一株梭鲈源无乳链球菌的分离、鉴定及分子分型分析

Identification,characterizations and molecular subtyping analysis of a Streptococcus agalactiae isolated from Lucioperca lucioperca

为查明梭鲈(Lucioperca lucioperca)凸眼病病原,从患病梭鲈病灶眼、肝、脾和肾分离纯化病原菌,进行了生理生化特性测定及16S rRNA基因序列分析,并开展人工感染试验,测定其血清型及MLST分型,并利用纸片扩散法进行药敏特性分析。结果显示:分离菌株(命名为SL1701株)为本次引发梭鲈病害的致病菌,Ib-ST261型。回归感染实验证实,分离得到的细菌SL1701对健康梭鲈具有非常强的致病性,4.5×105CFU/m L时可使80%受感染梭鲈死亡,并可复制出自然发病鱼的症状。菌株SL1701革兰氏染色为阳性链球菌,γ溶血,生理生化特性与普通无乳链球菌基本一致,16S rRNA基因序列与无乳链球菌(Streptococcus agalactiae)同源性最高,综合判定分离菌株为无乳链球菌(Streptococcus agalactiae)。药敏试验结果显示:分离菌株SL1701对氟苯尼考、氟罗沙星等17种抗生素高度敏感,对青霉素、磷霉素等8种抗生素耐药。结果表明,分离菌株SL1701为Ib-ST261型无乳链球菌,养殖时可选用氟苯尼考及氟罗沙星等药物进行防控。

Pathogens were isolated and purified from eyeball lesions, liver, spleen and kidney of Lucioperea lucioperca to find out the pathogen of bulging eyes disease on L. lucioperea. The pathogenic bacteria was identified by analyzing the physiology biochenfistry characteristics and 16S rRNA gene sequence, testing the molecular serolyping and multilocus sequence typing （MLST） , and carrying out the artificial infection experiment, meanwhile, the antinficrobial susceptibility was also tested by disc diffusion method. The results showed that strain SL1701 was the pathogen with strong pathogenicity to L lucio- perca, which could cause 80% mortality as the concentration of bacterium reached to 4.5 × 10^5 CFU/mL. The infected L. lucioperea has the sinfilar symptom with the spontaneous sick individuals. The isolates were all gram - positive and γ - hemolysis with the same physiological and biochemical characteristics of normal Streptococcus agalactiae. According to the 16S rRNA gene sequence analysising, the homology of strain SL1701 was close to S. agalactiae, which was the rare types of Ib - ST261 S. agalactiae. The antimicrobial susceptibility test results showed that strain SL1701 was susceptible to 17 agents including floffenicol, fleroxaein and other antibiotics. Meanwhile, it showed resistance to penicillin, fosfomycin and other 8 antibiotics. The results demonstrated that S. agalactiae was the pathogen causing high mortality on L lucioperca, and the disease could be prevented by using drugs such as floffenicol and fleroxacin in fisheries.