两种微藻改善虾池环境增强凡纳对虾抗病力的研究

人工引入波吉卵囊藻和微绿球藻于凡纳对虾养殖环境中,检测与凡纳对虾抗病力有关因子变化和测定主要水质因子,研究微藻生态调控对凡纳对虾抗病力的影响。结果表明,引入波吉卵囊藻和微绿球藻能改善养殖水体的水质,凡纳对虾的血细胞数目,血清蛋白的含量以及酚氧化酶、超氧化物歧化酶,溶菌酶,抗菌酶的活性都较对照组显提高。因此,微藻生态调控是防止对虾疾病的重要技术措施。     

叉鞭金藻固定化培养的初步研究

以海藻酸钙为载体,初步考察了CaCl2浓度、胶球大小、密度及初始细胞密度等条件：对叉鞭金藻固定化培养的影响,确定了该藻固定化生长的优化条件;当藻细胞密度大于10^6cells/ml,CaCl2浓度为0.15mol/L,在50ml培养液中加入150个微藻胶球时,藻细胞的生长量最大。与游离的叉鞭金藻相比,固定化叉鞭金藻生长速度慢,但生长周期长。     

一株饵料微藻与益生菌混合固定化培养条件的优化

目的:一株饵料微藻与益生菌混合固定化培养条件的优化.方法:在单因素实验基础上,采用U*6(64)均匀设计表,对海藻酸钠浓度、氯化钙浓度、藻的接种量和菌的接种量进行优化实验.并采用DPS软件对实验结果进行分析,以获得适宜的混合固定化条件;并按照此优化条件对混合固定化和单藻、单菌固定化胶球中微藻和益生菌的生长速率进行比较.结果:①获得适宜的混合固定化条件为:海藻酸钠浓度4%,氯化钙浓度1.2%.藻接种量为21.9889×102个细胞,菌接种量为58.7676×109个细胞;②混合固定化胶球中藻细胞的生长速率比单藻固定化胶球中的藻的生长速率提高了18.8%;菌的平均生长速率比单菌固定化胶球中的菌的平均生长速率提高了92.6%:结论:该实验首次对饵料微藻和海洋益生菌的混合同定化培养条件进行优化.发现藻菌混合固定化胶球中藻和菌的生长速率比单独固定化均有较明显提高.实验结果说明,在混合固定化的微环境体系中,特定的藻菌株具有相互促进生长的作用.实验结果为藻菌混合固定化技术的应用和发展提供了必要的实验数据和理论基础.     

微藻碳酸酐酶的特性及其环境调控

本文概述微藻碳酸酐酶的性质、种类、分布、分离纯化和环境调控的研究进展,并对未来有关微藻碳酸酐酶研究中需要探讨的问题作了展望。     

固定化海洋微藻对污水中Ni2+的吸附

采用海藻酸钠包埋小球藻和叉鞭金藻,制得含藻细胞的固定化胶球,用其对Ni^2 进行生物吸附,研究了固定化小球藻和固定化叉鞭金藻对污水中Ni^2 的吸附率。结果表明：对于同一种固定化微藻,处于对数生长中期时对Ni^2 吸附效果较好,且吸附过程主要在前4h完成;Ni^2 浓度越大,吸附率越高;固定化微藻比悬浮态微藻吸附率高;在相同的实验条件下,固定化小球藻比固定化叉鞭金藻吸附率高。     

