Skip to main content
Infection and Immunity logoLink to Infection and Immunity
. 1992 Nov;60(11):4633–4639. doi: 10.1128/iai.60.11.4633-4639.1992

Characterization of a toxin A-negative, toxin B-positive strain of Clostridium difficile.

D M Lyerly 1, L A Barroso 1, T D Wilkins 1, C Depitre 1, G Corthier 1
PMCID: PMC258212  PMID: 1398977

Abstract

This study was undertaken to examine toxin production by Clostridium difficile 8864, a naturally occurring isolate that has been reported to produce toxin B in the absence of toxin A. To date, this is the only strain of C. difficile reported to produce only one of the toxins. The results of our initial studies with antibodies against toxins A and B confirmed these observations. Toxin B from strain 8864 and from VPI strain 10463, a strain that produces high levels of both toxin A and toxin B, was purified to homogeneity by sequential anion-exchange chromatography on DEAE-Sepharose CL-6B, gel filtration on Ultrogel AcA22, and immunoadsorption chromatography, and their toxic activities were compared. Our results showed that toxin B from strain 8864 and toxin B from C. difficile VPI strain 10463 were comparable in their cytotoxic activities and that the 8864 toxin B was more lethal. In addition, we observed that toxin B from strain 8864 was weakly enterotoxic, which may explain the ability of this strain to cause intestinal disease in hamsters treated with antibiotics. Analysis with specific antibodies showed that the toxin B molecules from these strains were highly related but contained distinct epitopes. The results of hybridization studies with probes specific for the toxin B gene of VPI strain 10463 demonstrated differences between the toxin B genes of the two strains. In addition, probes specific for the toxin A gene of VPI strain 10463 showed that strain 8864 contains a region which shows identity with the 5' end of the toxin A gene but not the region of the gene which encodes a hydrophobic region and the repeating units.

Full text

PDF
4633

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Barroso L. A., Wang S. Z., Phelps C. J., Johnson J. L., Wilkins T. D. Nucleotide sequence of Clostridium difficile toxin B gene. Nucleic Acids Res. 1990 Jul 11;18(13):4004–4004. doi: 10.1093/nar/18.13.4004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Dove C. H., Wang S. Z., Price S. B., Phelps C. J., Lyerly D. M., Wilkins T. D., Johnson J. L. Molecular characterization of the Clostridium difficile toxin A gene. Infect Immun. 1990 Feb;58(2):480–488. doi: 10.1128/iai.58.2.480-488.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Dower W. J., Miller J. F., Ragsdale C. W. High efficiency transformation of E. coli by high voltage electroporation. Nucleic Acids Res. 1988 Jul 11;16(13):6127–6145. doi: 10.1093/nar/16.13.6127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Ehrich M., Van Tassell R. L., Libby J. M., Wilkins T. D. Production of Clostridium difficile antitoxin. Infect Immun. 1980 Jun;28(3):1041–1043. doi: 10.1128/iai.28.3.1041-1043.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  6. Laughon B. E., Viscidi R. P., Gdovin S. L., Yolken R. H., Bartlett J. G. Enzyme immunoassays for detection of Clostridium difficile toxins A and B in fecal specimens. J Infect Dis. 1984 May;149(5):781–788. doi: 10.1093/infdis/149.5.781. [DOI] [PubMed] [Google Scholar]
  7. Lima A. A., Lyerly D. M., Wilkins T. D., Innes D. J., Guerrant R. L. Effects of Clostridium difficile toxins A and B in rabbit small and large intestine in vivo and on cultured cells in vitro. Infect Immun. 1988 Mar;56(3):582–588. doi: 10.1128/iai.56.3.582-588.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Lyerly D. M., Krivan H. C., Wilkins T. D. Clostridium difficile: its disease and toxins. Clin Microbiol Rev. 1988 Jan;1(1):1–18. doi: 10.1128/cmr.1.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Lyerly D. M., Lockwood D. E., Richardson S. H., Wilkins T. D. Biological activities of toxins A and B of Clostridium difficile. Infect Immun. 1982 Mar;35(3):1147–1150. doi: 10.1128/iai.35.3.1147-1150.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Lyerly D. M., Phelps C. J., Toth J., Wilkins T. D. Characterization of toxins A and B of Clostridium difficile with monoclonal antibodies. Infect Immun. 1986 Oct;54(1):70–76. doi: 10.1128/iai.54.1.70-76.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Lyerly D. M., Phelps C. J., Wilkins T. D. Monoclonal and specific polyclonal antibodies for immunoassay of Clostridium difficile toxin A. J Clin Microbiol. 1985 Jan;21(1):12–14. doi: 10.1128/jcm.21.1.12-14.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lyerly D. M., Sullivan N. M., Wilkins T. D. Enzyme-linked immunosorbent assay for Clostridium difficile toxin A. J Clin Microbiol. 1983 Jan;17(1):72–78. doi: 10.1128/jcm.17.1.72-78.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Martinez R. D., Wilkins T. D. Purification and characterization of Clostridium sordellii hemorrhagic toxin and cross-reactivity with Clostridium difficile toxin A (enterotoxin). Infect Immun. 1988 May;56(5):1215–1221. doi: 10.1128/iai.56.5.1215-1221.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Matsudaira P. Sequence from picomole quantities of proteins electroblotted onto polyvinylidene difluoride membranes. J Biol Chem. 1987 Jul 25;262(21):10035–10038. [PubMed] [Google Scholar]
  15. Phelps C. J., Lyerly D. L., Johnson J. L., Wilkins T. D. Construction and expression of the complete Clostridium difficile toxin A gene in Escherichia coli. Infect Immun. 1991 Jan;59(1):150–153. doi: 10.1128/iai.59.1.150-153.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Popoff M. R. Purification and characterization of Clostridium sordellii lethal toxin and cross-reactivity with Clostridium difficile cytotoxin. Infect Immun. 1987 Jan;55(1):35–43. doi: 10.1128/iai.55.1.35-43.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Saghai-Maroof M. A., Soliman K. M., Jorgensen R. A., Allard R. W. Ribosomal DNA spacer-length polymorphisms in barley: mendelian inheritance, chromosomal location, and population dynamics. Proc Natl Acad Sci U S A. 1984 Dec;81(24):8014–8018. doi: 10.1073/pnas.81.24.8014. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  19. Sullivan N. M., Pellett S., Wilkins T. D. Purification and characterization of toxins A and B of Clostridium difficile. Infect Immun. 1982 Mar;35(3):1032–1040. doi: 10.1128/iai.35.3.1032-1040.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Infection and Immunity are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES