牙鲆盾纤毛虫病及防治技术研究

从牙鲆(Paralichthys oliuaceus)患病鱼体的溃烂组织中分离出一种盾纤毛虫,通过活体观察、碳酸银法和电镜观察,对其进行了形态学研究分析。鉴定该盾纤毛虫为贪食迈阿密虫(Miamiensis avidus Thompson & Moewus,1964)。这是国内关于贪食迈阿密纤毛虫感染养殖牙鲆的首次报道。同时本文在防治方法方面进行了初步研究,通过离体药敏实验表明:淡水、甲醛、双氧水对该纤毛虫有较好杀灭作用,而中草药的杀灭效果不理想。     

三甲氧苄氨嘧啶对养殖大菱鲆盾纤毛虫病治疗效果的研究

从感染纤毛虫的养殖大菱鲆上分离获得盾纤毛虫进行体外培养,通过药物抑制实验发现,三甲氧苄氨嘧啶(TMP)对盾纤毛虫的最小有效抑制浓度为25×10~(-6);TMP对大菱鲆的毒性实验表明,24小时半致死量为每千克鱼体内含药物量为0.35357 g,48小时半致死量为每千克鱼体内含药物量为0.33912 g,安全量为每千克鱼体内含药物量为0.09361 g;TMP对大菱鲆盾纤毛虫病的治疗实验结果显示,与对照组相比,注射药物或投喂药饵后,感染纤毛虫的大菱鲆成活率提高了50%～107.1%。     

1种牙鲆致病性纤毛虫的鉴定

报道了1种对牙鲆具致病性的纤毛虫,经活体观察和氨银法染色,对其形态学、口器结构、银线系统做了系统研究.参照Corliss分类系统,将这种纤毛虫确定为贪食迈阿密虫(Miamiensis avidu).查阅国内外相关文献,还未见贪食迈阿密虫能引起牙鲆发病的报道.     

莱州地区大菱鲆纤毛虫病流行病学调查

近年来伴随着大菱鲆养殖规模的扩大、养殖密度的增加以及品种种质的退化，病害问题日趋严重，已成为制约大菱鲆养殖业发展的重大障碍。由于纤毛虫病死亡率高，流行范围广，因而纤毛虫被认为是对海水养殖业危害较大的一类病原体。为了进一步了解该病在养殖中的危害，2008年3—7月对山东莱州地区养殖大菱鲆的纤毛虫病进行了流行病学调查研究，针对纤毛虫病新出现的一些流行病学特点进行了分析、总结，以期为大菱鲆纤毛虫病的预防及治疗提供参考。     

红顶虎头

大菱鲆(Scophthalmas maximus)适应低水温环境,抗逆性强,肉质鲜美,而且生长速度快.自引入我国10年来,在北方沿海地区已迅速发展成为一项特色产业,产生了很好的经济效益和社会效益.但是伴随着养殖范围的扩大和养殖密度的增加,以及品种的退化,大菱鲆的病害问题日趋严重.特别是由盾纤毛虫引起的病害最为严重,盾纤毛虫传染力强,危害性强,一旦发生就会给大菱鲆养殖业造成巨大损失.目前国内报道中治疗盾纤毛虫类病害的有效方法不多,在实际生产中,胡乱用药,用禁药的现象很普遍.因此如何筛选高效、低毒、低残留的无公害药物防治此病迫在眉睫.本文就中草药对盾纤毛虫的治疗效果作了初步研究,供广大养殖户参考.     

