大黄鱼(Pseudosciaena crocea)致病性维氏气单胞菌的分离鉴定与药敏特性研究

本研究从自然发病的养殖大黄鱼(Pseudosciaena crocea)肝脏处分离得到1株优势菌9L2,经形态学观察和传统生理生化鉴定,显示该菌为发酵型革兰氏阴性短杆菌,有动力,氧化酶阳性,水解赖氨酸及色氨酸等。利用Biolog细菌鉴定方法进行种属鉴定,确定9L2为维氏气单胞菌(Aeromonas veronii),相似性达100%。同时对其16S rDNA基因进行PCR扩增、测序和系统发育分析,结果表明该菌与气单胞菌属的菌株亲缘关系最近,与维氏气单胞菌(登录号为LMG13695)的16S rRNA基因序列相似性达99.79%。经人工腹腔注射感染吉富罗非鱼(Oreochromis niloticus)的试验结果表明,9L2对罗非鱼的半数致死剂量LD_(50)为1.08×10~3CFU/g。24种抗生素药敏试验结果表明,该菌株对诺氟沙星、新生霉素、呋喃妥因、头孢曲松等13种药物高度敏感,对洛美沙星、卡那霉素等5种药物中度敏感,对利福平、青霉素等6种药物产生耐药性。20种中药体外抑菌试验结果表明,其中11种对致病菌体外抑菌效果较好,平均抑菌浓度范围为0.11—7.20mg/mL。

ISOLATION AND IDENTIFICATION OF PATHOGENIC AEROMONAS VERONII FROM PSEUDOSCIAENA CROCEA

A dominant bacterial strain, named 9L2, was isolated from the liver of diseased Pseudosciaena crocea and identified by biochemical characterization, Biolog identification, and 16S rRNA gene sequence analysis. The bacteria was fermented, Gram-negative, motile, and oxidase-positive, could degrade lysine and tryptophan based on morphological, physiological, biochemical tests. The analysis of 16S rRNA gene sequence show that the organism and the type strain of Aeromonas veronii (LMG13695) formed consistently a monophyletic clade with a distant sequence similarity of 99.79%. According to the results, the bacterium was identified as A. veronii. Moreover, lethal dose and 50% trials were carried out to demonstrate the virulence of 9L2 in Oreochromis niloticus when administered at 1.08×10³CFU/g. Drug sensitivity test on 24 antimicrobial agents and 20 Chinese herbs showed that the strain were very sensitive to 13 agents (norfloxacin, novobiocin, nitrofurantoin, ceftriaxone, etc.), while mild or not sensitive to the rest; of 20 Chinese herbs, 11 herbs showed better antibacterial effect in vitro in average minimum inhibitory concentration ranging from 0.11 to 7.20mg/mL.