{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,1,20]],"date-time":"2026-01-20T02:04:53Z","timestamp":1768874693820,"version":"3.49.0"},"reference-count":48,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2021,9,7]],"date-time":"2021-09-07T00:00:00Z","timestamp":1630972800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32002331 and 31872526"],"award-info":[{"award-number":["32002331 and 31872526"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Natural Science Foundation of Jiangsu Province of China","award":["BK20190893"],"award-info":[{"award-number":["BK20190893"]}]},{"name":"Agricultural Science and Technology Independent Innovation Fund of Jiangsu Province","award":["CX(20)3091"],"award-info":[{"award-number":["CX(20)3091"]}]},{"name":"China Postdoctoral Science Foundation funded project","award":["2019M651984 and 2021T140579"],"award-info":[{"award-number":["2019M651984 and 2021T140579"]}]},{"name":"A Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD)","award":["2019"],"award-info":[{"award-number":["2019"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pharmaceuticals"],"abstract":"<jats:p>Recently, a novel efflux pump gene cluster called tmexCD1-toprJ1 and its variants have been identified, which undermine the antibacterial activity of tigecycline, one of the last remaining options effective against multidrug-resistant (MDR) Gram-negative bacteria. Herein, we report the potent synergistic effect of the non-steroidal anti-inflammatory drug benzydamine in combination with tigecycline at sub-inhibitory concentrations against various temxCD-toprJ-positive Gram-negative pathogens. The combination of benzydamine and tigecycline killed all drug-resistant pathogens during 24 h of incubation. In addition, the evolution of tigecycline resistance was significantly suppressed in the presence of benzydamine. Studies on the mechanisms of synergism showed that benzydamine disrupted the bacterial proton motive force and the functionality of this kind of novel plasmid-encoded resistance-nodulation-division efflux pump, thereby promoting the intracellular accumulation of tigecycline. Most importantly, the combination therapy of benzydamine and tigecycline effectively improved the survival of Galleria mellonella larvae compared to tigecycline monotherapy. Our findings provide a promising drug combination therapeutic strategy for combating superbugs carrying the tmexCD-toprJ gene.<\/jats:p>","DOI":"10.3390\/ph14090907","type":"journal-article","created":{"date-parts":[[2021,9,7]],"date-time":"2021-09-07T10:24:25Z","timestamp":1631010265000},"page":"907","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":15,"title":["Benzydamine Reverses TMexCD-TOprJ-Mediated High-Level Tigecycline Resistance in Gram-Negative Bacteria"],"prefix":"10.3390","volume":"14","author":[{"given":"Ziwen","family":"Tong","sequence":"first","affiliation":[{"name":"College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tianqi","family":"Xu","sequence":"additional","affiliation":[{"name":"College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tian","family":"Deng","sequence":"additional","affiliation":[{"name":"College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jingru","family":"Shi","sequence":"additional","affiliation":[{"name":"College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Zhiqiang","family":"Wang","sequence":"additional","affiliation":[{"name":"College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China"},{"name":"Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Joint International Research Laboratory of Agriculture and Agri-Product Safety, The Ministry of Education of China, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9622-6471","authenticated-orcid":false,"given":"Yuan","family":"Liu","sequence":"additional","affiliation":[{"name":"College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China"},{"name":"Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Joint International Research Laboratory of Agriculture and Agri-Product Safety, The Ministry of Education of China, Yangzhou University, Yangzhou 225009, China"},{"name":"Institute of Comparative Medicine, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2021,9,7]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1136.e1","DOI":"10.1016\/j.cell.2018.02.018","article-title":"Antibiotic resistance","volume":"172","author":"Yelin","year":"2018","journal-title":"Cell"},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Liu, Y., Tong, Z., Shi, J., Jia, Y., Yang, K., and Wang, Z. (2020). Correlation between exogenous compounds and the