[go: up one dir, main page]
More Web Proxy on the site http://driver.im/
Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

The role of toxin A and toxin B in Clostridium difficile infection

Nature volume 467, pages 711–713 (2010)Cite this article

Subjects

Abstract

Clostridium difficile infection is the leading cause of healthcare-associated diarrhoea in Europe and North America1,2. During infection, C. difficile produces two key virulence determinants, toxin A and toxin B. Experiments with purified toxins have indicated that toxin A alone is able to evoke the symptoms of C. difficile infection, but toxin B is unable to do so unless it is mixed with toxin A or there is prior damage to the gut mucosa3. However, a recent study indicated that toxin B is essential for C. difficile virulence and that a strain producing toxin A alone was avirulent4. This creates a paradox over the individual importance of toxin A and toxin B. Here we show that isogenic mutants of C. difficile producing either toxin A or toxin B alone can cause fulminant disease in the hamster model of infection. By using a gene knockout system5,6 to inactivate the toxin genes permanently, we found that C. difficile producing either one or both toxins showed cytotoxic activity in vitro that translated directly into virulence in vivo. Furthermore, by constructing the first ever double-mutant strain of C. difficile, in which both toxin genes were inactivated, we were able to completely attenuate virulence. Our findings re-establish the importance of both toxin A and toxin B and highlight the need to continue to consider both toxins in the development of diagnostic tests and effective countermeasures against C. difficile.

Access through your institution
Buy or subscribe

This is a preview of subscription content, access via your institution

Access options

Access through your institution

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Characterization of C. difficile toxin mutants.
Figure 2: In vitro cytotoxicity.
Figure 3: Virulence of C. difficile strains in hamsters.

Similar content being viewed by others

References

  1. Poutanen, S. M. & Simor, A. E. Clostridium difficile-associated diarrhea in adults. Can. Med. Assoc. J. 171, 51–58 (2004)

    Article  Google Scholar 

  2. Elliott, B., Chang, B. J., Golledge, C. L. & Riley, T. V. Clostridium difficile-associated diarrhoea. Intern. Med. J. 37, 561–568 (2007)

    Article  CAS  Google Scholar 

  3. Lyerly, D. M., Saum, K. E., MacDonald, D. K. & Wilkins, T. D. Effects of Clostridium difficile toxins given intragastrically to animals. Infect. Immun. 47, 349–352 (1985)

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Lyras, D. et al. Toxin B is essential for virulence of Clostridium difficile . Nature 458, 1176–1179 (2009)

    Article  ADS  CAS  Google Scholar 

  5. Heap, J. T., Pennington, O. J., Cartman, S. T., Carter, G. P. & Minton, N. P. The ClosTron: a universal gene knock-out system for the genus Clostridium . J. Microbiol. Methods 70, 452–464 (2007)

    Article  CAS  Google Scholar 

  6. Heap, J. T. et al. The ClosTron: Mutagenesis in Clostridium refined and streamlined. J. Microbiol. Methods 80, 49–55 (2010)

    Article  CAS  Google Scholar 

  7. Just, I. & Gerhard, R. Large clostridial cytotoxins. Rev. Physiol. Biochem. Pharmacol. 152, 23–47 (2004)

    Article  CAS  Google Scholar 

  8. Hussain, H. A., Roberts, A. P. & Mullany, P. Generation of an erythromycin-sensitive derivative of Clostridium difficile strain 630 (630Δerm) and demonstration that the conjugative transposon Tn916ΔE enters the genome of this strain at multiple sites. J. Med. Microbiol. 54, 137–141 (2005)

    Article  CAS  Google Scholar 

  9. Torres, J., Camorlinga-Ponce, M. & Munoz, O. Sensitivity in culture of epithelial cells from rhesus monkey kidney and human colon carcinoma to toxins A and B from Clostridium difficile . Toxicon 30, 419–426 (1992)

    Article  CAS  Google Scholar 

  10. Sebaihia, M. et al. The multidrug-resistant human pathogen Clostridium difficile has a highly mobile, mosaic genome. Nature Genet. 38, 779–786 (2006)

    Article  Google Scholar 

  11. Hachler, H., Berger-Bachi, B. & Kayser, F. H. Genetic characterization of a Clostridium difficile erythromycin-clindamycin resistance determinant that is transferable to Staphylococcus aureus . Antimicrob. Agents Chemother. 31, 1039–1045 (1987)

    Article  CAS  Google Scholar 

  12. O’Connor, J. R. et al. Construction and analysis of chromosomal Clostridium difficile mutants. Mol. Microbiol. 61, 1335–1351 (2006)

    Article  Google Scholar 

  13. Kim, P. H., Iaconis, J. P. & Rolfe, R. D. Immunization of adult hamsters against Clostridium difficile-associated ileocecitis and transfer of protection to infant hamsters. Infect. Immun. 55, 2984–2992 (1987)

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Voth, D. E. & Ballard, J. D. Clostridium difficile toxins: mechanism of action and role in disease. Clin. Microbiol. Rev. 18, 247–263 (2005)

    Article  CAS  Google Scholar 

  15. Du, T. & Alfa, M. J. Translocation of Clostridium difficile toxin B across polarized Caco-2 cell monolayers is enhanced by toxin A. Can. J. Infect. Dis. 15, 83–88 (2004)

