{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,24]],"date-time":"2026-10-24T01:20:05Z","timestamp":1792804805183,"version":"4.5.0"},"reference-count":51,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2025,4,17]],"date-time":"2025-04-17T00:00:00Z","timestamp":1744848000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Key R&amp;D Program of China","award":["2021YFA1300904"],"award-info":[{"award-number":["2021YFA1300904"]}]},{"name":"National Key R&amp;D Program of China","award":["2023YFF0713600"],"award-info":[{"award-number":["2023YFF0713600"]}]},{"name":"National Key R&amp;D Program of China","award":["NSFC 32300152"],"award-info":[{"award-number":["NSFC 32300152"]}]},{"name":"National Key R&amp;D Program of China","award":["SKLRD-Z-202414"],"award-info":[{"award-number":["SKLRD-Z-202414"]}]},{"name":"National Key R&amp;D Program of China","award":["SKLRD-Z-202301"],"award-info":[{"award-number":["SKLRD-Z-202301"]}]},{"name":"National Key R&amp;D Program of China","award":["2024A04J4273"],"award-info":[{"award-number":["2024A04J4273"]}]},{"name":"National Natural Science Foundation of China","award":["2021YFA1300904"],"award-info":[{"award-number":["2021YFA1300904"]}],"id":[{"id":"https:\/\/ror.org\/01h0zpd94","id-type":"ROR","asserted-by":"crossref"}]},{"name":"National Natural Science Foundation of China","award":["2023YFF0713600"],"award-info":[{"award-number":["2023YFF0713600"]}],"id":[{"id":"https:\/\/ror.org\/01h0zpd94","id-type":"ROR","asserted-by":"crossref"}]},{"name":"National Natural Science Foundation of China","award":["NSFC 32300152"],"award-info":[{"award-number":["NSFC 32300152"]}],"id":[{"id":"https:\/\/ror.org\/01h0zpd94","id-type":"ROR","asserted-by":"crossref"}]},{"name":"National Natural Science Foundation of China","award":["SKLRD-Z-202414"],"award-info":[{"award-number":["SKLRD-Z-202414"]}],"id":[{"id":"https:\/\/ror.org\/01h0zpd94","id-type":"ROR","asserted-by":"crossref"}]},{"name":"National Natural Science Foundation of China","award":["SKLRD-Z-202301"],"award-info":[{"award-number":["SKLRD-Z-202301"]}],"id":[{"id":"https:\/\/ror.org\/01h0zpd94","id-type":"ROR","asserted-by":"crossref"}]},{"name":"National Natural Science Foundation of China","award":["2024A04J4273"],"award-info":[{"award-number":["2024A04J4273"]}],"id":[{"id":"https:\/\/ror.org\/01h0zpd94","id-type":"ROR","asserted-by":"crossref"}]},{"name":"State Key Laboratory of Respiratory Disease","award":["2021YFA1300904"],"award-info":[{"award-number":["2021YFA1300904"]}],"id":[{"id":"https:\/\/ror.org\/04hja5e04","id-type":"ROR","asserted-by":"crossref"}]},{"name":"State Key Laboratory of Respiratory Disease","award":["2023YFF0713600"],"award-info":[{"award-number":["2023YFF0713600"]}],"id":[{"id":"https:\/\/ror.org\/04hja5e04","id-type":"ROR","asserted-by":"crossref"}]},{"name":"State Key Laboratory of Respiratory Disease","award":["NSFC 32300152"],"award-info":[{"award-number":["NSFC 32300152"]}],"id":[{"id":"https:\/\/ror.org\/04hja5e04","id-type":"ROR","asserted-by":"crossref"}]},{"name":"State Key Laboratory of Respiratory Disease","award":["SKLRD-Z-202414"],"award-info":[{"award-number":["SKLRD-Z-202414"]}],"id":[{"id":"https:\/\/ror.org\/04hja5e04","id-type":"ROR","asserted-by":"crossref"}]},{"name":"State Key Laboratory of Respiratory Disease","award":["SKLRD-Z-202301"],"award-info":[{"award-number":["SKLRD-Z-202301"]}],"id":[{"id":"https:\/\/ror.org\/04hja5e04","id-type":"ROR","asserted-by":"crossref"}]},{"name":"State Key Laboratory of Respiratory Disease","award":["2024A04J4273"],"award-info":[{"award-number":["2024A04J4273"]}],"id":[{"id":"https:\/\/ror.org\/04hja5e04","id-type":"ROR","asserted-by":"crossref"}]},{"name":"Guangzhou Institute of Respiratory Diseases","award":["2021YFA1300904"],"award-info":[{"award-number":["2021YFA1300904"]}]},{"name":"Guangzhou Institute of Respiratory Diseases","award":["2023YFF0713600"],"award-info":[{"award-number":["2023YFF0713600"]}]},{"name":"Guangzhou Institute of Respiratory Diseases","award":["NSFC 32300152"],"award-info":[{"award-number":["NSFC 32300152"]}]},{"name":"Guangzhou Institute of Respiratory Diseases","award":["SKLRD-Z-202414"],"award-info":[{"award-number":["SKLRD-Z-202414"]}]},{"name":"Guangzhou Institute of Respiratory Diseases","award":["SKLRD-Z-202301"],"award-info":[{"award-number":["SKLRD-Z-202301"]}]},{"name":"Guangzhou Institute of Respiratory