Isolation and identification of the pathogen in a case of Takifugu rubripes coinfected with parasites and bacteria

In August 2023, a significant outbreak occurred at a puffer fish farm in Dandong, Liaoning, China, affecting a large number of Takifugu rubripes . The affected fish displayed symptoms including fin ulcers and white spots, which were confirmed through clinical testing conducted at Dalian Ocean University. Microscopic examinations revealed a substantial presence of black dot-like unicellular parasites on the gills of the diseased fish. Further analyses, including methyl green-pyronine staining, optical microscopy, and biological scanning electron microscopy, facilitated the identification of the parasite as Amyloodinium ocellatum . Additionally, bacterial isolation and culture, Gram staining, physiological and biochemical testing, 16S rDNA sequence analysis, phylogenetic tree, and drug sensitivity experiments were performed on the organs of the affected fish. The results identified two dominant bacterial strains, Vibrio scophthalmi and Vibrio harveyi , present in the spleen and kidney of the infected fish. Antibiotic sensitivity tests indicated that V. scophthalmi is susceptible to antibiotics such as ciprofloxacin, florfenicol, and enrofloxacin, while V. harveyi showed sensitivity to ciprofloxacin, florfenicol, and ofloxacin. In conclusion, this case was determined to be a combined infection involving both parasites and bacteria. The findings of this study offer valuable insights for the prevention and control of epidemics affecting Takifugu rubripes . • Parasites and bacteria co-infected Takifugu rubripes. • Amyloodinium ocellatum was identified and analyzed. • Vibrio harveyi isolate had higher drug resistance.