罗氏沼虾(Macrobrachium rosenbergii)幼体新暴发病病原阴沟肠杆菌的分离鉴定  

IDENTIFICATION AND ISOLATION OF A NOVEL BACTERIAL PATHOGEN(ENTEROBACTER CLOACAE) FROM DISEASED LARVAE OF GIANT FRESHWATER PRAWN MACROBRACHIUM ROSENBERGII

作　　者：陈雪峰[1] 杨国梁[1] 高强[1] 夏正龙[1] 濮剑威[1] 慎佩晶[1] 黄振远[1] 

机构地区：[1]国家罗氏沼虾遗传育种中心农业部淡水渔业健康养殖重点实验室浙江省淡水水产遗传育种重点实验室浙江省淡水水产研究所,湖州313001

出　　处：《海洋与湖沼》2015年第6期1467-1477,共11页Oceanologia Et Limnologia Sinica

基　　金：国家科技支撑计划项目;2012BAD26B04;浙江省重大科技专项;2012C12907;浙江省淡水养殖重点科技创新团队项目;2010R50026-02;湖州市重大科技专项;2011ZD2005号;浙江省科研院所专项;2014F50002号

摘　　要：2010年,浙江湖州部分罗氏沼虾(Macrobrachium rosenbergii)育苗场在幼体培育阶段,暴发了幼体大规模发病死亡的新病害。从2个育苗场6个发病育苗池的发病幼体匀浆液中分离得到6株优势生长菌株(编号为NTH01、NTH02、NTH03、316D、316X、316C),菌落形态较为一致。取代表菌株NTH01,以浸泡方式进行人工回感,结果菌株NTH01能够导致幼体发病死亡,且症状与生产表现一致。经形态学观察、生理生化鉴定、16S r RNA与gyr B基因序列系统发育关系构建,鉴定菌株NTH01为阴沟肠杆菌(Enterobacter cloacae)。以阴沟肠杆菌omp A基因片段为靶序列,建立了PCR检测方法,对另外5株优势菌进行检测,结果该5株菌都是阴沟肠杆菌。2011年,对4个发病育苗场幼体与亲虾进行检测,结果发病幼体中分离的优势菌检测呈阳性,亲虾肝胰腺与肠道组织检测到该菌存在,但亲虾不表现病症。对30种抗生素的药敏试验结果表明,菌株NTH01对恩诺沙星、氧氟沙星、庆大霉素等6种抗生素敏感,对发病幼体进行针对性用药,效果明显。本试验结果表明,阴沟肠杆菌是导致罗氏沼虾育苗期间幼体大规模发病死亡的致病菌。In 2010, a new disease broke out in most Macrobrachium rosenbergii hatcheries in Huzhou, Zhejiang Province. The cumulative mortality of larvae was 70%—80%, resulting in great economic losses. Six dominant bacterium（NTH01, NTH02, NTH03, 316 D, 316 X, and 316C） were isolated from the homogenate of infected larvae. The morphology of isolated bacterium from Nutrient Agar was consistent. An artificial infection experiment was performed to test the pathogenicity of representative strain NTH01. The result shows that healthy larvae could be infected by strain NTH01. In morphological, physiological, and biochemical aspects, strain NTH01 was identified as Enterobacter cloacae. A PCR method established for E. cloacae detection was used to detect the other 5 dominant bacteria, showing a positive result. In 2011, samples of larvae and adult shrimp from 4 infected hatcheries were detected by the PCR method, the PCR showed a positive result in diseased larvae samples, but negative in healthy larvae. Although positive detection results were also found in hepatopancreas and gut, adult shrimp did not showed a diseased symptoms. Strain NTH01 was resistant to 30 kinds of commonly antibiotics, such as cefoxitin, enrofloxacin, amikacin, and so on. The results of drug sensitivity test show that enrofloxacin, ofloxacin, gentamicin, neomycin, doxycycline and aztreonam were most effective against the pathogenic bacterium. However, the treatment of E. cloacae infections is difficult and expensive. Therefore, a specific pathogen-free environment for larvae culture shall be constructed to prevent and control the disease caused by E. cloacae.

关 键 词：罗氏沼虾幼体 新病原 阴沟肠杆菌 分离鉴定 药敏试验 

分 类 号：Q523.8[生物学—生物化学] S917[农业科学—水产科学]