How to cite this article: Vet. Res. (2010) 41:21
DOI: 10.1051/vetres/2009069

Association between avian necrotic enteritis and Clostridium perfringens strains expressing NetB toxin

Anthony L. Keyburn^{1, 2, 3}, Xu-Xia Yan², Trudi L. Bannam², Filip Van Immerseel⁴, Julian I. Rood^{2, 3} and Robert J. Moore<sup>1, 2, 3

1</sup>  CSIRO Livestock Industries, Australian Animal Health Laboratory, Geelong, Victoria 3220, Australia
²  Australian Research Council Centre of Excellence in Structural and Functional Microbial Genomics, Department of Microbiology, Monash University, Clayton, Victoria 3800, Australia
³  Australian Poultry Cooperative Research Centre, Armidale, New South Wales 2315, Australia
⁴  Department of Pathology, Bacteriology and Avian Diseases, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, B-9890 Merelbeke, Belgium

Received 15 September 2009; accepted 20 November 2009; published online 25 November 2009

Abstract - A novel toxin, NetB, has recently been identified in virulent avian Clostridium perfringens isolates and shown to be an essential virulence factor in a clinical necrotic enteritis isolate. To assess whether NetB is more generally associated with avian necrotic enteritis isolates we have screened a range of C. perfringens strains from geographically diverse locations for both the presence and expression of the netB gene. Forty-four isolates were derived from necrotic enteritis disease cases from Australia, Belgium, Denmark and Canada and 55 isolates from healthy chickens from Australia and Belgium. The majority of strains isolated from necrotic enteritis-affected birds were netB positive (70%) and there was an absolute correlation between the presence of netB and in vitro expression of the NetB protein. Only two of the C. perfringens isolates from healthy chickens carried netB. Sequencing of the netB gene from 23 positive isolates showed that NetB is highly conserved, with only one predicted amino acid (A168T) difference, in six isolates, compared to the published sequence. This change did not alter the in vitro activity of the NetB toxin. The gene encoding the recently discovered TpeL toxin was also screened using PCR and only found in a small proportion of NetB-positive isolates from diseased birds. A selection of NetB-negative isolates, originating from diseased birds, was unable to cause disease in a necrotic enteritis induction model. This study provides further evidence that NetB is important in pathogenesis and advances our current understanding of C. perfringens virulence factors in avian necrotic enteritis.

Key words: Clostridium perfringens / necrotic enteritis / NetB toxin / virulence factor / TpeL

Corresponding author: rob.moore@csiro.au

© INRA, EDP Sciences 2009