香鱼(Plecoglossus altivelis)致病性类志贺邻单胞菌(Plesiomonas shigelloides)的分离与生化药敏鉴定*

香鱼(Plecoglossus altivelis)是名贵洄游性鱼类,由于高密度养殖等因素导致病害频发。类志贺邻单胞菌广泛存在于养殖水环境中,给香鱼养殖业造成严重经济损失。对患体表溃疡病的香鱼进行病原分离和鉴定,从患病鱼体表和肠道组织中分离到一株优势菌(NH21),其菌落低突,边缘光滑,表面湿润。革兰氏染色结果显示NH21为革兰氏阴性菌,菌形态为短杆状,端圆。经生化特性和16S rDNA序列分析鉴定为类志贺邻单胞菌(Plesiomonas shigelloides)。回归感染后,香鱼呈现与发病鱼相似的症状,皮肤、肌肉溃烂并伴有周围皮肤充血,组织切片可观察到病鱼肌纤维断裂,周围红细胞、炎性细胞浸润；鳃丝和鳃小片中红细胞增多；肝细胞核呈空泡,部分细胞核染色质边集；脾脏内包细胞增多,红细胞呈梭形；肠黏膜上皮细胞脱落。经计算其半致死量LD50为1.99×10⁷ CFU/mL。25种抗生素的药敏试验显示NH21菌株对阿奇霉素、头孢氨苄、新生霉素、多粘菌素B、头孢西丁、呋喃唑酮6种抗生素敏感。检出NH21菌株毒力基因9个,耐药基因8个。综合分析表明,分离菌株NH21为类志贺邻单胞菌属,是本次香鱼体表溃烂病病原菌,研究结果为香鱼出现类似病症的预防和治疗提供了基础资料。

Isolation, Biochemical Identification, and Antibiotic Sensitivity of the Pathogenetic Plesiomonas shigelloides isolated from Plecoglossus Altivelis

In this study, the pathogens were isolated and identified from sweetfish (Plecoglossus altivelis) with superficial ulcer disease. A dominant strain, NH21, was isolated from the body surface and intestinal tissues of the diseased fish. The colonies were low-protruding, with smooth edges and moist surfaces. Gram staining showed that NH21 was a Gram-negative bacterium with a short rod-like morphology and rounded ends. Biochemical characterization and 16S rDNA identification indicated that NH21 was Plesiomonas shigelloides. The pathogenicity of NH21 was investigated by recursive infection. The symptoms of P. altivelis artificially infected with strain NH21 were similar to naturally infected fish. The skin and muscles were ulcerated and accompanied by hyperemia of the surrounding skin. In tissue sections, the muscle fibers of the diseased fish were broken, and the surrounding red blood cells and inflammatory cells were infiltrated; red blood cells in the gill filaments and gill small pieces were observed increased; the nuclei of hepatocytes were vacuolated, and some nuclei of chromatin were bordered; the spleen enclosing cells were increased, and the red blood cells were spindle-shaped; the intestinal mucosal epithelial cells were shed. The LD50 was calculated to be 1.99×10⁷ CFU/mL. Drug susceptibility test showed that NH21 was sensitive to 6 antibiotics (azithromycin, cephalexin, novobiocin, polymyxin B, cefoxitin, and furazolidone). A total of nine virulence genes and eight drug resistance genes were detected by the detection of virulence genes and drug resistance genes. The comprehensive analysis showed that the isolated strain NH21, a subspecies of P. shigelloides, was the causative agent of the present sweetfish surface ulcer disease. The results of this study provided basic information for the prevention and treatment of similar diseases in sweetfish.