{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,7]],"date-time":"2026-10-07T06:49:38Z","timestamp":1791355778807,"version":"4.3.1"},"reference-count":37,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,10,7]],"date-time":"2026-10-07T00:00:00Z","timestamp":1791331200000},"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. Microbiol."],"abstract":"<jats:p>\n                    Community-acquired pneumonia (CAP) is a global public health burden with high morbidity, but timely diagnosis is hindered by limitations of conventional methods. We validated a novel multiplex real-time PCR (mRT-PCR) for CAP diagnosis in the emergency department, focusing on\n                    <jats:italic>Mycoplasma pneumoniae<\/jats:italic>\n                    (\n                    <jats:italic>M. pneumoniae<\/jats:italic>\n                    ). A retrospective cohort of 371 CAP patients (2019\u20132021) from the Chinese PLA General Hospital was enrolled. The panel targeted 85 pathogens (40 bacteria, 18 viruses, 17 fungi, 10 other) plus 8 resistance genes, with a turnaround time (TAT) under 4\u202fh and a cost of approximately US$100 per test. mRT-PCR showed performance comparable to mNGS for common pathogens (concordance &amp;gt;69% for bacteria\/fungi) and high concordance with the final clinical diagnosis. Notably, it detected\n                    <jats:italic>M. pneumoniae<\/jats:italic>\n                    in 36.7% (136\/371) of cases, outperforming conventional methods by identifying 204 exclusive positives (55.0% of cohort), with\n                    <jats:italic>M. pneumoniae<\/jats:italic>\n                    (40%) and viruses (30%) predominating. Polymicrobial infections occurred in 36.1% of cases;\n                    <jats:italic>M. pneumoniae<\/jats:italic>\n                    co-infections (38 cases) showed elevated neutrophils and reduced lymphocytes vs. single infections. An independent reference standard analysis excluding both index tests showed that 24.1% (210\/873) of calls were independently corroborated - high for influenza, SARS-CoV-2 and\n                    <jats:italic>M. pneumoniae<\/jats:italic>\n                    , lower for\n                    <jats:italic>S. pneumoniae<\/jats:italic>\n                    , consistent with the limited sensitivity of sputum culture. Unlike conventional syndromic panels such as FilmArray, which profile fewer targets without concurrent resistance-gene detection, this mRT-PCR combines broad coverage with rapid TAT and low cost, making it promising for timely CAP diagnosis, particularly in resource-limited settings and for targeted therapy.\n                  <\/jats:p>","DOI":"10.3389\/fmicb.2026.1979614","type":"journal-article","created":{"date-parts":[[2026,10,7]],"date-time":"2026-10-07T05:51:32Z","timestamp":1791352292000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":0,"title":["Clinical validation of a novel multiplex real-time PCR for diagnosing community-acquired pneumonia in the emergency department: a promising diagnostic tool for Mycoplasma pneumoniae"],"prefix":"10.3389","volume":"17","author":[{"given":"Liuyang","family":"Yang","sequence":"first","affiliation":[{"name":"Department of Laboratory Medicine, The First Medical Center, Chinese PLA General Hospital","place":["Beijing, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chaomin","family":"Guo","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, The First Medical Center, Chinese PLA General Hospital","place":["Beijing, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qiang","family":"Zhao","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, The First Medical Center, Chinese PLA General Hospital","place":["Beijing, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiyong","family":"Yang","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, The First Medical Center, Chinese PLA General Hospital","place":["Beijing, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Heng","family":"Zhang","sequence":"additional","affiliation":[{"name":"Department of Emergency, The First Medical Center, Chinese PLA