{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,3,24]],"date-time":"2026-03-24T11:56:35Z","timestamp":1774353395837,"version":"3.50.1"},"posted":{"date-parts":[[2026,3,19]]},"group-title":"Microbiology","reference-count":61,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,3,19]],"date-time":"2026-03-19T00:00:00Z","timestamp":1773878400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,3,19]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  DNA supercoiling is essential for the developmental cycle of\n                  <jats:italic>Chlamydia trachomatis<\/jats:italic>\n                  , yet its role in shaping antibiotic responses remains poorly understood. We investigated how the fluoroquinolone moxifloxacin (Mox) influenced\n                  <jats:italic>C. trachomatis<\/jats:italic>\n                  growth across developmental stages with its distinct supercoiling levels. Early Mox exposure completely halted bacterial growth, whereas treatment during mid-cycle produced enlarged, persistent forms and abolished formation of infectious progeny. These stage-specific outcomes coincided with inhibition of DNA replication, depletion of DNA gyrase, and transcriptional repression of\n                  <jats:italic>ompA<\/jats:italic>\n                  and\n                  <jats:italic>omcB<\/jats:italic>\n                  , accompanied by preserved or elevated expression of the stress-responsive\n                  <jats:italic>groESL1<\/jats:italic>\n                  operon. Mox also elicited compensatory downregulation of topoisomerase I (TopA), consistent with attempts to rebalance intracellular supercoiling. Together, these data demonstrate that fluoroquinolone susceptibility in\n                  <jats:italic>C. trachomatis<\/jats:italic>\n                  reflects stage-dependent supercoiling levels. Perturbation of supercoiling homeostasis drives developmental arrest and persistence phenotypes, highlighting coordinated gyrase-TopA activity as a key determinant of fluoroquinolone tolerance and a potential target for overcoming persistent infection.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>Importance<\/jats:title>\n                  <jats:p>\n                    <jats:italic>C. trachomatis<\/jats:italic>\n                    , a medically significant bacterial pathogen, can persist under antimicrobial pressure, complicating treatment strategy. This study links supercoiling homeostasis to fluoroquinolone tolerance, offering mechanistic insights into chlamydial adaptation to antibiotic stress and identifying potential targets to overcome persistence\u2014an urgent challenge in global reproductive health.\n                  <\/jats:p>\n                <\/jats:sec>","DOI":"10.64898\/2026.03.19.712910","type":"posted-content","created":{"date-parts":[[2026,3,20]],"date-time":"2026-03-20T04:00:24Z","timestamp":1773979224000},"source":"Crossref","is-referenced-by-count":0,"title":["Coordinated topoisomerase function shapes the fluoroquinolone response of\n                  <i>Chlamydia trachomatis<\/i>"],"prefix":"10.64898","author":[{"given":"Andrew","family":"Terrebonne","sequence":"first","affiliation":[{"name":"Department of Microbiology, Immunology, and Parasitology, Louisiana State University Health Sciences Center, New Orleans, LA 70112"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Caitlynn","family":"Diggs","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Immunology, and Parasitology, Louisiana State University Health Sciences Center, New Orleans, LA 70112"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3721-6839","authenticated-orcid":false,"given":"Scot P.","family":"Ouellette","sequence":"additional","affiliation":[{"name":"Department of Pathology, Microbiology, and Immunology, University of Nebraska Medical Center, Omaha, Nebraska, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Leiqiong","family":"Gao","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Immunology, and Parasitology, Louisiana State University Health Sciences Center, New Orleans, LA 70112"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yuk-Ching","family":"Tse-Dinh","sequence":"additional","affiliation":[{"name":"Biomolecular Sciences Institute, Department of Chemistry and Biochemistry, Florida International University, Miami, FL 33199, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9869-7317","authenticated-orcid":false,"given":"Li","family":"Shen","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Immunology, and Parasitology, Louisiana State University Health Sciences Center, New Orleans, LA 70112"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"54368","reference":[{"key":"2026032403150818000_2026.03.19.712910v1.1","unstructured":"WHO. 2021. Global progress report on HIV, viral hepatitis, and sexually transmitted infections 2021: accountability for the global health sector strategies,2016-2021.. Vol. 53."},{"key":"2026032403150818000_2026.03.19.712910v1.2","doi-asserted-by":"publisher","DOI":"10.1093\/infdis\/jiab610"},{"key":"2026032403150818000_2026.03.19.712910v1.3","first-page":"e21331","article-title":"The Role of Chlamydia Trachomatis in the Pathogenesis of Cervical Cancer","volume":"14","year":"2022","journal-title":"Cureus"},{"key":"2026032403150818000_2026.03.19.712910v1.4","doi-asserted-by":"publisher","DOI":"10.1002\/(SICI)1097-0215(20000101)85:1<35::AID-IJC6>3.0.CO;2-A"},{"key":"2026032403150818000_2026.03.19.712910v1.5","doi-asserted-by":"publisher","DOI":"10.1128\/iai.00339-23"},{"key":"2026032403150818000_2026.03.19.712910v1.6","doi-asserted-by":"crossref","first-page":"10","DOI":"10.1007\/s40588-016-0028-4","article-title":"Chlamydial Antibiotic Resistance and Treatment Failure in Veterinary and Human Medicine","volume":"3","year":"2016","journal-title":"Current Clinical Microbiology Reports"},{"key":"2026032403150818000_2026.03.19.712910v1.7","doi-asserted-by":"crossref","first-page":"1427","DOI":"10.2217\/fmb.10.96","article-title":"Antibiotic resistance in Chlamydiae","volume":"5","year":"2010","journal-title":"Future Microbiology"},{"key":"2026032403150818000_2026.03.19.712910v1.8","doi-asserted-by":"publisher","DOI":"10.1016\/j.femsre.2005.03.002"},{"key":"2026032403150818000_2026.03.19.712910v1.9","doi-asserted-by":"publisher","DOI":"10.1038\/nrmicro.2016.30"},{"key":"2026032403150818000_2026.03.19.712910v1.10","doi-asserted-by":"crossref","first-page":"1185571","DOI":"10.3389\/fcimb.2023.1185571","article-title":"Persistence of obligate intracellular pathogens: alternative strategies to overcome host-specific stresses","volume":"13","year":"2023","journal-title":"Front Cell Infect Microbiol"},{"key":"2026032403150818000_2026.03.19.712910v1.11","first-page":"000204","article-title":"Chlamydial clinical isolates show subtle differences in persistence phenotypes and growth in vitro","volume":"3","year":"2021","journal-title":"Access Microbiol"},{"key":"2026032403150818000_2026.03.19.712910v1.12","doi-asserted-by":"crossref","first-page":"3101","DOI":"10.3389\/fmicb.2018.03101","article-title":"Chlamydia Persistence: A Survival Strategy to Evade Antimicrobial Effects in-vitro and in-vivo","volume":"9","year":"2018","journal-title":"Front Microbiol"},{"key":"2026032403150818000_2026.03.19.712910v1.13","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.62.9.3705-3711.1994"},{"key":"2026032403150818000_2026.03.19.712910v1.14","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.42295"},{"key":"2026032403150818000_2026.03.19.712910v1.15","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2958.2006.05465.x"},{"key":"2026032403150818000_2026.03.19.712910v1.16","first-page":"e01464","article-title":"Clinical Persistence of <span class=\u201cnamed-content genus-species\u201d id=\u201cnamed-content-1\u201d>Chlamydia trachomatis<\/span> Sexually Transmitted Strains Involves Novel Mutations in the Functional \u03b1\u03b2\u03b2\u03b1 Tetramer of the Tryptophan Synthase Operon","volume":"10","year":"2019","journal-title":"mBio"},{"key":"2026032403150818000_2026.03.19.712910v1.17","doi-asserted-by":"publisher","DOI":"10.1093\/infdis\/jix155"},{"key":"2026032403150818000_2026.03.19.712910v1.18","doi-asserted-by":"publisher","DOI":"10.1093\/infdis\/jiag023"},{"key":"2026032403150818000_2026.03.19.712910v1.19","doi-asserted-by":"publisher","DOI":"10.1097\/OLQ.0000000000000489"},{"key":"2026032403150818000_2026.03.19.712910v1.20","first-page":"157","article-title":"Outbreak and persistence of Chlamydia pneumoniae infection in an Italian family","volume":"11","year":"2003","journal-title":"Infez Med"},{"key":"2026032403150818000_2026.03.19.712910v1.21","doi-asserted-by":"crossref","first-page":"e0031022","DOI":"10.1128\/jb.00310-22","article-title":"Diversity of \u03c3(66)-Specific Promoters Contributes to Regulation of Developmental Gene Expression in Chlamydia trachomatis","volume":"205","year":"2023","journal-title":"J Bacteriol"},{"key":"2026032403150818000_2026.03.19.712910v1.22","doi-asserted-by":"publisher","DOI":"10.1128\/msphere.00391-23"},{"key":"2026032403150818000_2026.03.19.712910v1.23","doi-asserted-by":"crossref","unstructured":"Soules KR , LaBrie SD , May BH , Hefty PS . 2020. Sigma 54-Regulated Transcription Is Associated with Membrane Reorganization and Type III Secretion Effectors during Conversion to Infectious Forms of Chlamydia trachomatis. mBio 11.","DOI":"10.1128\/mBio.01725-20"},{"key":"2026032403150818000_2026.03.19.712910v1.24","doi-asserted-by":"publisher","DOI":"10.1128\/JB.01083-08"},{"key":"2026032403150818000_2026.03.19.712910v1.25","first-page":"812448","article-title":"Robust Heat Shock Response in Chlamydia Lacking a Typical Heat Shock Sigma Factor","volume":"12","year":"2021","journal-title":"Front Microbiol"},{"key":"2026032403150818000_2026.03.19.712910v1.26","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1005125"},{"key":"2026032403150818000_2026.03.19.712910v1.27","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1207300109"},{"key":"2026032403150818000_2026.03.19.712910v1.28","doi-asserted-by":"publisher","DOI":"10.1128\/jb.00132-20"},{"key":"2026032403150818000_2026.03.19.712910v1.29","doi-asserted-by":"publisher","DOI":"10.1101\/gad.1784009"},{"key":"2026032403150818000_2026.03.19.712910v1.30","doi-asserted-by":"publisher","DOI":"10.1111\/mmi.13241"},{"key":"2026032403150818000_2026.03.19.712910v1.31","doi-asserted-by":"publisher","DOI":"10.1128\/JB.00431-08"},{"key":"2026032403150818000_2026.03.19.712910v1.32","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2958.1990.tb02064.x"},{"key":"2026032403150818000_2026.03.19.712910v1.33","doi-asserted-by":"publisher","DOI":"10.1016\/j.csbj.2019.07.013"},{"key":"2026032403150818000_2026.03.19.712910v1.34","doi-asserted-by":"crossref","first-page":"26","DOI":"10.1186\/s12860-019-0211-6","article-title":"DNA supercoiling and transcription in bacteria: a two-way street","volume":"20","year":"2019","journal-title":"BMC Molecular and Cell Biology"},{"key":"2026032403150818000_2026.03.19.712910v1.35","doi-asserted-by":"publisher","DOI":"10.3390\/cells13060553"},{"key":"2026032403150818000_2026.03.19.712910v1.36","first-page":"2022","article-title":"The balance between gyrase and topoisomerase I activities determines levels of supercoiling, nucleoid compaction, and viability in bacteria","volume":"13","year":"2023","journal-title":"Frontiers in Microbiology"},{"key":"2026032403150818000_2026.03.19.712910v1.37","doi-asserted-by":"crossref","first-page":"1097","DOI":"10.1021\/acsinfecdis.4c00128","article-title":"Gyrase and Topoisomerase IV: Recycling Old Targets for New Antibacterials to Combat Fluoroquinolone Resistance","volume":"10","year":"2024","journal-title":"ACS Infectious Diseases"},{"key":"2026032403150818000_2026.03.19.712910v1.38","doi-asserted-by":"crossref","unstructured":"Millanao AR , Mora AY , Villagra NA , Bucarey SA , Hidalgo AA . 2021. Biological Effects of Quinolones: A Family of Broad-Spectrum Antimicrobial Agents. Molecules 26.","DOI":"10.3390\/molecules26237153"},{"key":"2026032403150818000_2026.03.19.712910v1.39","doi-asserted-by":"crossref","first-page":"1354","DOI":"10.2174\/1389450121666200621193355","article-title":"Fluoroquinolones: Blessings Or Curses","volume":"21","year":"2020","journal-title":"Curr Drug Targets"},{"key":"2026032403150818000_2026.03.19.712910v1.40","doi-asserted-by":"crossref","unstructured":"Hooper DC , Jacoby GA . 2016. Topoisomerase Inhibitors: Fluoroquinolone Mechanisms of Action and Resistance. Cold Spring Harb Perspect Med 6.","DOI":"10.1101\/cshperspect.a025320"},{"key":"2026032403150818000_2026.03.19.712910v1.41","doi-asserted-by":"crossref","unstructured":"Bush NG , Diez-Santos I , Abbott LR , Maxwell A. 2020. Quinolones: Mechanism, Lethality and Their Contributions to Antibiotic Resistance. Molecules 25.","DOI":"10.3390\/molecules25235662"},{"key":"2026032403150818000_2026.03.19.712910v1.42","doi-asserted-by":"publisher","DOI":"10.1007\/s10156-004-0332-4"},{"key":"2026032403150818000_2026.03.19.712910v1.43","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.42.10.2474"},{"key":"2026032403150818000_2026.03.19.712910v1.44","doi-asserted-by":"publisher","DOI":"10.1093\/jac\/dkm447"},{"key":"2026032403150818000_2026.03.19.712910v1.45","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.44.12.3288-3297.2000"},{"key":"2026032403150818000_2026.03.19.712910v1.46","doi-asserted-by":"publisher","DOI":"10.1128\/jb.00190-25"},{"key":"2026032403150818000_2026.03.19.712910v1.47","doi-asserted-by":"publisher","DOI":"10.1128\/mbio.02584-23"},{"key":"2026032403150818000_2026.03.19.712910v1.48","doi-asserted-by":"crossref","first-page":"27244","DOI":"10.1038\/srep27244","article-title":"Quantifying promoter activity during the developmental cycle of Chlamydia trachomatis","volume":"6","year":"2016","journal-title":"Scientific Reports"},{"key":"2026032403150818000_2026.03.19.712910v1.49","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkac679"},{"key":"2026032403150818000_2026.03.19.712910v1.50","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1525047113"},{"key":"2026032403150818000_2026.03.19.712910v1.51","doi-asserted-by":"publisher","DOI":"10.1038\/msb4100135"},{"key":"2026032403150818000_2026.03.19.712910v1.52","doi-asserted-by":"publisher","DOI":"10.1128\/MR.58.4.686-699.1994"},{"key":"2026032403150818000_2026.03.19.712910v1.53","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.90.9.3998"},{"key":"2026032403150818000_2026.03.19.712910v1.54","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.2535394100"},{"key":"2026032403150818000_2026.03.19.712910v1.55","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0217753"},{"key":"2026032403150818000_2026.03.19.712910v1.56","doi-asserted-by":"publisher","DOI":"10.1007\/s12551-016-0238-2"},{"key":"2026032403150818000_2026.03.19.712910v1.57","doi-asserted-by":"publisher","DOI":"10.1038\/sj.embor.7400729"},{"key":"2026032403150818000_2026.03.19.712910v1.58","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.84.20.7024"},{"key":"2026032403150818000_2026.03.19.712910v1.59","first-page":"105","article-title":"Regulation of the genes for E","volume":"34","year":"1983","journal-title":"coli DNA gyrase: homeostatic control of DNA supercoiling. Cell"},{"key":"2026032403150818000_2026.03.19.712910v1.60","doi-asserted-by":"crossref","unstructured":"Greer M , Elnaggar JH , Taylor CM , Shen L. 2022. Mycoplasma decontamination in Chlamydia trachomatis culture: a curative approach. Pathog Dis 79.","DOI":"10.1093\/femspd\/ftab056"},{"key":"2026032403150818000_2026.03.19.712910v1.61","doi-asserted-by":"publisher","DOI":"10.1016\/j.mimet.2012.08.012"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.64898\/2026.03.19.712910","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,3,24]],"date-time":"2026-03-24T10:15:19Z","timestamp":1774347319000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.64898\/2026.03.19.712910"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,3,19]]},"references-count":61,"URL":"https:\/\/doi.org\/10.64898\/2026.03.19.712910","relation":{},"subject":[],"published":{"date-parts":[[2026,3,19]]},"subtype":"preprint"}}