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This study explored potential CS in clinical and experimentally evolved drug-resistant\n                  <jats:italic>Pseudomonas aeruginosa<\/jats:italic>\n                  (PA) and investigated underlying mechanisms. Whole-genome sequencing and RNA-seq were analyzed to identify genetic and transcriptional correlations.\n                  <jats:italic>In vitro<\/jats:italic>\n                  efficacies were assessed with co-and sequential-exposure regimens. Multiple CF isolates and experimentally evolved gentamycin (GEN) resistant strains consistently exhibited strong CS to novobiocin (NOV). Comparative genomics revealed\n                  <jats:italic>pmrB<\/jats:italic>\n                  gain-of-function mutations, which was further supported by transcriptomic signatures of pmrAB activation. Transcriptomic data suggests potential outer-membrane remodeling characterized by polyamine accumulation and compromised porin channel expression. Additionally, the reduction in proton motive force (PMF) further explains the possible mechanism underlying GEN resistance. As NOV efflux is PMF-dependent, this energetic deficit created a PMF-efflux mismatch, leading to hypersensitivity to NOV. Notably, sequential GEN\u2192NOV treatment effectively restricted the emergence of GEN resistant subpopulations. Overall, our data suggest GEN resistance in PA may arises through envelope remodeling and reduced PMF, which impairs efflux pumps and creates hypersensitivity to NOV. Exploiting this PMF-efflux mismatch with sequential treatment effectively restricted the emergence of GEN resistance.\n                <\/jats:p>","DOI":"10.64898\/2026.05.01.722319","type":"posted-content","created":{"date-parts":[[2026,5,6]],"date-time":"2026-05-06T04:40:17Z","timestamp":1778042417000},"source":"Crossref","is-referenced-by-count":0,"title":["Proton motive force mediated efflux mismatch drives gentamycin-novobiocin collateral sensitivity in\n                  <i>Pseudomonas aeruginosa<\/i>"],"prefix":"10.64898","author":[{"given":"Hafij Al","family":"Mahmud","sequence":"first","affiliation":[{"name":"Department of Biological Sciences, Texas Tech University, Lubbock, Texas, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"MD Siddiqur","family":"Rahman","sequence":"additional","affiliation":[{"name":"Department of Biological Sciences, Texas Tech University, Lubbock, Texas, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Erika","family":"Ortiz","sequence":"additional","affiliation":[{"name":"Department of Biological Sciences, Texas Tech University, Lubbock, Texas, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alex","family":"Luecke","sequence":"additional","affiliation":[{"name":"Department of Biological Sciences, Texas Tech University, Lubbock, Texas, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Amanda M.V.","family":"Brown","sequence":"additional","affiliation":[{"name":"Department of Biological Sciences, Texas Tech University, Lubbock, Texas, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Catherine","family":"Wakeman","sequence":"additional","affiliation":[{"name":"Department of Biological Sciences, Texas Tech University, Lubbock, Texas, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"54368","reference":[{"key":"2026050813250888000_2026.05.01.722319v1.1","doi-asserted-by":"crossref","unstructured":"Hasan M , Dawan J , Ahn J . 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