horizontal transfer of plasmid-borne antibiotic resistance genes. Microorganisms, 8.","DOI":"10.3390\/microorganisms8081211"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"2625","DOI":"10.1093\/jac\/dky262","article-title":"Proposal for assignment of allele numbers for mobile colistin resistance (mcr) genes","volume":"73","author":"Partridge","year":"2018","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"682","DOI":"10.1128\/CMR.05035-11","article-title":"Carbapenemases in Klebsiella pneumoniae and other enterobacteriaceae: An evolving crisis of global dimensions","volume":"25","author":"Tzouvelekis","year":"2012","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"470","DOI":"10.1093\/jac\/dki248","article-title":"Tigecycline","volume":"56","author":"Pankey","year":"2005","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"80","DOI":"10.3389\/fmicb.2019.00080","article-title":"Infections caused by carbapenem-resistant enterobacteriaceae: An update on therapeutic options","volume":"10","author":"Sheu","year":"2019","journal-title":"Front. Microbiol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1450","DOI":"10.1038\/s41564-019-0445-2","article-title":"Emergence of plasmid-mediated high-level tigecycline resistance genes in animals and humans","volume":"4","author":"He","year":"2019","journal-title":"Nat. Microbiol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"e02930-19","DOI":"10.1128\/mBio.02930-19","article-title":"Emergence of a plasmid-encoded resistance-nodulation-division efflux pump conferring resistance to multiple drugs, including tigecycline, in Klebsiella pneumoniae","volume":"11","author":"Lv","year":"2020","journal-title":"mBio"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1102","DOI":"10.1080\/22221751.2020.1768805","article-title":"Co-existence of a novel plasmid-mediated efflux pump with colistin resistance gene mcr in one plasmid confers transferable multidrug resistance in Klebsiella pneumoniae","volume":"9","author":"Sun","year":"2020","journal-title":"Emerg. Microbes Infect."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"e2000014","DOI":"10.1002\/bies.202000014","article-title":"Emerging high-level tigecycline resistance: Novel tetracycline destructases spread via the mobile tet(X)","volume":"42","author":"Fang","year":"2020","journal-title":"BioEssays"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"730","DOI":"10.1038\/nchembio.2376","article-title":"Plasticity, dynamics, and inhibition of emerging tetracycline resistance enzymes","volume":"13","author":"Park","year":"2017","journal-title":"Nat. Chem. Biol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1038\/s41467-018-02875-z","article-title":"Overcoming mcr-1 mediated colistin resistance with colistin in combination with other antibiotics","volume":"9","author":"Macnair","year":"2018","journal-title":"Nat. Commun."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"337","DOI":"10.1128\/CMR.00117-14","article-title":"The challenge of efflux-mediated antibiotic resistance in Gram-negative bacteria","volume":"28","author":"Li","year":"2015","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"523","DOI":"10.1038\/s41579-018-0048-6","article-title":"Multidrug efflux pumps: Structure, function and regulation","volume":"16","author":"Du","year":"2018","journal-title":"Nat. Rev. Microbiol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"e02229-20","DOI":"10.1128\/AAC.02229-20","article-title":"A novel transferable resistance-nodulation-division pump gene cluster, tmexCD2-toprJ2, confers tigecycline resistance in Raoultella ornithinolytica","volume":"65","author":"Wang","year":"2021","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Wang, Q., Peng, K., Liu, Y., Xiao, X., Wang, Z., and Li, R. (2021). Characterization of TMexCD3-TOprJ3, an RND-type efflux system conferring resistance to tigecycline in Proteus mirabilis, and its associated integrative conjugative element. Antimicrob. Agents Chemother., 65.","DOI":"10.1128\/AAC.02712-20"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"141","DOI":"10.1038\/s41579-018-0141-x","article-title":"Drug combinations: A strategy to extend the life of antibiotics in the 21st century","volume":"17","author":"Tyers","year":"2019","journal-title":"Nat. Rev. Genet."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"301","DOI":"10.1080\/1040841X.2019.1599813","article-title":"Antibiotic adjuvants: An alternative approach to overcome multi-drug resistant Gram-negative bacteria","volume":"45","author":"Liu","year":"2019","journal-title":"Crit. Rev. Microbiol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1902227","DOI":"10.1002\/advs.201902227","article-title":"Metformin restores tetracyclines susceptibility against multidrug resistant bacteria","volume":"7","author":"Liu","year":"2020","journal-title":"Adv. Sci."