{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,13]],"date-time":"2026-10-13T15:41:09Z","timestamp":1791906069317,"version":"4.3.4"},"reference-count":29,"publisher":"Oxford University Press (OUP)","issue":"5","license":[{"start":{"date-parts":[[2026,5,1]],"date-time":"2026-05-01T00:00:00Z","timestamp":1777593600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/academic.oup.com\/journals\/pages\/open_access\/funder_policies\/chorus\/standard_publication_model"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,5,1]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Terbinafine (TBF) is a first-line allylamine antifungal used to treat dermatophytosis caused by Trichophyton species, targeting squalene epoxidase (SQLE). The recent emergence of TBF-resistant Trichophyton rubrum (T. rubrum) strains represents a significant global clinical challenge. Although the SQLE Leu393Phe substitution is a commonly reported resistance mechanism in Japan, direct genetic evidence demonstrating its effect when introduced into a T. rubrum recipient strain has been lacking. In this study, we established the causal relationship between the SQLE Leu393Phe substitution and TBF resistance via targeted gene replacement in T. rubrum. A homologous recombination plasmid containing SQLE from the TBF-resistant T. rubrum strain BGUTR13, which harbours the 1177TTA\u2192TTC mutation encoding Leu393Phe, together with the neomycin phosphotransferase II (nptII) selection marker, was constructed and introduced into the TBF-susceptible strain T. rubrum CBS118892 \u0394ku80. The resulting mutant transformants exhibited a &amp;gt; 8000-fold increase in TBF minimum inhibitory concentration compared with the parental strain, whereas control transformants carrying nptII but lacking the SQLE Leu393Phe substitution remained susceptible. Structural modelling indicated that the Leu393Phe substitution causes substantial steric hindrance within the SQLE binding pocket, displacing TBF\u2019s aliphatic chain and disrupting key hydrophobic interactions. Molecular docking simulations predicted reduced TBF binding affinity (\u0394G changed from \u22126.3 to \u22120.8 kcal\/mol) following the mutation. These findings provide the first robust genetic evidence that the single SQLE Leu393Phe substitution is sufficient to confer high-level TBF resistance in T. rubrum. These findings highlight the importance of monitoring SQLE mutations in clinical isolates and provide a foundation for developing strategies to address antifungal resistance in dermatophytes.<\/jats:p>","DOI":"10.1093\/mmy\/myag044","type":"journal-article","created":{"date-parts":[[2026,5,7]],"date-time":"2026-05-07T11:45:02Z","timestamp":1778154302000},"source":"Crossref","is-referenced-by-count":1,"title":["Genetic confirmation of terbinafine resistance in\n                    <i>Trichophyton rubrum<\/i>\n                    mediated by the squalene epoxidase Leu393Phe mutation via targeted gene replacement"],"prefix":"10.1093","volume":"64","author":[{"ORCID":"https:\/\/orcid.org\/0009-0005-2459-5955","authenticated-orcid":false,"given":"Shusaku","family":"Suzuki","sequence":"first","affiliation":[{"name":"Graduate School of Health Care Science, Bunkyo Gakuin University , Tokyo 113-8668 ,","place":["Japan"]},{"name":"Japanese Red Cross Kanto-Koshinetsu Block Blood Center , Tokyo 135-8521 ,","place":["Japan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Masaki","family":"Ishii","sequence":"additional","affiliation":[{"name":"Research Institute of Pharmaceutical Sciences, Faculty of Pharmacy, Musashino University , Tokyo 202-8585 ,","place":["Japan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shinya","family":"Ohata","sequence":"additional","affiliation":[{"name":"Research Institute of Pharmaceutical