%0 Journal Article
%T Use of a Multiplex PCR for the Detection of Toxin-Encoding Genes netB and tpeL in Strains of Clostridium perfringens
%A Matthew A. Bailey
%A Kenneth S. Macklin
%A James T. Krehling
%J ISRN Veterinary Science
%D 2013
%R 10.1155/2013/865702
%X Some studies have shown that the NetB toxin may be an important virulence factor of Clostridium perfringens associated necrotic enteritis in poultry. Additionally, research has shown that strains of C. perfringens positive for both the netB gene and a second toxin-encoding gene, tpeL, appear to be more virulent than strains with only netB. In the past, detection of these genes has been performed relatively inefficiently using two single locus PCRs. This report describes a novel multiplex PCR developed to detect netB and tpeL simultaneously in C. perfringens strains isolated from cases of necrotic enteritis in broilers, providing a more efficient diagnostic tool in the screening of strains for these genes. 1. Introduction The disease necrotic enteritis (NE) is a major issue affecting the poultry industry, causing extensive economic loss through mortality, reduced bird performance, and carcass condemnation at slaughter [1, 2]. Caused by the bacterium Clostridium perfringens (CP), NE is characterized by necrotic lesions, primarily in the jejunum and ileum, which can vary in severity from thickened mucosa and multifocal ulceration in less severe cases to the formation of a greenish or yellowish pseudomembrane in the case of extensive mucosa inflammation and necrosis [3]. In the past, alpha-toxin produced by CP type A has been implicated as the primary virulence factor in NE pathogenesis [2, 4, 5]; however, the NetB toxin, encoded by the netB gene, was shown to be important for the virulence of certain strains [6, 7]. Another toxin, TpeL (encoded by the tpeL gene) [8], is a potential virulence factor as well. In a recent study, inoculation of broilers with strains positive for both tpeL and netB was associated with greater severity of gross lesions over strains with only netB [9]. Existing studies ascertaining the prevalence of netB and tpeL have been limited to certain geographic populations of CP, mainly in Australia, Belgium, Denmark, Sweden, and Canada [1, 7]. In the United States, studies on the prevalence of these genes have analyzed CP populations from New England, New York, and Pennsylvania [10]. The relatively small number of sampled CP populations underlines a need for more analysis to determine the importance of these genes on a worldwide scale. Additionally, detection of these genes has been performed relatively inefficiently using two single locus PCRs. This report describes a multiplex PCR developed to detect netB and tpeL simultaneously, providing a more efficient tool in screening CP strains for these genes. 2. Materials and Methods Strains
%U http://www.hindawi.com/journals/isrn.veterinary.science/2013/865702/