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Microbial Immunity and Vaccines

Salmonella enterica Serovar Typhimurium Expressing a Chromosomally Integrated Copy of the Bacillus anthracis Protective Antigen Gene Protects Mice against an Anthrax Spore Challenge

Helen S. Garmory, Richard W. Titball, Kate F. Griffin, Ulrike Hahn, Reinhard Böhm, Wolfgang Beyer
Helen S. Garmory
1Defence Science and Technology Laboratory, Salisbury SP4 0JQ
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  • For correspondence: hsgarmory@dstl.gov.uk
Richard W. Titball
1Defence Science and Technology Laboratory, Salisbury SP4 0JQ
2Department of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, London WC1E 7HT, United Kingdom
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Kate F. Griffin
1Defence Science and Technology Laboratory, Salisbury SP4 0JQ
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Ulrike Hahn
3Institute of Environmental and Animal Hygiene, University of Hohenheim, 70593 Stuttgart, Germany
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Reinhard Böhm
3Institute of Environmental and Animal Hygiene, University of Hohenheim, 70593 Stuttgart, Germany
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Wolfgang Beyer
3Institute of Environmental and Animal Hygiene, University of Hohenheim, 70593 Stuttgart, Germany
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DOI: 10.1128/IAI.71.7.3831-3836.2003
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  • FIG. 1.
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    FIG. 1.

    Construction of ZpVDL9.3PA83ec and ZpUTKmPA83ec.

  • FIG. 2.
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    FIG. 2.

    Western blot of auxotrophic S. enterica serovar Typhimurium cell lysates and culture supernatants separated by SDS-PAGE and probed with anti-PA monoclonal antibody. ZpVDL9.3PA83ec (plasmid-borne PA gene), ZpUTKmPA83ec (chromosomal PA gene), and Zoosaloral H (parent) were cultured overnight in Luria broth with appropriate antibiotic selection. Harvested cells and supernatants were normalized to similar CFU per milliliter and lysed in SDS-PAGE sample buffer. Lane M, Bio-Rad biotinylated SDS-PAGE standards, low range; lane 1, 1 mg of recombinant PA/ml; lane 2, Zoosaloral H cell lysate; lane 3, ZpVDL9.3PA83ec cell lysate; lane 4, ZpUTKmPA83ec cell lysate; lane 5, Zoosaloral H supernatant; lane 6, ZpVDL9.3PA83ec supernatant; lane 7, ZpUTKmPA83ec supernatant.

  • FIG. 3.
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    FIG. 3.

    IgG1 and IgG2a responses to PA determined by ELISA of serum samples following oral or i.v. immunization of mice. Groups of A/J mice (n = 10) were inoculated i.v. (105 CFU) (A) or orally (109 CFU, with a gavage needle) (B) with S. enterica serovar Typhimurium expressing chromosomally encoded PA83ec-′hlyA. Circles show individual values; mean values are indicated by horizontal lines. Results for mice with responses to PA below the limit of detection are not shown.

  • FIG. 4.
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    FIG. 4.

    Numbers of mice surviving challenge with 100 (A) or 1,000 (B) LD50s of B. anthracis STI spores over 14 days after oral (open circles) or i.v. (open squares) inoculation with S. enterica serovar Typhimurium expressing chromosomally encoded PA83ec-′hlyA. The survival of control mice inoculated with S. enterica serovar Typhimurium Zoosaloral H alone is also shown (closed triangles).

Tables

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  • TABLE 1.

    Codon usage in native PA83, E. coli codon-optimized PA83ec, and S. enterica serovar Typhimuriuma

    Amino acidCodonFrequency/1,000 codons in:
    PA83PA83ecSalmonella
    RCGA2.70.03.5
    CGC0.010.822.5
    CGG5.40.06.2
    CGU4.127.119.9
    AGA21.70.03.1
    AGG4.10.02.0
    LCUA4.10.04.6
    CUC2.71.39.9
    CUG1.458.449.4
    CUU8.214.910.5
    UUA51.610.813.4
    UUG9.52.711.7
    SUCA19.00.07.4
    UCC4.125.810.9
    UCG9.50.08.3
    UCU28.547.58.7
    AGC1.413.516.4
    AGU31.26.77.6
    TACA28.51.36.8
    ACC8.244.824.1
    ACG13.68.115.5
    ACU25.821.78.3
    PCCA13.68.16.5
    CCC4.11.36.0
    CCG8.224.423.7
    CCU12.22.77.7
    AGCA23.113.513.5
    GCC2.78.126.7
    GCG9.510.837.2
    GCU17.721.714.7
    GGGA25.80.07.4
    GGC2.721.734.3
    GGG14.90.011.3
    GGU4.124.419.6
    VGUA23.114.912.2
    GUC2.76.717.7
    GUG13.610.824.8
    GUU14.921.715.8
    KAAA59.847.536.7
    AAG17.129.812.0
    NAAC21.774.721.3
    AAU70.717.620.2
    QCAA35.312.212.8
    CAG5.428.531.0
    HCAC1.48.19.4
    CAU12.26.711.9
    EGAA51.644.838.4
    GAC8.236.621.7
    DGAG16.323.021.5
    GAU55.727.134.1
    YUAC8.227.112.6
    UAU29.910.817.6
    CUGC0.00.06.3
    UGU0.00.05.0
    FUUC6.820.315.3
    UUU25.812.222.5
    IAUA21.70.06.7
    AUC13.648.924.8
    AUU38.023.028.2
    MAUG10.910.925.5
    WUGG9.59.511.8
    • ↵ a Data for PA83 codon usage were derived from (Williamson et al., (8 July 2001, British Patent Office). Data for S. enterica serovar Typhimurium codon usage were derived from Tzschaschel et al. (28).

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Salmonella enterica Serovar Typhimurium Expressing a Chromosomally Integrated Copy of the Bacillus anthracis Protective Antigen Gene Protects Mice against an Anthrax Spore Challenge
Helen S. Garmory, Richard W. Titball, Kate F. Griffin, Ulrike Hahn, Reinhard Böhm, Wolfgang Beyer
Infection and Immunity Jul 2003, 71 (7) 3831-3836; DOI: 10.1128/IAI.71.7.3831-3836.2003

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Salmonella enterica Serovar Typhimurium Expressing a Chromosomally Integrated Copy of the Bacillus anthracis Protective Antigen Gene Protects Mice against an Anthrax Spore Challenge
Helen S. Garmory, Richard W. Titball, Kate F. Griffin, Ulrike Hahn, Reinhard Böhm, Wolfgang Beyer
Infection and Immunity Jul 2003, 71 (7) 3831-3836; DOI: 10.1128/IAI.71.7.3831-3836.2003
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KEYWORDS

anthrax
Anthrax Vaccines
Antigens, Bacterial
bacterial toxins
Salmonella Typhimurium
Vaccines, Synthetic

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