{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,12]],"date-time":"2026-10-12T22:02:50Z","timestamp":1791842570184,"version":"4.3.4"},"reference-count":36,"publisher":"American Society for Microbiology","issue":"15","license":[{"start":{"date-parts":[[2013,8,1]],"date-time":"2013-08-01T00:00:00Z","timestamp":1375315200000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["J Bacteriol"],"published-print":{"date-parts":[[2013,8]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n          <jats:p>\n            <jats:named-content xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" content-type=\"genus-species\" xlink:type=\"simple\">Listeria monocytogenes<\/jats:named-content>\n            is a Gram-positive, facultative intracellular pathogen capable of causing severe invasive disease with high mortality rates in humans. While previous studies have largely elucidated the bacterial and host cell mechanisms necessary for invasion, vacuolar escape, and subsequent cell-to-cell spread, the\n            <jats:named-content xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" content-type=\"genus-species\" xlink:type=\"simple\">L. monocytogenes<\/jats:named-content>\n            factors required for rapid replication within the restrictive environment of the host cell cytosol are poorly understood. In this report, we describe a differential fluorescence-based genetic screen utilizing fluorescence-activated cell sorting (FACS) and high-throughput microscopy to identify\n            <jats:named-content xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" content-type=\"genus-species\" xlink:type=\"simple\">L. monocytogenes<\/jats:named-content>\n            mutants defective in optimal intracellular replication. Bacteria harboring deletions within the identified gene\n            <jats:italic>menD<\/jats:italic>\n            or\n            <jats:italic>pepP<\/jats:italic>\n            were defective for growth in primary murine macrophages and plaque formation in monolayers of L2 fibroblasts, thus validating the ability of the screening method to identify intracellular replication-defective mutants. Genetic complementation of the\n            <jats:italic>menD<\/jats:italic>\n            and\n            <jats:italic>pepP<\/jats:italic>\n            deletion strains rescued the\n            <jats:italic>in vitro<\/jats:italic>\n            intracellular infection defects. Furthermore, the\n            <jats:italic>menD<\/jats:italic>\n            deletion strain displayed a general extracellular replication defect that could be complemented by growth under anaerobic conditions, while the intracellular growth defect of this strain could be complemented by the addition of exogenous menaquinone. As prior studies have indicated the importance of aerobic metabolism for\n            <jats:named-content xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" content-type=\"genus-species\" xlink:type=\"simple\">L. monocytogenes<\/jats:named-content>\n            infection, these findings provide further evidence for the importance of menaquinone and aerobic metabolism for\n            <jats:named-content xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" content-type=\"genus-species\" xlink:type=\"simple\">L. monocytogenes<\/jats:named-content>\n            pathogenesis. Lastly, both the\n            <jats:italic>menD<\/jats:italic>\n            and\n            <jats:italic>pepP<\/jats:italic>\n            deletion strains were attenuated during\n            <jats:italic>in vivo<\/jats:italic>\n            infection of mice. These findings demonstrate that the differential fluorescence-based screening approach provides a powerful tool for the identification of intracellular replication determinants in multiple bacterial systems.\n          <\/jats:p>","DOI":"10.1128\/jb.00210-13","type":"journal-article","created":{"date-parts":[[2013,5,18]],"date-time":"2013-05-18T06:08:34Z","timestamp":1368857314000},"page":"3331-3340","update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":13,"title":["A Differential Fluorescence-Based Genetic Screen Identifies Listeria monocytogenes Determinants Required for Intracellular Replication"],"prefix":"10.1128","volume":"195","author":[{"given":"Kyle J.","family":"Perry","sequence":"first","affiliation":[{"name":"Department of Microbiology and Immunobiology, Harvard Medical School, Boston, Massachusetts, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Darren E.","family":"Higgins","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Immunobiology, Harvard Medical School, Boston, Massachusetts, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"235","reference":[{"key":"e_1_3_3_2_2","doi-asserted-by":"publisher","DOI":"10.1128\/CMR.14.3.584-640.2001"},{"key":"e_1_3_3_3_2","doi-asserted-by":"crossref","first-page":"1597","DOI":"10.1083\/jcb.109.4.1597","article-title":"Actin filaments and the growth, movement, and spread of the intracellular bacterial parasite, Listeria monocytogenes.","volume":"109","author":"Tilney LG","year":"1989","unstructured":"TilneyLGPortnoyDA. 1989. 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