ABSTRACT
Clostridium perfringens causes necrotic enteritis in piglets, characterized by diarrhea, weight loss and death. The purpose of this study was to genotype C. perfringens strains isolated from diarrheic piglets by multiplex PCR, using specific primers for genes of alpha (cpa), beta (cpb), beta-2 (cpb2), epsilon (etx), enterotoxin (cpe) and iota (ia) toxins. Sixty-five samples from diarrheic piglets ranging from 7 to 36 days of age were studied. The material was seeded into egg-yolk agar with cicloserine. Colonies with suspicion of C. perfringens were subjected to Gram staining and biochemical identification. After certification for purity, bacteria were subcultivated in blood agar and one single colony was used as a DNA template for PCR amplification after direct bacterial lysis. No DNA purification procedure was used. Anaerobic Gram-positive bacilli were isolated from 59 out of 65 fecal samples tested. Among these, 27 were identified as C. perfringens and typified by multiplex PCR. Twenty-one (77.8%) of these isolates were type A, of which 5 had thecpb-2 gene. Five (18.5%) were type C and 4 of them contained the cpb-2 gene. One (3.7%) was type D, while no strains were positive for ia or cpe genes. This study demonstrated that Clostridium perfringens A is the most common type in feces of diarrheic piglets.
KEY WORDS
Clostridium perfringens
; necrotic enteritis; genotyping; swine; multiplex PCR