Clostridium Perfringens represent one of the natural inhabitant bacteria in the gastrointestinal tract of chickens. However, predisposing factors, mainly co-infection with Coccidia, represent a favorable condition for over growth of C. perfringens that induce necrotic enteritis (NE). The prevalence of C. perfringens in commercial broilers with enteric disease in Egypt was investigated via bacteriological isolation and identification and then confirmed by toxin typing by PCR. Furthermore, culture-independent quantitative real time PCR targeting alpha toxin gene was improved via reducing the effect of PCR inhibitors in the intestinal content (IC). Out of 61 broiler flocks, 40 were positive for C. perfringens with a higher incidence in winter (81.25%) and in 20-30 days old broilers (66.67%). All isolates belonged to C. perfringens type A with the absence of enterotoxin or Beta2 toxin genes. Pretreatment of the intestinal content with acetone improve the detection limit of quantitative real time PCR assay to 102 cells/gm which was highly correlated to the culture-based counting method using cooked meet media. C. perfringes type A widespread in broilers in Egypt and the extraction method used herein could be used for direct quantification of C. perfringens in the intestinal content of broiler chickens without prior enrichment.
Key words: Clostridium perfringens, Broilers, intestinal inhibitors, Necrotic Enteritis, Quantitative Real-Time PCR.
|
