In vitro and in vivo stability of recombinant plasmids in a vaccine strain of Salmonella enterica var. Typhimurium

Sarah J. Dunstan, Cameron P. Simmons, Richard A. Strugnell

Research output: Contribution to journalArticleResearchpeer-review

Abstract

This study examined the ability of different plasmid vectors encoding HC fragment, the non-toxic binding portion of tetanus toxin, to be stably retained by Salmonella enterica var. Typhimurium (Salmonella typhimurium) vaccine strain BRD509 and, upon immunisation, to induce an antibody response against the carried antigen. The HC fragment expression cassette containing the transcription/translation signals, HC fragment open reading frame and the downstream TrpA terminator, was excised from pTETtac4 and incorporated into the plasmids pIC20H, pBR322, pACYC184 and pRSF1010. The resulting constructs were transferred into attenuated S. typhimurium, BRD509, and the level of HC fragment expression was examined by Western blot analysis. The relative stability of each plasmid in S. typhimurium was determined in vitro in the absence of antibiotic selection, and in vivo following immunisation. The ability of each HC fragment-expressing strain to induce lipopolysaccharide- and tetanus toxoid-specific antibody responses was assayed by an enzyme-linked immunosorbent assay. These studies showed that all the vaccine vector constructs, except the S. typhimurium carrying the expression vector based on pIC20H, were able to elicit a high titre immune response. The level of tetanus toxoid-specific antibody induced by S. typhimurium directly correlated with the level of in vitro and in vivo stability of the HC fragment expression plasmid carried by the bacterium, and not with an increased copy number of the parent plasmid vector.

Original languageEnglish
Pages (from-to)111-119
Number of pages9
JournalFEMS Immunology and Medical Microbiology
Volume37
Issue number2-3
DOIs
Publication statusPublished - 15 Jul 2003
Externally publishedYes

Keywords

  • Plasmid stability
  • Salmonella typhimurium
  • Vaccine vector

Cite this

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title = "In vitro and in vivo stability of recombinant plasmids in a vaccine strain of Salmonella enterica var. Typhimurium",
abstract = "This study examined the ability of different plasmid vectors encoding HC fragment, the non-toxic binding portion of tetanus toxin, to be stably retained by Salmonella enterica var. Typhimurium (Salmonella typhimurium) vaccine strain BRD509 and, upon immunisation, to induce an antibody response against the carried antigen. The HC fragment expression cassette containing the transcription/translation signals, HC fragment open reading frame and the downstream TrpA terminator, was excised from pTETtac4 and incorporated into the plasmids pIC20H, pBR322, pACYC184 and pRSF1010. The resulting constructs were transferred into attenuated S. typhimurium, BRD509, and the level of HC fragment expression was examined by Western blot analysis. The relative stability of each plasmid in S. typhimurium was determined in vitro in the absence of antibiotic selection, and in vivo following immunisation. The ability of each HC fragment-expressing strain to induce lipopolysaccharide- and tetanus toxoid-specific antibody responses was assayed by an enzyme-linked immunosorbent assay. These studies showed that all the vaccine vector constructs, except the S. typhimurium carrying the expression vector based on pIC20H, were able to elicit a high titre immune response. The level of tetanus toxoid-specific antibody induced by S. typhimurium directly correlated with the level of in vitro and in vivo stability of the HC fragment expression plasmid carried by the bacterium, and not with an increased copy number of the parent plasmid vector.",
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In vitro and in vivo stability of recombinant plasmids in a vaccine strain of Salmonella enterica var. Typhimurium. / Dunstan, Sarah J.; Simmons, Cameron P.; Strugnell, Richard A.

In: FEMS Immunology and Medical Microbiology, Vol. 37, No. 2-3, 15.07.2003, p. 111-119.

Research output: Contribution to journalArticleResearchpeer-review

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T1 - In vitro and in vivo stability of recombinant plasmids in a vaccine strain of Salmonella enterica var. Typhimurium

AU - Dunstan, Sarah J.

AU - Simmons, Cameron P.

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N2 - This study examined the ability of different plasmid vectors encoding HC fragment, the non-toxic binding portion of tetanus toxin, to be stably retained by Salmonella enterica var. Typhimurium (Salmonella typhimurium) vaccine strain BRD509 and, upon immunisation, to induce an antibody response against the carried antigen. The HC fragment expression cassette containing the transcription/translation signals, HC fragment open reading frame and the downstream TrpA terminator, was excised from pTETtac4 and incorporated into the plasmids pIC20H, pBR322, pACYC184 and pRSF1010. The resulting constructs were transferred into attenuated S. typhimurium, BRD509, and the level of HC fragment expression was examined by Western blot analysis. The relative stability of each plasmid in S. typhimurium was determined in vitro in the absence of antibiotic selection, and in vivo following immunisation. The ability of each HC fragment-expressing strain to induce lipopolysaccharide- and tetanus toxoid-specific antibody responses was assayed by an enzyme-linked immunosorbent assay. These studies showed that all the vaccine vector constructs, except the S. typhimurium carrying the expression vector based on pIC20H, were able to elicit a high titre immune response. The level of tetanus toxoid-specific antibody induced by S. typhimurium directly correlated with the level of in vitro and in vivo stability of the HC fragment expression plasmid carried by the bacterium, and not with an increased copy number of the parent plasmid vector.

AB - This study examined the ability of different plasmid vectors encoding HC fragment, the non-toxic binding portion of tetanus toxin, to be stably retained by Salmonella enterica var. Typhimurium (Salmonella typhimurium) vaccine strain BRD509 and, upon immunisation, to induce an antibody response against the carried antigen. The HC fragment expression cassette containing the transcription/translation signals, HC fragment open reading frame and the downstream TrpA terminator, was excised from pTETtac4 and incorporated into the plasmids pIC20H, pBR322, pACYC184 and pRSF1010. The resulting constructs were transferred into attenuated S. typhimurium, BRD509, and the level of HC fragment expression was examined by Western blot analysis. The relative stability of each plasmid in S. typhimurium was determined in vitro in the absence of antibiotic selection, and in vivo following immunisation. The ability of each HC fragment-expressing strain to induce lipopolysaccharide- and tetanus toxoid-specific antibody responses was assayed by an enzyme-linked immunosorbent assay. These studies showed that all the vaccine vector constructs, except the S. typhimurium carrying the expression vector based on pIC20H, were able to elicit a high titre immune response. The level of tetanus toxoid-specific antibody induced by S. typhimurium directly correlated with the level of in vitro and in vivo stability of the HC fragment expression plasmid carried by the bacterium, and not with an increased copy number of the parent plasmid vector.

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