{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,21]],"date-time":"2026-09-21T22:06:28Z","timestamp":1790028388812,"version":"4.0.1"},"reference-count":35,"publisher":"American Society for Microbiology","issue":"12","license":[{"start":{"date-parts":[[2024,12,5]],"date-time":"2024-12-05T00:00:00Z","timestamp":1733356800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2024,12,5]],"date-time":"2024-12-05T00:00:00Z","timestamp":1733356800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"funder":[{"name":"Yunnan Major Scientific and Technological Project","award":["202202AG050013"],"award-info":[{"award-number":["202202AG050013"]}]},{"name":"Yunnan Provincial Department of Science and Technology Kunming Medical University Applied Basic Research Joint Project","award":["202001AY070001-104"],"award-info":[{"award-number":["202001AY070001-104"]}]},{"name":"Reserve Talents Project for Young and Middle-Aged Academic and Technical Leaders of Yunnan Province","award":["202305AC160021"],"award-info":[{"award-number":["202305AC160021"]}]},{"name":"Reserve Talent Program of Yunnan Provincial Health Commission","award":["H-2019044"],"award-info":[{"award-number":["H-2019044"]}]}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["Microbiol Spectr"],"published-print":{"date-parts":[[2024,12,5]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n          <jats:sec>\n            <jats:title\/>\n            <jats:p>\n              Tuberculosis (TB) remains a major global health problem, and there is an urgent need for rapid, sensitive, and easy-to-use diagnostic technologies to improve TB diagnosis. In this study, we developed the nested multi-enzyme isothermal rapid amplification (nestMIRA) assay for TB. We designed several pairs of primers and probes targeting the IS6110 sequence of\n              <jats:italic>Mycobacterium tuberculosis<\/jats:italic>\n              (Mtb) and performed combinatorial testing to optimize the performance of the TB nestMIRA assay. The reaction can be performed at a constant temperature of approximately 40\u00b0C and completed within 30 minutes in the same tube without opening the central cap. There has been no cross-reactivity with common\n              <jats:italic>non-tuberculous mycobacteria<\/jats:italic>\n              (NTB) and respiratory pathogens. The TB nestMIRA assay has a minimum detection limit of 5 copies\/\u03bcL for H37Rv genomic DNA and a limit of quantification of 100 CFU\/ml. To test the diagnostic performance of the TB nestMIRA assay, we conducted a 163-person clinical cohort study using comprehensive reference standards as the gold standard for clinical diagnosis. Our study showed that TB nestMIRA performed slightly better than GeneXpert MTB\/RIF (Xpert) (85.27% vs. 82.17%) and significantly better than culture (55.81%) and acid-fast bacillus (AFB) smear (38.76%). The TB nestMIRA assay offers speed, specificity, sensitivity, and convenience. We believe that it has the potential to be a rapid alternative for TB diagnosis, particularly in resource-limited settings.\n            <\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>IMPORTANCE<\/jats:title>\n            <jats:p>\n              In this study, we have successfully developed a method called nested multi-enzyme isothermal rapid amplification (nestMIRA) for the detection of\n              <jats:italic>Mycobacterium tuberculosis<\/jats:italic>\n              (Mtb). This method involves a two-step thermostatic amplification process in the same tube and can be read using fluorescence and lateral flow dipstick (LFD) assays. It is known to be rapid, specific, and highly sensitive. Our method has shown promising results in the detection of clinical specimens, and we believe that it can be a valuable tool for the rapid detection of Mtb in a clinical setting.\n            <\/jats:p>\n          <\/jats:sec>","DOI":"10.1128\/spectrum.00887-24","type":"journal-article","created":{"date-parts":[[2024,10,28]],"date-time":"2024-10-28T13:00:14Z","timestamp":1730120414000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":6,"title":["Rapid and sensitive detection of\n            <i>Mycobacterium tuberculosis<\/i>\n            using nested multi-enzyme isothermal rapid amplification in a single reaction"],"prefix":"10.1128","volume":"12","author":[{"given":"Yingchao","family":"Chang","sequence":"first","affiliation":[{"name":"Faculty of Life Science and Technology, Kunming University of Science and Technology","place":["Kunming, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mi","family":"Zhang","sequence":"additional","affiliation":[{"name":"Medical Laboratory, Yunnan Provincial Hospital of Infectious Disease","place":["Kunming, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gaowen","family":"Liu","sequence":"additional","affiliation":[{"name":"Yunnan Kecan Biotechnology Co., Ltd","place":["Kunming, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xinlin","family":"Wu","sequence":"additional","affiliation":[{"name":"Faculty of Life Science and Technology, Kunming University of Science and Technology","place":["Kunming, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qiaolu","family":"Yan","sequence":"additional","affiliation":[{"name":"Department of Respiratory and Critical Care Medicine, Dali Bai Autonomous Prefecture People's Hospital","place":["Dali, Yunnan, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Cuixian","family":"Yang","sequence":"additional","affiliation":[{"name":"Medical Laboratory, Yunnan