    Article  Google Scholar 

  16. Chaves-Olarte, E. et al. R-Ras glucosylation and transient RhoA activation determine the cytopathic effect produced by toxin B variants from toxin A-negative strains of Clostridium difficile . J. Biol. Chem. 278, 7956–7963 (2003)

    Article  CAS  Google Scholar 

  17. Torres, J. F. Purification and characterisation of toxin B from a strain of Clostridium difficile that does not produce toxin A. J. Med. Microbiol. 35, 40–44 (1991)

    Article  CAS  Google Scholar 

  18. Alfa, M. J. et al. Characterization of a toxin A-negative, toxin B-positive strain of Clostridium difficile responsible for a nosocomial outbreak of Clostridium difficile-associated diarrhea. J. Clin. Microbiol. 38, 2706–2714 (2000)

    CAS  PubMed  PubMed Central  Google Scholar 

  19. Stabler, R. A., Dawson, L. F., Phua, L. T. & Wren, B. W. Comparative analysis of BI/NAP1/027 hypervirulent strains reveals novel toxin B-encoding gene (tcdB) sequences. J. Med. Microbiol. 57, 771–775 (2008)

    Article  CAS  Google Scholar 

  20. Drudy, D., Fanning, S. & Kyne, L. Toxin A-negative, toxin B-positive Clostridium difficile . Int. J. Infect. Dis. 11, 5–10 (2007)

    Article  CAS  Google Scholar 

  21. Drudy, D., Harnedy, N., Fanning, S., Hannan, M. & Kyne, L. Emergence and control of fluoroquinolone-resistant, toxin A-negative, toxin B-positive Clostridium difficile . Infect. Control Hosp. Epidemiol. 28, 932–940 (2007)

    Article  Google Scholar 

  22. Heap, J. T., Pennington, O. J., Cartman, S. T. & Minton, N. P. A modular system for Clostridium shuttle plasmids. J. Microbiol. Methods 78, 79–85 (2009)

    Article  CAS  Google Scholar 

  23. Williams, D. R., Young, D. I. & Young, M. Conjugative plasmid transfer from Escherichia coli to Clostridium acetobutylicum . J. Gen. Microbiol. 136, 819–826 (1990)

    Article  CAS  Google Scholar 

  24. Sorg, J. A. & Sonenshein, A. L. Bile salts and glycine as cogerminants for Clostridium difficile spores. J. Bacteriol. 190, 2505–2512 (2008)

    Article  CAS  Google Scholar 

  25. Cartman, S. T. & Minton, N. P. A mariner-based transposon system for in vivo random mutagenesis of Clostridium difficile . Appl. Environ. Microbiol. 76, 1103–1109 (2010)

    Article  CAS  Google Scholar 

  26. Sambrook, J. & Russell, D. W. Molecular Cloning: A Laboratory Manual (Cold Spring Harbor Laboratory Press, 2001)

    Google Scholar 

Download references

Acknowledgements

S.A.K., S.T.C., A.C. and N.P.M. acknowledge the financial support of the UK Medical Research Council (G0601176). Support for M.L.K. was provided by the European Union (HEALTH-F3-2008-223585) and for J.T.H. by the Biotechnology and Biological Sciences Research Council (BBSRC) (BB/F003390/1). We thank C. von-Eichel Streiber for supplying the neutralizing antibody serums and Y. Mahida for the Vero cells. We are also grateful to A. Olling and R. Gerhard for methodological advice on western blots.

Author information

Author notes

  1. Sarah A. Kuehne and Stephen T. Cartman: These authors contributed equally to this work.

Authors and Affiliations

  1. Clostridia Research Group, Centre for Biomolecular Sciences, School of Molecular Medical Sciences, Nottingham Digestive Diseases Centre, NIHR Biomedical Research Unit, University of Nottingham, Nottingham, NG7 2RD, UK

    Sarah A. Kuehne, Stephen T. Cartman, John T. Heap, Michelle L. Kelly, Alan Cockayne & Nigel P. Minton

Authors
  1. Sarah A. Kuehne
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Stephen T. Cartman
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. John T. Heap
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Michelle L. Kelly
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Alan Cockayne
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Nigel P. Minton
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

The study was conceived by N.P.M. and designed by S.A.K., S.T.C. and J.T.H. Construction of mutants and in vitro characterization was carried out by S.A.K. In vivo work was carried out by S.T.C., M.L.K. and A.C. Analysis of data was carried out by S.A.K. and M.L.K. with assistance from S.T.C. and J.T.H. The manuscript was written by S.A.K. and S.T.C. with critical input from all other authors. Funding for the study was sourced by N.P.M. and A.C.

Corresponding author

Correspondence to Nigel P. Minton.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Figures

This file contains Supplementary Figure 1 with a legend. (PDF 669 kb)

PowerPoint slides

Rights and permissions

Reprints and permissions

About this article

Cite this article

Kuehne, S., Cartman, S., Heap, J. et al. The role of toxin A and toxin B in Clostridium difficile infection. Nature 467, 711–713 (2010). https://doi.org/10.1038/nature09397

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature09397

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing