A total of 1347 bacteria isolated from environments and intestinal tracts of Japanese flounder, Paralichthys olivaceus, along with feeds, were examined for the antibacterial activity against four fish pathogens. As a result, 142, 22, 20 and 17 isolates inhibited the growth of P. piscicida K-III, V. vulnificus RIMD 2219009, E. seriolicida ATCC 49156 and V. anguillarum ATCC 19264, respectively, and 8.3-30.9% of isolates belonging to Aeromonas, Bacillus, coryneforms, Pseudomonas, Streptococcus and Vibrio showed activities against one of the four target bacteria. Antibacterial producers were detected at low densities and occurrence in rearing water, bottom sands and feeds, while those accounted for 59% of vibrios isolated from fish intestines. This result strongly suggests that antibacterial producers could not control fish pathogen efficiently in aquaria of Japanese flounder.