title: Use of stabilized luciferase-expressing plasmids to examine in vivo-induced promoters in the Vibrio cholerae vaccine strain CVD 103-HgR.
abstract: Live, attenuated Vibrio cholerae vaccines can induce potent immune responses after only a single oral dose. The strategy of harnessing these strains to present antigens from heterologous pathogens to the mucosal immune system shows great promise. To fully realize this possibility, V. cholerae strains must be created that stably express antigens in vivo in sufficient quantity to generate an immune response. In vivo-induced promoters have been shown to increase the stability and immunogenicity of foreign antigens expressed from multicopy plasmids. We report the construction of a series of genetically stabilized plasmids expressing luciferase as a heterologous protein from the following in vivo-induced promoters: V. cholerae P(argC), P(fhuC) and P(vca1008), and Salmonella enterica serovar Typhi P(ompC). We demonstrate that several of these expression plasmids meet two critical criteria for V. cholerae live vector vaccine studies. First, the plasmids are highly stable in the V. cholerae vaccine strain CVD 103-HgR at low copy number, in the absence of selective pressure. Second, real-time bioluminescent imaging (BLI) demonstrates inducible in vivo expression of the promoters in the suckling mouse model of V. cholerae colonization. Moreover, the use of BLI allows for direct quantitative comparison of in vivo expression from four different promoters at various time points.
authors.affiliation: Department of Microbiology & Immunology, Center for Vaccine Development, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
authors.forename: Cara E
authors.initials: CE
authors.lastname: Morin
authors.forename: James B
authors.initials: JB
authors.lastname: Kaper
mesh_terms: D000818:Animals; D019677:Artificial Gene Fusion; D022121:Cholera Vaccines / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D017930:Genes, Reporter; D042822:Genomic Instability; D008156:Luciferases / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012485:Salmonella typhi / Q000235:genetics
chemical_list: D022121:Cholera Vaccines; C414990:cholera vaccine CVD 103-HgR; D008156:Luciferases
references.citation: Infect Immun. 2005 Feb;73(2):972-80
references.pmid: 15664940
references.citation: Vaccine. 1997 Apr-May;15(6-7):700-8
references.pmid: 9178472
references.citation: Infect Immun. 2003 Oct;71(10):5498-504
references.pmid: 14500467
references.citation: Immunol Lett. 2005 Mar 15;97(2):181-8
references.pmid: 15752556
references.citation: Infect Immun. 1992 Sep;60(9):3916-7
references.pmid: 1500200
references.citation: Methods Mol Biol. 2008;415:101-18
references.pmid: 18370150
references.citation: Microbiol Rev. 1986 Sep;50(3):314-52
references.pmid: 3534538
references.citation: Biotechnol Lett. 2008 Apr;30(4):571-9
references.pmid: 18008168
references.citation: Trends Microbiol. 2001 Aug;9(8):372-6
references.pmid: 11514219
references.citation: Dev Biol Stand. 1995;84:237-44
references.pmid: 7796960
references.citation: Infect Immun. 1996 Oct;64(10):4373-7
references.pmid: 8926115
references.citation: Vaccine. 1990 Dec;8(6):577-80
references.pmid: 2087879
references.citation: FEMS Microbiol Lett. 1990 Mar 1;56(1-2):149-54
references.pmid: 2332153
references.citation: Infect Immun. 1997 Jun;65(6):2127-35
references.pmid: 9169742
references.citation: Cell Host Microbe. 2007 Oct 11;2(4):264-77
references.pmid: 18005744
references.citation: Proc Natl Acad Sci U S A. 2003 Jul 8;100(14):8508-13
references.pmid: 12826608
references.citation: Proc Natl Acad Sci U S A. 2001 Jun 5;98(12):6889-94
references.pmid: 11391007
references.citation: Vaccine. 1996 Apr;14(6):545-52
references.pmid: 8782354
references.citation: Infect Immun. 2001 Dec;69(12):7493-500
references.pmid: 11705925
references.citation: Science. 2004 Feb 6;303(5659):851-3
references.pmid: 14764883
references.citation: Infect Immun. 2005 Dec;73(12):8167-78
references.pmid: 16299312
references.citation: Infect Immun. 2001 May;69(5):3150-8
references.pmid: 11292735
references.citation: Trends Microbiol. 2000 Apr;8(4):189-91
references.pmid: 10754579
references.citation: J Bacteriol. 2004 Aug;186(15):5167-71
references.pmid: 15262955
references.citation: Infect Immun. 2006 Sep;74(9):5391-6
references.pmid: 16926434
references.citation: Nature. 2002 Jun 6;417(6889):642-5
references.pmid: 12050664
references.citation: Biotechnol Adv. 1995;13(2):247-61
references.pmid: 14537822
references.citation: Infect Immun. 1992 Oct;60(10):4430-2
references.pmid: 1398956
references.citation: J Biotechnol. 2007 Jan 1;127(2):188-98
references.pmid: 16891024
references.citation: Mol Imaging Biol. 2008 Jan-Feb;10(1):54-61
references.pmid: 17994265
references.citation: J Infect Dis. 1992 Oct;166(4):837-41
references.pmid: 1527420
references.citation: Infect Immun. 1999 Dec;67(12):6341-5
references.pmid: 10569747
references.citation: Vaccine. 2000 Aug 15;19(1):75-85
references.pmid: 10924789
references.citation: Mol Microbiol. 1998 Feb;27(4):797-805
references.pmid: 9515705
references.citation: Infect Immun. 2008 Oct;76(10):4422-30
references.pmid: 18663003
references.citation: Infect Immun. 2005 Aug;73(8):4488-93
references.pmid: 16040959
references.citation: Infect Immun. 2000 Mar;68(3):1171-5
references.pmid: 10678922
references.citation: Proc Natl Acad Sci U S A. 2007 Nov 13;104(46):18229-34
references.pmid: 17986616
references.citation: Nat Protoc. 2008;3(4):629-36
references.pmid: 18388945
references.citation: Proc Natl Acad Sci U S A. 2007 Oct 16;104(42):16669-74
references.pmid: 17925449
references.citation: Infect Immun. 2007 Apr;75(4):1827-34
references.pmid: 17145938
references.citation: Infect Immun. 1998 Apr;66(4):1648-53
references.pmid: 9529093
references.citation: J Bacteriol. 2000 Apr;182(8):2350-3
references.pmid: 10735886
references.citation: EcoSal Plus. 2004 Dec;1(1):
references.pmid: 26443366
references.citation: Infect Immun. 2004 Dec;72(12):7096-106
references.pmid: 15557633
references.citation: Infect Immun. 1999 Dec;67(12):6424-33
references.pmid: 10569759
references.citation: Infect Immun. 2003 Mar;71(3):1352-60
references.pmid: 12595452
references.citation: Mol Microbiol. 2002 Mar;43(6):1471-91
references.pmid: 11952899
references.citation: Mol Genet Genomics. 2005 Sep;274(2):180-8
references.pmid: 16047200
references.citation: Infect Immun. 2008 Sep;76(9):4145-51
references.pmid: 18591230