养殖鱼类链球菌病病原的分离鉴定及其16S rDNA分析

从患病的海水网箱养殖鱼中分离到5株链球菌,分别命名为HD-1、HD-2、HD-3、HD-4和HD-50。常规生化结果表明,其中4株即HD-1、HD-2、HD-3和HD-4为海豚链球菌(Streptococcus iniae);另外1株即HD-50为停乳链球菌(Streptococcus dysgalactiae)。分别对上述分离菌株的16S rDNA进行PCR扩增和测序,并与NC-BI中收录的其它链球菌的16S rDNA序列一起进行聚类分析并构建了系统发生树,确定其分类地位。分子分析结果与生化鉴定结果一致。攻毒实验表明以上4株S.iniae均能引起罗非鱼发病,呈现典型的链球菌病症状,并能从发病的罗非鱼脑、心、肝肾和血液中成功回收S.iniae。以上生化和分子生物学鉴定以及罗非鱼攻毒实验结果表明养殖鱼类链球菌病的主要病原为S.iniae,这与国外有关鱼类链球菌病病原的研究报道一致。

Isolation, Identification and Characteristics of 16S rDNA Gene Sequences of the Pathogens Responsible for the Streptococcicosis in Cultured Fish

Five strains of Streptococcus,designated HD-1,HD-2,HD-3,HD-4 and HD-50,were isolated from diseased seawater farmed fish in Guangdong Province.HD-1,HD-2,HD-3 and HD-4 were identified,according to their physiological and biochemical characteristics,as Streptococcus iniae,and HD-50 as Streptococcus dysgalactiae.The 16S rDNA of these isolates were amplified and sequenced,and the sequences were aligned with those of other Streptococci retrieved from NCBI.The result of the alignment and clustering analysis is in accordance with the result of physiological and biochemical identification of the isolates.Typical symptoms of streptococcicosis and death were showed after healthy tilapia(Oreochromis niloticus) challenged i.p.with these Streptococcus iniae isolates,and the bacteria were successfully recovered from the brain,liver,kidney,spleen and blood of the infected tilapia.It is concluded that Streptococcus iniae is the primary pathogenic agent of streptococcicosis in farmed fish,which is in accordance with the related reports from other countries in the world.