Identification And Genotyping Of Clostridium Perfringens Isolates And Enterotoxigenic Clostridium Perfringens Isolates With Multiplex PCR

Clostridium perfringens is a gram-positive, endospore-forming anaerobe that has long been recognized as a significant cause of both histoxic and gastrointestinal (GI) diseases in human and domestic animals. This bacterium was wildly distributed in soil, water resource, human and animal GI tract. C. perfringens isolates are commonly classified in five types (type A to E), depending upon their abilitis to express four lethal toxins (alpha, beta, epsilon, and iota toxin). The classical typing method is based on mouse lethality test and sero-protection with neutralizing antibodies raised against crude culture supematants of each representative C. perfringens toxin-type. The serum neutralizing test for typing of C. perfringens are time-consuming and complex-handle. In previous study, a series of multiplex PCRs were developed by Qiyi Wen et al to genotype, sub-genotype different types of C. perfringens and to locate cpe genesih CPE positive type A C. perfringens. In this study, we tested these multiplex PCRs, and the results showed that these PCRs could be efficiently used to genotype, sub-genotype different types of C. perfringens and to locate cpe genes in CPE positive type A C. perfringens.In this study, 1311 fecal samples were collected from people, pigs, goats and cattle, and C. perfringens were isolated from these samples. These isolates were identified by biochemical test and the multiplex PCR reaction. Application of conventional culture method combined with the developed genotyping assay to investigate C. perfringens from above fecal samples, and 5.11 C. perfringens isolates were isolated from those fecal samples and all C. perfringens isolates were identified by biochemical test andgenotyped by the multiplex PCR assay. A total of C. perfringens isolates were genotyped with this assay, shown that all of C. perfringens isolates were found to be type A C. perfringens while 24 isolates belonged to type C C. perfringens. During this study, a cpe positive type A C. perfringens isolate was found, and the isolate harbored the chromosomal cpe after the latter assay..