Molecular epidemiologic factors contributing to quinolone resistance in clinical multidrug-resistant Klebsiella pneumoniae isolates from Shanghai, China

Molecular epidemiologic factors contributing to quinolone resistance in clinical multidrug-resistant Klebsiella pneumoniae isolates from Shanghai, China | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research article Molecular epidemiologic factors contributing to quinolone resistance in clinical multidrug-resistant Klebsiella pneumoniae isolates from Shanghai, China Yan Wang, Guoping Cai, Jinan Zhang, Xiaogang Xu, Hongzhou Lu This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-492410/v2 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 07 Nov, 2022 Read the published version in Journal of Medical Microbiology → Version 2 posted You are reading this latest preprint version Show more versions Abstract Background The soaring quinolone-resistance rate of Klebsiella pneumoniae, a common pathogen in immunocompromised individuals, has seriously undermined the wide applications of antimicrobials of this class. This study aimed to investigate the emerging key contributors to quinolone-resistance in multidrug resistant K. pneumoniae (MDR-KP) isolates from a clinical setting with continuing point-source infection outbreaks in Shanghai, China. Results Between January and March 2017, a total of 34 K. pneumoniae isolates, including 30 carbapenem-resistant K. pneumoniae (CRKP), were selected and characterized from a teaching hospital participating in an ongoing Bacterial Resistance Surveillance Project in Shanghai, China. Two predominant high-risk CRKP clones, ST11-wzi64 and ST15-wzi19/wzi24, caused three point-source nosocomial outbreaks in intensive care unit and/or neurosurgery department potentially by respiratory-route, promoting the co-selection and evolution of multidrug-resistant determinants. Multiple quinolone resistance-determining region (QRDR) mutations occurred in isolates of ST15 (S83F, D87A; S80I), ST11 (S83I, D87G; S80I), and ST218 (D87A; S80I). Plasmid-mediated quinolone resistance determinants, qnrS1, aac(6’)-Ib-cr, oqxAB, were detected in 32 (94.1%) isolates alone or in combination, spreading accompanied with β-lactamases (mainly, KPC-2-type carbapenemase and CTX-M-type extended-spectrum β-lactamase), 16S rRNA methylases (ArmA and RmtB), and putrescine ABC transporter permease (PotI) variants, independently of QRDR-mutations. AcrR, AcrAB transcriptional repressor, was insertion-inactivated by IS5-transposase in isolates of ST11. Thirteen ompK36 variants associated with specific ST (n=7) and wzi-allele (n=9) clustered into 10 (sub)lineages in the phylogenetic tree possibly affecting the MDR phenotype and the infection outcome of isolates. Isolates of ST11, ST15, and ST218 had frameshift disruptions in OmpK35 coupled with specific GD-insertion at position 134-135 in OmpK36, all showing distinct microevolution clusters of ompK36 genotypes. Seven quinolone-susceptible isolates kept the porin genes integral, including two each CRKPs of ST13-wzi74 (carbapenemase KPC-2 and NDM-1-coproducers) and ST65-wzi72. Conclusions Under selective pressures, accumulation of mutations of three types (QRDR, AcrR, OmpK36/OmpK35) and acquisition of resistance-conferring genes has been continuously contributing to quinolone-resistance in clinical MDR-KP isolates, reinforcing the importance of ongoing epidemiologic surveillance on the evolution and transmission of these isolates. Our findings provided detailed mechanistic analyses and epidemiologic implications for further infection control and antibiotic stewardship initiatives. General Microbiology Applied & Industrial Microbiology Klebsiella pneumoniae Quinolone resistance Carbapenem resistance Multidrug resistance Molecular epidemiology Phylogenetic analysis Shanghai Figures Figure 1 Figure 2 Figure 3 Full Text Supplementary Files YanWangSupplementaryMaterial.pdf Cite Share Download PDF