Analysis of pathogenicity and drug resistance of ST520 Aeromonas veronii from Lithobates catesbeiana producing MBLs

Lithobates catesbeiana (American bullfrog) is extensively cultivated in China, valued by consumers for its tender texture, palatable taste, and abundant nutritional components.This study aimed to identify the etiological agent causing skin hemorrhage and ascites in diseased L. catesbeiana collected from a bullfrog farm in Jingzhou, Hubei Province. A dominant bacterium was isolated from the intestinal tissue of diseased L. catesbeiana. Its pathogenicity and drug resistance were analyzed through morphological observation, physiological and biochemical identification, 16S rRNA gene sequencing, combined with MLST analysis, virulence gene detection, pathogenicity experiments, drug resistance gene detection, and drug susceptibility testing. The isolated strain was a Gram-negative bacterium, and 16S rRNA gene sequence alignment revealed highest similarity with A. veronii (MK584925.1), confirming it as A. veronii isolated from L. catesbeiana and designated W0525. MLST analysis determined that the isolated strain was of the ST520 type. The strain carried five virulence genes: aer, act, alt, ahyB, and exu, and exhibited β-hemolytic activity. In the artificial infection experiments, the deceased L. catesbeiana showed clinical signs similar to natural infection, including skin redness and swelling, abdominal ascites, and gastrointestinal inflammatory lesions. The median lethal dose (LD₅₀) of the isolate to L. catesbeiana was 5.62×10⁸ CFU/mL. The isolate carried five resistance genes: bla-TEM, bla-CphA, aadA1, aadA2, and tetA. It demonstrated strong biofilm-forming ability and was resistant to 16 antibiotics, including penicillin and amoxicillin, but highly sensitive to the traditional Chinese medicine Caesalpinia sappan (Sappan wood). Strain W0525 was identified as a metallo-β-lactamase (MBL)-producing multidrug-resistant strain, and its bla-CphA gene clustered with bla-CphA4 in phylogenetic analysis. This study successfully isolated and characterized an MDR A. veronii strain from diseased L. catesbeiana, providing critical insights into its pathogenic mechanisms and antimicrobial resistance profile. These findings establish a theoretical foundation and empirical data for the development of targeted prevention and control strategies against A. veronii infections in bullfrog aquaculture.