DIAGNOSIS OF CLOSTRIDIUM-PERFRINGENS TYPE-C ENTERITIS IN PIGS USING ADNA AMPLIFICATION TECHNIQUE (PCR)

Clostridium perfringens type C, which produces alpha- and beta-toxin, causes severe haemorrhagic and necrotic enteritis in animals and human s. A polymerase-chain-reaction (PCR) assay was developed for the speci fic detection of the genes encoding alpha-, beta-, epsilon- and entero toxin of C. perfringens for rapid typing of C. perfringens strains, an d especially for the identification of type C strains. Both the alpha- and beta-toxin genes were detected directly in porcine C. perfringens type C cultures and also in type B and type C collection strains to a sensitivity of 10(3) cells without purification of the DNA. The alpha -toxin gene was detected in all types of C. perfringens. The epsilon-t oxin gene was found in type B and type D, and the enterotoxin gene in some type A strains. Nine other species of Clostridium and a variety o f intestinal pathogenic bacteria showed no signal for these toxin gene s in this PCR assay. The alpha- and beta-toxin genes PCR assay were us ed to identify C. perfringens strains isolated from intestinal content s of 36 necropsied piglets that had suddenly died or died after premon itory signs of diarrhoea. At necropsy, 20 piglets showed necrotizing e nteritis (15 acute and 5 chronic cases) and were suspected to have suf fered from a C. perfringens type C infection. All of them had C. perfr ingens which gave a positive PCR signal for alpha and beta-toxin genes , and, hence, were identified as type C strains. From the 16 other pig lets with lesions other than necrotizing enteritis, C. perfringens str ains with the alpha-toxin gene, but no beta-toxin gene, were isolated. The necropsy findings and the anamnesis showed a very good correlatio n with the PCR identification of toxin genes. It was therefore conclud ed that the PCR-based toxin gene examination is a good alternative to the time-consuming, less specific, and more expensive mouse neutraliza tion test in the routine diagnostic laboratory.