Enhanced virulence of Acinetobacter johnsonii at low temperatures induces acute immune response and systemic infection in American bullfrogs (Aquarana catesbeiana)

Acinetobacter johnsonii is a denitrifying bacterium commonly used as an environmental probiotic in wastewater treatment. However, research on its potential pathogenicity to animals is limited. During an epidemiological survey conducted from 2022 to 2024 at bullfrog farms in Guangdong Province, China, multiple strains were isolated from diseased bullfrogs during the low-temperature season. Three representative strains were selected for virulence testing, which showed high virulence to bullfrogs. Molecular identification confirmed these strains as A. johnsonii. One strain was named NW220314S and chosen for further study. Artificial infection through different routes revealed that the mortality rate of bullfrogs infected with this strain was higher at 15°C compared to 28°C. Additionally, the expression of at least 11 virulence genes was significantly higher at 15°C. Pathological examinations of bullfrogs infected with A. johnsonii showed systemic infection with extensive infiltration of inflammatory factors in organs, muscles, and skin. Immune-related gene expression analysis indicated a rapid and intense inflammatory response in bullfrogs post-infection. Our findings uncovered the novel phenomenon of the pathogenicity of A. johnsonii in bullfrogs under low-temperature conditions, warning of the potential threat of A. johnsonii to amphibian populations and the risks associated with its use in various environmental applications.