大鲵脾脏结节症病原菌的分离鉴定及药敏试验

Isolation, identification and drug sensitivity test of the pathogenic bacteria from the Chinese giant salamander with splenic tuberosity

为探究重庆地区中国大鲵(Andrias davidianus)脾脏结节症的病因及防控措施,本研究对发病大鲵进行了临床剖检及病理组织学观察,并采用微生物学方法从其肝脏、脾脏中分离病原菌,通过人工感染试验确定分离菌株的致病性,并对菌株的基本形态、理化特性、分子特征及药物敏感性等进行了系统研究。结果显示,患病大鲵脾脏有大量白色结节,质脆,病理切片发现脾组织有出血及丝网状纤维素分布,淋巴细胞部分消失;肝细胞肿胀、变性。从肝脏中分离到1 株病原菌,命名为KNL-1。人工感染试验发现,该菌对试验鲫鱼和大鲵均有较强的致病力,试验动物死亡率高达100%,大鲵发病症状与临床自然发病症状一致,确认为该病的致病菌。综合分离菌的16SrDNA 序列测定、培养特性、生化试验及BIOFOSUN-GN 系统鉴定结果,确定KNL-1 为杀鲑气单胞菌杀日本鲑亚种(Aeromonas salmonicida ssp. masoucida)。药敏试验结果显示,该菌对头孢氨苄、卡那霉素、头孢他啶、头孢曲松、先锋霉素、新霉素、庆大霉素、左氧氟沙星、米诺环素、丁胺卡那霉素和氟苯尼考高度敏感;对氨苄西林、麦迪霉素和羧苄西林耐药。

The aim of this study was to confirm the causation of the splenic tuberosity of the Chinese giant salamander. Samples including liver and spleen were taken and conducted with histopathological and bacteriological examinations. The isolates were identified by morphological, biochemical and molecular characteristics. Pathogenicity tests were performed on the healthy crucians and Chinese giant salamanders. Antibiotic susceptibility test was conducted with 16 antibiotics by the Kirby-Bauer disc diffusion method. Necropsy results showed that abundant white tuberosities were observed in the spleen of the animals. Histological analysis indicated that hemorrhage, reticular fiber and vanish of partial lymphocytes were in the spleen,while swelling and degeneration were found in the hepatocyte. The isolate (named KNL1) was presented with highly virulence to the crucians and Chinese giant salamanders,suggesting that it is the key factor for the disease. According to the morphological, biochemical and molecular identification results, KNL1 was phylogenetically related to Aeromonas salmonicida ssp. masoucida. Drug sensitivity test indicated that KNL1 was sensitive to cephalexin, kanamycin, ceftazidime, ceftazidime, cephalosporin, neomycin, gentamicin, levofloxacin, minocycline, amikacin sulphate and florfenicol, while it was resistant to ampicillin, midecamycin and carbenicillin.