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Infection and Immunity, April 2003, p. 1784-1793, Vol. 71, No. 4
0019-9567/03/$08.00+0     DOI: 10.1128/IAI.71.4.1784-1793.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Expression of Clostridium difficile Toxins A and B and Their Sigma Factor TcdD Is Controlled by Temperature

Sture Karlsson,1,2 Bruno Dupuy,3 Kakoli Mukherjee,4 Elisabeth Norin,2 Lars G. Burman,1 and Thomas Åkerlund1*

Department of Molecular Epidemiology and Biotechnology, Swedish Institute for Infectious Disease Control, S-17182 Solna,1 Microbiology and Tumor Biology Center, Karolinska Institute, S-17177 Stockholm, Sweden,2 Unité de Génétique Moléculaire Bactérienne, Institut Pasteur, 75724 Paris Cedex 15, France,3 AstraZeneca Research Foundation India, Malleswaram, 560003 Bangalore, India4

Received 26 August 2002/ Returned for modification 30 October 2002/ Accepted 30 December 2002

Growth temperature was found to control the expression of toxins A and B in Clostridium difficile VPI 10463, with a maximum at 37°C and low levels at 22 and 42°C in both peptone yeast (PY) and defined media. The up-regulation of toxin A and B mRNA and protein levels upon temperature upshift from 22 to 37°C followed the same kinetics, showing that temperature control occurred at the level of transcription. Experiments with Clostridium perfringens using gusA as a reporter gene demonstrated that both toxin gene promoters were temperature controlled and that their high activity at 37°C was dependent on the alternative sigma factor TcdD. Furthermore, tcdD was found to be autoinduced at 37°C. Glucose down-regulated all these responses in the C. perfringens constructs, similar to its impact on toxin production in C. difficile PY broth cultures. C. difficile proteins induced at 37°C and thus coregulated with the toxins by temperature were demonstrated by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and identified as enzymes involved in butyric acid production and as electron carriers in oxidation-reduction reactions. The regulation of toxin production in C. difficile by temperature is a novel finding apparently reflecting an adaptation of the expression of its virulence to mammalian hosts.

* Corresponding author. Mailing address: Swedish Institute for Infectious Disease Control, Department of Molecular Epidemiology and Biotechnology, S-171 82 Solna, Sweden. Phone: 46 8 4572467. Fax: 46 8 302566. E-mail: Thomas.Akerlund{at}smi.ki.se.

Editor: J. T. Barbieri

Infection and Immunity, April 2003, p. 1784-1793, Vol. 71, No. 4
0019-9567/03/$08.00+0     DOI: 10.1128/IAI.71.4.1784-1793.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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