{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,15]],"date-time":"2026-10-15T13:04:49Z","timestamp":1792069489559,"version":"4.4.0"},"reference-count":33,"publisher":"Microbiology Society","issue":"11","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2006,11,1]]},"abstract":"<jats:p>This paper is the first detailed description of the development and use of new genetic tools specifically for the safe manipulation of highly pathogenic<jats:italic>Francisella tularensis<\/jats:italic>subsp.<jats:italic>tularensis<\/jats:italic>. Most of these tools are also demonstrated to work with other<jats:italic>F. tularensis<\/jats:italic>subspecies. Kanamycin and hygromycin resistance determinants that function as genetic markers in<jats:italic>F. tularensis<\/jats:italic>subsp.<jats:italic>tularensis<\/jats:italic>strain Schu and sets of episomal shuttle vectors that are either unstable or stably maintained in the absence of selection were developed. In addition, the<jats:italic>hyg<\/jats:italic>gene, expressed from the<jats:italic>F. tularensis groESL<\/jats:italic>promoter, was successfully used as a marker for transposon mutagenesis. This work also includes the development of<jats:italic>sacB<\/jats:italic>-based suicide plasmids expressing kanamycin resistance that can be used for electroporation-mediated allelic exchange of unmarked mutations in Schu and the<jats:italic>F. tularensis<\/jats:italic>live vaccine strain (LVS). Using these plasmids, the two predicted<jats:italic>\u03b2<\/jats:italic>-lactamase genes,<jats:italic>blaA<\/jats:italic>and<jats:italic>blaB<\/jats:italic>, in Schu and LVS were deleted. Only the \u0394<jats:italic>blaB1<\/jats:italic>mutants had increased susceptibility to ampicillin, and this phenotype was complemented by a plasmid expressing<jats:italic>blaB<\/jats:italic><jats:sup>+<\/jats:sup>. The results suggest that the<jats:italic>\u03b2<\/jats:italic>-lactam antibiotic resistance phenotype of Schu and LVS is likely due to only one of the two<jats:italic>\u03b2<\/jats:italic>-lactamase genes present and that ampicillin resistance can be used as an additional selectable marker in<jats:italic>\u03b2<\/jats:italic>-lactamase deletion mutants. The collection of tools presented in this report will be helpful for the genetic analyses of<jats:italic>F. tularensis<\/jats:italic>subsp.<jats:italic>tularensis<\/jats:italic>pathogenesis.<\/jats:p>","DOI":"10.1099\/mic.0.29121-0","type":"journal-article","created":{"date-parts":[[2006,10,30]],"date-time":"2006-10-30T19:52:12Z","timestamp":1162237932000},"page":"3425-3435","source":"Crossref","is-referenced-by-count":80,"title":["Genetic tools for highly pathogenic Francisella tularensis subsp. tularensis"],"prefix":"10.1099","volume":"152","author":[{"given":"Eric D.","family":"LoVullo","sequence":"first","affiliation":[{"name":"University of Rochester Medical Center, 601 Elmwood Ave, Box 672, Rochester, NY 14642, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lani A.","family":"Sherrill","sequence":"additional","affiliation":[{"name":"University of Rochester Medical Center, 601 Elmwood Ave, Box 672, Rochester, NY 14642, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lanyn L.","family":"Perez","sequence":"additional","affiliation":[{"name":"University of Rochester Medical Center, 601 Elmwood Ave, Box 672, Rochester, NY 14642, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Martin S.","family":"Pavelka","sequence":"additional","affiliation":[{"name":"University of Rochester Medical Center, 601 Elmwood Ave, Box 672, Rochester, NY 14642, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"345","reference":[{"key":"R1","volume-title":"Current Protocols in Molecular Biology","author":"Ausubel","year":"1987"},{"key":"R2","doi-asserted-by":"crossref","first-page":"212","DOI":"10.1128\/JCM.22.2.212-215.1985","article-title":"Antimicrobial susceptibility testing of Francisella tularensis with a modified Mueller-Hinton broth","volume":"22","author":"Baker","year":"1985","journal-title":"J Clin Microbiol"},{"key":"R3","doi-asserted-by":"crossref","first-page":"63","DOI":"10.1111\/j.1574-6968.2000.tb08874.x","article-title":"Isolation and characterization of Francisella novicida mutants defective in lipopolysaccharide biosynthesis","volume":"182","author":"Cowley","year":"2000","journal-title":"FEMS Microbiol Lett"},{"key":"R4","doi-asserted-by":"crossref","first-page":"2763","DOI":"10.1001\/jama.285.21.2763","article-title":"Tularemia as a biological weapon: medical and public health management","volume":"285","author":"Dennis","year":"2001","journal-title":"JAMA"},{"key":"R5","first-page":"1","article-title":"Using the EZ\u200a:\u200a:\u200aTN transposome for transposon mutagenesis in Mycobacterium smegmatis","volume":"7","author":"Derbyshire","year":"2000","journal-title":"Epicentre Forum"},{"key":"R6","doi-asserted-by":"crossref","first-page":"415","DOI":"10.4049\/jimmunol.87.4.415","article-title":"Prophylactic effectiveness of live and killed tularemia vaccines. 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