养殖日本囊对虾副溶血弧菌的分离与鉴定  

Isolation and Identification of Vibrio parahaemolyticus from Cultured Marsupenaeus japonicus

作　　者：王攀攀[1] 王军[1] 林瑶琼 郑锦斌 乔莹[1] 陈信忠[2] 毛勇[1] 

机构地区：[1]厦门大学海洋与地球学院,厦门361005 [2]厦门出入境检验检疫局检验检疫技术中心,厦门361026

出　　处：《热带生物学报》2015年第4期374-380,共7页Journal of Tropical Biology

基　　金：现代农业虾产业技术体系专项(CARS-47);厦门市南方海洋中心项目(14CZY033HJ07);福建省海洋高新产业发展专项(闽海洋高新[2014]12号)

摘　　要：从患病的日本囊对虾(Marsupenaeus japonicus)的肝胰腺、胃和肠组织中分离到2株细菌MJBX-1和MJBX-2。人工感染实验结果表明,2株分离菌浓度为10~cfu·m L^(-1)时,均可致健康日本囊对虾100%死亡;在10~6cfu·m L^(-1)浓度时,均可导致约90%的健康日本囊对虾死亡;该分离菌可使日本囊对虾致病,病状与养殖条件下的相似。MJBX-1和MJBX-2菌的主要特性为:革兰氏阴性,形状为弧状、杆状、丝状,无芽孢,大多有1个明显的弧形,在LB培养基上为透明或半透明的圆形菌落,在TCBS培养基上为绿色菌落。能在含2%,4%,6%,8%Na Cl的LB培养基上生长,但在含0,10%Na Cl的LB培养基上不生长。鉴定结果表明,2株分离菌均为副溶血性弧菌(Vibrio parahaemolyticus)。药敏实验结果表明,分离菌对氟苯尼考、诺氟沙星、呋喃妥因、链霉素、庆大霉素、卡那霉素等6种药物敏感,但对阿莫西林和四环素不敏感。Two strains （ MJBX-1 and MJBX-2） of bacteria were isolated from the hepatopancreas, stomach and intestines of infected Marsupenaeus japonicus, and used for immersion challenge of M. japonicus to study their in- fectivity. These isolates induced 100% mortality of M. japonicus immersion challenged at the concentration of 107cfu·mL-1, and 90% mortality at 106cfu·mL-1 Subsequent bacterial isolation from the challenged M. japonicus showed the same bacterial colony type as in the field samples. These isolates were gram-negative, arc shaped , rhabditiform or filamentous, without spore-forming, but mostly with an obvious arc shape. They formed transparent or translucent round colonies on the LB solid medium and green colonies on the TCBS medium. In the meantime, they could grow at the culture medium containing 2%, 4%, 6% or 8% of NaCL, but not 0% or 10% of NaCL. The isolated strains were preliminarily identified by using the matrix assisted laser desorption/i- onization time of flight mass spectrometry （MALDI-TOF-MS） through comparison of the mass spectrometry be- tween the isolated strains and the standard strain. Moreover, their sequences were analyzed by using the 16S rRNA to produce phylogenetic tree. The MALDI-TOF-MS and the 16S rRNA analysis showed that the isolated strains MJBX-1 and MJBX-2 belonged to Vibrio parahaemolyticus. The antibiotic sensitivity test showed that MJBX-1 and MJBX-2 were sensitive to Florfenicol, Norfloxacin, Macrodantin, Gentamicin, Streptomycin and Kanamycin, but not to Amoxicillin and Tetracycline. This study will provide scientific reference for accurate, quick identification and control of the disease of M. japonicus caused by V. parahaemolyticus.

关 键 词：日本囊对虾 基质辅助激光解吸/电离飞行时间质谱 16S RRNA 副溶血弧菌 药敏实验 

分 类 号：S945.45[农业科学—水产养殖]