1887

Abstract

The objective of this study was to compare full binary toxin loci (CDTloc) sequences from a collection of isolates in an effort to further understand the regulation of the binary toxin (CdtAB) and its putative regulator (CdtR). Sequences from different ribotypes and toxinotypes were analysed phylogenetically and for polymorphisms, non-sense mutations, promoter features and signal sequences. Expression of , which was also representative of expression, was measured by quantitative PCR (qPCR). Several consensus promoter features and various polymorphisms were identified including a non-sense mutation identified in a ribotype 078 gene that is predicted to result in a severely truncated protein. Despite this mutation, expression was still detected by qPCR. Dendrograms based on total sequences indicated that isolates belonging to the same ribotype shared the greatest similarity within the binary toxin locus. Although is thought to be involved in regulation of expression, a non-sense mutation did not inhibit expression of , suggesting that either the truncated protein is functional or another regulator of the binary toxin exists.

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2011-08-01
2024-04-19
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References

  1. Bidet P., Barbut F., Lalande V., Burghoffer B., Petit J. C. 1999; Development of a new PCR-ribotyping method for Clostridium difficile based on ribosomal RNA gene sequencing. FEMS Microbiol Lett 175:261–266 [View Article][PubMed]
    [Google Scholar]
  2. Bouvet P. J., Popoff M. R. 2008; Genetic relatedness of Clostridium difficile isolates from various origins determined by triple-locus sequence analysis based on toxin regulatory genes tcdC, tcdR, and cdtR . J Clin Microbiol 46:3703–3713 [View Article][PubMed]
    [Google Scholar]
  3. Carter G. P., Lyras D., Allen D. L., Mackin K. E., Howarth P. M., O’Connor J. R., Rood J. I. 2007; Binary toxin production in Clostridium difficile is regulated by CdtR, a LytTR family response regulator. J Bacteriol 189:7290–7301 [View Article][PubMed]
    [Google Scholar]
  4. Dailey D., Schloemer R. 1988; Identification and characterization of Clostridium difficile promoter element that is functional in Escherichia coli. . Gene 70:343–350 [CrossRef]
    [Google Scholar]
  5. Gasteiger E., Hoogland C., Gattiker A., Duvaud S., Wilkins M. R., Appel R. D., Bairoch A. 2005; Protein identification and analysis tools on the ExPASy Server. In The Proteomics Protocols Handbook pp. 571–607 Edited by Walker J. M. Totowa, NJ: Humana Press; [View Article]
    [Google Scholar]
  6. Gerber M., Walch C., Löffler B., Tischendorf K., Reischl U., Ackermann G. 2008; Effect of sub-MIC concentrations of metronidazole, vancomycin, clindamycin and linezolid on toxin gene transcription and production in Clostridium difficile . J Med Microbiol 57:776–783 [View Article][PubMed]
    [Google Scholar]
  7. Govind R., Vediyappan G., Rolfe R. D., Dupuy B., Fralick J. A. 2009; Bacteriophage-mediated toxin gene regulation in Clostridium difficile . J Virol 83:12037–12045 [View Article][PubMed]
    [Google Scholar]
  8. Griffiths D., Fawley W., Kachrimanidou M., Bowden R., Crook D. W., Fung R., Golubchik T., Harding R. M., Jeffery K. J. M. et al. 2010; Multilocus sequence typing of Clostridium difficile . J Clin Microbiol 48:770–778 [View Article][PubMed]
    [Google Scholar]
  9. Gülke I., Pfeifer G., Liese J., Fritz M., Hofmann F., Aktories K., Barth H. 2001; Characterization of the enzymatic component of the ADP-ribosyltransferase toxin CDTa from Clostridium difficile . Infect Immun 69:6004–6011 [View Article][PubMed]
    [Google Scholar]
  10. Harley C. B., Reynolds R. P. 1987; Analysis of E. coli promoter sequences. Nucleic Acids Res 15:2343–2361 [View Article][PubMed]
    [Google Scholar]
  11. He M., Sebaihia M., Lawley T. D., Stabler R. A., Dawson L. F., Martin M. J., Holt K. E., Seth-Smith H. M., Quail M. A. et al. 2010; Evolutionary dynamics of Clostridium difficile over short and long time scales. Proc Natl Acad Sci U S A 107:7527–7532 [View Article][PubMed]
