2种青虾运输方法的比较试验

在保持水温15~17℃条件下,采用添加与未添加噬菌蛭弧菌的尼龙袋充氧法和添加与未添加噬菌蛭弧菌的网格增氧法运输青虾(Macrobrachium nipponensis),统计运输时间分别为4、6、8、10、12、16、20、24、283、2 h的青虾存活率。结果表明,长时间运输时,采用网格增氧法运输所获得的存活率比采用尼龙袋充氧法运输的高,而且噬菌蛭弧菌溶液对运输存活率的提高有明显的改善作用。     

细菌QJ2对对虾病原性哈维氏弧菌的拮抗作用

对气味黄杆菌(Flavobacteria ordoratum)QJ2及其去细胞上清液(CFS)对对虾病原性哈维氏弧菌(Virio harveyi)Z3G2、E022的拮抗作用进行了研究.QJ2的CFS对病原菌表现显著的拮抗性能,在6h内使数量为105 ～106cfu@mL-1的病原菌Z3G2、E022均降为0.经热、酸碱、蛋白酶处理的CFS,抑菌活性均有不同程度下降,结果为经60℃处理60min的CFS,经6h使Z3G2细菌数降为0,经15min使E022数量降为最低,为初始菌数的 63.0%经100℃处理10min,CFS使Z3G2和E022数量分别在1h和15min降为最低,各为初始菌数的23.7%和 72.1%;经pH4作用30min,CFS使Z3G2和E022数量分别在15min和1h降为最低,各为初始菌数的88.3%和 15.9%,而经pH12处理30min后,CFS没有使病原菌数下降,但使病原菌以比对照组细菌低的速率缓慢上升; 经蛋白酶K于37℃作用60min后,CFS在15min时使Z3G2和E022数量降为最低,各为初始菌数的15.0%和 66.1%.在人工水体中,QJ2活菌对病原菌也具有显著的拮抗性能,在24h内使Z3G2、E022数量下降到最低值,各下降了1.6、2.4 Log1o数量级.结果表明QJ2产生的抗性物质为细菌素,对病原性哈维氏弧菌具有良好的拮抗作用.     

环境因子对发光细菌的生长及日本对虾仔虾感染死亡率的影响

以水温、盐度、pH、COD、NH3－N为环境因子，哈维氏弧菌（Vibro harveyi)感染日本对虾（Penaeus japonicus）仔虾，试验进行3d。结果表明，水温、温度、pH和COD对发光细菌的生长和日本对虾仔虾的感染死亡率具有明显的促进作用。在水温30℃，盐度32．8，pH7．654和COD10．2mg／L时，发光细菌在试验结束时的数量比初始时分别增加了11．2倍，6．5倍和7．9倍；日本对虾仔虾的死亡率比对照组分别增加了60％，45％，43％和50％。NH3－N对发光细菌的促生长作用不明显，但其质量浓度1．2－1．6mg/L时，能增加日本对虾仔虾对发光病的感染率和死亡率。     

Presence of snapper, seabass, and siganid inhibits growth of luminous bacteria in a simulated shrimp culture system

The antibacterial effect of the presence of Tilapia hornorum against luminous bacteria in shrimp culture has been reported. This study investigates how the presence of commercially valued marine species such as seabass, snapper and siganid affect the growth of luminous bacteria in shrimp culture water. Results showed that luminous bacterial count of water stocked with seabass, siganid and snapper are significantly lower than those without fish. Therefore this study has demonstrated that seabass, siganid and snapper are alternative species for culture with shrimp to control or inhibit the growth of luminous bacteria in shrimp ponds.     

九孔鲍肠道及其养殖水体中弧菌产酶能力的分析

从汕尾健生鲍鱼场养殖水体和九孔鲍肠道中共分离筛选到25株弧菌,其中14株来自成鲍肠道,11株来自养殖水体,并对它们产多种胞外酶的能力进行了比较分析。试验结果表明:78.6%的肠道弧菌能分泌蛋白酶,21.4%能分泌脂肪酶,64.3%能分泌卵磷脂酶或淀粉酶,92.9%能分泌明胶酶;水体中的弧菌63.6%能分泌蛋白酶,54.5%能分泌脂肪酶或明胶酶,72.7%能分泌卵磷脂酶,100%能分泌淀粉酶。同时筛选出5株胞外酶分泌能力超强的菌株,除分泌蛋白酶能力最强的弧菌(Bh14)来自鲍鱼肠道外,其余分泌其它4种胞外酶能力最强者(Sh05、Sh03、Sh11、Sh08)均来自养殖水体。研究还发现来自水体的弧菌分泌多种胞外酶能力强于来自肠道的弧菌。由此提示,应用有益微生物以改善鲍鱼肠道中的菌群结构,可能会提高鲍鱼饲料利用率。     

Interactions of microorganisms isolated from gilthead sea bream, Sparus aurata L., on Vibrio harveyi, a pathogen of farmed Senegalese sole, Solea senegalensis (Kaup)

