Protective immune responses of recombinant outer membrane proteins OmpF and OmpK of Aeromonas hydrophila in European eel (Anguilla anguilla)

Outer membrane proteins (Omps) of Gram‐negative bacteria have been proven to be efficient subunit vaccines against bacteriosis. In this study, OmpF and OmpK of Aeromonas hydrophila were expressed, and their immune protective effects in European eel (Anguilla anguilla) were evaluated. The genomic DNA of A. hydrophila 322A was used as a template, and two kinds of prokaryotic expression plasmids, pET‐32a‐OmpF and pET‐32a‐OmpK, were constructed. Recombinant OmpF protein (r‐OmpF) and r‐OmpK were purified and were proven to have antigenicity by Western‐blot analysis. r‐OmpF and r‐OmpK were used as immunogens to immunize European eel by intraperitoneal injection. The mRNA expression of 6 immune‐related genes (IgM, IL‐10, IRF3, IRF7, LysG4 and HexB) in the liver tissues of eels at 1 hr, 3 hr, 6 hr, 12 hr, 24 hr, 72 hr and 10 days postimmunization was analysed by real‐time PCR. At 30 dpi, the serum antibody response was measured by ELISA. Fish were attacked at 15 dpi by live 322A to assess the protective immunity of r‐OmpF and r‐OmpK. All the six tested genes responded to r‐OmpF or r‐OmpK vaccination at varying degrees. The serum antibody titre of r‐OmpF‐ and r‐OmpK‐immunized groups was 1:1,600 and 1:3,200 respectively. In addition, r‐OmpF gave 35.5% of the relative immune protection rate to European eels, while r‐OmpK gave 70.0%. By analysing the protective immunity and the regulatory role in the immune‐related gene expression of the two recombinant proteins that were studied, it was found that r‐OmpK was a potential vaccine candidate against A. hydrophila.     

同位素稀释液质联用法测定鱼体中氯丙嗪的残留

建立了高效液相色谱-串联质谱(HPLC/MS/MS)分析鱼体中氯丙嗪残留的方法.样品用乙腈提取,MCX固相萃取柱净化,氨化甲醇洗脱.在正离子模式下以电喷雾串联质谱仪进行测定,采用内标法定量,氘代氯丙嗪为内标物.以欧洲鳗鲡、鳜鱼和大口黑鲈为研究对象,在1.0、5.0、25.0μg/kg添加水平下,方法平均回收率为90.1...     

澳洲长鳍鳗染色体组型的初步研究

以澳洲长鳍鳗(Anguilla reinhardtii)头肾组织为材料,采用腹腔注射植物血细胞凝集素(PHA)、秋水仙素和空气干燥法制备染色体标本,进行染色体组型分析.结果显示:澳洲长鳍鳗染色体为38条,核型公式为2n=14m+6sm+18t,NF=58;未发现随体、次级缢痕及异形染色体.     

Migratory history of the giant mottled eel (Anguilla marmorata) in the Bonin Islands of Japan

Chino N, Arai T. Migratory history of the giant mottled eel ( Anguilla marmorata ) in the Bonin Islands of Japan.
Ecology of Freshwater Fish 2010: 19: 19–25. © 2009 John Wiley & Sons A/S
Abstract –  In order to elucidate the migratory history and habitat use of the giant mottled eel Anguilla marmorata, in the Bonin Islands of Japan, strontium (Sr) and calcium (Ca) concentrations in otoliths were examined by X-ray electron microprobe analysis. The X-ray intensity map and line analyses showed three fluctuation patterns: (i) freshwater residence, (ii) continuous residence in brackish water, and (iii) residence in freshwater after recruitment, while returning to brackish water. Based on these results and previous studies, the migratory behaviours of A. marmorata appear to differ in each habitat in response to inter- and intra-specific competition. The results suggest that A. marmorata has a flexible pattern of migration, with an ability to adapt to various habitats and salinities.     

日本鳗鲡胶原蛋白和小清蛋白的过敏原性

以日本鳗鲡皮与肌肉组织为研究对象,采用碱溶、酸溶、盐析、冻干等方法纯化得到胶原蛋白,采用加热、饱和硫酸铵分级盐析、DEAE-Sepharose阴离子交换层析等方法纯化得到两种亚型的小清蛋白(PV-Ⅰ和PV-Ⅱ),纯化的目标蛋白经动物特异性抗体的免疫印迹实验确证。酶联免疫吸附测定和免疫印迹分析结果显示,纯化的胶原蛋白和小清蛋白分别与鱼类过敏患者阳性血清发生特异性反应,且二者之间无免疫交叉反应。体外模拟胃液消化实验和SDS-PAGE分析结果显示,胶原蛋白和小清蛋白均具有较高的消化稳定性。结果提示,日本鳗胶原蛋白和小清蛋白具有较高的消化稳定性和免疫原性,二者可引发不同患者的IgE介导特异性超敏反应。     

