{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,20]],"date-time":"2026-10-20T21:58:33Z","timestamp":1792533513731,"version":"4.5.0"},"reference-count":41,"publisher":"MDPI AG","issue":"17","license":[{"start":{"date-parts":[[2020,8,25]],"date-time":"2020-08-25T00:00:00Z","timestamp":1598313600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Aminoglycoside antibiotics are powerful bactericidal therapeutics that are often used in the treatment of critical Gram-negative systemic infections. The emergence and global spread of antibiotic resistance, however, has compromised the clinical utility of aminoglycosides to an extent similar to that found for all other antibiotic-drug classes. Apramycin, a drug candidate currently in clinical development, was suggested as a next-generation aminoglycoside antibiotic with minimal cross-resistance to all other standard-of-care aminoglycosides. Here, we analyzed 591,140 pathogen genomes deposited in the NCBI National Database of Antibiotic Resistant Organisms (NDARO) for annotations of apramycin-resistance genes, and compared them to the genotypic prevalence of carbapenem resistance and 16S-rRNA methyltransferase (RMTase) genes. The 3-N-acetyltransferase gene aac(3)-IV was found to be the only apramycin-resistance gene of clinical relevance, at an average prevalence of 0.7%, which was four-fold lower than that of RMTase genes. In the important subpopulation of carbapenemase-positive isolates, aac(3)-IV was nine-fold less prevalent than RMTase genes. The phenotypic profiling of selected clinical isolates and recombinant strains expressing the aac(3)-IV gene confirmed resistance to not only apramycin, but also gentamicin, tobramycin, and paromomycin. Probing the structure\u2013activity relationship of such substrate promiscuity by site-directed mutagenesis of the aminoglycoside-binding pocket in the acetyltransferase AAC(3)-IV revealed the molecular contacts to His124, Glu185, and Asp187 to be equally critical in binding to apramycin and gentamicin, whereas Asp67 was found to be a discriminating contact. Our findings suggest that aminoglycoside cross-resistance to apramycin in clinical isolates is limited to the substrate promiscuity of a single gene, rendering apramycin best-in-class for the coverage of carbapenem- and aminoglycoside-resistant bacterial infections.<\/jats:p>","DOI":"10.3390\/ijms21176133","type":"journal-article","created":{"date-parts":[[2020,8,25]],"date-time":"2020-08-25T10:48:24Z","timestamp":1598352504000},"page":"6133","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":37,"title":["Epidemiologic, Phenotypic, and Structural Characterization of Aminoglycoside-Resistance Gene aac(3)-IV"],"prefix":"10.3390","volume":"21","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-5839-2404","authenticated-orcid":false,"given":"Michel","family":"Plattner","sequence":"first","affiliation":[{"name":"Institute of Medical Microbiology, University of Zurich, 8006 Zurich, Switzerland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marina","family":"Gysin","sequence":"additional","affiliation":[{"name":"Institute of Medical Microbiology, University of Zurich, 8006 Zurich, Switzerland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Klara","family":"Haldimann","sequence":"additional","affiliation":[{"name":"Institute of Medical Microbiology, University of Zurich, 8006 Zurich, Switzerland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5788-321X","authenticated-orcid":false,"given":"Katja","family":"Becker","sequence":"additional","affiliation":[{"name":"Institute of Medical Microbiology, University of Zurich, 8006 Zurich, Switzerland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9205-0384","authenticated-orcid":false,"given":"Sven N.","family":"Hobbie","sequence":"additional","affiliation":[{"name":"Institute of Medical Microbiology, University of Zurich, 8006 Zurich, Switzerland"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2020,8,25]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"318","DOI":"10.1016\/S1473-3099(17)30753-3","article-title":"Discovery, research, and development of new antibiotics: The WHO priority list of antibiotic-resistant bacteria and tuberculosis","volume":"18","author":"Tacconelli","year":"2018","journal-title":"Lancet Infect. Dis."