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Link to original content: https://doi.org/10.1038/nature07822
Toxin B is essential for virulence of Clostridium difficile | Nature
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Toxin B is essential for virulence of Clostridium difficile

Nature volume 458, pages 1176–1179 (2009)Cite this article

Abstract

Clostridium difficile is the leading cause of infectious diarrhoea in hospitals worldwide, because of its virulence, spore-forming ability and persistence1,2. C. difficile-associated diseases are induced by antibiotic treatment or disruption of the normal gastrointestinal flora3,4. Recently, morbidity and mortality resulting from C. difficile-associated diseases have increased significantly due to changes in the virulence of the causative strains and antibiotic usage patterns1,2,5,6. Since 2002, epidemic toxinotype III NAP1/027 strains1,2, which produce high levels of the major virulence factors, toxin A and toxin B, have emerged. These toxins have 63% amino acid sequence similarity7 and are members of the large clostridial glucosylating toxin family, which are monoglucosyltransferases that are pro-inflammatory, cytotoxic and enterotoxic in the human colon8,9,10. Inside host cells, both toxins catalyse the transfer of glucose onto the Rho family of GTPases, leading to cell death8,11. However, the role of these toxins in the context of a C. difficile infection is unknown. Here we describe the construction of isogenic tcdA and tcdB (encoding toxin A and B, respectively) mutants of a virulent C. difficile strain and their use in the hamster disease model to show that toxin B is a key virulence determinant. Previous studies showed that purified toxin A alone can induce most of the pathology observed after infection of hamsters with C. difficile8,9,12 and that toxin B is not toxic in animals unless it is co-administered with toxin A, suggesting that the toxins act synergistically12. Our work provides evidence that toxin B, not toxin A, is essential for virulence. Furthermore, it is clear that the importance of these toxins in the context of infection cannot be predicted exclusively from studies using purified toxins, reinforcing the importance of using the natural infection process to dissect the role of toxins in disease.

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Figure 1: Genetic organization of the C. difficile PaLoc and analysis of toxin mutants.
Figure 2: Comparative analysis of toxin production by wild-type and mutant C. difficile strains.
Figure 3: Virulence of C. difficile wild-type and mutant strains in hamsters.

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Acknowledgements

Research at Monash University was supported by Program Grant 284214 from the Australian National Health and Medical Research Council, funding from the ARC Centre of Excellence in Structural and Functional Microbial Genomics and grant AI057637 from the United States National Institute of Allergy and Infectious Diseases. S.J., D.N.G. and G.V. were supported by Merit Review Grants from the United States Department of Veterans Affairs Research Service. We thank D. Lyerly, K. Aktories and C. von-Eichel Streiber for providing toxin-A-specific and toxin-B-specific antibodies, K. Nagaro and A. Cheknis for assistance with the hamster experiments, V. K. Viswanathan for providing intestinal epithelial cell lines, E. Hartland for providing the HT29 cell line, and M. Merrigan for adherence assays.

Author Contributions D.L., J.R.O’C., G.P.C., V.A. and J.I.R. designed the genetic and molecular aspects of the research, which were carried out by D.L., P.M.H. and G.P.C. S.J., S.P.S., J.R.O’C. and D.N.G. planned and developed the animal experiments, which were carried out by S.P.S. and J.R.O’C. Tissue culture assays were designed, planned and carried out by D.L., G.P.C., R.P., T.P. and G.V. All authors were involved in data analysis and interpretation. The manuscript was primarily written by D.L. and J.I.R. but all authors had very significant input into its content and reviewed and edited the manuscript.

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Authors and Affiliations

  1. Australian Bacterial Pathogenesis Program and,,

    Dena Lyras, Jennifer R. O’Connor, Pauline M. Howarth, Glen P. Carter, Tongted Phumoonna, Rachael Poon, Vicki Adams & Julian I. Rood

  2. Department of Microbiology, Australian Research Council Centre of Excellence in Structural and Functional Microbial Genomics, Monash University, Victoria 3800, Australia,

    Rachael Poon & Julian I. Rood

  3. Medical Service and Research Service, Hines VA Hospital, Hines, Illinois 60141, USA, and Infectious Disease Section, Stritch School of Medicine, Loyola University Chicago, Maywood, Illinois 60153, USA ,

    Jennifer R. O’Connor, Susan P. Sambol, Gayatri Vedantam, Stuart Johnson & Dale N. Gerding

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Lyras, D., O’Connor, J., Howarth, P. et al. Toxin B is essential for virulence of Clostridium difficile. Nature 458, 1176–1179 (2009). https://doi.org/10.1038/nature07822

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