Status: Published Journal Publication published 07 Nov, 2022 Read the published version in Journal of Medical Microbiology → Version 2 posted You are reading this latest preprint version Show more versions Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-492410","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research article","associatedPublications":[],"authors":[{"id":34619797,"identity":"7ebbcf4c-8e61-4379-9051-e095b582dc8f","order_by":0,"name":"Yan Wang","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA0UlEQVRIiWNgGAWjYDADA+bDBw58qCBJC1ta4sEZZ0jTkmN8mLeFGJXHzx5+zVNzh8GcjefDAd4GBnl+sQMEtJzJS7PmOfaMwbKNd8MByR0MhjNnJ+DXYnYgx8yYh+0wg8H93g0HDM8wJBjcJqTl/Bugln9ALcd4HhxIbCNGy40c48e8bWAtDAcOEqPF/sYbM8a5fSAtbAYHG85IEPaLZH+O8Yc330BamB9//lNhI88vTUALELBJ8TAw1DdAOBIElYMA88cfRKkbBaNgFIyCEQsAb8hLOcttJuMAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0002-8457-5287","institution":"Fudan University","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Yan","middleName":"","lastName":"Wang","suffix":""},{"id":34619798,"identity":"4401d625-e032-4ef4-8b54-fdcfc110c127","order_by":1,"name":"Guoping Cai","email":"","orcid":"","institution":"Fudan University Jinshan Hospital","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Guoping","middleName":"","lastName":"Cai","suffix":""},{"id":34619799,"identity":"5fca6914-9bbc-43b0-9098-ee504560aa0c","order_by":2,"name":"Jinan Zhang","email":"","orcid":"","institution":"Shanghai Jiankang Medical University","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Jinan","middleName":"","lastName":"Zhang","suffix":""},{"id":34619800,"identity":"ea5b1c0d-bde5-4907-9863-352a169da509","order_by":3,"name":"Xiaogang Xu","email":"","orcid":"","institution":"Fudan University Huashan Hospital Institute of Antibiotics","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Xiaogang","middleName":"","lastName":"Xu","suffix":""},{"id":34619801,"identity":"2d730bb2-2d33-46be-9165-0bf7fe088b06","order_by":4,"name":"Hongzhou Lu","email":"","orcid":"","institution":"Fudan University Affiliated Public Health Clinical Center: Shanghai Public Health Clinical Center","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Hongzhou","middleName":"","lastName":"Lu","suffix":""}],"badges":[],"createdAt":"2021-05-03 17:52:38","currentVersionCode":2,"declarations":"","doi":"10.21203/rs.3.rs-492410/v2","doiUrl":"https://doi.org/10.21203/rs.3.rs-492410/v2","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1099/jmm.0.001583","type":"published","date":"2022-11-08T00:00:00+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":10600989,"identity":"c38b2dd1-2483-4ceb-bb22-83487c2e8a6a","added_by":"auto","created_at":"2021-06-14 18:06:28","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":82851,"visible":true,"origin":"","legend":"Phylogenetic analysis based on the coding sequences of ompK36 genes for K. pneumoniae isolates. The dendrogram was constructed using neighbor-joining method by MEGA program version 7.0. Bootstrap resampling (1000 replications) was used, and bootstrap values ≥70% were shown. Scale bar represents nt substitutions per site. Reference strains were named in reference to the accession-number, strain name, country of origin, isolation-year, and MLST. ★denotes K. pneumoniae isolates in the present study (GenBank accession number MK341466-MK341478) with its common name and ST shown in bold. The ompK36 for a Raoultella planticola isolate (CP026047) was taken as an out-group. The red box indicates the phylogenetic cluster position of isolates ST11 (Lineage IV).","description":"","filename":"Fig1.png","url":"https://assets-eu.researchsquare.com/files/rs-611050/v1/8f1a904ce47d9b9442374605.png"},{"id":10600991,"identity":"3ca0d67a-4e13-4994-8f73-b0536ba4eb47","added_by":"auto","created_at":"2021-06-14 18:06:28","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":931064,"visible":true,"origin":"","legend":"Comparison and alignment of Ompk36 porin