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"4S","DOI":"10.1046\/j.1365-2672.92.5s1.13.x","article-title":"Exploiting current understanding of antibiotic action for discovery of new drugs","volume":"92","author":"Chopra","year":"2002","journal-title":"J. Appl. Microbiol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"a027029","DOI":"10.1101\/cshperspect.a027029","article-title":"Aminoglycosides: An Overview","volume":"6","author":"Krause","year":"2016","journal-title":"Cold Spring Harb. Perspect. Med."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"151","DOI":"10.1016\/j.drup.2010.08.003","article-title":"Aminoglycoside modifying enzymes","volume":"13","author":"Ramirez","year":"2010","journal-title":"Drug Resist. Updates"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"980","DOI":"10.1021\/acsinfecdis.8b00001","article-title":"Plazomicin Retains Antibiotic Activity against Most Aminoglycoside Modifying Enzymes","volume":"4","author":"Cox","year":"2018","journal-title":"ACS Infect. Dis."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"523","DOI":"10.1016\/j.idc.2016.02.011","article-title":"Aminoglycoside Resistance: The Emergence of Acquired 16S Ribosomal RNA Methyltransferases","volume":"30","author":"Doi","year":"2016","journal-title":"Infect. Dis. Clin. North Am."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"4","DOI":"10.1038\/ja.2017.117","article-title":"Destination of aminoglycoside antibiotics in the \u201cpost-antibiotic era\u201d","volume":"71","author":"Takahashi","year":"2018","journal-title":"J. Antibiot."},{"key":"ref_8","unstructured":"(2020, August 25). World Health Organization Model List of Essential Medicines, 21st List, 2019. Geneva: World Health Organization. Available online: https:\/\/apps.who.int\/iris\/bitstream\/handle\/10665\/325771\/WHO-MVP-EMP-IAU-2019.06-eng.pdf?ua=1."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"10984","DOI":"10.1073\/pnas.1204073109","article-title":"Dissociation of antibacterial activity and aminoglycoside ototoxicity in the 4-monosubstituted 2-deoxystreptamine apramycin","volume":"109","author":"Matt","year":"2012","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"329","DOI":"10.1038\/s41429-020-0291-8","article-title":"Antibiotics in the clinical pipeline in October 2019","volume":"73","author":"Butler","year":"2020","journal-title":"J. Antibiot."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"944","DOI":"10.1093\/jac\/dky546","article-title":"In vitro activity of apramycin against multidrug-, carbapenem- and aminoglycoside-resistant Enterobacteriaceae and Acinetobacter baumannii","volume":"74","author":"Juhas","year":"2019","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"69","DOI":"10.1128\/AAC.14.1.69","article-title":"Enzymatic modification of aminoglycoside antibiotics: 3-N-acetyltransferase with broad specificity that determines resistance to the novel aminoglycoside apramycin","volume":"14","author":"Davies","year":"1978","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"373","DOI":"10.1128\/AAC.01124-10","article-title":"Novel apramycin resistance gene apmA in bovine and porcine methicillin-resistant Staphylococcus aureus ST398 isolates","volume":"55","author":"Fessler","year":"2011","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"803","DOI":"10.1093\/jac\/20.6.803","article-title":"AAC(1): A new aminoglycoside-acetylating enzyme modifying the Cl aminogroup of apramycin","volume":"20","author":"Lovering","year":"1987","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"4401","DOI":"10.1128\/AAC.00926-07","article-title":"Novel plasmid-mediated 16S rRNA m1A1408 methyltransferase, NpmA, found in a clinically isolated Escherichia coli strain resistant to structurally diverse aminoglycosides","volume":"51","author":"Wachino","year":"2007","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"915","DOI":"10.1099\/00221287-133-4-915","article-title":"Cloning of Aminoglycoside-Resistance Determinants from Streptomyces Tenebrarius and Comparison with Related Genes from Other Actinomycetes","volume":"133","author":"Skeggs","year":"1987","journal-title":"Microbiology"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"297","DOI":"10.1111\/j.1574-6968.2004.tb09547.x","article-title":"Use of a novel allele of the Escherichia coli aacC4 aminoglycoside resistance gene as a genetic marker in mycobacteria","volume":"234","author":"Consaul","year":"2004","journal-title":"FEMS Microbiol. Lett."