{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,25]],"date-time":"2026-08-25T23:49:38Z","timestamp":1787701778854,"version":"build-2784847793"},"reference-count":26,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2023,10,20]],"date-time":"2023-10-20T00:00:00Z","timestamp":1697760000000},"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><jats:title>Background<\/jats:title><jats:p>Traditional drug susceptibility testing cannot be performed in clinical laboratories due to the slow-growing characteristics of <jats:italic>Mycoplasma genitalium<\/jats:italic> when cultured <jats:italic>in vitro<\/jats:italic>. Sanger sequencing is the standard method for detecting drug resistance-associated mutations. It has been used in some laboratories to guide the choice of macrolide antibiotics for <jats:italic>Mycoplasma genitalium<\/jats:italic> infected patients. Furthermore, resistance to fluoroquinolone has become another emerging clinical challenge.<\/jats:p><\/jats:sec><jats:sec><jats:title>Objective<\/jats:title><jats:p>Sequencing analysis can detect unknown mutations, but it is time-consuming, requires professional analytical skills and the appropriate testing equipment. The main objective of this study was to establish a nested real-time PCR method for the simultaneous detection of <jats:italic>23S rRNA<\/jats:italic> and <jats:italic>parC<\/jats:italic> genotypes in relation to the macrolide and fluoroquinolone resistance.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>105 MG-positive samples and 27 samples containing other pathogens were used for validation. The limit of the nested real-time PCR detection was 500 copies\/reaction and there was no cross-reaction with <jats:italic>Ureaplasma urealyticum<\/jats:italic>, <jats:italic>Mycoplasma hominis<\/jats:italic>, <jats:italic>Chlamydia trachomatis<\/jats:italic>, <jats:italic>Neisseria gonorrhoeae<\/jats:italic>, <jats:italic>Human papillomavirus<\/jats:italic>, <jats:italic>Herpes simplex virus<\/jats:italic>, <jats:italic>Candida albicans<\/jats:italic> and <jats:italic>Ureaplasma parvum<\/jats:italic>, but the <jats:italic>23S rRNA<\/jats:italic> assay cross-reacted with <jats:italic>Mycoplasma pneumoniae<\/jats:italic>. Compared with sequencing results, the sensitivity of <jats:italic>23S rRNA<\/jats:italic> was 100% (95% CI; 93.3 -100), the specificity was 94.3% (95% CI; 79.4 - 99.0), the overall consistency was 98% (95% CI; 92.5 - 99.7) and <jats:italic>kappa<\/jats:italic> value was 0.96 (<jats:italic>P<\/jats:italic> &amp;lt; 0.001); the sensitivity of <jats:italic>parC<\/jats:italic> was 100% (95% CI; 93.4 - 100), the specificity was 89.7% (95% CI; 71.5 - 97.3) and the overall consistency was 96.9% (95% CI; 90.7 - 99.2) with a <jats:italic>kappa<\/jats:italic> value of 0.92 (<jats:italic>P<\/jats:italic> &amp;lt; 0.001).<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusions<\/jats:title><jats:p>The results of this sensitive and rapid alternative for identifying resistant genotypes of <jats:italic>Mycoplasma genitalium<\/jats:italic> are intuitive and easy to interpret, especially for mixed MG populations. Although the relevant <jats:italic>23S rRNA<\/jats:italic> primers need further adjustment, this reliable method would provide an effective diagnostic tool for the selection of antibiotics in clinical practice.<\/jats:p><\/jats:sec>","DOI":"10.3389\/fcimb.2023.1271392","type":"journal-article","created":{"date-parts":[[2023,10,20]],"date-time":"2023-10-20T13:12:16Z","timestamp":1697807536000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":7,"title":["Detection of macrolide and fluoroquinolone resistance-associated 23S rRNA and parC mutations in Mycoplasma genitalium by nested real-time PCR"],"prefix":"10.3389","volume":"13","author":[{"given":"Wenyin","family":"He","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ying","family":"Yuan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Junyu","family":"Liang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xuejiao","family":"Fan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lei","family":"Li","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xingfei","family":"Pan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2023,10,20]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","first-page":"1228","DOI":"10.1093\/cid\/ciu1162","article-title":"Macrolide resistance and azithromycin failure in a Mycoplasma genitalium-infected cohort and response of azithromycin failures to alternative antibiotic regimens","volume":"60","author":"Bissessor","year":"2015","journal-title":"Clin. 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