大菱鲆与牙鲆盾纤类纤毛虫病的防治技术

盾纤类纤毛虫病是一种对大菱鲆与牙鲆危害很大的寄生虫性的疾病,2000～2002年山东省文登市的几家养殖场先后发生过此病,引起苗种及商品鱼大量死亡,死亡率高达70%以上,危害很大。现将我们掌握的情况介绍如下。一、病原喙突拟梭虫(Paralembusrostratus)是盾纤类纤毛虫(Scuticociliatidciliate)的一种,为兼性寄生纤毛虫,以细菌和有机碎屑为食,在水中自由生活,亦可侵入寄主体内营寄生生活,虫体为葵花籽状,体被均匀一致的纤毛,大小20～40微米。二、症状患病鱼摄食量减少,在育苗池或养殖池中分布散乱,且常有不安地狂动、狂游。患病的大菱鲆商品鱼…     

大菱鲆纤毛虫病发病机理及中药治疗技术研究

通过研究大菱鲆纤毛虫病的发病机理,开发中药治疗大菱鲆纤毛虫病,通过完善药物组方、用药方法、临床验证,近三年来使治疗的效率达到80%以上,同时,完全符合无公害养殖要求和食品安全规定。     

大鹏澳浮游纤毛虫群落与环境因子间的关系

为了解大鹏澳浮游纤毛虫时空分布特征及与环境因子间的关系,于2014年冬季、春季、夏季和秋季对大鹏澳海域进行了浮游纤毛虫的调查,并采用多元分析方法研究了环境因子对其的影响。结果显示,共检出纤毛虫31种,隶属于4纲、8目、18属。纤毛虫平均丰度为1 487个·L-1(10~5 040个·L-1),以砂壳目(Tintinnida)、急游目(Stombidiid)和环毛目(Cyclotrichida)为主。浮游纤毛虫群落结构时空差异较大。浮游纤毛虫丰度在春季最高,冬季最低;种类在夏季最多,春季最少。此外,在鱼类网箱养殖区浮游纤毛虫丰度最高,而在王母河入海口最低。通过冗余分析(redundancy analysis,RDA)发现,大鹏澳浮游纤毛虫群落结构差异主要与水体营养盐水平、温度和叶绿素a含量有关,且在不同季节有不同的主要影响因子。     

防治盾纤毛虫病药物筛选实验

对目前我国牙鲆、大菱鲆养殖过程中普遍发生的一种重要疾病——盾纤毛虫病的防治药物进行了筛选及临床实验。实验表明,甲醛(400 mg/L以上)和硫酸铜(淡水条件下)有较好杀除效果,而淡水浸泡、双氧水、高锰酸钾等则无效。     

大菱鲆与牙鲆盾纤毛虫病的防治技术

盾纤类纤毛虫是一种危害大菱鲆与牙鲆的寄生虫性疾病,2000年~2002年,山东省文登市的几家养殖场先后发生此病,引起苗种及商品鱼的大量死亡,死亡率高达70%以上,危害较大。一、病原喙突拟梭虫(Paralembusrostratus),是盾纤类纤毛虫(Scuticociliatid ciliate)的一种,为兼性寄生纤毛虫,以细菌和有机碎屑为食,在水中自由生活,亦可侵入寄主体内营寄生生活。虫体为葵化籽状,体被均匀一致的纤毛,大小20um~40um。二、症状患病鱼体摄食量减少,在育苗池或养殖池中分布散乱,且常有不安地狂动、狂游。患病的大菱鲆商品鱼体色发黑或发红,眼球突出,鳃盖及吻…     

斜带石斑鱼肝细胞分离及原代培养方法的建立

本研究以斜带石斑鱼肝细胞为实验对象,在不同培养条件下进行原代培养,旨在探讨稳定可靠的斜带石斑鱼肝细胞分离及原代培养方法。采用组织块分离法和胰蛋白酶(含EDTA)消化法分离肝细胞,并通过密度梯度离心法分离纯化肝细胞,细胞悬液于DMEM/F-12、M199和L-15培养液中培养;细胞活力及数量采用血球计数板计数,并通过MTT法测定细胞增殖率;同时,测定不同时间培养上清液中乳酸脱氢酶(LDH)活性、白蛋白(ALB)和尿素氮(BUN)的含量,以分析肝细胞生长状态。结果表明,组织块方法不适于斜带石斑鱼肝细胞的培养,未见细胞从组织块中迁出,而胰蛋白酶消化法获得良好稳定的培养效果,细胞产量达到1.6×108个/g肝重,活细胞数达到95%;L-15培养基细胞生长明显优于DMEM/F-12和M199培养基;启动原代培养的48~72 h阶段肝细胞生长代谢旺盛,培养上清液中LDH活性显著降低,ALB和BUN含量显著升高。结果显示,0.25%的胰蛋白酶常温消化法适合斜带石斑鱼肝细胞的分离,斜带石斑鱼肝细胞原代培养的最适培养基为L-15培养基,肝细胞在启动原代培养的48~72 h生长代谢旺盛。     