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"48","DOI":"10.1093\/jac\/dkq408","article-title":"Activity of aminoglycosides, including ACHN-490, against carbapenem-resistant Enterobacteriaceae isolates","volume":"66","author":"Livermore","year":"2011","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Hu, Y., Liu, L., Zhang, X., Feng, Y., and Zong, Z. (2017). In Vitro Activity of Neomycin, Streptomycin, Paromomycin and Apramycin against Carbapenem-Resistant Enterobacteriaceae Clinical Strains. Front. Microbiol., 8.","DOI":"10.3389\/fmicb.2017.02275"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"188","DOI":"10.1016\/j.diagmicrobio.2017.03.006","article-title":"In vitro Apramycin Activity against multidrug-resistant Acinetobacter baumannii and Pseudomonas aeruginosa","volume":"88","author":"Kang","year":"2017","journal-title":"Diagn. Microbiol. Infect. Dis."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"168","DOI":"10.1016\/j.diagmicrobio.2018.05.018","article-title":"Evaluation of apramycin activity against methicillin-resistant, methicillin-sensitive, and vancomycin-intermediate Staphylococcus aureus clinical isolates","volume":"92","author":"Truelson","year":"2018","journal-title":"Diagn. Microbiol. Infect. Dis."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Kang, A.D., Smith, K.P., Berg, A.H., Truelson, K.A., Eliopoulos, G.M., McCoy, C., and Kirby, J.E. (2018). Efficacy of Apramycin against Multidrug-Resistant Acinetobacter baumannii in the Murine Neutropenic Thigh Model. Antimicrob. Agents Chemother., 62.","DOI":"10.1128\/AAC.02585-17"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1311","DOI":"10.1093\/jac\/dkz012","article-title":"Evaluation of apramycin against spectinomycin-resistant and -susceptible strains of Neisseria gonorrhoeae","volume":"74","author":"Riedel","year":"2019","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Brennan-Krohn, T., and Kirby, J.E. (2019). Synergistic Combinations and Repurposed Antibiotics Active against the Pandrug-Resistant Klebsiella pneumoniae Nevada Strain. Antimicrob. Agents Chemother., 63.","DOI":"10.1128\/AAC.01374-19"},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Galani, I., Nafplioti, K., Adamou, P., Karaiskos, I., Giamarellou, H., and Souli, M. (2019). Nationwide epidemiology of carbapenem resistant Klebsiella pneumoniae isolates from Greek hospitals, with regards to plazomicin and aminoglycoside resistance. BMC Infect. Dis., 19.","DOI":"10.1186\/s12879-019-3801-1"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"9","DOI":"10.1089\/mdr.2019.0075","article-title":"Dissemination of International Clone II Acinetobacter baumannii Strains Coproducing OXA-23 Carbapenemase and 16S rRNA Methylase ArmA in Athens, Greece","volume":"26","author":"Nafplioti","year":"2020","journal-title":"Microb. Drug Resist."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Hao, M., Shi, X., Lv, J., Niu, S., Cheng, S., Du, H., Yu, F., Tang, Y.-W., Kreiswirth, B.N., and Zhang, H. (2020). In vitro Activity of Apramycin Against Carbapenem-Resistant and Hypervirulent Klebsiella pneumoniae Isolates. Front. Microbiol., 11.","DOI":"10.3389\/fmicb.2020.00425"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"439","DOI":"10.1016\/j.diagmicrobio.2016.09.002","article-title":"Evaluation of apramycin activity against carbapenem-resistant and -susceptible strains of Enterobacteriaceae","volume":"86","author":"Smith","year":"2016","journal-title":"Diagn. Microbiol. Infect. Dis."