General Hospital, Medical School of Chinese PLA","place":["Beijing, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yongzhi","family":"Zhai","sequence":"additional","affiliation":[{"name":"Department of Emergency, The First Medical Center, Chinese PLA General Hospital, Medical School of Chinese PLA","place":["Beijing, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yanning","family":"Ma","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, The First Medical Center, Chinese PLA General Hospital","place":["Beijing, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2026,10,7]]},"reference":[{"key":"ref1","doi-asserted-by":"publisher","first-page":"2050","DOI":"10.4081\/monaldi.2021.2050","article-title":"Association between neutrophil to lymphocyte ratio and mortality among community acquired pneumonia patients: a meta-analysis","volume":"92","author":"Alzoubi","year":"2021","journal-title":"Monaldi Arch. Chest Dis."},{"key":"ref2","doi-asserted-by":"publisher","first-page":"123","DOI":"10.1186\/s40168-021-01067-0","article-title":"Pre-digest of unprotected DNA by Benzonase improves the representation of living skin bacteria and efficiently depletes host DNA","volume":"9","author":"Amar","year":"2021","journal-title":"Microbiome"},{"key":"ref3","doi-asserted-by":"publisher","first-page":"2114","DOI":"10.1093\/bioinformatics\/btu170","article-title":"Trimmomatic: a flexible trimmer for Illumina sequence data","volume":"30","author":"Bolger","year":"2014","journal-title":"Bioinformatics"},{"key":"ref4","doi-asserted-by":"publisher","first-page":"1320","DOI":"10.1111\/crj.12674","article-title":"Diagnosis and treatment of community-acquired pneumonia in adults: 2016 clinical practice guidelines by the Chinese thoracic society, Chinese Medical Association","volume":"12","author":"Cao","year":"2018","journal-title":"Clin. Respir. J."},{"key":"ref5","volume-title":"Performance Standards for Antimicrobial Susceptibility Testing. 33rd ed. CLSI Supplement M100","year":"2023"},{"key":"ref6","doi-asserted-by":"publisher","first-page":"3001","DOI":"10.1007\/s00431-024-05519-1","article-title":"Challenges in the treatment of pediatric Mycoplasma pneumoniae pneumonia","volume":"183","author":"Ding","year":"2024","journal-title":"Eur. J. Pediatr."},{"key":"ref7","doi-asserted-by":"publisher","first-page":"1171","DOI":"10.1007\/s10096-019-03526-x","article-title":"Comparison of Unyvero P55 pneumonia cartridge, in-house PCR and culture for the identification of respiratory pathogens and antibiotic resistance in bronchoalveolar lavage fluids in the critical care setting","volume":"38","author":"Gadsby","year":"2019","journal-title":"Eur. J. Clin. Microbiol. Infect. Dis."},{"key":"ref8","doi-asserted-by":"publisher","first-page":"2386636","DOI":"10.1080\/07853890.2024.2386636","article-title":"Laboratory diagnosis and treatment of Mycoplasma pneumoniae infection in children: a review","volume":"56","author":"Gao","year":"2024","journal-title":"Ann. Med."},{"key":"ref9","doi-asserted-by":"publisher","DOI":"10.7754\/Clin.Lab.2018.181042","article-title":"Neutrophil-to-lymphocyte ratio in adult community-acquired pneumonia patients correlates with unfavorable clinical outcomes","volume":"65","author":"Ge","year":"2019","journal-title":"Clin. Lab."},{"key":"ref10","doi-asserted-by":"publisher","first-page":"115","DOI":"10.1038\/s41591-020-1105-z","article-title":"Rapid pathogen detection by metagenomic next-generation sequencing of infected body fluids","volume":"27","author":"Gu","year":"2020","journal-title":"Nat. Med."},{"key":"ref11","doi-asserted-by":"publisher","first-page":"668","DOI":"10.1080\/1040841x.2019.1681933","article-title":"mNGS in clinical microbiology laboratories: on the road to maturity","volume":"45","author":"Han","year":"2019","journal-title":"Crit. Rev. Microbiol."},{"key":"ref12","doi-asserted-by":"publisher","first-page":"1407","DOI":"10.1056\/nejm197312272892607","article-title":"Fever of undetermined origin","volume":"289","author":"Jacoby","year":"1973","journal-title":"N. Engl. J. Med."