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Distinguishing\n              <jats:italic>M. ulcerans<\/jats:italic>\n              from related species, including\n              <jats:italic>Mycobacterium marinum<\/jats:italic>\n              , poses challenges with respect to making accurate identifications. In this study, we developed a rapid and simple identification method based on mycobacterial lipid profiles and used matrix-assisted laser desorption\/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to analyze the lipid profiles of\n              <jats:italic>M. ulcerans<\/jats:italic>\n              (\n              <jats:italic>n<\/jats:italic>\n              = 35) and\n              <jats:italic>M. marinum<\/jats:italic>\n              (\n              <jats:italic>n<\/jats:italic>\n              = 19) isolates. Bacterial colonies pre-cultured on 2% Ogawa egg slants for 2 months were collected, and total lipids were extracted using an MBT Lipid Xtract kit. Spectra were obtained in negative ion mode using a MALDI Biotyper Sirius system, with ClinProTools v3.0 being used to analyze the spectra based on the application of three algorithms (genetic algorithm [GA], supervised neural network [SNN], and quick classifier [QC)]). Cross-validation was performed using a 20% leave-out set randomly selected from the samples. Models generated using GA, SNN, and QC showed cross-validation values of 100%, 100%, and 97.9%, respectively, and all algorithms achieved 100% recognition capability values. Our findings indicate that MALDI-TOF analysis of lipid profiles can accurately differentiate two mycobacterial species (\n              <jats:italic>M. ulcerans<\/jats:italic>\n              and\n              <jats:italic>M. marinum<\/jats:italic>\n              ) that are difficult to distinguish using conventional protein-targeting methods.\n            <\/jats:p>\n            <jats:sec>\n              <jats:title>IMPORTANCE<\/jats:title>\n              <jats:p>\n                Buruli ulcer, caused by\n                <jats:italic>Mycobacterium ulcerans<\/jats:italic>\n                , is a neglected tropical disease. However, distinguishing\n                <jats:italic>M. ulcerans<\/jats:italic>\n                from related species, including\n                <jats:italic>Mycobacterium marinum<\/jats:italic>\n                , presents certain challenges. In this study, we demonstrate the utility of a rapid yet simple method for differentiating isolates of these mycobacteria based on their lipid profiles using matrix-assisted laser desorption\/ionization time-of-flight mass spectrometry. This new approach can accurately identify species that are otherwise difficult to distinguish using conventional techniques. This represents a significant diagnostic advance for clinical laboratories, in that it enables a more rapid and precise identification, thereby leading to earlier treatment initiation and more appropriate treatment regimens for infections caused by these bacteria.\n              <\/jats:p>\n            <\/jats:sec>\n          <\/jats:sec>","DOI":"10.1128\/jcm.01400-24","type":"journal-article","created":{"date-parts":[[2025,1,27]],"date-time":"2025-01-27T14:00:45Z","timestamp":1737986445000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":2,"title":["A rapid and simple MALDI-TOF MS lipid profiling method for differentiating\n            <i>Mycobacterium ulcerans<\/i>\n            from\n            <i>Mycobacterium marinum<\/i>"],"prefix":"10.1128","volume":"63","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-5962-8295","authenticated-orcid":true,"given":"Takeshi","family":"Komine","sequence":"first","affiliation":[{"name":"Department of Mycobacteriology, Leprosy Research Center, National Institute of Infectious Diseases","place":["Higashimurayama, Japan"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2143-1160","authenticated-orcid":true,"given":"Hanako","family":"Fukano","sequence":"additional","affiliation":[{"name":"Department of Mycobacteriology, Leprosy Research Center, National Institute of Infectious Diseases","place":["Higashimurayama, Japan"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4061-6910","authenticated-orcid":true,"given":"Mitsunori","family":"Yoshida","sequence":"additional","affiliation":[{"name":"Department of Mycobacteriology, Leprosy Research Center, National Institute of Infectious Diseases","place":["Higashimurayama, Japan"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yuji","family":"Miyamoto","sequence":"additional","affiliation":[{"name":"Department of Mycobacteriology, Leprosy Research Center, National Institute of Infectious Diseases","place":["Higashimurayama, Japan"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Makoto","family":"Nakaya","sequence":"additional","affiliation":[{"name":"Department of Mycobacteriology, Leprosy Research Center, National Institute of Infectious Diseases","place":["Higashimurayama, Japan"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Azumi","family":"Fujinaga","sequence":"additional","affiliation":[{"name":"Application Department, Microbiology & Diagnostics MID Division, Bruker Japan K.K.","place":["Yokohama, Japan"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kohei","family":"Doke","sequence":"additional","affiliation":[{"name":"Application Department, Microbiology & Diagnostics MID Division, Bruker Japan K.K.","place":["Yokohama, Japan"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yoshihiko","family":"Hoshino","sequence":"additional","affiliation":[{"name":"Department of Mycobacteriology, Leprosy Research Center, National Institute of Infectious Diseases","place":["Higashimurayama, Japan"]}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"235","reference":[{"key":"e_1_3_3_2_2","doi-asserted-by":"publisher","DOI":"10.1016\/S1473-3099(06)70464-9"},{"key":"e_1_3_3_3_2","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.01041-07"},{"key":"e_1_3_3_4_2","doi-asserted-by":"publisher","DOI":"10.1093\/bmb\/ldp046"},{"key":"e_1_3_3_5_2","doi-asserted-by":"publisher","DOI":"10.3201\/eid2512.190427"},{"key":"e_1_3_3_6_2","doi-asserted-by":"publisher","DOI":"10.3201\/eid2712.203182"},{"key":"e_1_3_3_7_2","doi-asserted-by":"crossref","unstructured":"Suzuki K Luo Y Miyamoto Y Murase C Mikami-Sugawara M Yotsu RR. 2019. 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