来自鳗鲡的二株益生菌适宜培养基的研究

用正交试验法,通过研究培养基基础成分、主要添加成分和无机盐对养殖鳗鲡(Anguilla japonica)肠道益生菌枯草芽孢杆菌(A40209CDC4)和少动鞘氨醇单胞菌(A31009NA)的生长影响,确定了优化的培养基。菌株A31009NA的优化配方为:蛋白胨1.0%,牛肉膏0.1%,食盐0.5%,酵母膏0.5%,葡萄糖1.0%,硫酸铵0.05%,MgSO4.7H2O 0.1 g/L,CuSO4.5 H2O 0.15 mg/L,MnSO4.H2O 1.5 mg/L,CoCl.H2O 0.15 mg/L;A40209CDC4优化培养基配方为:蛋白胨1.0%,牛肉膏0.1%,食盐1.0%,酵母膏0.5%,硫酸铵0.05%,MgSO4.7 H2O 0.1 g/L,MnSO4.H2O 1.5 mg/L,CoCl.H2O 0.15 mg/L,CaCl20.02 g/L。优化培养基与普通营养肉汤(蛋白胨1.0%,牛肉膏0.3%,食盐0.5%)培养对照试验结果表明,优化培养基有效地提高了细菌产量。     

The effect of probiotic bacteria in culture media using organic fertilizer for population density,biomass production and nutrient quality of Phronima sp. as natural feed

Phronima sp. is a natural feed with high nutrient content, and it can be an alternative substitute for Artemia. The existence of probiotic bacteria in culture media aims to improve water quality through biodegradation, maintain microbial balance and control pathogenic bacteria, so that nutrients in culture media can be utilized by Phronima sp. This study aimed to determine the effect of fermented organic fertilizer by probiotic bacteria in mass culture media on the population density, biomass production and nutrient content of Phronima sp. and to find the best culture media composition for mass culture of Phronima sp. The study was conducted with a completely randomized design with five treatments and three replications. The treatments were as follows: A: 0% chicken manure, 50% rice bran, 50% tofu waste; B: 25% chicken manure, 37.5% rice bran, 37.5% tofu waste; C: 50% chicken manure, 25% rice bran, 25% tofu waste; D: 75% chicken manure, 12.5% rice bran, 12.5% tofu waste and E: 100% chicken manure. The test animals were Phronima sp., and during cultivation, fertilization was carried out every 3 days. Population density and biomass were observed, and proximate analysis, amino acid profile analysis and fatty acid profile analysis were performed. The highest population density was at treatment B, with a peak population of 98 individuals/L that occurred on the 16th day of maintenance culture, and the highest biomass of 0.51 g was at the same treatment. The highest nutrient content was obtained at treatment B, for which the highest protein proximate analysis value was 58.90%, the proportion of the fatty acids comprised of eicosapentaenoic acid was 7.53%, and lysine amino acids were found at 44.16 ppm.     

红螯螯虾XO-SG神经细胞的原代培养

本研究采用酶解的方法获得红螯螯虾眼柄XO-SG复合体的单个神经细胞,并依据显微观察对XO-SG神经细胞进行分类,同时利用Leibovitz's L-15等作为基础培养基离体培养解离的神经细胞。目的是建立虾类XO-SG神经细胞的分类标准并确定合适的培养条件,便于在体内外开展神经内分泌系统的调控研究。结果显示,根据神经细胞形态特点,红螯螯虾XO-SG神经内分泌细胞分为6种类型,不同类型的细胞在细胞大小以及显微结构上存在明显差异;建立了红螯螯虾眼柄XO-SG神经元的体外原代培养方法,细胞在改良的L-15培养基中存活状态良好,原代培养的神经细胞可以在体外存活14 d。离体培养过程中,不同类型神经细胞的再生速率存在差异,部分细胞在第2天就出现再生的轴突或树突。再生过程基本可以持续7 d,随后细胞开始萎缩凋亡。本研究为红螯螯虾眼柄神经细胞的分类以及利用神经细胞在体外开展虾类的内分泌代谢和调节机制研究提供了基础依据和研究平台。     

