Isolation, identification and drug resistance analysis of Acinetobacter johnsonii from American bullfrog (Rana catesbeiana) tadpole

American bullfrogs (Rana catesbeiana) are a globally distributed invasive anuran species. They are extensively farmed in aquaculture facilities, particularly in countries like China, where they are highly valued as a protein-rich delicacy. In early March 2023, an outbreak of a serious infectious disease occurred in a bullfrog breeding farm in Yubei District, Chongqing, China. To elucidate the cause of the disease and find treatment options, this study isolated predominant bacteria from the liver and intestine tissues of diseased bullfrog tadpole. Morphological observation, physiological and biochemical experiments, 16S rDNA and rpoB sequencing, and phylogenetic analysis were employed to identify the bacteria. To ascertain the pathogenic mechanism of these dominant bacteria, artificial infection experiments, histopathological observations, virulence gene detection, growth characteristic studies, and drug sensitivity testing were conducted. The results showed that a predominant strain isolated from the liver and intestine of the diseased bullfrog tadpoles, LT202303, was identified as A. johnsonii. It harbored a trio of virulence genes, OmpA, Omp34, and OmpTsx, exhibited strong salt tolerance and the ability to survive within a wide pH range, and had a half-lethal concentration of 6.8×10⁶ CFU/mL. Clinical symptoms included liver redness and swelling with large white nodules, and a transparent yellowing of the intestine. Histopathological observations indicated significant inflammation and focal necrosis in bullfrog tadpole liver and intestine. Drug sensitivity testing revealed that the isolated strain LT202303 is a β-lactamase-producing multidrug-resistant bacterium. In summary, A. johnsonii has high pathogenicity to R. catesbeiana tadpoles, causing pathological damage to multiple organ tissues including the liver and intestine, ultimately resulting in diseases or even death of bullfrog tadpoles. This study reports, for the first time, an outbreak of R. catesbeiana tadpole infectious disease caused by A. johnsonii infection and provides theoretical references for the diagnosis and prevention of this disease.