{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,2]],"date-time":"2026-09-02T01:13:30Z","timestamp":1788311610431,"version":"build-2803163510"},"reference-count":46,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2025,6,9]],"date-time":"2025-06-09T00:00:00Z","timestamp":1749427200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Cell. Infect. Microbiol."],"abstract":"<jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>\n                      Although several World Health Organization\u2013endorsed targeted next-generation sequencing (tNGS) assays exist for tuberculosis (TB) drug-resistance detection, their target selection and diagnostic accuracy vary widely. In this study, we developed a novel tNGS workflow (the TB Pro assay) and evaluated its performance in identifying\n                      <jats:italic>Mycobacterium<\/jats:italic>\n                      species and predicting drug resistance.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>\n                      The TB Pro assay was validated for identifying 10\n                      <jats:italic>Mycobacterium tuberculosis<\/jats:italic>\n                      complex (MTBC) and 39 nontuberculous mycobacterial (NTM) species, as well as predicting resistance to 4 first-line and 13 second-line anti-TB drugs. The limit of detection (LOD) was determined using 11 reference strains spiked in sputum. The prediction of resistance to anti-tuberculous drugs\/drug classes was compared with phenotypic drug susceptibility testing (pDST) and whole-genome sequencing (WGS) using 435 clinical isolates.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      The assay demonstrated high sensitivity with a calculated LOD of 3.0 CFU\/ml for MTB and 1.4\u201316.2 CFU\/ml for most NTMs, except for\n                      <jats:italic>Mycobacterium intracellulare<\/jats:italic>\n                      with 117.9 CFU\/ml. Using pDST as the reference standard, the sensitivity of the TB Pro assay for the detection of resistance ranged from 74.3% (ethambutol) to 94.4% (rifampicin), with specificity values &amp;gt;98% for all drugs. Compared with WGS, the sensitivity of the TB Pro assay was over 98.0% for all drugs except pyrazinamide (66.7%), and the specificity values were all nearly 100.0%. Directly on sputum, the TB Pro assay showed 100% agreement with smear- and culture-positive sputum specimens.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>The TB Pro assay represents a sensitive and specific solution for simultaneous mycobacterial identification and comprehensive drug-resistance profiling, performing robustly on both cultured isolates and direct clinical specimens.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.3389\/fcimb.2025.1584237","type":"journal-article","created":{"date-parts":[[2025,6,9]],"date-time":"2025-06-09T01:25:40Z","timestamp":1749432340000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":7,"title":["A comprehensive evaluation of a novel targeted-sequencing workflow for Mycobacterium species identification and anti-tuberculosis drug-resistance detection"],"prefix":"10.3389","volume":"15","author":[{"given":"Xichao","family":"Ou","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shaojun","family":"Pei","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hongru","family":"Li","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhonghua","family":"Qin","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Richard","family":"Anthony","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jichun","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zexuan","family":"Song","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ruida","family":"Xing","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lixia","family":"Zhang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chong","family":"Teng","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hui","family":"Xia","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yang","family":"Zhou","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yuanyuan","family":"Song","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yang","family":"Zheng","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shengfen","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bing","family":"Zhao","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yanlin","family":"Zhao","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2025,6,9]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0277145","article-title":"High proportion of RR-TB and mutations conferring RR outside of the RRDR of the rpoB gene detected in GeneXpert MTB\/RIF assay positive pulmonary tuberculosis cases, in Addis Ababa, Ethiopia","volume":"17","author":"Akalu","year":"2022","journal-title":"PloS One"},{"key":"B2","doi-asserted-by":"publisher","first-page":"2495","DOI":"10.1128\/JCM.00128-10","article-title":"Evaluation of the analytical performance of the Xpert MTB\/RIF assay","volume":"48","author":"Blakemore","year":"2010","journal-title":"J. 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