铜锈微囊藻两种表型的生长生理特性及毒素组成比较分析

从滇池蓝藻水华中分离得到的铜锈微囊藻群体在实验室无机营养中解聚成单细胞,结果表明,群体微囊藻的生长速度明显低于单细胞微囊藻;前者具明显可见的胞外酸性多糖胶鞘,而单细胞则几乎没有;按常规方法分析比较两种细胞形态的毒性大小和毒素组成,发现群体微囊藻主要含有三种微囊藻毒素的异构体,而单细胞以MCLR为主;且单细胞微囊藻的毒性约为群体的10倍.二者的LDH和PGM同工酶酶谱也有差异.本研究为解释毒素的合成和调控机理提供了新的证据.       

底泥中微囊藻复苏和生长特性的研究

研究了微囊藻群体从底泥中释放进入水体的过程及这一过程与水体温度、光照及营养盐的关系 ,并比较了底泥和水体中微囊藻群体的生长特性。同时 ,比较了温度对经低温 (4℃ )处理的和处于对数期的Microcystis.sp .94 0的叶绿素荧光强度的影响。结果表明 ,在 15℃ ,30 μEm-2 s-1光照条件下 ,底泥中的微囊藻群体复苏开始启动 ,并于15d后开始上升到水体中。研究表明 ,存在于底泥中的微囊藻群体从底泥中迁移至上层水体的最适条件为 2 0℃ ,30 μEm-2 s-1。分别培养底泥微囊藻群体和同时期水体中的微囊藻群体 ,研究它们的生长特性 ,发现底泥中的微囊藻群体生长的最适温度为 2 0℃ ,光照强度为 30 μEm-2 s-1,与同期水体中的微囊藻群体生长条件相似。经低温 (4℃ )处理的微囊藻群体和生长周期处于对数期的微囊藻群体叶绿素荧光的影响的实验中 ,作者发现在 2 0℃和 2 5℃时 ,两种经过不同处理的微囊藻群体都随着时间增加而增长。但是 ,在 10℃和 15℃时 ,低温处理的微囊藻群体的叶绿素荧光随着时间增加而增长 ,而处于对数期的微囊藻群体的叶绿素荧光随着时间增加而降低。这表明长期处于低温和黑暗环境中的微囊藻细胞的光系统Ⅱ未受到严重的损伤 ,当环境转变有利于生长时 ,微囊藻细胞的光系统Ⅱ恢复活性。本研究结     

光强对微囊藻群体形态的影响及其生理机制研究简

为了探讨光照对微囊藻形态的影响,研究了6株不同种的群体微囊藻在不同光强下群体形态的变化及其响应机制。研究发现,随着光强的增加,6株群体微囊藻的群体尺寸变大。当光强为80—200μmol/(m2·s)时,群体微囊藻DH-M1和DC-M2的比生长速率显著增大,而另4株在高光强下比生长速率无显著性差异;对多糖含量分析发现,高光强对群体微囊藻TH-M2、DC-M1、FACHB1174和FACHB1027胞外及胶被多糖的分泌与释放有显著的促进效果,而DH-M1和DC-M2多糖含量增加不明显。对于不同的微囊藻株,高光强促进群体形态变化的作用机理不同:光饱和点低的微囊藻是通过分泌大量的胞外及胶被多糖使群体尺寸变大,而光饱和点高的微囊藻是通过生长来促进群体尺寸的增大。此外,对产毒藻株在不同光强下的毒素基因表达及胞内毒素测定发现,高光强组的群体微囊藻mcyB和mcyD表达量升高,且胞内微囊藻毒素含量增加显著,推测微囊藻毒素也可能是影响微囊藻群体形态及大小的作用因子之一。     

光强对微囊藻群体形态的影响及其生理机制研究

为了探讨光照对微囊藻形态的影响,研究了6株不同种的群体微囊藻在不同光强下群体形态的变化及其响应机制。研究发现,随着光强的增加,6株群体微囊藻的群体尺寸变大。当光强为80200 mol/（m2s）时,群体微囊藻DH-M1和DC-M2的比生长速率显著增大,而另4株在高光强下比生长速率无显著性差异;对多糖含量分析发现,高光强对群体微囊藻TH-M2、DC-M1、FACHB1174和FACHB1027胞外及胶被多糖的分泌与释放有显著的促进效果,而DH-M1和DC-M2多糖含量增加不明显。对于不同的微囊藻株,高光强促进群体形态变化的作用机理不同:光饱和点低的微囊藻是通过分泌大量的胞外及胶被多糖使群体尺寸变大,而光饱和点高的微囊藻是通过生长来促进群体尺寸的增大。此外,对产毒藻株在不同光强下的毒素基因表达及胞内毒素测定发现,高光强组的群体微囊藻mcyB和mcyD表达量升高,且胞内微囊藻毒素含量增加显著,推测微囊藻毒素也可能是影响微囊藻群体形态及大小的作用因子之一。       

群体和单细胞微囊藻对短期高光胁迫的生理响应

为了探讨不同形态的微囊藻（Microcystis）对光的耐受能力及其应对机制,研究比较了短期高光强条件下群体微囊藻和单细胞微囊藻的生理响应,结果表明,在高光强胁迫下,群体和单细胞微囊藻的叶绿素含量、最大电子传递速率（ETR_max）均降低,但与单细胞微囊藻相比,群体微囊藻的下降幅度较小;在高光强胁迫下,群体微囊藻的过氧化氢酶（CAT）与超氧化物歧化酶（SOD）的活性均显著增加,而单细胞微囊藻只有CAT活性增加;在短期高光胁迫下,群体微囊藻的死亡率没有显著变化。这些结果表明群体微囊藻比单细胞微囊藻能耐受更高的光强,也暗示了群体微囊藻在野外高光强条件下更具竞争优势。     

鲢、鲤和鲫肝细胞原代培养

微囊藻毒素(Microcystins)是一类淡水水体中危害很严重的生物毒素(Biotoxin),由微囊藻毒素所引发的环境问题及其对人类健康的危害正日益受到科学家的关注1.已知微囊藻毒素作用的靶器官为肝脏,以往的研究多集中在微囊藻毒素对动物肝脏组织的损伤,如口服或腹腔注射毒素,引起肝组织结构破坏、肝出血甚至肝坏死,但用整体实验动物或器官研究微囊藻毒素毒理学较难深入,因此建立毒理学实验模型十分重要.肝脏作为动物体内最重要的解毒器官,是研究微囊藻毒素毒理学的主要对象.一般毒理学实验都采用肝脏原代培养细胞,因为原代培养细胞生理生化及遗传特性稳定,适于研究外界毒物的毒性、毒理及肝细胞对毒物的应答和解毒机理.本实验通过对鲢(Hypophthalmichthy molitrix Carier et Valencienines)、鲤(Cyprinus carpio Linnaeus)和鲫(Carassius auratus Linnaeus)肝脏原代细胞培养,以建立稳定的毒理学实验模型,为微囊藻毒素毒理学研究奠定基础.       

蓝藻胞外多聚物生物合成、群体形成与微囊藻水华

有毒微囊藻水华在太湖、巢湖和滇池等饮用水源地频繁暴发, 对居民健康和水产养殖等构成严重威胁, 亟需开发新技术加以有效控制和利用。在水华暴发时, 蓝藻大量分泌胞外多聚物而形成细胞群体, 是蓝藻水华发生的关键和前提。蓝藻群体中胶质状胞外多聚物由胞外多糖、蛋白质和其他生物大分子组成, 对其结构、功能和生物合成途径研究了解仍然有限。生物信息学和比较基因组学分析发现微囊藻和其他多种蓝藻中编码大量的具有称之为PEP-CTERM结构域的潜在胞外蛋白质, 这些潜在的蛋白质可能通过特殊的分选系统分泌到细胞表面, 与胞外多糖相互作用形成结构更复杂的胞外多聚物, 介导细胞群体的形成和水华发生。亟需建立微囊藻遗传操作技术, 深入揭示胞外多聚物生物合成和群体形成的分子机制, 寻找控制蓝藻胞外多聚物的组装和分泌及群体形成的关键靶点, 将有助于揭示蓝藻水华形成机理及开发新型控藻技术。     

群体和单细胞微囊藻对短期温度变化的生理响应

为了探索短期温度变化对群体微囊藻和单细胞微囊藻的影响, 在室内受控模拟条件下研究了在10℃、25℃和35℃三个温度梯度下, 群体和单细胞微囊藻对短期温度变化的生理响应。研究表明: 与对照组25℃相比, 在10℃培养下, 微囊藻叶绿素浓度显著降低, SOD活性和死亡率均显著增加。与群体微囊藻相比, 在10℃下单细胞微囊藻叶绿素浓度显著下降, F_v/F_m下降, SOD活性显著增加。在35℃培养下, 单细胞微囊藻叶绿素浓度上升, 死亡率和SOD活性增加, 而群体微囊藻则呈现出叶绿素浓度和死亡率降低, CAT活性增加。结果表明短期的温度变化影响了群体和单细胞微囊藻生理机制, 与单细胞微囊藻相比, 群体更能适应短期的温度胁迫, 导致其更具优势。     

