一株克氏原螯虾病原——霍乱弧菌的分离鉴定  

Isolation and identification of Vibrio cholerae, a pathogen of Procambarus Clarkii

作　　者：鹿豪洁 李凯 吕锦贤 王永杰[1,2] 杨明树 LU Haojie;LI Kai;LYU Jinxian;WANG Yongjie;YANG Mingshu(College of Food Science and Technology,Shanghai Ocean University,Shanghai,201306;Laboratory of Quality and Safety Risk Assessment for Aquatic Products on Storage and Preservation(Shanghai),Ministry of Agriculture and Rural Affairs,Shanghai 201306;College of Food Science and Engineering,Hainan Tropical Ocean University,Sanya 572022)

机构地区：[1]上海海洋大学食品学院,上海201306 [2]农业农村部水产品质量安全贮藏保鲜风险评估实验室(上海),上海201306 [3]海南热带海洋学院食品科学与工程学院,三亚572022

出　　处：《安徽农业大学学报》2024年第3期441-449,共9页Journal of Anhui Agricultural University

基　　金：国家自然科学基金项目(41876195)资助。

摘　　要：从尾扇边缘溃烂的克氏原螯虾中分离到一株优势菌LK-18,并对该菌株进行种属鉴定和致病性分析。通过菌落形态观察、16S rRNA测序、血清型分析及基因特性鉴定对细菌的类别进行初步判断,并采用浸泡感染、肌肉注射和腹腔注射等方式进行人工感染实验对细菌的致病性进行评估。经16S rRNA测序分析和4个管家基因(atp A、pyr H、rec A、gyr B)串联分析,该菌株最终被鉴定并命名为VibriocholeraeLK-18。V. choleraeLK-18经PCR鉴定不属于O1和O139血清型,但含hly毒力基因和几丁质分解相关的几丁质酶(chitinase,Chi)基因。人工感染实验虽未能重现烂尾症状,但从克氏原螯虾烂尾组织中分离的V. choleraeLK-18具有致病性,能够引起克氏原螯虾的死亡。研究结果表明霍乱弧菌LK-18是一种致病菌,不仅扩大了非O1/O139群霍乱弧菌的感染宿主范围,同时预警霍乱弧菌有导致克氏原螯虾养殖业暴发疾病的可能,因此要提前做好该疾病的预防工作,从而避免霍乱弧菌感染带来的损失。A dominant strain,LK-18,was isolated from the lesion tissue of the tail fan of Procambarus clarkia,and its species identification and pathogenicity analysis of the strain was conducted.Preliminary bacteria classification was determined through observation of colony morphology,16S rRNA sequencing,serotype analysis and gene characterization.The pathogenicity of LK-18 was assessed using artificial infection tests,which included immersion infection,intramuscular injection and intraperitoneal injection.After 16S rRNA sequencing analysis and four housekeeping gene(ATP synthase alpha,atpA;uridylate kinase gene,pyrH;recombinase A gene,recA;DNA gyrase subunit B gene,gyrB)concatenation analysis,the strain was ultimately identified and named as V.cholerae LK-18.V.cholerae LK-18 was identified by PCR as not belonging to the O1 and O139 serotypes but containing ctx virulence gene and chitinase(Chi)gene related to chitin degradation.Although the artificial infection tests failed to reproduce the symptoms of rotting tail,V.cholerae LK-18 isolated from the rotting tail tissue could cause mortality in P.clarkii,suggesting its potential pathogenicity.The results of this study indicated that V.cholerae LK-18 is a pathogenic bacteria,which not only broadens the known infected host range of O1/O139 group V.cholerae,but also raises concerns about the potential for disease outbreaks in the P.clarkii aquaculture industry.Therefore,it is necessary to take preventive measures in advance to mitigate the risks of V.cholerae infection and to avoid associated losses.

关 键 词：克氏原螯虾 致病菌 霍乱弧菌(Vibrio cholerae) 分离鉴定 

分 类 号：S945.4[农业科学—水产养殖]