一株感染深水网箱养殖许氏平鲉的 病原菌分离与鉴定

Isolation, Identification, and Biological Characteristics of a Pathogenic Bacterial Strain from Cage-Cultured Black Rockfish (Sebastes schlegelii)

中文关键词: 许氏平鲉  皮肤溃疡  轮虫弧菌  耐药性

英文关键词: Sebastes schlegelii  Skin ulcer  Vibrio rotiferianus  Drug resistance

中文摘要:

      2016年7月，山东省长岛县深水网箱养殖许氏平鲉(Sebastes schlegeli)暴发严重皮肤溃疡症。作者对病鱼进行病原菌分离。通过形态学观察、常规生理生化试验、gyrB和16S rDNA基因克隆测序等方法对分离菌株进行鉴定。在病灶溃疡处分离到一株绝对优势菌BZ01，该菌株在TSB固体培养基上呈半透明菌落，在TCBS选择性培养基上菌落呈绿色。透射电镜观察为短棒状，具有单根极生鞭毛。人工回接感染证明，该菌株对许氏平鲉具有较强的致病力，可以引起皮肤溃疡等症状，且与自然感染症状一致，其LD50为2.07×106 CFU/ml。通过gyrB和16S rDNA基因序列测定并构建系统发育树显示，菌株BZ01与弧菌属同源性最高，并在系统发育树中与轮虫弧菌(Vibrio rotiferianus)聚为一枝，结合形态及生理生化表型测定结果，将该菌株鉴定为轮虫弧菌(V. rotiferianus)。药敏试验结果显示，该菌株对四环素类、喹诺酮类、香豆素类、肽酰转移酶类高度敏感，而对大环内酯类、多肽类、磺胺类、β-内酰胺类、氨基糖苷类中度敏感或不敏感。

英文摘要:

      In July 2016, a severe disease outbreak occurred in offshore-cage cultured black rockfish (Sebastes schlegelii) in Changdao County, Shangdong Province, which resulted in significant mortality. The main symptom of diseased rockfish was skin ulcers on the body. In order to determine the cause of the disease, pathogenic bacteria were isolated from diseased fish and identified by various methods including morphological observation, traditional physiological and biochemical tests, and gyrB gene sequence alignment. A dominant bacterial strain was isolated from ulcerative skin and was marked as BZ01. The strain formed a subtransparent colony on TSB solid medium plate and a green colony on TCBS selective medium plate. Its shape was short and rod-like with a single polar flagellum when observed by transmission electron microscopy (TEM). An artificial infection experiment was carried out to verify its pathogenicity to black rockfish. The results demonstrated that this strain of BZ01 had strong pathogenicity to black rockfish and could cause death. The artificial infected black rockfishes showed the same symptoms as those of natural infection. The value of LD50 was calculated as 2.07106 CFU/ml. Analysis of the gyrB gene sequence suggested that the strain of BZ01 was most closely related to Vibrio. In gyrB gene sequencing, it clustered into Vibrio rotiferianus in the phylogenetic tree with 100% confidence. Based on these results, this strain was identified as V. rotiferianus. At the same time, the drug sensitivity test showed that this strain of BZ01 was highly sensitive to tetracyclines (tetracycline, doxycycline), quinolones (norfloxacin, ofloxacin, fleroxacin, nalidixic acid), coumarin (streptonivicin), and peptidyl transferase (florfenicol), while medium sensitive or resistant to macrolides, peptides, sulfonamides, β-lactams, and aminoglycosides. This is the first report that V. rotiferianus could infect offshore-cage cultured rockfish in China, which would provide an effective reference for disease control in cultured S. schlegelii.