The virulence of the representative isolates VP66 (trh+, ureC+, T3SS1+, T3SS2β+, T6SS2+) and VP80 (T3SS1+, T6SS1+, T6SS2+) were further assessed in zebrafish and mouse infection model in vivo, and the tested isolates were shown to be lethal to both zebrafish and mice. These results suggest that crayfish may serve as a carrier of V. parahaemolyticus in freshwater, and that some isolates may have the potential to cause foodborne disease in humans.