regions from the representative of K. pneumoniae sample and reference isolates in Figure 1. The differentiated regions included Loops (L), beta-strands (B), turns (T) and alpha-helix (H). The consensus protein sequence was established employing the most common aa at each position in the present study. Amino acids are represented by standard single-letter codes and deletion in a site is indicated by a dash (-); those sites identical to the consensus sequence are indicated with dots. The numbers along the sequence indicated aa positions. Each sequence was designated as in the phylogenetic tree. The red box indicates the substitutions and insertions in loops of L3,L4,L5,L6,L7, and L8 for ST11 strains.","description":"","filename":"Fig2.png","url":"https://assets-eu.researchsquare.com/files/rs-611050/v1/b68b1500427ebf1b2622a7e3.png"},{"id":10600993,"identity":"7e0285a2-c917-45f4-aff5-afe749d99b1b","added_by":"auto","created_at":"2021-06-14 18:06:28","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":44895,"visible":true,"origin":"","legend":"MLST and wzi-genotyping of 30 K. pneumonia dinical isolates. The dendrogram of wzi-alleles (0474) was built using neighbor-joining method by MEGA program version 7.0. Bootstrap resampling (1000 replications/ was used, and bootstrap values 770% were shown. Scale bar represents nt substitutions per site. Each known wzi-allele number is followed by the corresponding capsular (K) type, with dots separating several K types indicating cross-reactions. The three main branches (A, B, and C) are labeled. The representative wzi sequences (GenBank accession number MK301453-MK301061) of sample isolates from the present study and their identical wzi-alleles in the same cluster were shown in bold. 0 denotes the new wzi variants without known allele-number. The common names of isolates were followed by ward of origin, isolation-date, and MLST. ND, Neurosurgery Department; PW, Pediatric Ward.","description":"","filename":"Fig3.png","url":"https://assets-eu.researchsquare.com/files/rs-611050/v1/371ec5891a1f9b23be7b7335.png"},{"id":29151180,"identity":"927d2181-5390-4174-b6e6-3fc6d078b081","added_by":"auto","created_at":"2022-11-16 18:06:52","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":952966,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-492410/v2/6e019079-a4a2-4282-8b15-aafc673e43df.pdf"},{"id":10600995,"identity":"e47728c8-47ae-4bc7-8517-f81d6a818869","added_by":"auto","created_at":"2021-06-14 18:06:28","extension":"pdf","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":716594,"visible":true,"origin":"","legend":"","description":"","filename":"YanWangSupplementaryMaterial.pdf","url":"https://assets-eu.researchsquare.com/files/rs-611050/v1/f1ad730ba7f64ac0a77676b6.pdf"}],"financialInterests":"","formattedTitle":"\u003cp\u003eMolecular epidemiologic factors contributing to quinolone resistance in clinical multidrug-resistant \u003cem\u003eKlebsiella pneumoniae\u003c/em\u003e isolates from Shanghai, China\u003c/p\u003e","fulltext":[{"header":"Full Text","content":"This preprint is available for \u003ca href='/article/rs-611050/latest.pdf' target='_blank'\u003edownload as a PDF\u003c/a\u003e."}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Klebsiella pneumoniae, Quinolone resistance, Carbapenem resistance, Multidrug resistance, Molecular epidemiology, Phylogenetic analysis, Shanghai ","lastPublishedDoi":"10.21203/rs.3.rs-492410/v2","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-492410/v2","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eBackground\u003c/p\u003e\u003cp\u003eThe soaring quinolone-resistance rate of Klebsiella pneumoniae, a common pathogen in immunocompromised individuals, has seriously undermined the wide applications of antimicrobials of this class. This study aimed to investigate the emerging key contributors to quinolone-resistance in multidrug resistant K. pneumoniae (MDR-KP) isolates from a clinical setting