{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,23]],"date-time":"2026-10-23T20:27:24Z","timestamp":1792787244298,"version":"4.5.0"},"reference-count":43,"publisher":"American Society for Microbiology","issue":"9","license":[{"start":{"date-parts":[[2026,9,22]],"date-time":"2026-09-22T00:00:00Z","timestamp":1790035200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,9,22]],"date-time":"2026-09-22T00:00:00Z","timestamp":1790035200000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"funder":[{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01 AI132154"],"award-info":[{"award-number":["R01 AI132154"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Department of Education and Training | Australian Research Council","award":["DP250102726, DE230100356"],"award-info":[{"award-number":["DP250102726, DE230100356"]}],"id":[{"id":"https:\/\/ror.org\/05mmh0f86","id-type":"ROR","asserted-by":"crossref"}]},{"name":"Department of Education and Training | Australian Research Council","award":["FT220100564"],"award-info":[{"award-number":["FT220100564"]}],"id":[{"id":"https:\/\/ror.org\/05mmh0f86","id-type":"ROR","asserted-by":"crossref"}]},{"name":"Department of Education and Training | Australian Research Council","award":["IM230100534"],"award-info":[{"award-number":["IM230100534"]}],"id":[{"id":"https:\/\/ror.org\/05mmh0f86","id-type":"ROR","asserted-by":"crossref"}]},{"DOI":"10.13039\/501100000925","name":"National Health and Medical Research Council","doi-asserted-by":"publisher","award":["APP2025937"],"award-info":[{"award-number":["APP2025937"]}],"id":[{"id":"10.13039\/501100000925","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["mSystems"],"published-print":{"date-parts":[[2026,9,22]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:sec>\n                    <jats:title\/>\n                    <jats:p>\n                      Polymyxins are often last-resort antibiotics against high-priority Gram-negative pathogens, particularly\n                      <jats:italic toggle=\"yes\">Acinetobacter baumannii<\/jats:italic>\n                      . However, most mechanistic studies to date have overlooked the heterogeneity within bacterial populations during polymyxin treatment. Using time-lapse imaging and propidium iodide (PI) staining, we observed that a subset of PI-positive (PI\n                      <jats:sup>+<\/jats:sup>\n                      ) cells, which are traditionally considered non-viable, were capable of regrowth. This unexpected scenario promoted further investigation into the distinct metabolic responses of PI\n                      <jats:sup>+<\/jats:sup>\n                      or PI-negative (PI\n                      <jats:sup>\u2212<\/jats:sup>\n                      ) subpopulations following polymyxin exposure. By combining a synthetic fluorescent polymyxin probe, FADDI-043, with fluorescence-activated cell sorting (FACS), we isolated PI\n                      <jats:sup>+<\/jats:sup>\n                      and PI\n                      <jats:sup>\u2212<\/jats:sup>\n                      cells and profiled their metabolic responses. PI\n                      <jats:sup>+<\/jats:sup>\n                      cells exhibited increased levels of phosphatidylethanolamine, likely to compensate for the severe membrane damage by polymyxins. In contrast, PI\n                      <jats:sup>\u2212<\/jats:sup>\n                      cells, where polymyxin interacted with bacterial membranes without causing extensive damage, demonstrated broader metabolic adaptations. Notably, arginine metabolism was uniquely upregulated in the PI\n                      <jats:sup>\u2212<\/jats:sup>\n                      group, and exogenous arginine supplementation conferred protection against polymyxin treatment. Collectively, this is the first study to demonstrate subpopulation-specific metabolic responses to polymyxins, highlighting dynamic and heterogeneous bacterial adaptations. Our findings underscore the importance of single-cell analysis to unravel antibiotic mechanisms and may inform novel metabolic reprogramming strategies to enhance antibiotic efficacy and minimize resistance emergence.\n                    <\/jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE<\/jats:title>\n                      <jats:p>\n                        Multidrug-resistant\n                        <jats:italic toggle=\"yes\">Acinetobacter baumannii<\/jats:italic>\n                        is designated a World Health Organization \u201ccritical priority\u201d pathogen, and polymyxins remain the few effective treatment options, particularly in low- and middle-income countries. However, polymyxin heteroresistance poses a major global clinical challenge, and its mechanistic basis remains poorly defined. Most antimicrobial studies rely on population-level measurements, obscuring the biological consequences of phenotypic heterogeneity. Here, we demonstrate that a subset of polymyxin-treated, propidium iodide-positive (PI\n                        <jats:sup>+<\/jats:sup>\n                        )\n                        <jats:italic toggle=\"yes\">A. baumannii<\/jats:italic>\n                        cells, typically classified as non-viable, retain the capacity to regrow. By isolating PI\n                        <jats:sup>+<\/jats:sup>\n                        and PI-negative (PI\n                        <jats:sup>\u2212<\/jats:sup>\n                        ) subpopulations, we uncover distinct metabolic adaptations under polymyxin exposure: PI\n                        <jats:sup>+<\/jats:sup>\n                        cells exhibit elevated phosphatidylethanolamine levels, whereas the PI\n                        <jats:sup>\u2212<\/jats:sup>\n                        cells activate their arginine metabolism. These findings reveal an unrecognized layer of metabolic heterogeneity underlying polymyxin exposure. Our work challenges conventional interpretations of viability assays and highlights the importance of subpopulation-resolved analyses for identifying metabolic vulnerabilities to enhance polymyxin efficacy while limiting resistance emergence.\n                      <\/jats:p>\n                    <\/jats:sec>\n                  <\/jats:sec>","DOI":"10.1128\/msystems.00261-26","type":"journal-article","created":{"date-parts":[[2026,8,7]],"date-time":"2026-08-07T13:00:13Z","timestamp":1786107613000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["Metabolic rewiring of bacterial subpopulations governs polymyxin responses in\n                    <i>Acinetobacter baumannii<\/i>"],"prefix":"10.1128","volume":"11","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-0861-2437","authenticated-orcid":true,"given":"Mei-Ling","family":"Han","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/02bfwt286","id-type":"ROR","asserted-by":"publisher"}],"name":"Infection Program and Department of Microbiology, Biomedicine Discovery Institute, Monash University","place":["Clayton, Australia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhi Ying","family":"Kho","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/02bfwt286","id-type":"ROR","asserted-by":"publisher"}],"name":"Infection Program and Department of Microbiology, Biomedicine Discovery Institute, Monash University","place":["Clayton, Australia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xingjian","family":"Wang","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/02bfwt286","id-type":"ROR","asserted-by":"publisher"}],"name":"Infection Program and Department of Microbiology, Biomedicine Discovery Institute, Monash University","place":["Clayton, Australia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jinxin","family":"Zhao","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/02bfwt286","id-type":"ROR","asserted-by":"publisher"}],"name":"Infection Program and Department of Microbiology, Biomedicine Discovery Institute, Monash University","place":["Clayton, Australia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ben","family":"Finnin","sequence":"additional","affiliation":[{"name":"Cell Therapy Pty Ltd","place":["Melbourne, Australia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7342-3782","authenticated-orcid":true,"given":"Yan","family":"Zhu","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/042pyga86","id-type":"ROR","asserted-by":"publisher"}],"name":"Systems Biology Centre, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences","place":["Tianjin, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christopher K.","family":"Barlow","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05t8s3y29","id-type":"ROR","asserted-by":"publisher"}],"name":"Monash Proteomics and Metabolomics Platform, Biomedicine Discovery Institute, Monash University","place":["Clayton, Australia"]},{"id":[{"id":"https:\/\/ror.org\/05t8s3y29","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Biochemistry, Monash Biomedicine Discovery Institute, Monash University","place":["Clayton, Australia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7497-7082","authenticated-orcid":true,"given":"Darren J.","family":"Creek","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05t8s3y29","id-type":"ROR","asserted-by":"publisher"}],"name":"Monash Proteomics and Metabolomics Platform, Biomedicine Discovery Institute, Monash University","place":["Clayton, Australia"]},{"id":[{"id":"https:\/\/ror.org\/04n35qp23","id-type":"ROR","asserted-by":"publisher"}],"name":"Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University","place":["Parkville, Australia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tony","family":"Velkov","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/02bfwt286","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Pharmacology, Monash Biomedicine Discovery Institute, Monash University","place":["Clayton, Australia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mohammad A. 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