Diseases","award":["2024A04J4273"],"award-info":[{"award-number":["2024A04J4273"]}]},{"name":"Guangzhou Science and Technology Plan-Youth Doctoral \u201cSail\u201d Project","award":["2021YFA1300904"],"award-info":[{"award-number":["2021YFA1300904"]}]},{"name":"Guangzhou Science and Technology Plan-Youth Doctoral \u201cSail\u201d Project","award":["2023YFF0713600"],"award-info":[{"award-number":["2023YFF0713600"]}]},{"name":"Guangzhou Science and Technology Plan-Youth Doctoral \u201cSail\u201d Project","award":["NSFC 32300152"],"award-info":[{"award-number":["NSFC 32300152"]}]},{"name":"Guangzhou Science and Technology Plan-Youth Doctoral \u201cSail\u201d Project","award":["SKLRD-Z-202414"],"award-info":[{"award-number":["SKLRD-Z-202414"]}]},{"name":"Guangzhou Science and Technology Plan-Youth Doctoral \u201cSail\u201d Project","award":["SKLRD-Z-202301"],"award-info":[{"award-number":["SKLRD-Z-202301"]}]},{"name":"Guangzhou Science and Technology Plan-Youth Doctoral \u201cSail\u201d Project","award":["2024A04J4273"],"award-info":[{"award-number":["2024A04J4273"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>The emergence of multidrug-resistant and extensively drug-resistant Mycobacterium tuberculosis strains poses serious challenges to global tuberculosis control, highlighting the urgent need to elucidate the mechanisms underlying multidrug resistance. In this study, we screened for spontaneous bortezomib (BTZ)-resistant Mycobacterium smegmatis (Msm) mutants and identified a strain, Msm-R1-2, exhibiting 16- and 64-fold increases in minimum inhibitory concentrations (MICs) to BTZ and linezolid (LZD), respectively, compared to the parental strain. Whole-genome sequencing revealed resistance-associated mutations in two functionally distinct genes: MSMEG_1380, encoding a transcriptional regulator involved in efflux pump expression, and MSMEG_0965, encoding a porin protein. CRISPR-Cpf1-assisted gene knockout and editing experiments confirmed that single mutations in either MSMEG_1380 or MSMEG_0965 caused low-level resistance (4-fold MIC increase) to BTZ and LZD, while dual mutations conferred resistance levels comparable to Msm-R1-2, with 16- and 64-fold increases in MICs for BTZ and LZD, respectively. An ethidium bromide accumulation assay demonstrated that mutations in MSMEG_0965 reduce cell wall permeability, contributing to multidrug resistance. Furthermore, quantitative real-time PCR showed that mutations in MSMEG_1380 upregulate the mmpS5-mmpL5 efflux system. Together, these dual mechanisms function synergistically: restricted drug entry combined with enhanced drug efflux confers robust multidrug resistance. These findings provide novel insights into the evolutionary mechanisms of resistance in mycobacteria.<\/jats:p>","DOI":"10.3390\/ijms26083779","type":"journal-article","created":{"date-parts":[[2025,4,17]],"date-time":"2025-04-17T00:48:46Z","timestamp":1744850926000},"page":"3779","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Dual Mutations in MSMEG_0965 and MSMEG_1380 Confer High-Level Resistance to Bortezomib and Linezolid by Both Reducing Drug Intake and Increasing Efflux in Mycobacterium smegmatis"],"prefix":"10.3390","volume":"26","author":[{"ORCID":"https:\/\/orcid.org\/0009-0007-6753-7135","authenticated-orcid":false,"given":"Han","family":"Zhang","sequence":"first","affiliation":[{"name":"School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei 230000, China"},{"name":"State Key Laboratory of Respiratory Disease, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"Guangdong-HongKong-Macau Joint Laboratory of Infectious Respiratory Diseases, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"China-New Zealand Belt and Road Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8045-3098","authenticated-orcid":false,"given":"Cuiting","family":"Fang","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Respiratory Disease, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"Guangdong-HongKong-Macau Joint Laboratory of Infectious Respiratory Diseases, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"China-New Zealand Belt and Road Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4021-4519","authenticated-orcid":false,"given":"Buhari","family":"Yusuf","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Respiratory Disease, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"Guangdong-HongKong-Macau Joint Laboratory of Infectious Respiratory Diseases, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"China-New