鰤鱼诺卡氏菌培养条件及培养基的优化

对鰤鱼诺卡氏菌（Nocardia seriolea）ZJ0503的增菌培养基和生长条件进行优化。研究了温度、盐度、初始pH对鰤鱼诺卡氏菌生长的影响,并通过单因素试验对培养基的碳源、氮源和无机盐成分进行了筛选,采用正交实验法对培养基各主要成分的添加量进行了优化。结果表明,鰤鱼诺卡氏菌最适宜生长条件为温度25 ℃、盐度5、pH 6.5±0.2;经筛选,鰤鱼诺卡氏菌培养基中最佳碳源是葡萄糖,最佳氮源是酵母粉,促生长作用最强的2种无机盐是磷酸氢二钾（K2HPO4）和氯化钙（CaCl2）;确立了培养基优化配方为葡萄糖20 g·L-1,酵母粉15 g·L-1,K2HPO4 0.75 g·L-1,CaCl 20.2 g·L-1（单独灭菌）,氯化钠（NaCl2）5 g·L-1,pH 6.5±0.2。     

Potential of mixed culture of freshwater prawn (Macrobrachium rosenbergii) and self-recruiting small species mola (Amblypharyngodon mola) in rotational rice–fish/prawn culture systems in Bangladesh

The effects of different stocking densities of freshwater prawn (Macrobrachium rosenbergii) on its growth and production in relation to the presence of small self‐recruiting species mola (Amblypharyngodon mola) were investigated in modified rice fields after rice harvest at Mymensingh, Bangladesh. Keeping the stocking density of mola fixed at 20 000 ha^?1 in each treatment, four densities of freshwater prawn (treatments) were maintained: 10 000, 15 000, 20 000 and 25 000 ha^?1 respectively. The rice plots were limed (CaCO₃) and fertilized with urea, triple super phosphate and cowdung regularly. The prawns were fed daily with commercial pellets. Water quality parameters such as temperature, dissolved oxygen, transparency, pH, total alkalinity, inorganic nitrogen (nitrate, nitrite and total ammonia), chlorophyll a and orthophosphate were determined fortnightly. Numerical analysis of plankton communities was performed monthly. All water quality parameters were found to be within the suitable range for freshwater prawn culture, except high temperature, in the peak summer months. There was a homogenous abundance of plankton communities in all treatments throughout the experimental period. Mola started to breed in the second month of the original stocking with partial harvesting after the second month and continued until the final harvest. The total production of mola ranged between 124 and 152 kg ha^?1 during the 4‐month culture period. The average prawn survival ranged from 49% to 57% without any significant difference among treatments. Freshwater prawn production ranged from 294 to 596 kg ha^?1 with significantly higher production in the treatment where 20 000 ha^?1 freshwater prawn were stocked. This treatment also resulted in a higher net profit margin (74%), indicating that stocking at a combination of 20 000 ha^?1 freshwater prawn and 20 000 ha^?1 mola could be the optimum proposition for prawn–mola culture in modified rain‐fed rice fields after rice cultivation.     

