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Expression of diphtheria toxin in Streptococcus mutans and induction of toxin-neutralizing antisera

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Abstract:

The nontoxic full-length diphtheria toxin (DTX), fragment A (DTA), and fragment B (DTB) were each genetically fused to the major surface protein antigen P1 (SpaP) of Streptococcus mutans. Repeated attempts to express the recombinant DTX and DTB in the live oral vaccine candidate Streptococcus gordonii were unsuccessful, whereas DTA could be readily expressed in this bacterium. However, the recombinant DTX, DTB, and DTA could be expressed in the related oral bacterium S. mutans. Western blotting and enzyme-linked immunosorbant assay (ELISA) using anti-DTX and anti-P1 antibodies demonstrated the expression of the three fusion proteins in S. mutans. Mouse antisera raised against the recombinant S. mutans recognized the native DTX in Western immunoblotting. The antibodies raised against S. mutans expressing the recombinant DTX and DTA neutralized the cytotoxicity of the native toxin in a Vero cell assay, but the neutralization titers were relatively low. The potential of using S. gordonii as a live vaccine against diphtheria faces major challenges in the expression of DTX in this organism and in the induction of high-titer toxin-neutralizing antibodies.Key words: diphtheria toxin, Streptococcus mutans, Streptococcus gordonii.

La toxine diphtérique non toxique de pleine longueur (DTX), le fragment A (DTA) et le fragment B (DTB) ont été chacun génétiquement fusionnés à l'antigène protéique majeur de surface P1 (SpaP) de Streptococcus mutans. Des tentatives répétées pour exprimer la DTX et la DTB recombinantes dans le candidat pour un vaccin oral vivant Streptococcus gordonii ont échoué alors que la DTA a pu être aisément exprimée dans cette bactérie. Toutefois, les DTX, DTB et DTA recombinantes ont pu être exprimées chez la bactérie orale apparentée S. mutans. Un immunobuvardage de type Western et un ELISA utilisant des anticorps dirigés contre DTX et P1 ont démontré l'expression des trois protéines de fusion chez S. mutans. Des antisérums de souris développés contre le S. mutans recombinant ont reconnu la DTX native dans un immunobuvardage de type Western. Les anticorps développés contre S. mutans exprimant la DTX et la DTA recombinantes ont neutralisé la cytotoxicité de la toxine native dans un essai sur cellules Vero, mais les titres de neutralisation étaient relativement bas. Le potentiel d'utilisation de S. gordonii en tant que vaccin vivant contre la diphtérie fait face à des obstacles majeurs dans l'expression de la DTX par cet organisme et dans l'induction des titres élevés d'anticorps neutralisant la toxine.Mots clés : toxine diphtérique, Streptococcus mutans, Streptococcus gordonii.[Traduit par la Rédaction]

Document Type: Research Article

Publication date: September 1, 2005

More about this publication?
  • Published since 1954, this monthly journal contains new research in the field of microbiology including applied microbiology and biotechnology; microbial structure and function; fungi and other eucaryotic protists; infection and immunity; microbial ecology; physiology, metabolism and enzymology; and virology, genetics, and molecular biology. It also publishes review articles and notes on an occasional basis, contributed by recognized scientists worldwide.
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