},{"key":"ref13","doi-asserted-by":"publisher","first-page":"415","DOI":"10.1056\/NEJMoa1500245","article-title":"Community-acquired pneumonia requiring hospitalization among U.S. adults","volume":"373","author":"Jain","year":"2015","journal-title":"N. Engl. J. Med."},{"key":"ref14","doi-asserted-by":"publisher","first-page":"102","DOI":"10.1186\/s12859-020-3429-6","article-title":"A big data approach to metagenomics for all-food-sequencing","volume":"21","author":"Kobus","year":"2020","journal-title":"BMC Bioinformatics"},{"key":"ref15","doi-asserted-by":"publisher","first-page":"639","DOI":"10.3238\/arztebl.2014.0639","article-title":"The role of multiplex PCR in respiratory tract infections in children","volume":"111","author":"Krause","year":"2014","journal-title":"Dtsch. Arztebl. Int."},{"key":"ref16","doi-asserted-by":"publisher","first-page":"1754","DOI":"10.1093\/bioinformatics\/btp324","article-title":"Fast and accurate short read alignment with burrows-wheeler transform","volume":"25","author":"Li","year":"2009","journal-title":"Bioinformatics"},{"key":"ref17","doi-asserted-by":"publisher","first-page":"205","DOI":"10.3389\/fcimb.2018.00205","article-title":"Detection of pulmonary infectious pathogens from lung biopsy tissues by metagenomic next-generation sequencing","volume":"8","author":"Li","year":"2018","journal-title":"Front. Cell. Infect. Microbiol."},{"key":"ref18","first-page":"1089","article-title":"Study on epidemic characteristics and etiology of community acquired pneumonia in Guangzhou from 2009 to 2012","volume":"47","author":"Liu","year":"2013","journal-title":"Zhonghua Yu Fang Yi Xue Za Zhi"},{"key":"ref19","doi-asserted-by":"publisher","first-page":"107144","DOI":"10.1016\/j.rmed.2023.107144","article-title":"Blood heparin-binding protein and neutrophil-to-lymphocyte ratio as indicators of the severity and prognosis of community-acquired pneumonia","volume":"208","author":"Meng","year":"2023","journal-title":"Respir. Med."},{"key":"ref20","doi-asserted-by":"publisher","first-page":"e45","DOI":"10.1164\/rccm.201908-1581ST","article-title":"Diagnosis and treatment of adults with community-acquired pneumonia. An official clinical practice guideline of the American Thoracic Society and Infectious Diseases Society of America","volume":"200","author":"Metlay","year":"2019","journal-title":"Am. J. Respir. Crit. Care Med."},{"key":"ref21","doi-asserted-by":"publisher","first-page":"831","DOI":"10.1101\/gr.238170.118","article-title":"Laboratory validation of a clinical metagenomic sequencing assay for pathogen detection in cerebrospinal fluid","volume":"29","author":"Miller","year":"2019","journal-title":"Genome Res."},{"key":"ref22","doi-asserted-by":"publisher","first-page":"799","DOI":"10.1513\/AnnalsATS.202008-1002OC","article-title":"Persistent post-COVID-19 interstitial lung disease. An observational study of corticosteroid treatment","volume":"18","author":"Myall","year":"2021","journal-title":"Ann. Am. Thorac. Soc."},{"key":"ref23","doi-asserted-by":"publisher","first-page":"56","DOI":"10.1016\/j.ctrv.2015.11.007","article-title":"The impact of multidisciplinary team meetings on patient assessment, management and outcomes in oncology settings: a systematic review of the literature","volume":"42","author":"Pillay","year":"2016","journal-title":"Cancer Treat. Rev."},{"key":"ref24","doi-asserted-by":"publisher","first-page":"1896","DOI":"10.1016\/j.chest.2020.05.598","article-title":"Treatment of community-acquired pneumonia in immunocompromised adults: a consensus statement regarding initial strategies","volume":"158","author":"Ramirez","year":"2020","journal-title":"Chest"},{"key":"ref25","doi-asserted-by":"publisher","first-page":"e0162323","DOI":"10.1128\/jcm.01623-23","article-title":"Overview of changes in the clinical and laboratory standards institute performance standards for antimicrobial susceptibility testing: M100 32nd and 33rd editions","volume":"63","author":"Schuetz","year":"2025","journal-title":"J. Clin. Microbiol."