养殖澳洲宝石鱼迟缓爱德华氏菌的分离鉴定及致病基因的检测  

Identification and detection of virulence gene of the pathogenic bacteria Edwardsiella tarda in cultured Scortum barcoo

作　　者：叶旭红[1,2,3] 林先贵[1,2] 王一明[1,2] 

机构地区：[1]中国科学院南京土壤研究所土壤与农业可持续发展国家重点实验室,南京210008 [2]中国科学院南京土壤研究所-香港浸会大学土壤与环境联合开放实验室,南京210008 [3]中国科学院研究生院,北京100049

出　　处：《淡水渔业》2010年第1期50-54,共5页Freshwater Fisheries

基　　金：国家自然科学基金(20607024);国家"十一五"科技支撑计划(2006BAD10B05);农业科技成果转化资金项目(2006C40038)

摘　　要：从患病澳洲宝石鱼体内分离到一株致病菌(编号Et4),对该菌的生化特性及致病基因进行检测,并进行了药敏和人工感染实验。结果显示:菌株Et4的生化鉴定结果与迟缓爱德华氏菌(Edwardsiella tarda)标准菌株(AY77513)一致;其16S rRNA序列,经同源性比对与迟缓爱德华氏菌核苷酸相似度最高,达99%;根据迟缓爱德华氏菌的致病基因Ⅲ型分泌系统装置蛋白esaV基因序列设计引物,进行PCR扩增,得到708 bp序列,该序列与迟缓爱德华氏菌的esaV基因序列相似性达99.3%,说明菌株Et4具有esaV基因。菌株Et4对环丙沙星等较敏感,对其它药物中度或不敏感,具有较强的致病力(LD50=3.74×104CFU/mL),从病鱼中可以重新分离出此菌。综合形态学、生化特性、16S rDNA序列及致病基因序列鉴定其为迟缓爱德华氏菌。Bacterial strain Et4 was isolated from diseased Jade perch （Scortum barcoo）. The biochemical characteristics of the strain Et4 and the virulence gene were detected, and carried out sensitivity and challenge tests. The results indicated that： the strain Et4 was Edwardsiella tarda with 99% reliability resulted by API ID32E. The 16S rRNA gene sequence analysis revealed that the strain Et4 exhibited the highest level of similarity of 99% identity with E. tarda compared with GenBank database. Thus, the strain Et4 was identified as E. tarda. The T3SS apparatus gene esaV is one of the primary virulence genes of E. tarda. A 708 bp fragment of the strain was amplified by PCR amplification using the specific primers designed according to the esaV gene sequences of E. tarda. It was found that the sequences similarity of the fragment and esaV gene of E. tarda was 99. 3%, indicating that the strain Et4 as a virulent strain possessed esaV gene. The strain Et4 was shown sensitive to some antibiotics, such as ciprofloxacin. Challenge test proved that the strain Et4 had higher virulence （ LD50 = 3.74 × 10^4 CFU/mL） , and it also can be isolated from diseased fish. Composite morphology, biochemical characteristics, 16SrDNA sequences and virulence gene sequence identified the strain Et4 as E. tarda.

关 键 词：澳洲宝石鱼(Scortum barcoo) 迟缓爱德华氏菌(Edwardsiella tarda) 16S RRNA基因 esaV基因 

分 类 号：S941.42[农业科学—水产养殖] Q93-331[农业科学—水产科学]