Identification and characterization of non-O1/O139 Vibrio cholerae isolated from farmed tadpoles of the black-spotted frog (Pelophylax nigromaculatus).

Vibrio cholerae is traditionally recognised as the causative agent of cholera in humans; however, increasing evidence indicates that non-O1/non-O139 strains are widely distributed in aquatic environments and can infect aquatic animals. In this study, we report the first isolation and characterisation of V. cholerae from diseased Pelophylax nigromaculatus tadpoles exhibiting severe intestinal symptoms during an outbreak in a freshwater farm in Huanggang, China. The isolates were identified through 16S rRNA and gyrB gene sequencing, and further confirmed by species-specific genes (lolB and dnaJ) and phenotypic traits, including sucrose fermentation and sodium-independent growth. Whole-genome analysis revealed multiple virulence-associated genes, such as AHA_3493 (haemolysin III), wbkC (LPS biosynthesis), ricA (intracellular survival), and lbtC (siderophore-mediated iron uptake), as well as antibiotic resistance genes including crp, qnrVC5, and two chromosomal copies of catB9. These findings highlight the pathogenic potential of non-O1/non-O139 V. cholerae in amphibians and its adaptation to freshwater environments. This study expands the known host range of V. cholerae and highlights the need for early surveillance in amphibian aquaculture.