Toxinotyping of Clostridium perfringens in Poultry from Necrotic Enteritis Cases

Clostridium perfringens is well known causative agent of necrotic enteritis in poultry and is mainly caused by Type A toxin. NetB toxin is found to be one of the newly emerging virulent toxin gene which is also responsible for necrotic enteritis. The present study was carried out to characterize and to type the different toxins associated with C. perfringens in NE cases of poultry. For the present study total 125 samples were collected from poultry birds, out of which 50 samples were of intestinal content from diseased birds, 50 cloacal swabs and 25 intestinal content from healthy birds. These samples were further processed for isolation, identification, and toxinotyping of Clostridium perfringens isolates. Onisolation of C. perfringens on blood agar total 43 isolates were found positive showing a pattern of double hemolysis on blood agar. The positive isolates of C. perfringens were further confirmed by using 16S rRNA species specific PCR. After confirmation isolates were processed for toxinotyping mainly targeting cpa, cpb and cpb2 toxins by using multiplex PCR. On toxinotyping it was found that NE in poultry birds were mainly caused by C. perfringens type A. On virulent gene detection of netB toxin, total 4 isolates were found positive for netB toxin. This study pointed out that C. perfringens type A is responsible for development of NE in poultry along with net B toxin which is a new key virulent factor. Further studies of netB toxoid and C. perfringens type A for vaccine production could minimize the clostridial problems in broiler farms.