{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,17]],"date-time":"2026-09-17T13:12:15Z","timestamp":1789650735648,"version":"4.0.0"},"reference-count":79,"publisher":"American Society for Microbiology","issue":"9","license":[{"start":{"date-parts":[[2026,9,17]],"date-time":"2026-09-17T00:00:00Z","timestamp":1789603200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,9,17]],"date-time":"2026-09-17T00:00:00Z","timestamp":1789603200000},"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":["R21AI180326"],"award-info":[{"award-number":["R21AI180326"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["J Bacteriol"],"published-print":{"date-parts":[[2026,9,17]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:sec>\n                    <jats:title\/>\n                    <jats:p>\n                      <jats:italic toggle=\"yes\">Staphylococcus aureus<\/jats:italic>\n                      is a leading cause of biofilm-associated infections, in which communities of bacterial cells are encased in an extracellular matrix composed of polysaccharides, proteins, and extracellular DNA (eDNA) that protect bacteria from host immune defense and antibiotics. Despite their importance, the mechanisms by which matrix components are released from bacterial cells and incorporated into the biofilm matrix remain poorly understood. Using a drip-flow biofilm system, we showed that membrane vesicles (MVs) were associated with the biofilm matrix formed by\n                      <jats:italic toggle=\"yes\">S. aureus<\/jats:italic>\n                      clinical isolate MN8. Proteomic analysis of biofilm matrix proteins and purified MVs showed that biofilm-derived MVs carried cytoplasmic, membrane, and extracellular proteins that closely resembled the protein composition of the biofilm matrix, but differed significantly from MVs produced by planktonic cultures. Biofilm-derived MVs carried significantly higher levels of DNA than MVs from planktonic cultures, and MV-associated DNA was sensitive to DNase treatment, suggesting that eDNA is primarily associated with the MV surface. Although strain MN8 is known to form polysaccharide-dependent biofilms, exogenously added DNase or proteinase K significantly impaired biofilm formation and integrity. Importantly, supplementation with biofilm-derived MVs, but not MVs from planktonic cultures, significantly restored biofilm formation in enzyme-treated static cultures. Together, these findings provide evidence that\n                      <jats:italic toggle=\"yes\">S. aureus<\/jats:italic>\n                      MVs are generated within biofilms, and that these MVs serve as an important resource of matrix components and contribute to biofilm formation.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>IMPORTANCE<\/jats:title>\n                    <jats:p>\n                      Extracellular membrane vesicles (MVs) are important mediators of intercellular communication and have been implicated in the bacterial physiology and pathogenesis. MVs in fungi and gram-negative bacteria mediate key biofilm processes, such as formation and structural maintenance. However, MV production and function in biofilm formation in gram-positive bacteria have remained largely unexplored. Here, we report for the first time the purification and characterization of MVs derived from\n                      <jats:italic toggle=\"yes\">Staphylococcus aureus<\/jats:italic>\n                      biofilms. Our studies demonstrate that\n                      <jats:italic toggle=\"yes\">S. aureus<\/jats:italic>\n                      MVs are important components of the biofilm matrix that contribute to biofilm formation by serving as carriers of key matrix components. This work advances our limited understanding of MVs in gram-positive bacteria and reveals a previously unrecognized mechanism contributing to\n                      <jats:italic toggle=\"yes\">S. aureus<\/jats:italic>\n                      biofilm formation.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1128\/jb.00210-26","type":"journal-article","created":{"date-parts":[[2026,8,7]],"date-time":"2026-08-07T15:01:36Z","timestamp":1786114896000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["Extracellular membrane vesicles\u2014previously unrecognized components of\n                    <i>Staphylococcus aureus<\/i>\n                    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