{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,22]],"date-time":"2026-09-22T13:24:36Z","timestamp":1790083476451,"version":"4.0.1"},"reference-count":26,"publisher":"Oxford University Press (OUP)","issue":"1","license":[{"start":{"date-parts":[[2025,12,24]],"date-time":"2025-12-24T00:00:00Z","timestamp":1766534400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/academic.oup.com\/pages\/standard-publication-reuse-rights"}],"funder":[{"name":"National Key R&D Program","award":["2022YFD1800400"],"award-info":[{"award-number":["2022YFD1800400"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,1,6]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:sec>\n                    <jats:title>Objectives<\/jats:title>\n                    <jats:p>To evaluate the CRISPR\u2013Cas9 and CRISPR interference (CRISPRi) systems as an antibiotic re-sensitization strategy for reversing multidrug resistance in a clinical Escherichia coli isolate.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Materials and methods<\/jats:title>\n                    <jats:p>The CRISPR\u2013Cas9 system was applied for precise deletion of the acrB gene from clinical E. coli isolate GP53 and homologous recombination (HR) was provided for accurate repairs of double-strand breaks. An arabinose-inducible CRISPRi system was developed and optimized using fluorescent reporter strain GH01. Multiple guide RNAs (gRNAs) targeting acrB were designed, and the most effective gRNA was selected based on its transcriptional suppression of gene acrB. The minimum inhibitory concentrations (MICs) of selected antibiotics in GP53\u0394acrB, CRISPRi strains, WT and WT combined with efflux pump inhibitor PA\u03b2N were evaluated.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>The CRISPR\u2013Cas9 system precisely deleted the acrB gene in clinical E. coli isolate GP53 with 11.46% knockout efficiency. The constructed arabinose-inducible CRISPRi system effectively repressed fluorescent protein expression in strain GH01. Although dCas9 expression increased with L-arabinose concentration, the transcriptional repression efficiency of the target gene under 1\u2005mM induction reached a significant inhibitory level. The CRISPRi system targeting gene acrB exhibited 44.9%, 5.4% and 23.5% inhibition rates on the transcriptional levels with 1\u2005mM L-arabinose for three distinct gRNAs. Both the knockout and CRISPRi strains successfully restored susceptibility of the multidrug-resistant E. coli GP53 to quinolones and tetracyclines, outperforming the effect of PA\u03b2N combination therapy.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>In this study, CRISPR-based systems effectively reversed multidrug resistance in a clinical E. coli isolate, advancing the applications of CRISPR systems in controlling bacterial multidrug resistance.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1093\/jac\/dkaf442","type":"journal-article","created":{"date-parts":[[2025,11,25]],"date-time":"2025-11-25T12:55:32Z","timestamp":1764075332000},"source":"Crossref","is-referenced-by-count":2,"title":["Harnessing CRISPR\u2013Cas9 and CRISPRi systems to reverse antibiotic resistance in a clinical multidrug-resistant\n                    <i>Escherichia coli<\/i>\n                    isolate"],"prefix":"10.1093","volume":"81","author":[{"given":"Ruonan","family":"Zhao","sequence":"first","affiliation":[{"name":"Guangdong Provincial Key Laboratory of Veterinary Pharmaceutics Development and Safety Evaluation, College of Veterinary Medicine, South China Agricultural University , Guangzhou 510642 ,","place":["China"]},{"name":"National Risk Assessment Laboratory for Antimicrobial Resistance of Animal Original Bacteria, College of Veterinary Medicine, South China Agricultural University , Guangzhou 510642 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Peng","family":"Wan","sequence":"additional","affiliation":[{"name":"Guangdong Provincial Key Laboratory of Veterinary Pharmaceutics Development and Safety Evaluation, College of Veterinary Medicine, South China Agricultural University , Guangzhou 510642 ,","place":["China"]},{"name":"National Risk Assessment Laboratory for Antimicrobial Resistance of Animal Original Bacteria, College of Veterinary Medicine, South China Agricultural University , Guangzhou 510642 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Honghao","family":"Huang","sequence":"additional","affiliation":[{"name":"Guangdong Provincial Key Laboratory of Veterinary Pharmaceutics Development and Safety Evaluation, College of Veterinary Medicine, South China Agricultural University , Guangzhou 510642 ,","place":["China"]},{"name":"National Risk Assessment Laboratory for Antimicrobial Resistance of Animal Original Bacteria, College of Veterinary Medicine, South China Agricultural University , Guangzhou 510642 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qi","family":"Li","sequence":"additional","affiliation":[{"name":"Guangdong Provincial Key Laboratory of Veterinary Pharmaceutics Development and Safety Evaluation, College of Veterinary Medicine, South China Agricultural University , Guangzhou 510642 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