A total of 74 strains were isolated from the intestines of seven freshwater fish and 10 water samples, and identified to the species level by the conventional method based on morphological, physiological and biochemical properties, and DNA-DNA hybridization method. Achromopeptidase treatment was found to be useful for the recovery of DNAs from Plesiomonas shigelloides cells because the amount of DNA obtained with this treatment was 3.1–7.2 times greater than with lysozyme. All the strains were identified as P. shigelloides on the basis of the mol% G + C contents of DNAs and homology values against the type strain of this microorganism, along with morphological, physiological, and biochemical properties. The P. shigelloides was detected in 29 out of 51 fish specimens and four out of 10 water samples. Black bass, Japanese eel, ayu, and tilapia harbored high densities of P. shigelloides, ranging from 104–108 CFU/g, with high frequencies of occurrence (73–100%). These results strongly suggested that the microplate hybridization method is an excellent procedure for the ecological study of P. shigelloides.

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