistant to INH. There were 11 MTB clinical isolates detected with mutation in the inhA promoter (8.73%) and one isolate with ndh mutation, and all these isolates were cross-resistant to INH and Eto. There were 23 MTB clinical isolates detected with ethA mutations (18.25%) and 40 isolates with mshA mutations (31.75%), in which Eto-susceptible and -resistant isolates were detected. The diagnostic sensitivity, specificity and accuracy of inhA promoter tests for detection of cross-resistance to INH and Eto were 29.73% (95%CI: 16.44%-47.17%), 100.00% (95%CI: 87.36%-100.00%) and 63.38% (95%CI: 51.76%-73.63%) in MTB clinical isolates. Conclusions The prevalence of INH resistance is high in Eto-resistant MTB clinical isolates. Mutation in the inhA promoter region correlates with the cross-resistance to INH and Eto in MTB clinical isolates, and detection of mutation in the inhA promoter may be feasible to detect the cross-resistance to INH and Eto in MTB clinical isolates.
[1] World Health Organization.Global tuberculosis report2020[R/OL].[2022-06-07].https://www.who.int/publications/i/item/9789240013131.pdf. [2] LI B Y, SHI W P,ZHOU C M,et al.Rising challenge of multidrug-resistant tuberculosis in China:a predictive study using Markov modeling[J/OL].Infect Dis Poverty,2020,9(3):57-64.[2022-06-07].https://doi.org/10.1186/s40249-020-00682-7. [3] World Health Organization.WHO consolidated guidelines on drug-resistant tuberculosis treatment[M].Geneva:World Health Organization,2019. [4] LIU Q,YANG D,QIU B,et al.Drug resistance gene mutations and treatment outcomes in MDR-TB:a prospective study in Eastern China[J/OL].PLoS Negl Trop Dis,2021,15(1)[2022-06-07].https://doi.org/10.1371/journal.pntd.0009068. [5] 宋艳华,高孟秋,李琦.结核分枝杆菌对乙硫异烟胺/丙硫异烟胺耐药的机制及其增敏剂研究进展[J].中国防痨杂志,2020,42(2):173-177. SONG Y H,GAO M Q,LI Q.Research progress on the mechanism of drug resistance of Mycobacterium tuberculosis to ethionamide/pthionamide and ethionamide boosters[J].Chin J Antitubercul,2020,42(2):173-177. [6] VILCHÈZE C,JACOBS W R,Jr.Resistance to isoniazid and ethionamide in Mycobacterium tuberculosis:genes,mutations,and causalities[J/OL].Microbiol Spectr, 2014,2(4)[2022-06-07].https://doi.org/10.1128/microbiolspec.MGM2-0014-2013. [7] World Health Organization.Policy guidance on drug-susceptibility testing(DST)of second-line antituberculosis drugs[M].Geneva:World Health Organization,2008. [8] SOMERVILLE W,THIBERT L,SCHWARTZMAN K,et al.Extraction of Mycobacterium tuberculosis DNA:a question of containment[J].J Clin Microbiol,2005,43(6):2996-2997. [9] ISLAM M M, TAN Y,HAMEED H M A, et al.Detection of novel mutations associated with independent resistance and cross-resistance to isoniazid and prothionamide in Mycobacterium tuberculosis clinical isolates[J/OL].Clin Microbiol Infect, 2019, 25(8)[2022-06-07].https://doi.org/10.1016/j.cmi.2018.12.008. [10] 全国第五次结核病流行病学抽样调查技术指导组,全国第五次结核病流行病学抽样调查办公室.2010年全国第五次结核病流行病学抽样调查报告[J].中国防痨杂志,2012,8(34):485-508. Technical Guidance Group of the Fifth National TB Epidemiological Survey,the Office of the Fifth National TB Epidemiological Survey.The fifth national tuberculosis epidemiological survey in 2010[J].Chin J Antitubercul,2012,8(34):485-508 [11] 宋艳华,王桂荣,霍凤敏,等.耐多药和广泛耐药MTB的inhA基因突变与对丙硫异烟胺耐药的相关性分析[J].中国防痨杂志,2018,40(8):821-824. SONG Y H,WANG G R,HUO F M,et al.Correlation analysis between inhA gene mutation and protionamide-resistance in multidrug resistance and extensively drug-resistant Mycobacterium tuberculosis[J].Chin J Antitubercul,2018,40(8):821-824. [12] HALDER S K,ELMA F.In silico identification of novel chemical compounds with antituberculosis activity for the inhibition of InhA and EthR proteins from Mycobacterium tuberculosis[J/OL].J Clin Tuber Other Mycobacterial Dis,2021,24[2022-06-07].https://doi.org/10.1016/j.jctube.2021.100246. [13] ZHANG Y,YEW W W.Mechanisms of drug resistance in Mycobacterium tuberculosis:update 2015[J].Int J Tuberc Lung Dis,2015,19(11):1276-1289. [14] MIESEL L,WEISBROD T R,MARCINKEVICIENE J A,et al.NADH dehydrogenase defects confer isoniazid resistance and conditional lethality in Mycobacterium smegmatis[J].J Bacteriol,1998,180(9):2459-2467. [15] ROHDE K H,SORCI L.The prospective synergy of antitubercular drugs with NAD biosynthesis inhibitors[J/OL].Front Microbiol,2021,11[2022-06-07].https://doi.org/10.3389/fmicb.2020.634640. [16] VILCHÈZE C,AV-GAY Y,ATTARIAN R,et al.Mycothiol biosynthesis is essential for ethionamide susceptibility in Mycobacterium tuberculosis[J].Mol Microbiol,2008,69(5):1316-1329. [17] PINHATA J M W,BRANDAO A P,MENDES F D F,et al.Correlating genetic mutations with isoniazid phenotypic levels of resistance in Mycobacterium tuberculosis isolates from patients with drug-resistant tuberculosis in a high burden setting[J/OL].Eur J Clin Microbiol Infect Dis,2021,40(12):2551-2561[2022-06-07].https://doi.org/10.1007/s10096-021-04316-0. [18] DE SOUZA V C,ANTUNES D,SANTOS L H,et al.Insights into the mechanism of ethionamide resistance in Mycobacterium tuberculosis through an in silico structural evaluation of EthA and mutants identified in clinical isolates[J/OL].Catalysts,2020,10(5)[2022-06-07].https://doi.org/10.3390/catal10050543.
