Molecular Epidemiological Study Of Domestic Animal On Clostridium Perfringens In Shandong Province

According to the alpha toxin gene sequences of Clostridium welchii published on Gene Bank and publications, a pair of primes were designed to amplify a 325bp specific alpha toxin fragments, A kind of survey method for Clostridium welchii with PCR was established. It has good specification because it only can amplify specific fragments of Clostridium welchii, none of other strains such as Salmonella, E-coil, Enterococcus and Staphylococcus can be amplified. It possesses very high sensitivity because the system can detect 100 CFU/ml diluted template. This technique has been proved to be quick, simple and convenient detecting method . In addition, the detecting result to alpha toxin gene from the tissue of inoculated rat, diseased rabbit shows that the positive ratio of hepatic tissue is the highest, and the other such as spleen, kidney, heart, intestines take in order place.According to these toxin genes sequences published on Gene Bank and publications, four pairs of specific primes to alpha, beta, epsilon and jota toxin gene of Clostridium welchii were designed .The specific fragments were amplified by multiplex PCR. None of other strains such as Salmonella, E-coil, Enterococcus and Staphylococcus can be amplified in the same response system. The method showed good specification. The result of sensitivity experiment shows that it can detect 5×10~5 CFU/ml diluted template. It showed that Multi-PCR method is high sensitive for pathogenic diagnosis, and can be applied to clinical examination.A pair of primes were designed to amplify a 325 bp specific alpha toxin fragments, The PCR product was labeled with digoxigenin as DNA probe for detection of Clostridium welchii . The hybridization assay result showed the reference strain of the five serotypes Clostridium welchii was positive. No cross-hybridization was detected with DNA from other bacterial species. The sensitivity result showed that as few as 5×10~3 Clostridium welchii can be detected by DIG-labeled probe. The detection result for tissue sample showed that Clostridium welchii can be found in spleen, kidney, heart, intestines etc. The high sensitivity and specificity of the DIG-labeled probe will make it useful in field diagnostic work and...