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To differentiate pathogenic and nonpathogenic Edwardsiella tarda strains based on the detection of type III secretion system (T3SS) gene using polymerase chain reaction (PCR).Primers were designed to amplify Edw. tarda T3SS component gene esaV, catalase gene katB, haemolysin gene hlyA and 16S rRNA gene as an internal positive control. Genomic DNAs were extracted using a commercial isolation kit from 36 Edw. tarda strains consisting of 18 pathogenic and 18 nonpathogenic strains, and 50 ng of each DNA was used as the template for PCR amplification. PCR was performed with a thermocycler (TaKaRa TP600) in a 25-μl volume. Products of esaV were detected in all pathogenic strains, but not in nonpathogenic strains; katB was detected in all pathogenic strains and one of nonpathogenic strains; hlyA was not detected in any strains.The detection of esaV gene can be used for the assessment of pathogenic Edw. tarda strains.The strategy using T3SS gene as the virulence indicator provides a useful tool for the clinical assessment of pathogenic Edw. tarda strains and prediction of edwardsiellosis risk in fish culture environments.