{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,6,24]],"date-time":"2026-06-24T05:55:59Z","timestamp":1782280559439,"version":"3.54.5"},"posted":{"date-parts":[[2025,10,28]]},"group-title":"Microbiology","reference-count":82,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,10,28]],"date-time":"2025-10-28T00:00:00Z","timestamp":1761609600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/www.biorxiv.org\/about\/FAQ#license"}],"funder":[{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"crossref","award":["R21AI125475"],"award-info":[{"award-number":["R21AI125475"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"crossref","award":["R21AI163580"],"award-info":[{"award-number":["R21AI163580"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"crossref","award":["R01AI175106"],"award-info":[{"award-number":["R01AI175106"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/100007249","name":"University of Minnesota","doi-asserted-by":"crossref","award":["Research and Innovation Office Grant-In-Aid Bridge Award 766054"],"award-info":[{"award-number":["Research and Innovation Office Grant-In-Aid Bridge Award 766054"]}],"id":[{"id":"10.13039\/100007249","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2025,10,28]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  <jats:italic>Mycobacterium tuberculosis<\/jats:italic>\n                  uses several ESX type VII protein secretion systems for pathogenesis.\n                  <jats:italic>M. tuberculosis<\/jats:italic>\n                  ESX-5 is poorly characterized because it is essential for growth in standard lab culture conditions. To circumvent ESX-5 essentiality, we made an\n                  <jats:italic>M. tuberculosis<\/jats:italic>\n                  strain in which the central ESX-5 membrane component EccD\n                  <jats:sub>5<\/jats:sub>\n                  can be conditionally depleted. Here, we use this strain to demonstrate that\n                  <jats:italic>M. tuberculosis<\/jats:italic>\n                  requires the ESX-5 secretion system to grow using specific carbon sources\n                  <jats:italic>in vitro<\/jats:italic>\n                  , to grow in cultured macrophages, and to replicate and disseminate in aerosol-infected mice.\n                  <jats:italic>M. tuberculosis<\/jats:italic>\n                  requires ESX-5 to use glycerol or glucose as the sole carbon source. Use of glycerol and glucose also depends on the outer membrane protein PPE51. We show that\n                  <jats:italic>M. tuberculosis<\/jats:italic>\n                  requires ESX-5 activity for outer membrane export and surface exposure of PPE51. Expression of the outer membrane porin MspA enabled growth of ESX-5 deficient\n                  <jats:italic>M. tuberculosis<\/jats:italic>\n                  on glycerol, suggesting that the main function of ESX-5\n                  <jats:italic>in vitro<\/jats:italic>\n                  is to export nutrient transporters to the outer membrane. Importantly, depletion of EccD\n                  <jats:sub>5<\/jats:sub>\n                  in acutely infected mice resulted in clearance of\n                  <jats:italic>M. tuberculosis<\/jats:italic>\n                  from lung tissues, demonstrating the critical importance of ESX-5 activity during infection. Our findings suggest that ESX-5 promotes\n                  <jats:italic>M. tuberculosis<\/jats:italic>\n                  pathogenesis by mediating export of outer membrane proteins that enable nutrient acquisition.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>Importance<\/jats:title>\n                  <jats:p>\n                    <jats:italic>Mycobacterium tuberculosis<\/jats:italic>\n                    ESX type VII secretion systems play important roles in pathogenesis, but the functions of ESX-5 are not well characterized because it is essential for growth in standard lab culture conditions. We used a strain that conditionally expresses a central membrane component of the ESX-5 secretion apparatus to determine how ESX-5 impacts growth in lab cultures and in a mouse infection model. We found that\n                    <jats:italic>M. tuberculosis<\/jats:italic>\n                    requires ESX-5 to grow using several carbon sources and to grow in the lungs of infected mice. Inhibiting production of the ESX-5 secretion system in mice also led to clearance of\n                    <jats:italic>M. tuberculosis<\/jats:italic>\n                    from lung tissues. Our results demonstrate that the\n                    <jats:italic>M. tuberculosis<\/jats:italic>\n                    ESX-5 system is a critical virulence factor and suggest that ESX-5 is a strong candidate for anti-tubercular drug development.\n                  <\/jats:p>\n                <\/jats:sec>","DOI":"10.1101\/2025.10.28.685107","type":"posted-content","created":{"date-parts":[[2025,10,29]],"date-time":"2025-10-29T03:00:13Z","timestamp":1761706813000},"source":"Crossref","is-referenced-by-count":1,"title":["The\n                  <i>Mycobacterium tuberculosis<\/i>\n                  ESX-5 secretion system enables carbon source utilization and growth in mice"],"prefix":"10.64898","author":[{"given":"Alisha M.","family":"Block","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rashmi Ravindran","family":"Nair","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Virginia","family":"Meikle","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Parker C.","family":"Wiegert","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dylan W.","family":"White","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Leanne","family":"Zhang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4068-8092","authenticated-orcid":false,"given":"Michael","family":"Niederweis","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8404-0250","authenticated-orcid":false,"given":"Anna 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