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"278","DOI":"10.1016\/j.jgar.2018.10.020","article-title":"Aminoglycoside-modifying enzyme and 16S ribosomal RNA methyltransferase genes among a global collection of Gram-negative isolates","volume":"16","author":"Costello","year":"2019","journal-title":"J. Glob. Antimicrob. Resist."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"792","DOI":"10.1089\/mdr.2017.0078","article-title":"Prevalence and Molecular Characterization of New Delhi Metallo-Beta-Lactamases in Multidrug-Resistant Pseudomonas aeruginosa and Acinetobacter baumannii from India","volume":"24","author":"Rahman","year":"2018","journal-title":"Microb. Drug Resist."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"278","DOI":"10.1016\/j.ijantimicag.2018.03.016","article-title":"High prevalence of 16S rRNA methyltransferases among carbapenemase-producing Enterobacteriaceae in the UK and Ireland","volume":"52","author":"Taylor","year":"2018","journal-title":"Int. J. Antimicrob. Agents"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"106019","DOI":"10.1016\/j.ijantimicag.2020.106019","article-title":"Co-occurrence of aminoglycoside and \u03b2-lactam resistance mechanisms in aminoglycoside- non-susceptible Escherichia coli isolated in the Zurich area, Switzerland","volume":"56","author":"Bodendoerfer","year":"2020","journal-title":"Int. J. Antimicrob. Agents"},{"key":"ref_33","first-page":"170","article-title":"Optimal treatment of campylobacter dysentery","volume":"13","author":"Eiland","year":"2008","journal-title":"J. Pediatr. Pharmacol. Ther."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"398","DOI":"10.1001\/jama.2014.17103","article-title":"Diagnosis and Treatment of Clostridium difficile in Adults","volume":"313","author":"Bagdasarian","year":"2015","journal-title":"JAMA"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"179","DOI":"10.1007\/BF00327434","article-title":"Genes for gentamicin-(3)-N-acetyltransferases III and IV: I. Nucleotide sequence of the AAC(3)-IV gene and possible involvement of an IS140 element in its expression","volume":"193","author":"Pilz","year":"1984","journal-title":"Mol. Gen. Genet."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"16275","DOI":"10.1021\/bi051777d","article-title":"The Kinetic Mechanism of AAC(3)-IV Aminoglycoside Acetyltransferase from Escherichia coli \u2020","volume":"44","author":"Magalhaes","year":"2005","journal-title":"Biochemistry"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"1131","DOI":"10.1093\/nar\/gkq810","article-title":"Design, construction and characterization of a set of insulated bacterial promoters","volume":"39","author":"Davis","year":"2011","journal-title":"Nucleic Acids Res."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"235","DOI":"10.1093\/nar\/28.1.235","article-title":"The Protein Data Bank","volume":"28","author":"Berman","year":"2000","journal-title":"Nucleic Acids Res."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"1605","DOI":"10.1002\/jcc.20084","article-title":"UCSF Chimera\u2014A visualization system for exploratory research and analysis","volume":"25","author":"Pettersen","year":"2004","journal-title":"J. Comput. Chem."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"10218","DOI":"10.1021\/acs.jmedchem.8b01393","article-title":"Encoding of Promiscuity in an Aminoglycoside Acetyltransferase","volume":"61","author":"Kumar","year":"2018","journal-title":"J. Med. Chem."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"eaas8667","DOI":"10.1126\/sciadv.aas8667","article-title":"A low-barrier hydrogen bond mediates antibiotic resistance in a noncanonical catalytic triad","volume":"4","author":"Kumar","year":"2018","journal-title":"Sci. Adv."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/21\/17\/6133\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T10:06:33Z","timestamp":1760177193000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/21\/17\/6133"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,8,25]]},"references-count":41,"journal-issue":{"issue":"17","published-online":{"date-parts":[[2020,9]]}},"alternative-id":["ijms21176133"],"URL":"https:\/\/doi.org\/10.3390\/ijms21176133","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,8,25]]}}}