Efectos de extractos de una cianófita (Nostoc muscorum) sobre Sclerotinia sclerotiorum en plántulas de lechuga

p.1-4

New aerophytic morphospecies of Nostoc (Cyanobacteria) from São Paulo State, Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Desarrollo de la metodología para cuantificar la actividad de OCT, arginasa y arginina deaminasa en las células de "Nostoc" PCC 73101

Universidad de Las Palmas de Gran Canaria. Suficiencia investigadora

(Table 1) Effects of desiccation on photochemical processes and nitrogenase activity in Nostoc commune s.l. colonies

The effects of desiccation on photochemical processes and nitrogenase activity were evaluated in Nostoc commune s.l. colonies in situ from a wet thufur meadow at Petuniabukta, Billefjorden, Central Svalbard, during the 2009 arctic summer. The colonies were collected in the fully hydrated state, and were subjected to slow desiccation at ambient temperatures (5 - 8°C) and low light (30 - 80 µmol/m**2/s). For each colony the weight, area, photochemical performance, and nitrogenase activity were determined at the beginning, as well as on every day during the first four days of the experiment; thereafter, on every second day until desiccation was complete. The photochemical performance was evaluated from variable chlorophyll fluorescence parameters (FV/FM, Phi(PSII) , qP, and NPQ), and the nitrogenase activity was estimated by an acetylene-ethylene reduction assay. A significant decrease in the photochemically active area was recorded from the third day, when the colony had lost approximately 40% of its original weight indicating some changes in the extracellular matrix, and stopped on the 14th to 18th day. No effects of the desiccation on the main photochemical parameters (FV/FM, Phi(PSII), qP) were observed up to the sixth to eighth days of desiccation. Slightly lower values of FV/FM and Phi(PSII) recorded in fully-hydrated colonies could be caused by impaired diffusion of CO2 into cells. The steep reduction of photochemical activity occurred between the eighth and tenth day of the experiment, when the colony had lost approximately 80% of its fully-hydrated weight. The nitrogenase activity was highest on the first day, probably due to improved diffusion of N2 into cells, then declined, but was detectable until the sixth day of the experiment. Since Nostoc commune s.l. colonies were capable of photosynthesis and nitrogen fixation to the level of ca. 60% of its fully-hydrated weight, even partly-hydrated colonies contribute substantially to carbon and nitrogen cycling in the High Arctic wet meadow tundra ecosystem.

Evolution and detection of cyanobacterial hepatotoxin synthetase genes

Mass occurrences (blooms) of cyanobacteria are common in aquatic environments worldwide. These blooms are often toxic, due to the presence of hepatotoxins or neurotoxins. The most common cyanobacterial toxins are hepatotoxins: microcystins and nodularins. In freshwaters, the main producers of microcystins are Microcystis, Anabaena, and Planktothrix. Nodularins are produced by strains of Nodularia spumigena in brackish waters. Toxic and nontoxic strains of cyanobacteria co-occur and cannot be differentiated by conventional microscopy. Molecular biological methods based on microcystin and nodularin synthetase genes enable detection of potentially hepatotoxic cyanobacteria. In the present study, molecular detection methods for hepatotoxin-producing cyanobacteria were developed, based on microcystin synthetase gene E (mcyE) and the orthologous nodularin synthetase gene F (ndaF) sequences. General primers were designed to amplify the mcyE/ndaF gene region from microcystin-producing Anabaena, Microcystis, Planktothrix, and Nostoc, and nodularin-producing Nodularia strains. The sequences were used for phylogenetic analyses to study how cyanobacterial mcy genes have evolved. The results showed that mcy genes and microcystin are very old and were already present in the ancestor of many modern cyanobacterial genera. The results also suggested that the sporadic distribution of biosynthetic genes in modern cyanobacteria is caused by repeated gene losses in the more derived lineages of cyanobacteria and not by horizontal gene transfer. Phylogenetic analysis also proposed that nda genes evolved from mcy genes. The frequency and composition of the microcystin producers in 70 lakes in Finland were studied by conventional polymerase chain reaction (PCR). Potential microcystin producers were detected in 84% of the lakes, using general mcyE primers, and in 91% of the lakes with the three genus-specific mcyE primers. Potential microcystin-producing Microcystis were detected in 70%, Planktothrix in 63%, and Anabaena in 37% of the lakes. The presence and co-occurrence of potential microcystin producers were more frequent in eutrophic lakes, where the total phosphorus concentration was high. The PCR results could also be associated with various environmental factors by correlation and regression analyses. In these analyses, the total nitrogen concentration and pH were both associated with the presence of multiple microcystin-producing genera and partly explained the probability of occurrence of mcyE genes. In general, the results showed that higher nutrient concentrations increased the occurrence of potential microcystin producers and the risk for toxic bloom formation. Genus-specific probe pairs for microcystin-producing Anabaena, Microcystis, Planktothrix, and Nostoc, and nodularin-producing Nodularia were designed to be used in a DNA-chip assay. The DNA-chip can be used to simultaneously detect all these potential microcystin/nodularin producers in environmental water samples. The probe pairs detected the mcyE/ndaF genes specifically and sensitively when tested with cyanobacterial strains. In addition, potential microcystin/nodularin producers were identified in lake and Baltic Sea samples by the DNA-chip almost as sensitively as by quantitative real-time PCR (qPCR), which was used to validate the DNA-chip results. Further improvement of the DNA-chip assay was achieved by optimization of the PCR, the first step in the assay. Analysis of the mcy and nda gene clusters from various hepatotoxin-producing cyanobacteria was rewarding; it revealed that the genes were ancient. In addition, new methods detecting all the main producers of hepatotoxins could be developed. Interestingly, potential microcystin-producing cyanobacterial strains of Microcystis, Planktothrix, and Anabaena, co-occurred especially in eutrophic and hypertrophic lakes. Protecting waters from eutrophication and restoration of lakes may thus decrease the prevalence of toxic cyanobacteria and the frequency of toxic blooms.