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1038\/s42003-020-0877-5","article-title":"Anti-HIV agent azidothymidine decreases Tet(X)-mediated bacterial resistance to tigecycline in Escherichia coli","volume":"3","author":"Liu","year":"2020","journal-title":"Commun. Biol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"889","DOI":"10.1111\/anae.14224","article-title":"Topical benzydamine hydrochloride for prevention of postoperative sore throat in adults undergoing tracheal intubation for elective surgery: A systematic review and meta-analysis","volume":"73","author":"Kuriyama","year":"2018","journal-title":"Anaesthesia"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"462","DOI":"10.1016\/j.apsb.2019.11.004","article-title":"Benzydamine inhibits osteoclast differentiation and bone resorption via down-regulation of interleukin-1 expression","volume":"10","author":"Son","year":"2020","journal-title":"Acta Pharm. Sin. B"},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Yang, X., Ye, L., Chan, E.W.-C., Zhang, R., and Chen, S. (2021). Characterization of an IncFIB\/IncHI1B plasmid encoding efflux pump TMexCD1-TOprJ1 in a clinical tigecycline- and carbapenem-resistant Klebsiella pneumoniae strain. Antimicrob. Agents Chemother., 65.","DOI":"10.1128\/AAC.02340-20"},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Wan, M., Gao, X., Lv, L., Cai, Z., and Liu, J.-H. (2021). IS 26 mediates the acquisition of tigecycline resistance gene cluster tmexCD1-toprJ1 by IncHI1B-FIB plasmids in Klebsiella pneumoniae and Klebsiella quasipneumoniae from food market sewage. Antimicrob. Agents Chemother., 65.","DOI":"10.1128\/AAC.02178-20"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"4910","DOI":"10.7150\/thno.56205","article-title":"Drug repurposing for next-generation combination therapies against multidrug-resistant bacteria","volume":"11","author":"Liu","year":"2021","journal-title":"Theranostics"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"42","DOI":"10.3389\/fmicb.2019.00042","article-title":"Variation in mutant prevention concentrations","volume":"10","author":"Gianvecchio","year":"2019","journal-title":"Front. Microbiol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"7235","DOI":"10.1021\/bi9826299","article-title":"Mechanism of interaction of different classes of cationic antimicrobial peptides with planar bilayers and with the cytoplasmic membrane of Escherichia coli","volume":"38","author":"Wu","year":"1999","journal-title":"Biochemistry"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"688","DOI":"10.1016\/j.cell.2020.01.021","article-title":"A deep learning approach to antibiotic discovery","volume":"180","author":"Stokes","year":"2020","journal-title":"Cell"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"544","DOI":"10.1016\/j.chom.2020.02.002","article-title":"Proton motive force disruptors block bacterial competence and horizontal gene transfer","volume":"27","author":"Domenech","year":"2020","journal-title":"Cell Host Microbe"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1168","DOI":"10.1016\/j.chembiol.2013.07.006","article-title":"Collapsing the proton motive force to identify synergistic combinations against Staphylococcus aureus","volume":"20","author":"Farha","year":"2013","journal-title":"Chem. Biol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"322","DOI":"10.1111\/j.1572-0241.2005.40310.x","article-title":"Pyogenic liver abscess with a focus on Klebsiella pneumoniae as a primary pathogen: An emerging disease with unique clinical characteristics","volume":"100","author":"Lederman","year":"2005","journal-title":"Am. J. Gastroenterol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"597","DOI":"10.1016\/S1473-3099(10)70143-2","article-title":"Emergence of a new antibiotic resistance mechanism in India, Pakistan, and the UK: A molecular, biological, and epidemiological study","volume":"10","author":"Kumarasamy","year":"2010","journal-title":"Lancet Infect. Dis."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"161","DOI":"10.1016\/S1473-3099(15)00424-7","article-title":"Emergence of plasmid-mediated colistin resistance mechanism MCR-1 in animals and human beings in China: A microbiological and molecular biological study","volume":"16","author":"Liu","year":"2016","journal-title":"Lancet Infect. Dis."