中国水产科学Issue(5)：1044-1054,11.DOI:10.3724/SP.J.1118.2015.15127
我国罗非鱼源新型无乳链球菌的分离、鉴定及其分子特征
Isolation, identification, and molecular characteristics of a new geno-type of Streptococcus agalactiae from cultured tilapia in China
摘要
Abstract
A Streptococcosis outbreak in cultured tilapia caused by Streptococcus agalactiae occurred in Wenchang, Hainan Province, China in 2014. In this study, the moribund tilapias were collected from different farms for bac-teria isolation. Most of the moribund tilapias displayed typical clinical signs, such as melanosis, corneal opacity, and swimming abnormalities. A total of 19 isolates (TC-1, TC-2, BL1441–BL1448, and WT1451–WT1459) were isolated from the liver, kidney, spleen, eye, and brain of the diseased tilapia;then, these isolates were identified by morphological observation, physiological, and biochemical characteristics, and sequence analysis of the 16S rRNA gene. All of the isolates were identified as S. agalactiae, and the WT1451–WT1459 strains grew slowly and could not utilize trehalose and ribose. In addition, TC-1, TC-2, and BL1441–BL1448 isolates were positive forβ-hemolysis on sheep’s blood agar plates, but WT1451–WT1459 isolates did not exhibit hemolysis. The genetic characteristics of these isolates were analyzed by multilocus sequence typing (MLST), molecular serotyping, and virulence-related gene sequencing. The results indicated that TC-1, TC-2, and BL1441–BL1448 isolates were common types of Ia-ST7 S. agalactiae in tilapia in China. However, the WT1451–WT1459 isolates were rare types of Ib-ST261 S. agalactiae in tilapia worldwide. PCR revealed that the virulence-related genes bac, bca, bibA, cfb, hylB, iagA, fbsB, and cylE were present in TC-1, TC-2, and BL1441–BL1448 isolates, and the virulence-related genotype of these strains was bac+-bca+-bibA+-cfb+-hylB+-iagA+-fbsB+-lmb–-scpB–-cylE+-gbs20186–. However, the WT1451–WT1459 isolates were positive for bibA, cfb, hylB, iagA, fbsB, and gbs20186, and the virulence-related genotype was bac–-bca–-bibA+-cfb+-hylB+-iagA+-fbsB+-lmb–-scpB–-cylE–-gbs20186+. Despite the fact that the number of the virulence-related genes of the WT1451 strain was less than that of the BL1441 strain, the former was more virulent than the latter in tilapia. In this study, the virulence tests were conducted by intraperitoneal in-jection of the BL1441 and WT1451 strains in tilapia. The results of the challenge experiments showed that the WT1451 strain possessed stronger pathogenicity to tilapia than that of BL1441 strain. The WT1451 strain caused 85%mortality in tilapia at a dose of 4.5×103 CFU/mL. The BL1441 strain showed weak virulence in tilapia, with cumulative mortality of 65%at a dose of 4.5×108 CFU/mL postchallenge. In summary, the 19 S. agalactiae strains isolated from tilapia in Wenchang were divided into two genotypes based on physiological and biochemical char-acteristics, 16S rRNA gene sequences, hemolysis activity, molecular serotyping, and PCR screening of viru-lence-related genes. Although reports on the Ib-ST261 S. agalactiae strains in tilapia are rare worldwide, these strains were highly virulent in tilapia. Therefore, it is very important to obtain data on the epidemiology of Ib-ST261 S. agalactiae strains in tilapia in China. To the best of our knowledge, this is the first report on the hy-pervirulent Ib-ST261 strain of S. agalactiae, which was isolated from tilapia in China. This study provides a framework for the exploration of epidemiological analysis, vaccine development, prevention, and treatment of S. agalactiae in tilapia.关键词
罗非鱼/无乳链球菌/分子分型/致病性Key words
tilapia/Streptococcus agalactiae/molecular typing/pathogenicity分类
农业科技引用本文复制引用
张德锋,刘礼辉,任燕,李宁求,林强,潘厚军,石存斌,吴淑勤..我国罗非鱼源新型无乳链球菌的分离、鉴定及其分子特征[J].中国水产科学,2015,(5):1044-1054,11.基金项目
国家科技支撑计划项目(2012BAD25B02) （2012BAD25B02）
现代农业产业技术体系建设专项资金项目(CARS-46) （CARS-46）
