Production of a complete binary toxin (actin-specific ADP-ribosyltransferase) by Clostridium difficile CD196

A Clostridium difficile isolate was found to produce an actin-specific ADP-ribosyltransferase (CDT) homologous to the enzymatic components of Clostridium perfringens iota toxin and Clostridium spiroforme toxin (M. R. Popoff, E. J. Rubin, D. M. Gill, and P. Boquet, Infect. Immun. 56:2299-2306, 1988). The CDT locus from C. difficile CD196 was cloned and sequenced. It contained two genes (cdtA and cdtB) which display organizations and sequences similar to those of the iota toxin gene. The deduced enzymatic (CDTa) and binding (CDTb) components have 81 and 84% identity, respectively, with the corresponding components of iota toxin. CDTa and CDTb induced actin cytoskeleton alterations similar to those caused by other clostridial binary toxins. The lower level of production of binary toxin by CD196 than of iota toxin by C. perfringens was related to a lower transcript level, possibly due to a promoter region different from that of iota toxin genes. The cdtA and cdtB genes have been detected in 3 of 24 clinical isolates examined, and cdtB alone was found in 2 additional strains. One strain (in addition to CD196) was shown by Western blotting to produce CDTa and CDTb. These results indicate that some C. difficile strains synthesize a binary toxin that could be an additional virulence factor.

[1]  W. N. Burnette,et al.  "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. , 1981, Analytical biochemistry.

[2]  L. Simpson.  The binary toxin produced by Clostridium botulinum enters cells by receptor-mediated endocytosis to exert its pharmacologic effects. , 1989, The Journal of pharmacology and experimental therapeutics.

[3]  P. Boquet,et al.  Characterization of Clostridium perfringens Iota-toxin genes and expression in Escherichia coli , 1995, Infection and immunity.

[4]  C DeLisi,et al.  The detection and classification of membrane-spanning proteins. , 1985, Biochimica et biophysica acta.

[5]  M. Wilm,et al.  The Enterotoxin from Clostridium difficile (ToxA) Monoglucosylates the Rho Proteins(*) , 1995, The Journal of Biological Chemistry.

[6]  J. Vandekerckhove,et al.  Analysis of the catalytic site of the actin ADP‐ribosylating Clostridium perfringens iota toxin , 1996, FEBS letters.

[7]  R. Considine,et al.  Cellular and molecular actions of binary toxins possessing ADP-ribosyltransferase activity. , 1991, Toxicon : official journal of the International Society on Toxinology.

[8]  M. Popoff,et al.  Evidence that Arg‐295, Glu‐378, and Glu‐380 are active‐site residues of the ADP‐ribosyltransferase activity of iota toxin , 1996, FEBS letters.

[9]  P. Boquet,et al.  Transient expression of RhoA, -B, and -C GTPases in HeLa cells potentiates resistance to Clostridium difficile toxins A and B but not to Clostridium sordellii lethal toxin , 1995, Infection and immunity.

[10]  P. Boquet,et al.  Purification of the Clostridium spiroforme binary toxin and activity of the toxin on HEp-2 cells , 1989, Infection and immunity.

[11]  K. Aktories,et al.  Mechanisms of the cytopathic action of actin‐ADP‐ribosylating toxins , 1992, Molecular microbiology.

[12]  M. Mann,et al.  Glucosylation of Rho proteins by Clostridium difficile toxin B , 1995, Nature.

[13]  P. Boquet,et al.  Clostridium spiroforme toxin is a binary toxin which ADP-ribosylates cellular actin. , 1988, Biochemical and biophysical research communications.

[14]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[15]  E. Rubin,et al.  Actin-specific ADP-ribosyltransferase produced by a Clostridium difficile strain , 1988, Infection and immunity.

[16]  C. von Eichel-Streiber,et al.  Nucleotide sequence of Clostridium difficile toxin A. , 1990, Nucleic acids research.

[17]  A. Pantosti,et al.  Diarrhoea associated with toxigenic Clostridium spiroforme. , 1986, The Journal of infection.

[18]  L. A. Barroso,et al.  Nucleotide sequence of Clostridium difficile toxin B gene , 1990, Nucleic Acids Res..