Sciences, Faculty of Pharmacy, Musashino University , Tokyo 202-8585 ,","place":["Japan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yoko","family":"Mano","sequence":"additional","affiliation":[{"name":"Graduate School of Health Care Science, Bunkyo Gakuin University , Tokyo 113-8668 ,","place":["Japan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nobuhiko","family":"Furuya","sequence":"additional","affiliation":[{"name":"Graduate School of Health Care Science, Bunkyo Gakuin University , Tokyo 113-8668 ,","place":["Japan"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"286","published-online":{"date-parts":[[2026,5,7]]},"reference":[{"issue":"1","key":"2026052004413649700_bib1","first-page":"66","article-title":"Large-scale epidemiological survey on the actual situation and latent morbidity of tinea pedis and tinea unguium (Foot Check 2023)","volume":"42","author":"Hata","year":"2025","journal-title":"Jpn J Clin Dermatol"},{"issue":"3","key":"2026052004413649700_bib2","doi-asserted-by":"publisher","first-page":"177","DOI":"10.1265\/jjh.72.177","article-title":"Epidemiological study on Trichophyton disseminating from the feet of the elderly","volume":"72","author":"Suzuki","year":"2017","journal-title":"Nihon Eiseigaku Zasshi"},{"issue":"6","key":"2026052004413649700_bib3","doi-asserted-by":"publisher","first-page":"462","DOI":"10.7883\/yoken.JJID.2018.090","article-title":"Molecular epidemiological analysis of the spreading conditions of Trichophyton in long-term care facilities in Japan","volume":"71","author":"Suzuki","year":"2018","journal-title":"Jpn J Infect Dis"},{"issue":"8","key":"2026052004413649700_bib4","first-page":"612","article-title":"Relationship between foot problems, fall experience and fear of falling among Japanese community-dwelling elderly","volume":"57","author":"Harada","year":"2010","journal-title":"Nihon Koshu Eisei Zasshi"},{"issue":"4","key":"2026052004413649700_bib5","doi-asserted-by":"publisher","first-page":"85","DOI":"10.3314\/mmj.23-00008","article-title":"2021 Epidemiological survey of dermatomycoses in Japan","volume":"64","author":"Nakamura","year":"2023","journal-title":"Med Mycol J"},{"issue":"5","key":"2026052004413649700_bib6","doi-asserted-by":"publisher","first-page":"472","DOI":"10.3390\/antibiotics14050472","article-title":"Terbinafine resistance in Trichophyton rubrum and Trichophyton indotineae: a literature review","volume":"14","author":"Gupta","year":"2025","journal-title":"Antibiotics (Basel)"},{"issue":"3","key":"2026052004413649700_bib7","doi-asserted-by":"publisher","first-page":"151","DOI":"10.4265\/bio.23.151","article-title":"Discovery of terbinafine low-susceptibility Trichophyton rubrum strain in Japan","volume":"23","author":"Suzuki","year":"2018","journal-title":"Biocontrol Sci"},{"issue":"4","key":"2026052004413649700_bib8","doi-asserted-by":"publisher","first-page":"564","DOI":"10.1111\/1346-8138.15745","article-title":"Epidemiological study of terbinafine-resistant dermatophytes isolated from Japanese patients","volume":"48","author":"Hiruma","year":"2021","journal-title":"J Dermatol"},{"issue":"8","key":"2026052004413649700_bib9","doi-asserted-by":"publisher","first-page":"1068","DOI":"10.1111\/1346-8138.16780","article-title":"Epidemiological study of antifungal-resistant dermatophytes isolated from Japanese patients","volume":"50","author":"Hiruma","year":"2023","journal-title":"J Dermatol"},{"issue":"7","key":"2026052004413649700_bib10","doi-asserted-by":"publisher","first-page":"e00115","DOI":"10.1128\/AAC.00115-17","article-title":"Terbinafine resistance of Trichophyton clinical isolates caused by specific point mutations in the squalene epoxidase gene","volume":"61","author":"Yamada","year":"2017","journal-title":"Antimicrob Agents Chemother"},{"issue":"2","key":"2026052004413649700_bib11","doi-asserted-by":"publisher","first-page":"150","DOI":"10.3390\/jof8020150","article-title":"Increasing terbinafine resistance in Danish Trichophyton isolates 2019\u20132020","volume":"8","author":"Astvad","year":"2022","journal-title":"J Fungi (Basel)"},{"issue":"10","key":"2026052004413649700_bib12","doi-asserted-by":"publisher","first-page":"e01126","DOI":"10.1128\/AAC.01126-19","article-title":"Emerging terbinafine resistance in Trichophyton: clinical characteristics, squalene epoxidase gene mutations, and a reliable EUCAST method for detection","volume":"63","author":"Saunte","year":"2019","journal-title":"Antimicrob Agents Chemother"},{"issue":"1","key":"2026052004413649700_bib13","doi-asserted-by":"publisher","first-page":"96","DOI":"10.1186\/s13568-024-01755-8","article-title":"An efficient gene targeting system using \u0394ku80 and functional analysis of Cyp51A in Trichophyton rubrum","volume":"14","author":"Ishii","year":"2024","journal-title":"AMB Expr"},{"key":"2026052004413649700_bib14","article-title":"Reference method for broth dilution antifungal susceptibility testing of filamentous fungi; approved standard","volume-title":"CLSI document M38-A2","author":"Clinical and Laboratory Standards Institute","year":"2008","edition":"2nd ed."},{"issue":"9","key":"2026052004413649700_bib15","doi-asserted-by":"publisher","first-page":"2629","DOI":"10.1128\/JCM.36.9.2629-2633.1998","article-title":"Phylogenetic classification of Trichophyton mentagrophytes complex strains based on DNA sequences of nuclear ribosomal internal transcribed spacer 1 regions","volume":"36","author":"Makimura","year":"1998","journal-title":"J Clin Microbiol"},{"issue":"5","key":"2026052004413649700_bib16","doi-asserted-by":"publisher","first-page":"645","DOI":"10.1007\/s00438-002-0780-4","article-title":"Efficient disruption of a polyketide synthase gene (pks1) required for melanin synthesis through Agrobacterium-mediated transformation of Glarea lozoyensis","volume":"268","author":"Zhang","year":"2003","journal-title":"Mol Genet Genomics"},{"issue":"11","key":"2026052004413649700_bib17","doi-asserted-by":"publisher","first-page":"e00598","DOI":"10.1128\/aac.00598-25","article-title":"Synergistic effects of Cyp51 isozyme-specific azole antifungal agents on fungi with multiple cyp51 isozyme genes","volume":"69","author":"Ishii","year":"2025","journal-title":"Antimicrob Agents Chemother"},{"issue":"7873","key":"2026052004413649700_bib18","doi-asserted-by":"publisher","first-page":"583","DOI":"10.1038\/s41586-021-03819-2","article-title":"Highly accurate protein structure prediction with AlphaFold","volume":"596","author":"Jumper","year":"2021","journal-title":"Nature"},{"issue":"1","key":"2026052004413649700_bib19","doi-asserted-by":"publisher","first-page":"97","DOI":"10.1038\/s41467-018-07928-x","article-title":"Structure and inhibition mechanism of the catalytic domain of human squalene epoxidase","volume":"10","author":"Padyana","year":"2019","journal-title":"Nat Commun"},{"issue":"1","key":"2026052004413649700_bib20","doi-asserted-by":"publisher","first-page":"e14232","DOI":"10.2196\/14232","article-title":"Molecular docking using Chimera and AutoDock Vina software for nonbioinformaticians","volume":"1","author":"Butt","year":"2020","journal-title":"JMIR Bioinform Biotechnol"},{"issue":"6","key":"2026052004413649700_bib21","doi-asserted-by":"publisher","first-page":"741","DOI":"10.1016\/j.jiac.2022.02.010","article-title":"A deletion mutation in the amino acid sequence of squalene epoxidase in terbinafine-resistant Trichophyton rubrum","volume":"28","author":"Kano","year":"2022","journal-title":"J Infect Chemother"},{"issue":"8","key":"2026052004413649700_bib22","doi-asserted-by":"publisher","first-page":"936","DOI":"10.1111\/myc.13286","article-title":"Itraconazole resistance of Trichophyton rubrum mediated by the ABC transporter TruMDR2","volume":"64","author":"Yamada","year":"2021","journal-title":"Mycoses"},{"issue":"1","key":"2026052004413649700_bib23","doi-asserted-by":"publisher","first-page":"21","DOI":"10.3314\/mmj.20-00014","article-title":"ATP-binding cassette (ABC) transporter proteins in highly terbinafine-resistant strains of Trichophyton indotineae (formerly Trichophyton interdigitale)","volume":"62","author":"Kano","year":"2021","journal-title":"Med Mycol J"},{"issue":"6","key":"2026052004413649700_bib24","doi-asserted-by":"publisher","first-page":"947","DOI":"10.1007\/s11046-020-00455-8","article-title":"Trichophyton indotineae sp. nov.: a new highly terbinafine-resistant anthropophilic dermatophyte species","volume":"185","author":"Kano","year":"2020","journal-title":"Mycopathologia"},{"issue":"5","key":"2026052004413649700_bib25","doi-asserted-by":"publisher","first-page":"e02522","DOI":"10.1128\/AAC.02522-17","article-title":"Mutation in the squalene epoxidase gene of Trichophyton interdigitale and Trichophyton rubrum associated with allylamine resistance","volume":"62","author":"Rudramurthy","year":"2018","journal-title":"Antimicrob Agents Chemother"},{"issue":"7","key":"2026052004413649700_bib26","doi-asserted-by":"publisher","first-page":"477","DOI":"10.1111\/myc.12772","article-title":"High terbinafine resistance in Trichophyton interdigitale isolates in Delhi, India harbouring mutations in the squalene epoxidase gene","volume":"61","author":"Singh","year":"2018","journal-title":"Mycoses"},{"issue":"1","key":"2026052004413649700_bib27","doi-asserted-by":"publisher","first-page":"18612","DOI":"10.1038\/s41598-025-03300-4","article-title":"Computational analysis of missense mutations in squalene epoxidase associated with terbinafine resistance in clinically reported dermatophytes","volume":"15","author":"Mahmood","year":"2025","journal-title":"Sci Rep"},{"issue":"2","key":"2026052004413649700_bib28","doi-asserted-by":"publisher","first-page":"455","DOI":"10.1021\/ci100403b","article-title":"Detailed mechanism of squalene epoxidase inhibition by terbinafine: structural and docking analysis","volume":"51","author":"Nowosielski","year":"2011","journal-title":"J Chem Inf Model"},{"issue":"5","key":"2026052004413649700_bib29","doi-asserted-by":"publisher","first-page":"e0160923","DOI":"10.1128\/aac.01609-23","article-title":"The Ptk2-Pma1 pathway enhances tolerance to terbinafine in Trichophyton rubrum","volume":"68","author":"Ishii","year":"2024","journal-title":"Antimicrob Agents Chemother"}],"container-title":["Medical Mycology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/academic.oup.com\/mmy\/advance-article-pdf\/doi\/10.1093\/mmy\/myag044\/68244524\/myag044.pdf","content-type":"application\/pdf","content-version":"am","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/mmy\/article-pdf\/64\/5\/myag044\/68244524\/myag044.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/mmy\/article-pdf\/64\/5\/myag044\/68244524\/myag044.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,5,20]],"date-time":"2026-05-20T08:41:44Z","timestamp":1779266504000},"score":1,"resource":{"primary":{"URL":"https:\/\/academic.oup.com\/mmy\/article\/doi\/10.1093\/mmy\/myag044\/8671962"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,5,1]]},"references-count":29,"journal-issue":{"issue":"5","published-print":{"date-parts":[[2026,5,1]]}},"URL":"https:\/\/doi.org\/10.1093\/mmy\/myag044","relation":{},"ISSN":["1369-3786","1460-2709"],"issn-type":[{"value":"1369-3786","type":"print"},{"value":"1460-2709","type":"electronic"}],"subject":[],"published-other":{"date-parts":[[2026,5]]},"published":{"date-parts":[[2026,5,1]]},"article-number":"myag044"}}