日本鳗鲡腐皮病病原菌的分离及鉴定

从患腐皮病的日本鳗鲡(Anguilla japonica)体表溃烂处分离到1株病原菌(322A),对其进行了人工感染实验和生理生化分析,测定了该菌株的16S rRNA基因序列和促旋酶(gyrase)B亚单位gyrB基因序列,并分别构建系统发育树。结果显示:感染实验证实菌株322A具有致病性;生理生化分析鉴定该菌株属于气单胞菌属(Aero-monassp.);16S rRNA基因分析显示,该菌株与气单胞菌属细菌的同源性均在99%~100%,构建的系统树显示,菌株322A与嗜水气单胞菌(A.hydrophila(FJ462702))亲缘关系最近;gyrB基因分析表明,该菌株与A.hydrophila种内序列的相似性为96%~98%,种间序列的相似性为94%~95%,构建的系统树结果显示,该菌株与A.hydrophila(FJ608553、FJ608552、AF208259)聚为一个分支。综合上述实验结果,菌株322A可鉴定为嗜水气单胞菌(A.hydrophila)。     

几种环保型饲料添加剂对欧洲鳗鲡消化酶活性的影响

选取健康均匀的欧洲鳗苗1000尾,分为5组,每组2个重复,分别在基础料中添加4种环保型饲料添加剂,分析了4种环保型饲料添加剂对欧洲鳗鲡消化酶活性的影响。结果表明,1号饲料添加剂对于提高欧洲鳗鲡消化酶整体效果最好,对中草药原料的配伍进行选择优化,适当添加半胱胺,能显著提高欧洲鳗鲡幼鱼消化酶活性。     

莫桑比克鳗鲡养殖生物学主要特性研究

对莫桑比克鳗鲡（Anguillamor morata）玻璃鳗经形态学鉴定品种后，开展了主要养殖生物学特性研究。耐受高温和低温分别为39℃和7℃，适宜生存水温15℃～33℃；耐受盐度与水温相关，水温30℃致死盐度2．9％，适宜盐度小于1．1％，20℃致死盐度2．7％，适宜盐度小于1．8％，10℃致死盐度2．1％，适宜盐度小于2．0％；耐受pH值与水温相关，30℃致死pH值分别为3．5和11．0，适宜pH值6．0—7．0，20℃致死pH值分别为4．0和11．0，适宜pH值6．0～8．0，10℃致死pH值分别为3．5和10．5，适宜pH值5．5—8．5。常用药物S高聚碘、二氧化氯、三氯异氰尿酸、苯扎溴铵、溴氯海因、甲醛、敌百虫的安全浓度高于或接近常用浓度，硫酸铜、溴氰菊脂乳油、氯氰菊脂、甲苯咪唑的安全浓度低于常用浓度。水体中非离子态氨的安全浓度为2．51mg／L，亚硝酸氮的安全浓度为55．12mg／L。     

欧洲鳗鲡三类苗养殖技术初探

本试验应用膨化饲料养殖欧洲鳗鲡（Anguilla anguilla）三类苗。经过110d的养殖，两个试验组的平均体重分别达到16．13g和15．39g，相对增重率分别为151．1％和142．4％。日均增重量分别为88．3mg和82．2mg。平行养殖试验结果显示存活率分别为97．7％和98．5％，饲料转化率分别为59．1％和58．4％。整个养殖过程基本不换水，完全不用任何药物。     

超低温冷冻对日本鳗鲡精子酶活性的影响

研究超低温冷冻保存(-196℃)对日本鳗鲡精子内总ATP酶、肌酸激酶(CK)、琥珀酸脱氢酶(SDH)、乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷胱甘肽还原酶(GR)等酶活性的影响。运用试剂盒分别测定了冷冻前后日本鳗鲡精子内酶活性的变化。结果表明,经过超低温冷冻保存后,日本鳗鲡精子的活力下降,精子内GR活性显著升高(P<0.05),酶活性从冻前的358.52±45.65 U/L上升到646.30±70.30 U/L;其它几种酶的活性均显著下降(P<0.05),总ATP酶、CK和SDH的活性分别从冻前的3.14±0.61 U/ml、17.10±3.51 U/ml和32±5.94 U/ml下降到1.83±0.43 U/ml、7.33±1.74 U/ml和21±1.41 U/ml,LDH、SOD和CAT活性分别从冻前的2 266.67±313.25 U/L、220.47±32.94 U/ml和48.51±5.94U/ml下降到1 195.91±198.51 U/L、84.16±22.11 U/ml和21.8±4.14 U/ml。超低温冷冻对日本鳗鲡精子酶活性和精子活力均有较大影响。