虾池常见微藻的光照强度、温度和盐度适应性

通过Smith生态位宽度指数和Pianka生态位重叠指数分析了虾池常见微藻种群(啮蚀隐藻、新月菱形藻、微绿球藻和蛋白核小球藻)在光照强度、温度和盐度资源上的生态位宽度和生态位重叠特征.结果表明:新月菱形藻和蛋白核小球藻具有较大的生态位宽度值,啮蚀隐藻和微绿球藻的生态位宽度值则相对较小.蛋白核小球藻和微绿球藻在光照强度、温度和盐度资源上的生态位重叠值均为最大,啮蚀隐藻在各资源与其他微藻的重叠值最小.新月菱形藻与蛋白核小球藻适应光照强度的范围较广.当水温达16.9℃,可定向培育新月菱形藻;当水温达25℃,可定向培育新月菱形藻和啮蚀隐藻;当水温达30℃时,可定向培育新月菱形藻、蛋白核小球藻和微绿球藻.养殖水体盐度处于9～26,可引入蛋白核小球藻与微绿球藻;处于9～17.5,应引入啮蚀隐藻;高盐水体,应引入新月菱形藻.蛋白核小球藻和微绿球藻在光照强度、温度和盐度资源上的生态位重叠值均为最大,因此微藻定向培育,不可同时引入蛋白核小球藻与微绿球藻.     

微藻高效基因枪转化方法的建立

基因枪法是外源基因导入微藻细胞的重要手段。然而,发展至今,微藻细胞基因枪转化效率一直偏低（10~50个转化子/μg DNA）,高价低效的转化方法阻碍了基于高通量转化子的基因功能分析。为了提高基因枪的转化效率,本研究以三角褐指藻为材料,从抗生素选择培养基的改良,微载体的选择、制备、包埋、点膜和轰击参数的优化,以及受体细胞的处理等方面进行了系统研究。结果显示,采用50%海水盐度f/2培养基可以提高博来霉素的效价,f/2固体培养基中2216E营养物质的加入能缩短1/3的平板筛选时间。微载体制备应选择对金（钨）粉没有吸附作用的离心管,制备量/管应少于3.5 mg。微载体轰击量每次大约为0.75 mg,过量将会造成一个轰击死亡圈,过少将导致轰击成本上升。当轰击间距A为6.35 mm,间距B为11 mm,间距C为6 cm时,可以获得最多的转化细胞。109个受体细胞铺成较厚的多细胞层能显著提高转化效率。经过上述优化与改进,本研究将现有文献报道的转化效率提高了4.7~30倍,达到295 ± 60个转化子/μg DNA。该方法除适用于三角褐指藻外,也可广泛应用于其他微藻（杜氏盐藻、小球藻）的基因枪转化研究,可以为微藻基因工程研究提供快速,高效和可靠的操作技术。     

微藻与细菌作用关系的研究进展

藻类是水生环境中的初级生产者,它的生长常常伴随着细菌并受菌的影响。有研究者指出藻类和细菌有着密不可分的关系。一些研究表明与藻相关的主体细菌是特定的细菌群体,特别是α-变形菌频繁地发现,说明这类菌可能能够开启和维持共生关系。最近的研究提出了营养物质交换是菌藻共生的基础,这类相关化合物是复杂的和特定的分子,可能参与信号处理和监控作用,而不只是被动扩散。同时,这种作用很明显不是静态的,它的开启和终止可能是对环境和发育的响应。需要指出的是明确菌藻关系的作用机理还有待于进一步的深入研究,本篇综述结合新提出的理论,对细菌与微藻作用关系的研究进展进行总结,概括了微藻与菌的作用关系(进化关系,营养依赖,代谢互补和协作生物合成),这种作用关系涉及到的菌的分类(膜菌和藻际微环境菌,促生菌PGPB和溶藻菌)以及菌藻作用的应用(废水处理和生物燃料生产)的情况,并对菌藻关系的未来发展做了展望。     

汞对固定化斜生栅藻净化污水及其生理特征的影响

研究了在不同Ｈｇ浓度条件下,固定化斜生栅藻对污水中氨氮、正磷酸盐和Ｈｇ的净化效率及其生理特征变化,结果表明,固定藻的净化效率和对Ｈｇ毒性的抗性明显高于悬浮藻。０．２ｍｇ．Ｌ＾－１Ｈｇ对固定藻的净化效率无影响,并对ＰＯ＾３－４－Ｐ的吸收有刺激促进作用,而悬浮藻的开始下降;从０．８－１．４ｍｇ．Ｌ＾－１,随着Ｈｇ浓度的增加,固定藻对氨氮和正磷酸盐的净化效率逐渐下降,而悬浮藻的则急剧下降,固定藻对汞的去     

固定化对微生物生理变化的影响

固定化微生物技术近年来得到了广泛的应用研究,然而,人们关于固定化对微生物生理特性影响方面的知识还很缺乏,文献报道不多且比较零散,严重滞后于固定化微生物的应用研究。综述了固定化对微生物生理特性方面的影响,包括固定化对微生物生长速率、对微生物活性、对有毒物质的耐受性以及对细胞中DNARNA总量变化等方面的影响。     

氧化塘中细菌种群组成动态

对武汉地区氧化塘中的细菌数量、细菌种类、种群组成、优势种及群落多样性进行了初步研究。从氧化塘中分离的细菌经鉴定属于12个属,主要有假单胞菌属(Pseudomonas)、棒杆菌属(corynebacterium)、无色杆菌属(Achromobacter)和微杆菌属(Microbacterium),其它菌属是芽孢杆菌属(Bacillus)、色杆菌属(Chromobacterium)、短杆菌属(Brevibacterium)、葡萄球菌属(Staphylococcus)、微球菌属(Micrococcus)、黄杆菌属(Flavobacterium)、埃希氏菌属(Escherichia)和链球菌属(Streptococcus)。在5个小塘中,细菌数量、种群组成和群落多样性指数都有不同,这与各塘的污染程度有关。由此,多样性指数可用于监测氧化塘的净化效果。     

红藻糖苷对超低温冻存微藻的保护作用

为研究红藻糖苷对超低温冻存微藻细胞的保护作用,研究将3种不同的微藻置于含10% DMSO和不同浓度红藻糖苷的冻存液中,冻存并解冻后,以流式细胞仪检测细胞存活率,测定复养后藻株的生长曲线及相关生理参数。结果显示,由于冷冻损伤,冻存后各种藻细胞的生长速率、细胞密度及生理指标都显著性下降,而红藻糖苷协同DMSO能够显著增加细胞的存活率,尤其15%红藻糖苷能将紫球藻存活率提升20%（P0.05）;生长曲线得到明显改善;且对PSII最大光能转化效率也有显著性提高（P0.05）。总体结果来看,红藻糖苷对超低温冻存微藻,特别是紫球藻具有明显的保护作用,且效果强于蔗糖。     

3种海洋赤潮微藻蛋白质和核酸合成对UV-B辐射增强的响应

应用同位素标志法, 研究了赤潮异弯藻 (Heterosigmaakashiwo) 、亚历山大藻 (Alexandriumtamarense ) 和中肋骨条藻 (Skeletonemacostatum) 核酸和蛋白质合成对紫外线B波段 (UV_B, 2 80~ 32 0nm) 辐射增强的响应变化。结果表明 :1) 按照由高到低的顺序, 3种海洋赤潮微藻对UV_B辐射增强的敏感性依次是赤潮异弯藻、亚历山大藻和中肋骨条藻。 2 ) UV_B辐射增强抑制赤潮异弯藻的生长和脱氧核糖核酸 (DNA) 的合成, 而低剂量的UV_B辐射对中肋骨条藻和亚历山大藻的生长与DNA的合成表现出刺激作用, 高剂量则表现出抑制作用。 3) 随着UV_B辐射的增强, 3种海洋赤潮微藻核糖核酸 (RNA) 和蛋白质的合成速度下降, 其中赤潮异弯藻合成速度的下降幅度明显大于中肋骨条藻和亚历山大藻, 表明赤潮异弯藻RNA和蛋白质的合成对UV_B辐射增强的敏感性高于中肋骨条藻和亚历山大藻。     

CELL‐CELL INTERACTION IN THE EUKARYOTE‐PROKARYOTE MODEL OF THE MICROALGAE CHLORELLA VULGARIS AND THE BACTERIUM AZOSPIRILLUM BRASILENSE IMMOBILIZED IN POLYMER BEADS1

