Abstract：To study the isolation rate and toxin type of Clostridium perfringens in freshwarter fishes. METHODS: Four hundred and twenty fresh intestinal content samples (not including intestinal tissues) of freshwater fishes caught from one water reservoir were examined bacteriologically for the occurrence of C. perfringens. Isolates were examined by polymerase chain reaction (PCR) for genes encoding the four lethal toxins (α, β, ε and ι) for classification into toxin types and for genes encoding enterotoxin and the novel β2 toxin for further subclassification. RESULTS: C. perfringens could be isolated in 75 intestinal contents samples (17.9%) from freshwater fish, 58 strains (77.3%) were C. perfringens toxin type C (α and β toxin positive), 13 strains (17.3%) were toxin type A (α toxin positive) and 4 strains (5.3%) were toxin type B (α, β and ε toxin positive). In addition, the gene encoding for β2 toxin was found in all the isolates. These amplified toxin gene fragment were cloned and sequenced and compared with standard strains, the identity varied from 98.15% to 99.29%. CONCLUSION: This is the first report of C. perfringens α, β, ε, β2 toxins in freshwater fish and of β, ε toxins in fish in general, and is the first discovery that the β2 toxin could be detected in strains of type B. The origin of this bacterium and its importance to human food poisoning in freshwater fish is discussed.
