摘要
以养殖刺参"腐皮综合征"的重要致病菌——灿烂弧菌Vibrio splendidus的16S～23S间隔区序列,设计特异性引物,用PCR方法制备地高辛(DIG)标记探针,建立了原位杂交技术检测感染刺参体内灿烂弧菌的技术方法,并利用该方法对人工感染刺参和健康刺参各组织进行检测。结果显示,感染刺参的体壁结缔组织、肌肉组织、肠粘膜上皮、辐水管等组织的原位杂交检测呈阳性,而与健康刺参组织无交叉反应。在感染组织中,阳性信号(显色)强弱清晰,能准确反映出灿烂弧菌的侵染部位及感染程度,这为探明灿烂弧菌的感染途径、感染病程等致病机理研究奠定了基础,也为养殖刺参疾病预防和健康管理提供了技术支撑。
The skin ulcer syndrome is a serious disease causing massive mortality and eco- nomic losses in cultured sea cucumber Apostichopus japonicus. Vibrio splendidus was identi- fied as one of the most severely pathogeneses of this disease. In the present study, a DIG-la- beled V. splendidus DNA probe based on the sequence of the 16S- 23S rDNA intragenic spac- ers was synthesized and was used to establish in situ hybridization for detecting the bacterium inA. japonicus. V. splendidus-positive cells were observed in connective tissue, muscle tissue, epithelium of intestine and water-vacular tissues in artificial infected A. japonicus, while no cross-reaction was observed in tissues of healthy sea cucumber. The positive signals were clear in infected tissues, which could reveal the location and degree of the infection. Therefore, this established approach is useful to indicate the infection route and the pathogenesis of V. splendi- dus infection, which will provide basic data for disease control and animal health management.
出处
《渔业科学进展》
CSCD
北大核心
2013年第2期114-118,共5页
Progress in Fishery Sciences
基金
科研院所技术开发研究专项(2011EG34219)
山东省农业良种工程重大课题“速生、抗病、耐高温刺参良种选育”
国家自然科学基金项目(31202016)
“十二五”863项目(2012AA10A412)
青岛市科技计划项目(11-1-11-hy)共同资助
关键词
刺参
腐皮综合征
灿烂弧菌PCR
原位杂交
Apostichopus japonicus
Skin ulcer syndrome
Vibro splendidus
PCR
In situ hybridization