Diluted rotifer, Brachionus plicatilis (Muller), extract as culture medium stimulates hatching of red sea bream, Pagrus major (Temminck & Schlegel)

Hatching rates and hatching times (50% hatch) of red sea bream, Pagrus major (Temminck & Schlegel), eggs in culture media that were prepared to dilute rotifer Brachionus plicatilis Muller extract at 4000-, 6000-. 8000- and 10 000-fold with artificial sea water (σ₁₅, = 24.0) were investigated. Although differences in hatching rate between the culture media and control artificial sea water were negligible, hatching time was significantly shorter in media at 4000- and 6000-fold dilutions than in media at 8000- and 10 000-fold dilutions and in the control artificial sea water, when the eggs immediately post spawning were incubated. The hatching rates of eggs at three developmental stages, immediately post-spawning, gastrula stage and heart-beat initiation stage, incubated respectively in the medium at 6000-fold dilution and the control, did not differ either, yet the hatching times at each egg stage in the medium were significantly shorter than that in the control. These results suggest that hatching enzyme secretion from the hatching glands is promoted to perceive chemical stimuli in the water-soluble fraction of rotifer extract via the embryo's chemoreceptors, from the heart-beat initiation stage to immediately before hatching, thus shortening the hatching time.     

栉孔扇贝心脏细胞的体外培养

体外培养细胞对研究生物的资源保护、功能基因及病害发生机理与防治等均具有重要意义。然而,目前海洋双壳贝类中可以长期存活的组织细胞是有限的。本研究采用植块法启动栉孔扇贝(Chlamys farreri)心脏细胞的原代培养,通过优化培养基的方法,建立了可使其长期存活的原代培养体系。根据组织块迁出细胞的数量和细胞存活时间,确定L-15是3个培养基(L-15,M199,L-15+M199)中最适宜栉孔扇贝心脏细胞的基础培养基。通过三因子三水平正交实验,得出栉孔扇贝心脏细胞适宜的添加物组合:L-15基础培养基中添加5%胎牛血清、50 mmol/L牛磺酸和6 mmol/L Ca~(2+)。原代培养产物中以心肌细胞为主要的细胞类型,其中部分心肌细胞在原代培养14 d内可进行节奏性搏动;部分细胞可局部形成心肌束和肌管样结构;心肌细胞在体外存活时间达2个月。本研究将为栉孔扇贝基础生物学和功能基因的研究提供细胞平台。     

斑点叉尾[鱼回]头肾巨噬细胞分离及初步应用

通过Percoll密度梯度离心法分离斑点叉尾[鱼回]头肾巨噬细胞,经含2%胎牛血清的L-15培养液中培养,在光镜和电镜下观察其形态和超微结构;采用MTS法研究酵母细胞壁对巨噬细胞体外作用的合适浓度和最佳时间。结果表明:使用34%、51%的密度梯度分离斑点叉尾[鱼回]头肾细胞,经过3~12小时培养,巨噬细胞纯化率95%以上;分离的头肾巨噬细胞具有巨噬细胞的形态特征。酵母细胞壁对斑点叉尾[鱼回]的巨噬细胞最适的作用浓度为0-500pg/ml,作用时间为12小时。本研究为进一步探讨酵母细胞壁对鱼类免疫细胞的作用机理奠定了基础。     

Effect of Shrimp Stocking Density on Size-fractionated Phytoplankton and Ecological Groups of Ciliated Protozoa within Zero-water Exchange Shrimp Culture Systems

Zero‐water exchange shrimp culture systems are being investigated to reduce water and pathogens input and effluent output. The relative abundance and diversity of ciliates have been used as indicators of water quality and ecosystem dynamics in natural systems; however, their occurrence and role in shrimp production systems have not been investigated. The objectives of this research were to describe the succession of the ciliate community in a shrimp grow out system, to investigate the impact of increasing shrimp stocking density on the characteristics of the ciliated protozoa community, and to correlate this with shrimp performance. A growth trial with Litopenaeus vannamei was conducted outdoors with zero‐water exchange and three stocking densities (50, 75 and 100 shrimp/m²). In all treatments, free‐swimming ciliates were dominant in the early weeks, while substrate‐associated ciliates appeared later. The 100 shrimp/m² treatment exhibited a lower abundance of free‐swimming ciliates but higher abundance of hypotrichous ciliates in the second half of production and higher shrimp mortality and lower shrimp growth in the last 2 wk. These results provide a preliminary examination of the role of ciliates in shrimp culture systems and direction for future work in determining the role of protozoa as indicators of water quality.