with continuing point-source infection outbreaks in Shanghai, China. \u003c/p\u003e\u003cp\u003eResults\u003c/p\u003e\u003cp\u003eBetween January and March 2017, a total of 34 K. pneumoniae isolates, including 30 carbapenem-resistant K. pneumoniae (CRKP), were selected and characterized from a teaching hospital participating in an ongoing Bacterial Resistance Surveillance Project in Shanghai, China. Two predominant high-risk CRKP clones, ST11-wzi64 and ST15-wzi19/wzi24, caused three point-source nosocomial outbreaks in intensive care unit and/or neurosurgery department potentially by respiratory-route, promoting the co-selection and evolution of multidrug-resistant determinants. Multiple quinolone resistance-determining region (QRDR) mutations occurred in isolates of ST15 (S83F, D87A; S80I), ST11 (S83I, D87G; S80I), and ST218 (D87A; S80I). Plasmid-mediated quinolone resistance determinants, qnrS1, aac(6’)-Ib-cr, oqxAB, were detected in 32 (94.1%) isolates alone or in combination, spreading accompanied with β-lactamases (mainly, KPC-2-type carbapenemase and CTX-M-type extended-spectrum β-lactamase), 16S rRNA methylases (ArmA and RmtB), and putrescine ABC transporter permease (PotI) variants, independently of QRDR-mutations. AcrR, AcrAB transcriptional repressor, was insertion-inactivated by IS5-transposase in isolates of ST11. Thirteen ompK36 variants associated with specific ST (n=7) and wzi-allele (n=9) clustered into 10 (sub)lineages in the phylogenetic tree possibly affecting the MDR phenotype and the infection outcome of isolates. Isolates of ST11, ST15, and ST218 had frameshift disruptions in OmpK35 coupled with specific GD-insertion at position 134-135 in OmpK36, all showing distinct microevolution clusters of ompK36 genotypes. Seven quinolone-susceptible isolates kept the porin genes integral, including two each CRKPs of ST13-wzi74 (carbapenemase KPC-2 and NDM-1-coproducers) and ST65-wzi72. \u003c/p\u003e\u003cp\u003eConclusions\u003c/p\u003e\u003cp\u003eUnder selective pressures, accumulation of mutations of three types (QRDR, AcrR, OmpK36/OmpK35) and acquisition of resistance-conferring genes has been continuously contributing to quinolone-resistance in clinical MDR-KP isolates, reinforcing the importance of ongoing epidemiologic surveillance on the evolution and transmission of these isolates. Our findings provided detailed mechanistic analyses and epidemiologic implications for further infection control and antibiotic stewardship initiatives.\u003c/p\u003e","manuscriptTitle":"Molecular epidemiologic factors contributing to quinolone resistance in clinical multidrug-resistant Klebsiella pneumoniae isolates from Shanghai, China","msid":"","msnumber":"","nonDraftVersions":[{"code":2,"date":"2021-06-14 18:03:26","doi":"10.21203/rs.3.rs-492410/v2","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}},{"code":1,"date":"2021-05-06 13:53:40","doi":"10.21203/rs.3.rs-492410/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"92f7798c-0604-4402-86b5-eb8277d2ef3d","owner":[],"postedDate":"June 14th, 2021","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[{"id":5170754,"name":"General Microbiology"},{"id":5170755,"name":"Applied \u0026 Industrial Microbiology"}],"tags":[],"updatedAt":"2022-11-16T18:06:47+00:00","versionOfRecord":{"articleIdentity":"rs-492410","link":"https://doi.org/10.1099/jmm.0.001583","journal":{"identity":"journal-of-medical-microbiology","isVorOnly":true,"title":"Journal of Medical Microbiology"},"publishedOn":"2022-11-08 00:00:00","publishedOnDateReadable":"November 8th, 2022"},"versionCreatedAt":"2021-06-14 18:03:26","video":"","vorDoi":"10.1099/jmm.0.001583","vorDoiUrl":"https://doi.org/10.1099/jmm.0.001583","workflowStages":[]},"version":"v2","identity":"rs-492410","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-492410","identity":"rs-492410","version":["v2"]},"buildId":"7rjqhiLT3MXkJMwkYKINL","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}