Zealand Belt and Road Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"University of Chinese Academy of Sciences, Beijing 100000, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaoqing","family":"Zhu","sequence":"additional","affiliation":[{"name":"China-New Zealand Belt and Road Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shuai","family":"Wang","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Respiratory Disease, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"Guangdong-HongKong-Macau Joint Laboratory of Infectious Respiratory Diseases, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"China-New Zealand Belt and Road Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"University of Chinese Academy of Sciences, Beijing 100000, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"H. M. Adnan","family":"Hameed","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Respiratory Disease, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"Guangdong-HongKong-Macau Joint Laboratory of Infectious Respiratory Diseases, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"China-New Zealand Belt and Road Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"University of Chinese Academy of Sciences, Beijing 100000, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4131-3444","authenticated-orcid":false,"given":"Yamin","family":"Gao","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Respiratory Disease, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"Guangdong-HongKong-Macau Joint Laboratory of Infectious Respiratory Diseases, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"China-New Zealand Belt and Road Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"University of Chinese Academy of Sciences, Beijing 100000, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5647-6014","authenticated-orcid":false,"given":"Tianyu","family":"Zhang","sequence":"additional","affiliation":[{"name":"School of Basic Medical Sciences, Division of Life Science and Medicine, University of Science and Technology of China, Hefei 230000, China"},{"name":"State Key Laboratory of Respiratory Disease, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"Guangdong-HongKong-Macau Joint Laboratory of Infectious Respiratory Diseases, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"China-New Zealand Belt and Road Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510000, China"},{"name":"University of Chinese Academy of Sciences, Beijing 100000, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,4,17]]},"reference":[{"key":"ref_1","unstructured":"World Health Organization (2024). Global Tuberculosis Report 2024, World Health Organization."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"22","DOI":"10.1038\/s41572-024-00504-2","article-title":"Multidrug-resistant tuberculosis","volume":"10","author":"Dheda","year":"2024","journal-title":"Nat. Rev. Dis. Primers"},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"Zhang, X., Zhao, R., Qi, Y., Yan, X., Qi, G., and Peng, Q. (2024). The progress of Mycobacterium tuberculosis drug targets. Front. Med., 11.","DOI":"10.3389\/fmed.2024.1455715"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"e00253-15","DOI":"10.1128\/mBio.00253-15","article-title":"Target mechanism-based whole-cell screening identifies bortezomib as an inhibitor of caseinolytic protease in mycobacteria","volume":"6","author":"Moreira","year":"2015","journal-title":"mBio"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"416","DOI":"10.1128\/AAC.47.1.416-417.2003","article-title":"In vitro activities of linezolid against clinical isolates of Mycobacterium tuberculosis that are susceptible or resistant to first-line antituberculous drugs","volume":"47","author":"Bouza","year":"2003","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Ahmad, M.N., Aurooz, F., and Singh, V. (2025). Clp protease complex as a therapeutic target for tuberculosis. Bacterial Enzymes as Targets for Drug Discovery, Elsevier.","DOI":"10.1016\/B978-0-443-22222-1.00008-8"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1295","DOI":"10.1046\/j.1365-2958.2002.03242.x","article-title":"Ribosomal and non-ribosomal resistance to oxazolidinones: Species-specific idiosyncrasy of ribosomal alterations","volume":"46","author":"Sander","year":"2002","journal-title":"Mol. Microbiol."