Delineating the origins of the multidrug-resistant pathogens in ornamental fish farms by multilocus sequence typing and identification of a novel multidrug-resistant plasmid

To evaluate the overuse of antibiotics and to identify the origin of pathogens in the ornamental fish industry, we conducted a field investigation of three representative fish farms in Liaoning province, China. Drug-resistant pathogens in the fishponds and groundwater were isolated and subtyped by multilocus sequence typing (MLST). In total, 33 pathogenic strains, including Aeromonas veronii and five other pathogens, were isolated from diseased fish and from groundwater. MLST revealed that A. veronii obtained from diseased fish in three fish farms can be subtyped into four sequence types, which were also identified in the corresponding groundwater. All of the isolates obtained from diseased fish showed resistance to at least four antibiotics. Notably, Citrobacter freundii JY-17 exhibited resistance to the majority of the antibiotics and was a carrier of a megaplasmid with 15 drug resistance genes. PCR assays targeting beta-lactam, kanamycin, macrolide, phenicol, sulfonamide, and trimethoprim resistance genes in the pathogens from the diseased fish and groundwater were also conducted. The results revealed strong correlations between antibiotic treatment and increased antimicrobial resistance in fish pathogens. The results suggested that groundwater is the origin of the pathogens in ornamental fish. Antibiotic treatment of ornamental fish promoted the emergence of resistant pathogens.