Cell‐cell interaction in the eukaryote‐prokaryote model of the unicellular, freshwater microalga Chlorella vulgaris Beij. and the plant growth‐promoting bacterium Azospirillum brasilense, when jointly immobilized in small polymer alginate beads, was evaluated by quantitative fluorescence in situ hybridization (FISH) combined with SEM. This step revealed significant changes, with an increase in the populations of both partners, cluster (mixed colonies) mode of colonization of the bead by the two microorganisms, increase in the size of microalgae‐bacterial clusters, movement of the motile bacteria cells toward the immotile microalgae cells within solid matrix, and formation of firm structures among the bacteria, microalgae cells, and the inert matrix that creates a biofilm. This biofilm was sufficiently strong to keep the two species attached to each other, even after eliminating the alginate support. This study showed that the common structural phenotypic interaction of Azospirillum with roots of higher plants, via fibrils and sheath material, is also formed and maintained during the interaction of this bacterium with the surface of rootless single‐cell microalgae.     

EFFECTS OF NUTRIENT ENRICHMENT ON PRIMARY PRODUCTION AND BIOMASS OF SEDIMENT MICROALGAE IN A SUBTROPICAL SEAGRASS BED1

Eutrophication of coastal waters often leads to excessive growth of microalgal epiphytes attached to seagrass leaves; however, the effect of increased nutrient levels on sediment microalgae has not been studied within seagrass communities. A slow‐release NPK Osmocote fertilizer was added to sediments within and outside beds of the shoal grass Halodule wrightii, in Big Lagoon, Perdido Key, Florida. Gross primary production (GPP) and biomass (HPLC photopigments) of sediment microalgae within and adjacent to fertilized and control H. wrightii beds were measured following two 4‐week enrichment periods during June and July 2004. There was no effect of position on sediment microalgal GPP or biomass in control and enriched plots. However, nutrient enrichment significantly increased GPP in both June and July. These results suggest that sediment microalgae could fill some of the void in primary production where seagrass beds disappear due to excessive nutrient enrichment. Sedimentary chl a (proxy of total microalgal biomass) significantly increased only during the June enrichment period, whereas fucoxanthin (proxy of total diatom biomass) was not increased by nutrient enrichment even though its concentration doubled in the enriched plots in June.     

THE EFFECT OF SODIUM CHLORIDE ON CARBONIC ANHYDRASE ACTIVITY IN MARINE MICROALGAE1

The effects of NaCl on the internal and external carbonic anhydrase (CA) activity of several marine microalgae were studied. Unlike freshwater microalgae in which CA activity is generally inhibited by NaCl, marine microalgae exhibited considerable species-dependent variation when exposed to NaCl. CA activity in Phaeodactylum tricornutum, a diatom, was inhibited, whereas it was activated in the coccolithophorid Pleurochrysis carterae. CA activity in the chlorophyte Dunaliella primolecta was not significantly affected by NaCl. In Dunaliella salina and Dunaliella parva, NaCl inhibited external CA without affecting the internal activity, whereas in Chlorella vulgaris C-133 and Dunaliella peircei only the internal CA was inhibited. Internal CA of Dunaliella tertiolecta was not affected by NaCl, but the external enzyme was significantly enhanced. Salt substitution experiments revealed that chloride (Cl^-) is the ion affecting CA activity; the effect of Cl^- can be replaced by bromide ion. Cl^- affects external CA activity while also affecting the apparent affinity for inorganic carbon during photosynthesis. Microalgae whose internal CA activity was enhanced by Cl^- showed higher intracellular Cl^- concentrations than those species that were inhibited.     