},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Moreira, W., Santhanakrishnan, S., Dymock, B.W., and Dick, T. (2017). Bortezomib warhead-switch confers dual activity against mycobacterial caseinolytic protease and proteasome and selectivity against human proteasome. Front. Microbiol., 8.","DOI":"10.3389\/fmicb.2017.00746"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"478","DOI":"10.1021\/acsinfecdis.7b00231","article-title":"Targeting the proteostasis network for mycobacterial drug discovery","volume":"4","author":"Lupoli","year":"2018","journal-title":"ACS Infect. Dis."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1417","DOI":"10.1111\/j.1365-2958.2005.05036.x","article-title":"Structure of the Mycobacterium tuberculosis proteasome and mechanism of inhibition by a peptidyl boronate","volume":"59","author":"Hu","year":"2006","journal-title":"Mol. Microbiol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"e02307","DOI":"10.1128\/AAC.02307-16","article-title":"Towards selective mycobacterial ClpP1P2 inhibitors with reduced activity against the human proteasome","volume":"61","author":"Moreira","year":"2017","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1405","DOI":"10.1111\/j.1365-2958.2005.05035.x","article-title":"Mycobacterium tuberculosis prcBA genes encode a gated proteasome with broad oligopeptide specificity","volume":"59","author":"Lin","year":"2006","journal-title":"Mol. Microbiol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"2031","DOI":"10.1093\/jac\/dkaa136","article-title":"Systematic review of mutations associated with resistance to the new and repurposed Mycobacterium tuberculosis drugs bedaquiline, clofazimine, linezolid, delamanid and pretomanid","volume":"75","author":"Kadura","year":"2020","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Gan, W., Ng, H.F., and Ngeow, Y.F. (2023). Mechanisms of linezolid resistance in mycobacteria. Pharmaceuticals, 16.","DOI":"10.3390\/ph16060784"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"e01842-18","DOI":"10.1128\/AAC.01842-18","article-title":"Molecular analysis of linezolid-resistant clinical isolates of Mycobacterium abscessus","volume":"63","author":"Ye","year":"2019","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"459","DOI":"10.1002\/mlf2.12139","article-title":"EmbB and EmbC regulate the sensitivity of Mycobacterium abscessus to echinomycin","volume":"3","author":"He","year":"2024","journal-title":"mLife"},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Wang, S., Cai, X., Yu, W., Zeng, S., Zhang, J., Guo, L., Gao, Y., Lu, Z., Hameed, H.M.A., and Fang, C. (2022). Arabinosyltransferase C mediates multiple drugs intrinsic resistance by altering cell envelope permeability in Mycobacterium abscessus. Microbiol. Spectr., 10.","DOI":"10.1128\/spectrum.02763-21"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"e02289-24","DOI":"10.1128\/spectrum.02289-24","article-title":"GrcC1 mediates low-level resistance to multiple drugs in M. marinum, M. abscessus, and M. smegmatis","volume":"13","author":"Fang","year":"2025","journal-title":"Microbiol. Spectr."},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Nasiri, M.J., Haeili, M., Ghazi, M., Goudarzi, H., Pormohammad, A., Imani Fooladi, A.A., and Feizabadi, M.M. (2017). New insights in to the intrinsic and acquired drug resistance mechanisms in mycobacteria. Front. Microbiol., 8.","DOI":"10.3389\/fmicb.2017.00681"},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Richard, M., Guti\u00e9rrez, A.V., Viljoen, A.J., Ghigo, E., Blaise, M., and Kremer, L. (2018). Mechanistic and structural insights into the unique TetR-dependent regulation of a drug efflux pump in Mycobacterium abscessus. Front. Microbiol., 9.","DOI":"10.3389\/fmicb.2018.00649"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"106384","DOI":"10.1016\/j.micpath.2023.106384","article-title":"Overexpression of a membrane transport system MSMEG_1381 and MSMEG_1382 confers multidrug resistance in Mycobacterium smegmatis","volume":"185","author":"Salini","year":"2023","journal-title":"Microb. Pathog."