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However, the mechanisms underlying EV biogenesis remain incompletely understood. Using transmission electron microscopy, we demonstrate that X\u2010ray irradiation induces outward blebbing of the inner membrane in\n                    <jats:italic>Pseudomonas aeruginosa<\/jats:italic>\n                    PAO1 (\n                    <jats:italic>P. aeruginosa<\/jats:italic>\n                    PAO1), leading to the formation of outer\u2010inner membrane vesicles (OIMVs) through outer membrane pinching\u2010off. The endolysin Lys, which is negatively regulated by PrtR and positively regulated by PrtN, is essential for OIMV production. Lys translocates into the periplasmic space, where it disrupts the peptidoglycan layer, causing morphological changes from rod\u2010shaped to round cells and facilitating OIMV release. Furthermore, deletion of YciB, a protein crucial for inner membrane integrity, significantly increases OIMV production. In a murine model of acute pneumonia, OIMV immunisation significantly improves pulmonary bacterial clearance, reduces lung injury and enhances survival rates. Our findings reveal inner membrane blebbing as a novel mechanism of OIMV biogenesis in\n                    <jats:italic>P. aeruginosa<\/jats:italic>\n                    PAO1 under X\u2010ray irradiation and highlight the potential of OIMVs as promising vaccine candidates against\n                    <jats:italic>P. aeruginosa<\/jats:italic>\n                    infections.\n                  <\/jats:p>","DOI":"10.1002\/jev2.70151","type":"journal-article","created":{"date-parts":[[2025,8,19]],"date-time":"2025-08-19T00:20:57Z","timestamp":1755562857000},"source":"Crossref","is-referenced-by-count":6,"title":["X\u2010Ray Irradiation of\n                    <i>Pseudomonas aeruginosa<\/i>\n                    Induces Biogenesis of Outer\u2010Inner Membrane Vesicles With Potential as a Vaccine Against Acute Pneumonia"],"prefix":"10.1002","volume":"14","author":[{"given":"Li","family":"Zhang","sequence":"first","affiliation":[{"name":"State Key Laboratory of Biotherapy and Cancer Center 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