{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,12]],"date-time":"2026-08-12T13:37:20Z","timestamp":1786541840666,"version":"build-2736575974"},"reference-count":22,"publisher":"American Society for Microbiology","issue":"8","license":[{"start":{"date-parts":[[2026,8,12]],"date-time":"2026-08-12T00:00:00Z","timestamp":1786492800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,8,12]],"date-time":"2026-08-12T00:00:00Z","timestamp":1786492800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["J Clin Microbiol"],"published-print":{"date-parts":[[2026,8,12]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:sec>\n                    <jats:title\/>\n                    <jats:p>\n                      <jats:italic toggle=\"yes\">Shigella flexneri 2a<\/jats:italic>\n                      is the most common cause of shigellosis, a major public health concern in developing countries. Rapid and reliable diagnostic tools are critical for timely outbreak detection and management. Leveraging clustered regularly interspaced short palindromic repeats (CRISPR) technology, we developed a CRISPR-Cas12a-based assay for the rapid and specific detection of\n                      <jats:italic toggle=\"yes\">S. flexneri 2a<\/jats:italic>\n                      and validated its performance using stool specimens from patients. Two guide RNAs targeting the\n                      <jats:italic toggle=\"yes\">gtrII<\/jats:italic>\n                      and\n                      <jats:italic toggle=\"yes\">gtrX<\/jats:italic>\n                      genes, unique markers of the\n                      <jats:italic toggle=\"yes\">S. flexneri 2a<\/jats:italic>\n                      serotype, were designed to ensure specificity. Recombinase polymerase amplification (RPA) was coupled with Cas12a-mediated collateral cleavage for signal amplification, with detection by fluorescence or lateral flow. Analytical sensitivity, specificity, and clinical accuracy were compared with conventional PCR using purified DNA and 588 clinical stool specimens. The CRISPR-Cas12a assay achieved a detection limit of 10 copies\/\u00b5L, comparable to PCR, and showed 100% analytical specificity without cross-reactivity to other bacteria. The isothermal reaction operated at room temperature and was completed within 1 h. Both readouts allowed visual interpretation without specialized equipment. Clinical validation of the CRISPR-Cas12a assay demonstrated a diagnostic sensitivity of 95% and specificity of 98%, comparable to PCR when evaluated using the same clinical specimens. This study provides two key advances: it establishes a CRISPR-Cas12a assay specifically targeting\n                      <jats:italic toggle=\"yes\">S. flexneri<\/jats:italic>\n                      2a, the predominant serotype, and validates it using a large clinical cohort. The assay\u2019s simplicity, speed, and high diagnostic accuracy make it a valuable tool for clinical diagnostics and field-based surveillance in resource-limited settings.\n                    <\/jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE<\/jats:title>\n                      <jats:p>\n                        Rapid and accessible diagnostics are essential for effective management of infectious diseases such as shigellosis. We developed a clustered regularly interspaced short palindromic repeats (CRISPR)-Cas12a-based assay that specifically detects\n                        <jats:italic toggle=\"yes\">Shigella flexneri 2a<\/jats:italic>\n                        , the predominant serotype responsible for the global disease burden. This assay integrates isothermal amplification with CRISPR-mediated detection to achieve low-copy detection (10 copies\/\u00b5L) within 1 h, eliminating the need for complex instrumentation. Dual fluorescence and lateral-flow readouts enable flexible use in both clinical laboratories and low-resource settings. The method\u2019s simplicity, accuracy, and adaptability demonstrate the practical potential of CRISPR diagnostics for point-of-care applications. By enabling rapid, on-site identification of\n                        <jats:italic toggle=\"yes\">S. flexneri 2a<\/jats:italic>\n                        , this approach can significantly improve clinical diagnosis and strengthen public health responses to enteric pathogen outbreaks.\n                      <\/jats:p>\n                    <\/jats:sec>\n                  <\/jats:sec>","DOI":"10.1128\/jcm.01615-25","type":"journal-article","created":{"date-parts":[[2026,7,20]],"date-time":"2026-07-20T14:13:21Z","timestamp":1784556801000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["CRISPR-Cas12a assay for rapid and specific detection of\n                    <i>Shigella flexneri 2a<\/i>\n                    in clinical samples"],"prefix":"10.1128","volume":"64","author":[{"given":"Jipei","family":"Liao","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/04rq5mt64","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Pathology, University of Maryland School of Medicine","place":["Baltimore, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yun","family":"Su","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/04523zj19","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Medicine, Nanjing University of Chinese Medicine","place":["Nanjing, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-8824-0325","authenticated-orcid":true,"given":"Feng","family":"Jiang","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/04rq5mt64","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Pathology, University of Maryland School of 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