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Maslov, D.A., Shur, K.V., Vatlin, A.A., and Danilenko, V.N. (2020). MmpS5-MmpL5 transporters provide Mycobacterium smegmatis resistance to imidazo [1,2-b][1,2,4,5]tetrazines. Pathogens, 9.","DOI":"10.3390\/pathogens9030166"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"451","DOI":"10.1046\/j.1365-2958.2001.02394.x","article-title":"MspA provides the main hydrophilic pathway through the cell wall of Mycobacterium smegmatis","volume":"40","author":"Stahl","year":"2001","journal-title":"Mol. Microbiol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"46S","DOI":"10.1046\/j.1365-2672.92.5s1.7.x","article-title":"Cellular impermeability and uptake of biocides and antibiotics in gram-positive bacteria and mycobacteria","volume":"92","author":"Lambert","year":"2002","journal-title":"Symp. Ser. Soc. Appl. Microbiol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"3127","DOI":"10.1128\/AAC.00239-08","article-title":"Role of porins for uptake of antibiotics by Mycobacterium smegmatis","volume":"52","author":"Danilchanka","year":"2008","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"354","DOI":"10.1093\/femsre\/fux011","article-title":"Antimicrobial resistance in Mycobacterium tuberculosis: Mechanistic and evolutionary perspectives","volume":"41","author":"Gygli","year":"2017","journal-title":"FEMS Microbiol. Rev."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"1911","DOI":"10.1111\/j.1365-2958.1990.tb02040.x","article-title":"Isolation and characterization of efficient plasmid transformation mutants of Mycobacterium smegmatis","volume":"4","author":"Snapper","year":"1911","journal-title":"Mol. Microbiol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"800","DOI":"10.1128\/AAC.01189-07","article-title":"In vitro-selected linezolid-resistant Mycobacterium tuberculosis mutants","volume":"52","author":"Hillemann","year":"2008","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"e00947-17","DOI":"10.1128\/AEM.00947-17","article-title":"CRISPR-Cas12a-assisted recombineering in bacteria","volume":"83","author":"Yan","year":"2017","journal-title":"Appl. Environ. Microbiol."},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Gaurav, A., Bakht, P., Saini, M., Pandey, S., and Pathania, R. (2023). Role of bacterial efflux pumps in antibiotic resistance, virulence, and strategies to discover novel efflux pump inhibitors. Microbiology, 169.","DOI":"10.1099\/mic.0.001333"},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Sui, X., Guo, L., Bao, Z., Xian, M., and Zhao, G. (2025). Efflux pumps and porins enhance bacterial tolerance to phenolic compounds by inhibiting hydroxyl radical generation. Microorganisms, 13.","DOI":"10.3390\/microorganisms13010202"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1167","DOI":"10.1046\/j.1365-2958.2003.03662.x","article-title":"Mycobacterial porins\u2014New channel proteins in unique outer membranes","volume":"49","author":"Niederweis","year":"2003","journal-title":"Mol. Microbiol."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"844","DOI":"10.1111\/j.1365-2958.2009.06801.x","article-title":"Decreased outer membrane permeability protects mycobacteria from killing by ubiquitin-derived peptides","volume":"73","author":"Purdy","year":"2009","journal-title":"Mol. Microbiol."},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Rodrigues, L., Ramos, J., Couto, I., Amaral, L., and Viveiros, M. (2011). Ethidium bromide transport across Mycobacterium smegmatis cell-wall: Correlation with antibiotic resistance. BMC Microbiol., 11.","DOI":"10.1186\/1471-2180-11-35"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"P291","DOI":"10.1016\/S2213-2600(17)30079-6","article-title":"The epidemiology, pathogenesis, transmission, diagnosis, and management of multidrug-resistant, extensively drug-resistant, and incurable tuberculosis","volume":"5","author":"Dheda","year":"2017","journal-title":"Lancet Respir. Med."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"5655","DOI":"10.2147\/IDR.S375869","article-title":"Characterization of genetic variants associated with rifampicin resistance level in Mycobacterium tuberculosis clinical isolates collected in Guangzhou Chest Hospital, China","volume":"15","author":"Hameed","year":"2022","journal-title":"Infect. Drug Resist."