微囊藻细胞抽提物亚慢性暴露导致小鼠肝脏氧化应激

研究了微囊藻细胞抽提物亚慢性暴露对小鼠肝脏抗氧化系统的影响.采用腹腔注射进行连续染毒28d,染毒组剂量为3.3μg micmcystins/kg体重.结果显示,超氧化物歧化酶、过氧化氧酶、谷胱甘肽过氧化物酶在第4周时发生显著性升高,提示微囊藻细胞抽提物激活了小鼠肝脏抗氧化系统.谷胱甘肽-S-转移酶和对照组相比也显著提高,表明谷胱甘肽-S-转移酶作为解毒Ⅰ相酶加快了对肝脏微囊藻毒素的清除.脂质过氧化产物丙二醛也显著升高,说明抗氧化系统未能清除微囊藻细胞抽提物对小鼠肝脏的氧化损伤,导致了氧化应激的产生.结果表明低剂量微囊藻细胞抽提物长时间暴露能够导致小鼠肝脏氧化损伤.     

非生物因素对藻类胞外多聚糖含量影响

本文从研究铜绿微囊藻表型转换机制的需要出发,综述了非生物因素对藻类胞外多聚糖含量的影响.很多藻类的胞外多聚糖分泌量与环境中主要营养盐比例之间存在着一定的响应关系,在高碳氮比或高碳磷比条件下,即在氮不足或磷不足时,藻类光合作用固定的有机物质主要以不含氮磷的碳水化合物形式存在,胞内碳水化合物的过量累积导致其逐步向胞外转移释放,使得胞外多聚糖含量显著升高.在碳氮代谢不平衡的情况下,胞外多聚糖充当了接收过剩固定碳的汇.对于一些藻类,不同光谱、光强和光周期均可影响其胞外多聚糖的合成与分泌.温度对藻类胞外多聚糖的产生也有一定影响.由于胞外多聚糖在藻细胞相互粘结形成群体上发挥着重要作用,通过调控相关非生物因素使得铜绿微囊藻胞外多聚糖产量增加可能有助于在室内模拟铜绿微囊藻的群体形成.     

GENETIC VARIABILITY OF BRAZILIAN STRAINS OF THE MICROCYSTIS AERUGINOSA COMPLEX (CYANOBACTERIA/CYANOPHYCEAE) USING THE PHYCOCYANIN INTERGENIC SPACER AND FLANKING REGIONS (cpcBA)

The genetic and morphological variability among 15 Brazilian strains of Microcystis aeruginosa (Kütz.) Kütz. collected from four locations was examined and compared with several reference strains of M. aeruginosa , M. viridis (A. Br.) Lemm. and M. wesenbergii (Kom.) Kom. in Kondr. Brazilian strains were classified by morphological features and by comparison of the nucleotide sequences of the cpc BA intergenic spacer and flanking regions. Our results indicate that Brazilian strains classified as M. aeruginosa are phylogenetically diverse compared with reference strains of M. aeruginosa and that the current taxonomy underestimates genetic diversity within M. aeruginosa. The data also demonstrate that morphological criteria alone are inadequate to characterize Microcystis species. Although colonial characters were shown to vary considerably in culture, some genetic lineages demonstrated consistent cellular diameter ranges, indicating that cell size has value as a taxonomic character. The detection of six M. aeruginosa genotypes in a single water body indicates that morphological approaches can also seriously underestimate the diversity of Microcystis bloom populations.     

铜绿微囊藻对紫外辐射的生理代谢响应

采用紫外(UV)滤膜过滤日光UV以及紫外灯添加UV的方法,研究了UV辐射对铜绿微囊藻Microcystis aeruginosa单细胞藻株PCC7806和群体藻XW01生长及生理代谢的影响。结果显示,在室内条件下低剂量UV辐射可促进群体微囊藻XW01生长;室外条件下与滤除了UV的光照相比,含有UV的完全日光更有利于微囊藻生长;而相同的UV辐射强度均导致单细胞株死亡,群体株显示了较强的UV抗性;日光中的UV可促进XW01合成抗氧化相关的超氧化物歧化酶(SOD)和过氧化氢酶(CAT)、促进胞外多糖的产生并形成较大的群体、促进UV屏障物质类菌孢素氨基酸(MAAs)和伪枝藻素(Scy)积累。这些生理代谢的改变,消除了阳光辐射中UV对微囊藻的伤害。研究的结果提示,自然条件下阳光中的UV有助于群体微囊藻生长。     

The role of microcystins in maintaining colonies of bloom-forming Microcystis spp