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"711","DOI":"10.1038\/nrmicro1234","article-title":"Mechanisms of, and barriers to, horizontal gene transfer between bacteria","volume":"3","author":"Thomas","year":"2005","journal-title":"Nat. Rev. Microbiol."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"298","DOI":"10.1016\/j.ijmm.2013.02.001","article-title":"Plasmids and the spread of resistance","volume":"303","author":"Carattoli","year":"2013","journal-title":"Int. J. Med. Microbiol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"551","DOI":"10.1016\/j.coph.2012.07.005","article-title":"Adjuvant strategies for potentiation of antibiotics to overcome antimicrobial resistance","volume":"12","author":"Pieren","year":"2012","journal-title":"Curr. Opin. Pharmacol."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"e02574-18","DOI":"10.1128\/AAC.02574-18","article-title":"The Anthelmintic Drug Niclosamide synergizes with colistin and reverses colistin resistance in Gram-negative bacilli","volume":"63","author":"Domalaon","year":"2019","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"1802333","DOI":"10.1002\/advs.201802333","article-title":"Proanthocyanidin interferes with intrinsic antibiotic resistance mechanisms of Gram-negative bacteria","volume":"6","author":"Maisuria","year":"2019","journal-title":"Adv. Sci."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"1368","DOI":"10.1021\/acsinfecdis.8b00103","article-title":"New class of adjuvants enables lower dosing of colistin against Acinetobacter baumannii","volume":"4","author":"Minrovic","year":"2018","journal-title":"ACS Infect. Dis."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"141","DOI":"10.1021\/acsinfecdis.8b00288","article-title":"Using small-molecule adjuvants to repurpose azithromycin for use against Pseudomonas aeruginosa","volume":"5","author":"Hubble","year":"2019","journal-title":"ACS Infect. Dis."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"232","DOI":"10.1128\/MMBR.65.2.232-260.2001","article-title":"Tetracycline antibiotics: Mode of action, applications, molecular biology, and epidemiology of bacterial resistance","volume":"65","author":"Chopra","year":"2001","journal-title":"Microbiol. Mol. Biol. Rev."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"445","DOI":"10.1111\/j.1469-185X.1966.tb01501.x","article-title":"Chemiosmotic coupling in oxidative and photosynthetic phosphorylation","volume":"41","author":"Mitchell","year":"1966","journal-title":"Biol. Rev."},{"key":"ref_43","unstructured":"Clinical and Laboratory Standards Institute (2018). Performance Standards for Antimicrobial Susceptibility Testing, CLSI."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"2100749","DOI":"10.1002\/advs.202100749","article-title":"Plant natural flavonoids against multidrug resistant pathogens","volume":"8","author":"Song","year":"2021","journal-title":"Adv. Sci."},{"key":"ref_45","first-page":"1","article-title":"Resensitizing carbapenem- and colistin-resistant bacteria to antibiotics using auranofin","volume":"11","author":"Sun","year":"2020","journal-title":"Nat. Commun."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"439","DOI":"10.1128\/mr.51.4.439-457.1987","article-title":"Bacterial uptake of aminoglycoside antibiotics","volume":"51","author":"Taber","year":"1987","journal-title":"Microbiol. Rev."},{"key":"ref_47","doi-asserted-by":"crossref","unstructured":"Forster, S., Thumser, A.E., Hood, S.R., and Plant, N. (2012). Characterization of Rhodamine-123 as a tracer dye for use in in vitro drug transport assays. PLoS ONE, 7.","DOI":"10.1371\/journal.pone.0033253"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"1396","DOI":"10.1002\/jssc.201400152","article-title":"Quantitative analysis and pharmacokinetics study of tigecycline in human serum using a validated sensitive liquid chromatography with tandem mass spectrometry method","volume":"37","author":"Xie","year":"2014","journal-title":"J. Sep. Sci."}],"container-title":["Pharmaceuticals"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1424-8247\/14\/9\/907\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T06:58:15Z","timestamp":1760165895000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1424-8247\/14\/9\/907"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,9,7]]},"references-count":48,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2021,9]]}},"alternative-id":["ph14090907"],"URL":"https:\/\/doi.org\/10.3390\/ph14090907","relation":{},"ISSN":["1424-8247"],"issn-type":[{"value":"1424-8247","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,9,7]]}}}