| In May 2021, serious mortality of cultured Pelodiscus sinensis (Chinese softshell turtle) occurred in a recirculating aquaculture farm in Huzhou, Zhejiang Province and the infected turtles presented symptoms of perforations and ulcers. To identify the pathogen infecting the turtles and find effective inspection and prevention strategies for the disease, the dominant pathogenic strain was isolated from the liver, spleen, lung and kidney of diseased P. sinensis and labeled the AH0421 strain. Bacterial identification of the AH0421 strain was accomplished through gram staining, 16S rRNA sequencing and determination of the virulence factor, as well as morphological, physiological and biochemical characteristics. The AH0421 strain is rod-shaped and gram-negative, and phylogenetic tree analysis, based on 16S rRNA gene sequences, shows that the AH0421 strain clusters with a strain of Aeromonas hydrophila ATCC 7966 with homology of 99.66%. The AH0421 strain can utilize glucose, sucrose, and maltose and has O/129 tolerance. According to PCR amplification, the AH0421 strain possesses 11 virulence genes, including act, aerA, aha, ahh, ahp, ahpA, alt, ast, hlyA, lip, and ompA. Simultaneously, an artificial infection study was carried out on healthy P. sinensis by injecting different concentrations of a bacterial suspension of the AH0421 strain identified as A. hydrophila. P. sinensis (body mass, ~200 g) were randomly assigned to four treatment groups (6.8×107, 6.8×106, 6.8×105 and 6.8×104 CFU/mL) and a control group, with 10 P. sinensis in each group. The symptoms of the artificially infected turtles were identical to those of naturally infected turtles, and the concentration of strain AH0421 that was lethal for half of the test turtles (LC50) was 1.7×106 CFU/mL. Drug susceptibility of the AH0421 strain was tested by the paper extension and agar punch methods using antibiotics and Chinese herbal medicines. The results show that the A. hydrophila strain is highly sensitive to the antibiotics norfloxacin, doxycycline, enrofloxacin, neomycin, cephalosporin, cefixime and ceftriaxone, and to the extract of Schisandra chinensis. In conclusion, A. hydrophila (AH0421) was the pathogen responsible for the mortality observed in the P. sinensis recirculating aquaculture operation and cephalosporin antibiotics can be considered as therapeutic agents for the treatment of P. sinensis infected with A. hydrophila. |