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"853","DOI":"10.1099\/mic.0.26902-0","article-title":"The MspA porin promotes growth and increases antibiotic susceptibility of both Mycobacterium bovis BCG and Mycobacterium tuberculosis","volume":"150","author":"Mailaender","year":"2004","journal-title":"Microbiology"},{"key":"ref_38","doi-asserted-by":"crossref","unstructured":"Sonnenkalb, L., Carter, J., Spitaleri, A., Iqbal, Z., Hunt, M., Malone, K., Utpatel, C., Cirillo, D.M., Rodrigues, C., and Nilgiriwala, K.S. (2021). Deciphering bedaquiline and clofazimine resistance in tuberculosis: An evolutionary medicine approach. bioRxiv.","DOI":"10.1101\/2021.03.19.436148"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"e604","DOI":"10.1016\/S2666-5247(21)00175-0","article-title":"Genetic variants and their association with phenotypic resistance to bedaquiline in Mycobacterium tuberculosis: A systematic review and individual isolate data analysis","volume":"2","author":"Ismail","year":"2021","journal-title":"Lancet Microbe"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"447","DOI":"10.2147\/IDR.S239850","article-title":"Efflux pumps contribute to intrinsic clarithromycin resistance in clinical, Mycobacterium abscessus isolates","volume":"13","author":"Guo","year":"2020","journal-title":"Infect. Drug Resist."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"4296","DOI":"10.1038\/ncomms5296","article-title":"Opposing effects of target overexpression reveal drug mechanisms","volume":"5","author":"Palmer","year":"2014","journal-title":"Nat. Commun."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"1547","DOI":"10.1111\/jam.14478","article-title":"Recent updates on drug resistance in Mycobacterium tuberculosis","volume":"128","author":"Singh","year":"2020","journal-title":"J. Appl. Microbiol."},{"key":"ref_43","doi-asserted-by":"crossref","unstructured":"Gifford, D.R., Berr\u00edos-Caro, E., Joerres, C., Su\u00f1\u00e9, M., Forsyth, J.H., Bhattacharyya, A., Galla, T., and Knight, C.G. (2023). Mutators can drive the evolution of multi-resistance to antibiotics. PLoS Genet., 19.","DOI":"10.1371\/journal.pgen.1010791"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"e64116","DOI":"10.7554\/eLife.64116","article-title":"Multi-step vs. single-step resistance evolution under different drugs, pharmacokinetics, and treatment regimens","volume":"10","author":"Igler","year":"2021","journal-title":"eLife"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"34","DOI":"10.1186\/s13073-024-01289-5","article-title":"Detection of a historic reservoir of bedaquiline\/clofazimine resistance-associated variants in Mycobacterium tuberculosis","volume":"16","author":"Nimmo","year":"2024","journal-title":"Genome Med."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"2979","DOI":"10.1128\/AAC.00037-14","article-title":"Cross-resistance between clofazimine and bedaquiline through upregulation of MmpL5 in Mycobacterium tuberculosis","volume":"58","author":"Hartkoorn","year":"2014","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"e02199-23","DOI":"10.1128\/spectrum.02199-23","article-title":"Characterization of in vitro resistance to linezolid in Mycobacterium abscessus","volume":"11","author":"Negatu","year":"2023","journal-title":"Microbiol. Spectr."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"933","DOI":"10.1046\/j.1365-2958.1999.01472.x","article-title":"Cloning of the mspA gene encoding a porin from Mycobacterium smegmatis","volume":"33","author":"Niederweis","year":"1999","journal-title":"Mol. Microbiol."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"17827","DOI":"10.1074\/jbc.M800866200","article-title":"Rv1698 of Mycobacterium tuberculosis represents a new class of channel-forming outer membrane proteins","volume":"283","author":"Siroy","year":"2008","journal-title":"J. Biol. Chem."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"433","DOI":"10.1093\/jac\/45.4.433","article-title":"Interplay between the MexA-MexB-OprM multidrug efflux system and the outer membrane barrier in the multiple antibiotic resistance of Pseudomonas aeruginosa","volume":"45","author":"Li","year":"2000","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_51","unstructured":"Lewis, J.S., and Clinical and Laboratory Standards Institute (2023). Performance Standards for Antimicrobial Susceptibility Testing, Clinical and Laboratory Standards Institute."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/8\/3779\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T17:16:02Z","timestamp":1760030162000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/8\/3779"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,4,17]]},"references-count":51,"journal-issue":{"issue":"8","published-online":{"date-parts":[[2025,4]]}},"alternative-id":["ijms26083779"],"URL":"https:\/\/doi.org\/10.3390\/ijms26083779","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,4,17]]}}}