Four bacterial isolates from farmed gilthead sea bream, Sparus aurata, included in a previous study as members of the Vibrionaceae and Pseudomonodaceae and the genus Micrococcus, have been evaluated for their adhesive ability to skin and intestinal mucus of farmed Senegalese sole, Solea senegalensis, and their antagonistic effect on Vibrio harveyi, a pathogen of sole. These isolates showed higher adhesion to sole mucus than the pathogenic strains of V. harveyi assayed. Only two of the isolates showed antagonistic activity to V. harveyi. Interactions of the four isolates with V. harveyi in respect of adhesion to skin and intestinal mucus under exclusion, competition and displacement conditions were studied. Three isolates were able to reduce the attachment to skin and intestinal sole mucus of a pathogenic strain of V. harveyi under displacement and exclusion conditions, but not under competition conditions. The in vivo probiotic potential of isolate Pdp11 was assessed by oral administration followed by challenge with the pathogenic V. harveyi strain Lg14/00. A group of 50 Senegalese sole received a commercial diet supplemented with 10(8) cfu g(-1) of lyophilized Lg14/00 for 15 days. A second group of fish received a non-supplemented commercial diet. After challenge the mortality of the fish receiving the diet supplemented with the potential probiotic isolate was significantly lower than that in the fish receiving the non-supplemented commercial diet. This study has shown that the ability to interfere with attachment of pathogens, as well as the adhesion to host surfaces, are suitable criteria for selection of candidate probiotics for use in the culture of Senegalese sole.     

哈维氏弧菌的血清型与菌苗抗原性的关系

采用不同血清型哈维氏弧菌Vibrio hanwyi624、809和106等3个菌株，制备成福尔马林灭活菌苗，注射接种大黄鱼4周后，通过血清中凝集、交叉凝集抗体效价测定以及活菌攻毒试验，证明了3个菌株制备的菌苗抗原性存在差异。     

不同方法灭活的哈维氏弧菌菌苗对异育银鲫的免疫原性比较

将哈维氏弧菌（Vibrio harveyi)制备成福马尔林灭活菌苗（formalin killed V.harveyi,F-VH)，苯酚灭活菌苗（phenol killed V.harveyi,P-VH)和热灭活菌苗（heat killed V.harveyi,H-VH)，以注射法接种异育银鲫（Carassius auratus gibelio)后，通过测定受免鱼血清中凝集抗体效价，头肾及血液中吞噬细胞的吞噬活性和杀菌活性，对用不同方法制备的3种灭活菌苗的免疫原性进行了比较。结果表明，3种不同方法制备的V.harveyi菌苗对异育银鲫均显示出较强的免疫原性，其中以F－VH的免疫原性最强，P－VH其次，而H－VH的免疫原性较弱。说明用福尔马林灭活V.Harveyi较用苯酚和热灭活方法更有利于保护V.harveyi菌体的有效抗原。     

同步检测海水养殖动物5种病原菌的扩增子拯救多重PCR(Arm-PCR)方法的建立与应用

鳗弧菌(Vibrio anguillarum)、哈维氏弧菌(V.harveyi)、嗜水气单胞菌(Aeromonas hydrophila)、迟缓爱德华氏菌(Edwardsiella tarda)和副溶血弧菌(V.parahaemolyticus)是我国海水养殖动物5种重要的病原菌.本研究在分析了其致病因子基因的基础上,依据扩增子拯救多重PCR(Arm-PCR)的原理,设计了5套特异性的套式PCR引物,并在各内引物的5'端分别加上能被一对通用引物识别的标签序列;通过对套式PCR中影响扩增结果的引物混合物浓度、退火温度、Mg2+浓度、dNTPs浓度以及Taq DNA聚合酶浓度等5个反应参数的调整和优化,最终建立了一种能同步检测海水养殖动物5种常见病原菌的Arm-PCR方法.优化后的Arm-PCR方法第一步PCR反应体系为:10×PCR Buffer 5 μL(含20 mmol/L的Mg2+),dNTP(各2.5mmol/L)5μL,T的酶(2.5 U/μL)0.6 μL,10×Primer Mix(2 μmol/L)5 μL,DNA模板各1μL,灭菌双蒸馏水补足至50 μL,反应的退火温度为55℃.实验结果表明:对鳗弧菌、哈维氏弧菌、嗜水气单胞菌、迟缓爱德华氏菌和副溶血弧菌这5种病原菌,使用该方法可以在1支反应管内快速、同步进行检测,得到大小分别为144、190、266、315和371 bp的特异性产物,其检出灵敏度分别为1.745、1.847、16.000、28.126和369.900 pg细菌基因组DNA;该方法特异性强,与大肠杆菌(Escherichia coli)、溶藻弧菌(V.alginolyticus)、河豚毒素假交替单胞菌(Pseudoalteromonas tetraodonis、枯草芽孢杆菌(Bacillus subtilis)等其他细菌以及半滑舌鳎基因组DNA不产生交叉反应.2012年,应用该Arm-PCR方法对分离自病鱼的24个优势菌株进行了检测,确定出5株迟缓爱德华氏菌、3株嗜水气单胞菌、2株哈维氏弧菌及2株副溶血弧菌,证实该方法具有良好的可靠性和实用性.该方法不仅适用于海水养殖动物中致病性鳗弧菌、哈维氏弧菌、嗜水气单胞菌、迟缓爱德华氏菌、副溶血弧菌的快速、同步检测和病原流行情况调查,也为进一步开发相应的基因芯片检测方法打下了基础.