Histological and molecular characterization of Edwardsiella tarda infection in Siamese crocodile (Crocodylus siamensis) hatchlings

Abstract Edwardsiellosis is an important bacterial disease of fish, birds, reptiles and mammals. This study was conducted to describe the histopathology and etiology of the disease caused by Edwardsiella tarda in farmed hatchling Siamese crocodiles (Crocodylus siamensis) in Hainan, China. The main macroscopic lesions were congestion and hemorrhage of various internal organs, pericardial fluid and subcutaneous edema. Main histopathological lesions included necrotic hepatitis, hemorrhagic necrotizing nephritis, fibrinous pneumonia, and necrotic splenitis. E. tarda isolates were isolated from blood and various internal organs. Genotyping showed that isolates were positive for eight of ten investigated virulence genes (80%) and eleven of twenty antibiotic-resistance genes (55%), which were largely correlated phenotypically, except for erythromycin resistance. Moreover, tetracycline resistance genes were detected after two years of oxytetracycline use on the farm, but only tetA-harboring strains yielded high resistance. All isolates formed biofilm that increased under low concentration tetracycline stress. Zebrafish experimental infections revealed that all isolates were virulent, and that the most virulent isolate formed the highest amount of biofilm, rather than harboring the highest number of virulence and resistance genes. This study is the first to demonstrate the histopathology of edwardsiellosis among hatchling Siamese crocodiles, and characterize virulence factors and tetracycline resistance in E. tarda isolates from a farm that employed long-term prophylactic use of oxytetracycline. Biofilm was suggested as a key virulent factor in the pathogenesis of E. tarda.