蔗糖输入对凡纳滨对虾养殖系统真核微生物群落的影响

利用18S rRNA基因高通量测序, 研究蔗糖输入条件下凡纳滨对虾(Litopenaeus vannamei)养殖系统中水体与絮团的真核微生物群落组成及其相互作用关系。结果表明, 输入蔗糖能显著改变对虾养殖系统中真核微生物群落组成, 促进水体中纤毛虫和轮虫的生长, 对水体绿藻的过度生长有一定抑制作用。Mantel分析发现, 在第28天时, 蔗糖添加以及系统中的溶解氧、亚硝酸盐和磷酸盐对真核微生物群落结构变异有显著影响。利用共线性网络分析表明碳氮比的增加可以增强水体和絮团中不等鞭毛类、纤毛虫和轮虫的相互作用, 对养殖体系中藻类的过度繁殖具有一定的抑制作用。研究揭示了在蔗糖输入条件下凡纳滨对虾养殖系统中真核微生物群落的演替规律及絮团和水体微生物相互作用特征, 为生物絮团技术持续稳定的应用于对虾养殖系统提供理论支持。     

海水养虾池浮游动物对浮游植物牧食力的研究

利用常规显微镜直接计数法评估了海水养虾池浮游动物对浮游藻类的牧食力。测得虾池浮游动物对水柱浮游植物的总滤水率为８０．３８ｍｌ／（Ｌ·ｈ）,每个浮游动物的滤水率平均为１１．１３μｌ／（ｉｎｄ·ｈ）,总牧食率为１０．５０μｇ／（Ｌ·ｈ）,即１．２５ｎｇＣ／（ｉｎｄ·ｈ）。浮游动物对水柱原位主要浮游藻类的选食,以隐藻和舟形藻为最多,其次是扁藻和小环藻,６ｈ内对上述４种藻类的选食率分别为９２．６８％、７２．４６％、３６．２２％和３２．５６％,其相应的选择指数依次为０．７３３１、０．６７１７、０．４３４５和０．３９１３。对水柱总体而言,６ｈ内有３５．７２％的主要浮游藻类被选食了。     

ENDOGENOUS CYTOKININS IN THREE GENERA OF MICROALGAE FROM THE CHLOROPHYTA1

Nine axenic microalgal (Chlorophyta) strains from three genera (Protococcus, Chlorella, and Scenedesmus) were analyzed for endogenous cytokinins. Cytokinin‐like activity was detected using the excised cucumber cotyledon bioassay. Five strains showed no cytokinin‐like activity and four strains, low cytokinin‐like activity. Ethanolic extracts of the microalgae containing a mixture of deuterium‐labeled standards were purified using a combined DEAE‐Sephadex octadecysilica column and immunoaffinity column based on wide‐range specific mon‐oclonal antibodies and analyzed by HPLC linked to a micromass single quadrupole mass spectrometer with an electrospray interface and a photodiode array detector. There were similar trends in cytokinin profiles for the nine microalgal strains investigated, although concentrations did vary. Both isopentenyladenine and isopentenyladenosine were detected in all nine strains. cis‐Zeatin and cis‐zeatin riboside occurred at higher concentrations than the trans isomers, whereas trans‐zeatin‐O‐glucoside and trans‐zeatin riboside‐O‐glucoside were dominant over the cis isomers. Dihydrozeatin and its conjugates were not detected in any significant amounts. The aromatic benzyladenine always occurred at higher concentrations than benzyladenosine. The topolins were well represented with all three isomers (ortho, meta, and para) being detected, with ortho‐topolin and ortho‐topolin riboside occurring at higher concentrations than the other isomers. However, for the O‐glucosides, the meta isomers (meta‐topolin‐O‐glucoside and meta‐topolin riboside‐O‐glucoside) occurred at higher concentrations than the other isomers. No N‐glucosides were detected (isopentenyladenine‐9‐glucoside, zeatin‐9‐glucoside, dihydrozeatin‐9‐glucoside, benzyladenine‐9‐glucoside, ortho‐topolin‐9‐glucoside, and meta‐topolin‐9‐glucoside). Generally, zeatin and topolin conjugates were the dominant forms of isoprenoid and aromatic cytokinins, respectively. There was no distinct trend in the proportions of isoprenoid to aromatic cytokinins.