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Lipid-based matrix-assisted laser desorption\/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) is a promising tool for identifying pathogenic mycobacterial species. This study aims to define and use species-specific lipid profiles of members of the\n                      <jats:italic toggle=\"yes\">Mycobacterium tuberculosis<\/jats:italic>\n                      complex (MTBC) obtained by MALDI-TOF MS, to directly discriminate\n                      <jats:italic toggle=\"yes\">M. tuberculosis<\/jats:italic>\n                      from other members of the MTBC, such as\n                      <jats:italic toggle=\"yes\">Mycobacterium africanum<\/jats:italic>\n                      ,\n                      <jats:italic toggle=\"yes\">Mycobacterium bovis<\/jats:italic>\n                      , and\n                      <jats:italic toggle=\"yes\">M. bovis<\/jats:italic>\n                      bacillus Calmette-Guerin (BCG). Reference strains (\n                      <jats:italic toggle=\"yes\">M. tuberculosis<\/jats:italic>\n                      H37Rv,\n                      <jats:italic toggle=\"yes\">M. africanum<\/jats:italic>\n                      ,\n                      <jats:italic toggle=\"yes\">M. bovis<\/jats:italic>\n                      , and\n                      <jats:italic toggle=\"yes\">M. bovis<\/jats:italic>\n                      BCG) were grown in Middlebrook 7H11 media (supplemented with 10% oleic acid-albumin-dextrose-catalase growth supplement) and incubated for up to 6 weeks at 37\u00b0C to generate the large biomass (~10\n                      <jats:sup>9<\/jats:sup>\n                      bacteria) required for optimization, to assess reproducibility of the assay, and to set the reference lipid database. In clinical use, standard shorter culture periods would be sufficient. A blinded study was then performed using a collection of 46 mycobacterial clinical isolate strains composed of 30\n                      <jats:italic toggle=\"yes\">M<\/jats:italic>\n                      .\n                      <jats:italic toggle=\"yes\">tuberculosis<\/jats:italic>\n                      , 2\n                      <jats:italic toggle=\"yes\">M<\/jats:italic>\n                      .\n                      <jats:italic toggle=\"yes\">africanum<\/jats:italic>\n                      , 9\n                      <jats:italic toggle=\"yes\">M<\/jats:italic>\n                      .\n                      <jats:italic toggle=\"yes\">bovis<\/jats:italic>\n                      BCG, and 5\n                      <jats:italic toggle=\"yes\">M<\/jats:italic>\n                      .\n                      <jats:italic toggle=\"yes\">bovis<\/jats:italic>\n                      and grown under the same conditions. Cultured mycobacteria were heat-inactivated and loaded onto the matrix-assisted laser desorption target, followed by the addition of the matrix. Acquisition of the data was done using the negative ion mode. Using the species-specific glycolipid, sulfolipids,\n                      <jats:italic toggle=\"yes\">M. tuberculosis<\/jats:italic>\n                      was discriminated within MTBC using the MALDI-TOF process with a sensitivity and specificity of 86.7% (95% confidence interval [CI] 69.3\u201396.2) and 93.7% (95% CI 69.8\u201399.8), respectively. Direct detection of\n                      <jats:italic toggle=\"yes\">M. tuberculosis<\/jats:italic>\n                      within the MTBC based on mycobacterial lipid profiling provides a safe and accurate method, based on the detection of the sulfolipids, as a species-specific lipid biomarker of\n                      <jats:italic toggle=\"yes\">M. tuberculosis<\/jats:italic>\n                      .\n                    <\/jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE<\/jats:title>\n                      <jats:p>\n                        Tuberculosis remains a major infectious disease in humans and mammals, but one of the major challenges is to accurately discriminate\n                        <jats:italic toggle=\"yes\">M. tuberculosis<\/jats:italic>\n                        from the other mycobacterial species belonging to the MTBC. Here we report on a novel assay that can detect\n                        <jats:italic toggle=\"yes\">M. tuberculosis<\/jats:italic>\n                        directly within the MTBC. This approach is simple and relies on the detection of species-specific lipids by routine MALDI-TOF MS.\n                      <\/jats:p>\n                    <\/jats:sec>\n                  <\/jats:sec>","DOI":"10.1128\/spectrum.00356-25","type":"journal-article","created":{"date-parts":[[2025,7,22]],"date-time":"2025-07-22T13:00:33Z","timestamp":1753189233000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["Performance of direct detection of\n                    <i>Mycobacterium tuberculosis<\/i>\n                    within\n                    <i>Mycobacterium tuberculosis<\/i>\n                    complex by routine MALDI-TOF for diagnosis using species-specific lipid fingerprint"],"prefix":"10.1128","volume":"13","author":[{"given":"Bosco","family":"Cheong","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/041kmwe10","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Life Sciences, Faculty of Natural Sciences, Centre for Bacterial Resistance Biology, Imperial College London","place":["London, United Kingdom"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Changchunzi","family":"He","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/041kmwe10","id-type":"ROR","asserted-by":"publisher"}],"name":"Adult Infectious Diseases and Center for Bacterial Resistance Biology, Department of Infectious Diseases, Imperial College London Department of Infectious Disease","place":["London, United Kingdom"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ian","family":"Laurenson","sequence":"additional","affiliation":[{"name":"Scottish Mycobacteria Reference Laboratory","place":["Edinburgh, United Kingdom"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Pauline","family":"Claxton","sequence":"additional","affiliation":[{"name":"Scottish Mycobacteria Reference Laboratory","place":["Edinburgh, United Kingdom"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Markus","family":"Kostrzewa","sequence":"additional","affiliation":[{"name":"Bruker Daltonics GmbH & Co. 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