},{"key":"ref26","doi-asserted-by":"publisher","first-page":"155","DOI":"10.1007\/s10384-020-00794-5","article-title":"Practical use of multiplex and broad-range PCR in ophthalmology","volume":"65","author":"Sugita","year":"2021","journal-title":"Jpn. J. Ophthalmol."},{"key":"ref27","doi-asserted-by":"publisher","first-page":"e0042623","DOI":"10.1128\/jcm.00426-23","article-title":"Evaluation of the QIAstat-dx meningitis\/encephalitis panel, a multiplex PCR platform for the detection of community-acquired meningoencephalitis","volume":"61","author":"Sundelin","year":"2023","journal-title":"J. Clin. Microbiol."},{"key":"ref28","doi-asserted-by":"publisher","first-page":"834","DOI":"10.1002\/ajh.25829","article-title":"Hematological findings and complications of COVID-19","volume":"95","author":"Terpos","year":"2020","journal-title":"Am. J. Hematol."},{"key":"ref29","doi-asserted-by":"publisher","first-page":"834","DOI":"10.1016\/j.jinf.2022.03.021","article-title":"Multiplex PCR rapid testing for meningitis\/encephalitis","volume":"84","author":"Tseng","year":"2022","journal-title":"J. Infect."},{"key":"ref30","doi-asserted-by":"publisher","first-page":"R46","DOI":"10.1186\/gb-2014-15-3-r46","article-title":"Kraken: ultrafast metagenomic sequence classification using exact alignments","volume":"15","author":"Wood","year":"2014","journal-title":"Genome Biol."},{"key":"ref31","doi-asserted-by":"publisher","first-page":"j2471","DOI":"10.1136\/bmj.j2471","article-title":"Advances in the causes and management of community acquired pneumonia in adults","volume":"358","author":"Wunderink","year":"2017","journal-title":"BMJ"},{"key":"ref32","doi-asserted-by":"publisher","first-page":"3221","DOI":"10.1128\/jcm.43.7.3221-3226.2005","article-title":"Quantitative PCR assay using sputum samples for rapid diagnosis of pneumococcal pneumonia in adult emergency department patients","volume":"43","author":"Yang","year":"2005","journal-title":"J. Clin. Microbiol."},{"key":"ref33","doi-asserted-by":"publisher","first-page":"105307","DOI":"10.1016\/j.ebiom.2024.105307","article-title":"Enhancing lower respiratory tract infection diagnosis: implementation and clinical assessment of multiplex PCR-based and hybrid capture-based targeted next-generation sequencing","volume":"107","author":"Yin","year":"2024","journal-title":"EBioMedicine"},{"key":"ref34","doi-asserted-by":"publisher","first-page":"782","DOI":"10.1080\/23744235.2016.1193788","article-title":"Identification of pathogens by comprehensive real-time PCR versus conventional methods in community-acquired pneumonia in Japanese adults","volume":"48","author":"Yoshii","year":"2016","journal-title":"Infect Dis (Lond)"},{"key":"ref35","doi-asserted-by":"publisher","first-page":"585","DOI":"10.1186\/s12890-024-03380-4","article-title":"Significance of respiratory virus coinfection in children with Mycoplasma pneumoniae pneumonia","volume":"24","author":"Yu","year":"2024","journal-title":"BMC Pulm. Med."},{"key":"ref36","doi-asserted-by":"publisher","first-page":"e35287","DOI":"10.1016\/j.heliyon.2024.e35287","article-title":"Clinical impact of metagenomic next-generation sequencing of bronchoalveolar lavage fluids for the diagnosis of pulmonary infections in respiratory intensive care unit","volume":"10","author":"Zhang","year":"2024","journal-title":"Heliyon"},{"key":"ref37","first-page":"270","article-title":"PCR versus serology for diagnosing Mycoplasma pneumoniae infection: a systematic review & meta-analysis","volume":"134","author":"Zhang","year":"2011","journal-title":"Indian J. Med. Res."}],"container-title":["Frontiers in Microbiology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fmicb.2026.1979614\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,10,7]],"date-time":"2026-10-07T05:51:34Z","timestamp":1791352294000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fmicb.2026.1979614\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,10,7]]},"references-count":37,"alternative-id":["10.3389\/fmicb.2026.1979614"],"URL":"https:\/\/doi.org\/10.3389\/fmicb.2026.1979614","relation":{},"ISSN":["1664-302X"],"issn-type":[{"value":"1664-302X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,10,7]]},"article-number":"1979614"}}