Microcystis is a cosmopolitan genus of cyanobacteria and occurs in many different forms. Large surface blooms of the cyanobacterium are well known in eutrophic lakes throughout the globe. We evaluated the role of microcystins (MCs) in promoting and maintaining bloom-forming cell aggregates at environmentally relevant MC concentrations (0.25-10 μg l(-1)). MCs significantly enhanced Microcystis colony sizes. Colonial diameters in microcystin-RR (MC-RR)-treated cultures (at 1 μg l(-1)) were significantly larger than control colonies, by factors of 1.5, 2.6 and 2.7 in Microcystis wesenbergii DC-M1, M. ichthyoblabe TH-M1 and Microcystis sp. FACHB1027 respectively. Depletion of extracellular MC concentrations caused Microcystis colony size to decrease, suggesting that released MCs are intimately involved in the maintenance of Microcystis colonial size. MC-RR exposure did not influence Microcystis growth rate, but did significantly increase the production of extracellular polysaccharides (EPS). In addition, MC-RR exposure appeared to trigger upregulation of certain parts of four polysaccharide biosynthesis-related genes: capD, csaB, tagH and epsL. These results strongly indicate that induction of polysaccharides by MC-RR was the major mechanism through which MCs enhanced colony formation in Microcystis spp. Cellular release of MCs, therefore, may play a key role in the persistence of algal colonies and the dominance of Microcystis.     

An extracellular glycoprotein is implicated in cell-cell contacts in the toxic cyanobacterium Microcystis aeruginosa PCC 7806

Microcystins are the most common cyanobacterial toxins found in freshwater lakes and reservoirs throughout the world. They are frequently produced by the unicellular, colonial cyanobacterium Microcystis aeruginosa; however, the role of the peptide for the producing organism is poorly understood. Differences in the cellular aggregation of M. aeruginosa PCC 7806 and a microcystin-deficient Delta mcyB mutant guided the discovery of a surface-exposed protein that shows increased abundance in PCC 7806 mutants deficient in microcystin production compared to the abundance of this protein in the wild type. Mass spectrometric and immunoblot analyses revealed that the protein, designated microcystin-related protein C (MrpC), is posttranslationally glycosylated, suggesting that it may be a potential target of a putative O-glycosyltransferase of the SPINDLY family encoded downstream of the mrpC gene. Immunofluorescence microscopy detected MrpC at the cell surface, suggesting an involvement of the protein in cellular interactions in strain PCC 7806. Further analyses of field samples of Microcystis demonstrated a strain-specific occurrence of MrpC possibly associated with distinct Microcystis colony types. Our results support the implication of microcystin in the colony specificity of and colony formation by Microcystis.     

Different tolerances and responses to low temperature and darkness between waterbloom forming cyanobacterium <Emphasis Type="Italic">Microcystis</Emphasis> and a green alga <Emphasis Type="Italic">Scenedesmus</Emphasis>

The dynamics of planktonic cyanobacteria in eutrophicated freshwaters play an important role in formation of annual summer blooms, yet overwintering mechanisms of these water bloom forming cyanobacteria remain unknown. The responses to darkness and low temperature of three strains (unicellular Microcystis aeruginosa FACHB-905, colonial M. aeruginosa FACHB-938, and a green alga Scenedesmus quadricauda FACHB-45) were investigated in the present study. After a 30-day incubation under darkness and low temperature, cell morphology, cell numbers, chlorophyll a, photosynthetic activity (ETR_max and I _k), and malodialdehyde (MDA) content exhibited significant changes in Scenedesmus. In contrast, Microcystis aeruginosa cells did not change markedly in morphology, chlorophyll a, photosynthetic activity, and MDA content. The stress caused by low temperature and darkness resulted in an increase of the antioxidative enzyme-catalase (CAT) in all three strains. When the three strains re-grew under routine cultivated condition subjected to darkness and low temperature, specific growth rate of Scenedesmus was lower than that of Microcystis. Flow cytometry (FCM) examination indicated that two distinct types of metabolic response to darkness and low temperature existed in the three strains. The results from the present study reveal that the cyanobacterium Microcystis, especially colonial Microcystis, has greater endurance and adaptation ability to the stress of darkness and low temperature than the green alga Scenedesmus. Handling editor: D. Hamilton     

Simple method for a cell count of the colonial Cyanobacterium, Microcystis sp

The cell counting of colonial Microcystis spp. is a rather difficult and error-prone proposition, as this genus forms irregularly-shaped and irregularly-sized colonies, which are packed with cells. Thus, in order to facilitate a cell count, four methods of dividing the colonies into single cells were compared, including vortexing, sonication, TiO2 treatment, and boiling. As a result, the boiling method was determined to generate the greatest number of single cells from a colony, and all colonies were found to have divided completely after only 6 min of treatment. Furthermore, no significant cell destruction, which might alter the actual cell density, was detected in conjunction with the boiling method (P = 0.158). In order to compute the cell number more simply, the relationship between the colony size and the cell number was determined, via the boiling method. The colony volume, rather than the area or diameter was correlated more closely with the cell number (r2 = 0.727), thereby suggesting that the cell numbers of colonial Microcystis sp. can also be estimated effectively from their volumes.