A toxic approach to beta2-toxigenic Clostridium perfringens

Abstract

Clostridium perfringens is one of the most important causes of intestinal disease in animals and humans. Its virulence is attributed to the several toxins it can produce, including the beta2 toxin encoded by cpb2. In this thesis we studied the role of the beta2 toxin produced by C. perfringens in the development of intestinal disease. Results described in this thesis demonstrate that a possible role for beta2 toxin in the development of intestinal disease is still disputable. The development of a new primer set which recognizes both alleles of cpb2 and the development of a new test strategy for application of the newly developed PCR on fecal samples improved test sensitivity for the presence of cpb2. However, a relationship between cpb2-harbouring C. perfringens, beta2 toxin,and intestinal disease could not be proven in chickens and roe deer. Although cpb2-positive C. perfringens was abundantly present in the intestinal tract of roe deer, only two isolated strains were able to produce the beta2 toxin in vitro and the beta2 toxin was not demonstrated in the intestinal tract by immunohistochemistry in any of the animals. C. perfringens was isolated from both healthy and diseased laying hens with a newly described form of subclinical necrotic enteritis. No relationship could be determined between intestinal disease and the presence of beta2 toxin producing C. perfringens. However, focal necrotic lesions were only found in the intestinal tract of laying hens with a high watery intestinal content which may have played a predisposing factor in the development of disease providing substrate for beta2 toxin production by C. perfringens. A significant role for the beta2 toxin in the development of intestinal disease could not be established in an in vitro model in which the cytotoxic effect of supernatant of cpb2-positive C. perfringens cultures with and without beta2 toxin were compared. A proven role for the beta2 toxin in the development of intestinal disease or reproduction of the disease by other infection models has not been achieved yet. Therefore the role of the beta2 toxin in the development of intestinal disease remains at least questionable. Other virulence factors, the intestinal microbiota, and environmental circumstances might influence the significance of the intestinal presence of cpb2-harbouring C. perfringens in the development of intestinal disease. This hypothesis was supported by the finding of a reducing effect of Lactobacillus fermentum on beta2 toxin production by cpb2-harbouring C. perfringens. A lower transcription of cpb2 and a lower production of beta2 toxin were demonstrated with the presence of L. fermentum while the viability of C. perfringens was not affected. In this way, environmental influences might increase the role of cpb2-harbouring C. perfringens in the development of intestinal disease too. Articles on predisposing factors for the development of C. perfringens-associated intestinal disease in animals and human beings were reviewed and discussed. In vivo studies investigating both beta2 toxin and predisposing factors are necessary to definitely determine or exclude a potential role for beta2 toxin in the development of intestinal disease.