    [Google Scholar]
  12. Just I., Fritz G., Aktories K., Giry M., Popoff M. R., Boquet P., Hegenbarth S., von Eichel-Streiber C. 1994; Clostridium difficile toxin B acts on the GTP-binding protein Rho. J Biol Chem 269:10706–10712[PubMed]
    [Google Scholar]
  13. Just I., Selzer J., von Eichel-Streiber C., Aktories K. 1995; The low molecular mass GTP-binding protein Rho is affected by toxin A from Clostridium difficile . J Clin Invest 95:1026–1031 [View Article][PubMed]
    [Google Scholar]
  14. Karlsson S., Burman L. G., Åkerlund T. 2008; Induction of toxins in Clostridium difficile is associated with dramatic changes of its metabolism. Microbiology 154:3430–3436 [View Article][PubMed]
    [Google Scholar]
  15. Mani N., Lyras D., Barroso L., Howarth P., Wilkins T., Rood J. I., Sonenshein A. L., Dupuy B. 2002; Environmental response and autoregulation of Clostridium difficile TxeR, a sigma factor for toxin gene expression. J Bacteriol 184:5971–5978 [View Article][PubMed]
    [Google Scholar]
  16. Martin H., Willey B., Low D. E., Staempfli H. R., McGeer A., Boerlin P., Mulvey M., Weese J. S. 2008; Characterization of Clostridium difficile strains isolated from patients in Ontario, Canada, from 2004 to 2006. J Clin Microbiol 46:2999–3004 [View Article][PubMed]
    [Google Scholar]
  17. McDonald L., Killgore G., Thompson A., Owens R. Jr, Kazakova S., Sambol S., Johnson S., Gerding D. 2005; An epidemic, toxin gene-variant strain of Clostridium difficile . N Engl J Med 353:2433–2441 [View Article]
    [Google Scholar]
  18. Metcalf D., Sharif S., Weese J. S. 2010; Evaluation of candidate reference genes in Clostridium difficile for gene expression normalization. Anaerobe 16:439–443 [View Article][PubMed]
    [Google Scholar]
  19. Nikolskaya A. N., Galperin M. Y. 2002; A novel type of conserved DNA-binding domain in the transcriptional regulators of the AlgR/AgrA/LytR family. Nucleic Acids Res 30:2453–2459 [View Article][PubMed]
    [Google Scholar]
  20. Perelle S., Gibert M., Boquet P., Popoff M. R. 1993; Characterization of Clostridium perfringens iota-toxin genes and expression in Escherichia coli . Infect Immun 61:5147–5156[PubMed]
    [Google Scholar]
  21. Perelle S., Gibert M., Bourlioux P., Corthier G., Popoff M. R. 1997; Production of a complete binary toxin (actin-specific ADP-ribosyltransferase) by Clostridium difficile CD196. Infect Immun 65:1402–1407[PubMed]
    [Google Scholar]
  22. Pituch H., Kreft D., Obuch-Woszczatynski P., Wultańska D., Meisel-Mikołajczyk F., Łuczak M., van Belkum A. 2005; Clonal spread of a Clostridium difficile strain with a complete set of toxin A, toxin B, and binary toxin genes among Polish patients with Clostridium difficile-associated diarrhea. J Clin Microbiol 43:472–475 [View Article][PubMed]
    [Google Scholar]
  23. Schwan C., Stecher B., Tzivelekidis T., van Ham M., Rohde M., Hardt W. D., Wehland J., Aktories K. 2009; Clostridium difficile toxin CDT induces formation of microtubule-based protrusions and increases adherence of bacteria. PLoS Pathog 5:e1000626 [View Article]
    [Google Scholar]
  24. Song K. P., Faust C. 1998; Molecular analysis of the promoter region of the Clostridium difficile toxin B gene that is functional in Escherichia coli . J Med Microbiol 47:309–316 [View Article][PubMed]
    [Google Scholar]
  25. Stare B. G., Delmée M., Rupnik M. 2007; Variant forms of the binary toxin CDT locus and tcdC gene in Clostridium difficile strains. J Med Microbiol 56:329–335 [View Article][PubMed]
    [Google Scholar]
  26. Stubbs S., Rupnik M., Gibert M., Brazier J., Duerden B., Popoff M. 2000; Production of actin-specific ADP-ribosyltransferase (binary toxin) by strains of Clostridium difficile . FEMS Microbiol Lett 186:307–312 [View Article][PubMed]
    [Google Scholar]
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