Provincial Hospital of Infectious Disease","place":["Kunming, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0007-1262-2025","authenticated-orcid":false,"given":"Li","family":"Liu","sequence":"additional","affiliation":[{"name":"Faculty of Life Science and Technology, Kunming University of Science and Technology","place":["Kunming, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2718-0836","authenticated-orcid":false,"given":"Yue","family":"Feng","sequence":"additional","affiliation":[{"name":"Faculty of Life Science and Technology, Kunming University of Science and Technology","place":["Kunming, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1120-8870","authenticated-orcid":false,"given":"Xueshan","family":"Xia","sequence":"additional","affiliation":[{"name":"Yunnan Provincial Key Laboratory of Public Health and Biosafety, Kunming Medical University","place":["Kunming, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"235","reference":[{"key":"e_1_3_4_3_2","doi-asserted-by":"publisher","DOI":"10.4103\/ijabmr.ijabmr_348_23"},{"key":"e_1_3_4_4_2","doi-asserted-by":"publisher","DOI":"10.1155\/2022\/8078639"},{"key":"e_1_3_4_7_2","doi-asserted-by":"publisher","DOI":"10.1016\/S2666-5247(22)00087-8"},{"key":"e_1_3_4_8_2","doi-asserted-by":"publisher","DOI":"10.1111\/lam.13364"},{"key":"e_1_3_4_9_2","doi-asserted-by":"publisher","DOI":"10.1099\/jmm.0.000164"},{"key":"e_1_3_4_10_2","doi-asserted-by":"publisher","DOI":"10.1080\/22221751.2021.1959271"},{"key":"e_1_3_4_11_2","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.01528-08"},{"key":"e_1_3_4_12_2","doi-asserted-by":"publisher","DOI":"10.1007\/s10096-023-04626-5"},{"key":"e_1_3_4_13_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41429-023-00639-6"},{"key":"e_1_3_4_14_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.ab.2021.114341"},{"key":"e_1_3_4_15_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmolb.2021.763079"},{"key":"e_1_3_4_16_2","doi-asserted-by":"publisher","DOI":"10.1080\/22221751.2020.1793689"},{"key":"e_1_3_4_17_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.aca.2023.341263"},{"key":"e_1_3_4_18_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.diagmicrobio.2023.116067"},{"key":"e_1_3_4_20_2","volume-title":"Treatment of tuberculosis: guidelines, 4th Ed","author":"World Health Organization","year":"2010","unstructured":"World Health Organization. 2010. Treatment of tuberculosis: guidelines, 4th Ed. World Health Organization, Geneva."},{"key":"e_1_3_4_21_2","doi-asserted-by":"publisher","DOI":"10.1093\/cid\/ciw035"},{"key":"e_1_3_4_22_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2016.00030"},{"key":"e_1_3_4_23_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41564-023-01365-3"},{"key":"e_1_3_4_24_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0253235"},{"key":"e_1_3_4_25_2","doi-asserted-by":"publisher","DOI":"10.1002\/jcla.22655"},{"key":"e_1_3_4_26_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.mimet.2017.11.003"},{"key":"e_1_3_4_27_2","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.02214-10"},{"key":"e_1_3_4_28_2","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.01463-09"},{"key":"e_1_3_4_29_2","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.00128-10"},{"key":"e_1_3_4_30_2","doi-asserted-by":"publisher","DOI":"10.1128\/mBio.00812-17"},{"key":"e_1_3_4_31_2","doi-asserted-by":"publisher","DOI":"10.1186\/s12929-023-00932-4"},{"key":"e_1_3_4_32_2","doi-asserted-by":"publisher","DOI":"10.3389\/fbioe.2023.1233353"},{"key":"e_1_3_4_33_2","doi-asserted-by":"publisher","DOI":"10.1016\/S1473-3099(17)30691-6"},{"key":"e_1_3_4_34_2","doi-asserted-by":"publisher","DOI":"10.1080\/22221751.2019.1664939"},{"key":"e_1_3_4_35_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.jinf.2021.04.032"},{"key":"e_1_3_4_36_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2023.1117085"},{"key":"e_1_3_4_37_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmoldx.2020.04.212"},{"key":"e_1_3_4_38_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.talanta.2023.124645"},{"key":"e_1_3_4_39_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.jhazmat.2023.131487"},{"key":"e_1_3_4_40_2","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2023.1172035"}],"container-title":["Microbiology Spectrum"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/journals.asm.org\/doi\/pdf\/10.1128\/spectrum.00887-24","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/journals.asm.org\/doi\/pdf\/10.1128\/spectrum.00887-24","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2024,12,5]],"date-time":"2024-12-05T14:07:58Z","timestamp":1733407678000},"score":1,"resource":{"primary":{"URL":"https:\/\/journals.asm.org\/doi\/10.1128\/spectrum.00887-24"}},"subtitle":[],"editor":[{"given":"Vittal Prakash","family":"Ponraj","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2024,12,5]]},"references-count":35,"journal-issue":{"issue":"12","published-print":{"date-parts":[[2024,12,5]]}},"alternative-id":["10.1128\/spectrum.00887-24"],"URL":"https:\/\/doi.org\/10.1128\/spectrum.00887-24","relation":{},"ISSN":["2165-0497"],"issn-type":[{"value":"2165-0497","type":"electronic"}],"subject":[],"published":{"date-parts":[[2024,12,5]]},"assertion":[{"value":"2024-04-08","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2024-09-03","order":2,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2024-10-28","order":3,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}],"article-number":"e00887-24"}}