An evaluation of the trophic status of the Shen Reservoir by an analysis of phytoplankton composition and water characteristics

In an effort to evaluate the production potential of an artificial impoundment, the phytoplankton of the Shen reservoir was sampled from November 1981 to June 1982 at three stations during three periods of distinct seasonal hydrographic characteristics. The samples were subsampled and quantified. Most of the phytoplankton were identified to the species level. There were in all 53 species comprising Chlorophyceae contributing 36.70% with species of Volvox, Pediastrum, Closterium, Staurodesmus and Ankistrodesmus as dominant species in this group. The Cyanophyceae contributed 30.00% with species of Microcystis, Nostoc , and Oscillatoria as the dominant species. An analysis of temporal and spatial changes in composition and abundance of the various groups showed that these were influenced by water temperature, sampling period and station. Based on the trophic status of the most abundant species, the composition of the phytoplankton is indicative of a tropical reservoir with a moderate productivity for fish culture

Constraining the interpretation of 2-methylhopanoids through genetic and phylogenetic methods

Hopanoids are a class of sterol-like lipids produced by select bacteria. Their preservation in the rock record for billions of years as fossilized hopanes lends them geological significance. Much of the structural diversity present in this class of molecules, which likely underpins important biological functions, is lost during fossilization. Yet, one type of modification that persists during preservation is methylation at C-2. The resulting 2-methylhopanoids are prominent molecular fossils and have an intriguing pattern over time, exhibiting increases in abundance associated with Ocean Anoxic Events during the Phanerozoic. This thesis uses diverse methods to address what the presence of 2-methylhopanes tells us about the microbial life and environmental conditions of their ancient depositional settings. Through an environmental survey of hpnP, the gene encoding the C-2 hopanoid methylase, we found that many different taxa are capable of producing 2-methylhopanoids in more diverse modern environments than expected. This study also revealed that hpnP is significantly overrepresented in organisms that are plant symbionts, in environments associated with plants, and with metabolisms that support plant-microbe interactions; collectively, these correlations provide a clue about the biological importance of 2-methylhopanoids. Phylogenetic reconstruction of the evolutionary history of hpnP revealed that 2-methylhopanoid production arose in the Alphaproteobacteria, indicating that the origin of these molecules is younger than originally thought. Additionally, we took genetic approach to understand the role of 2-methylhopanoids in Cyanobacteria using the filamentous symbiotic Nostoc punctiforme. We found that hopanoids likely aid in rigidifying the cell membrane but do not appear to provide resistance to osmotic or outer membrane stressors, as has been shown in other organisms. The work presented in this thesis supports previous findings that 2-methylhopanoids are not biomarkers for oxygenic photosynthesis and provides new insights by defining their distribution in modern environments, identifying their evolutionary origin, and investigating their role in Cyanobacteria. These efforts in modern settings aid the formation of a robust interpretation of 2-methylhopanes in the rock record.

藻胆体的光谱特性和光能传递的研究

藻胆体在低浓度磷酸缓冲溶液中发生解离，我们通过藻胆体在解离过程中荧光发射光谱的变化研究藻胆体中藻胆蛋白之间的光能传递． 1．发菜(Nostoc flagelliforme)藻胆体在0.9M磷酸缓冲溶液中较稳定，其77K荧光发射光谱中只有一个荧光峰F686，属于别藻蓝蛋白-B的荧光峰。当藻胆体在低浓度缓冲溶液中时，荧光峰除了686nm，还出现F648和F666肩，而且F648先于F666肩出现．这说明C-藻蓝蛋白(F'648)所捕获的光能已不能全部传给别藻蓝蛋白-B，并说明藻蓝蛋白与别藻蓝蛋白之间的断裂先于别藻蓝蛋白与别藻蓝蛋白-B之间的断裂。当进一步解离时，主峰仍位于648nm，次峰位于686nm．而666nm荧光肩消失，说明C-藻蓝蛋白所捕获的光能已不能传给别藻蓝蛋白，但能传给别藻蓝蛋白-B．我们因此提出在该藻胆体中藻胆蛋白之间的光能传递途径如下： C一藻红蛋白一C-藻蓝蛋白一别藻蓝蛋白_-别藻蓝蛋白-B 在藻胆体的结构方面，我们提出一部分C-藻蓝蛋白与别藻蓝蛋白相连接，另一部分与别藻蓝蛋白-B相连接． 2．聚球藻( synechococcus leopoliensis 625)藻胆体在0.6M，0.3M和0.1M磷酸缓冲液中解离时，其77K荧光光谱中只出现别藻蓝蛋白-B(F'684)和C-藻蓝蛋白(F655)的荧光峰消长变化，没有出现别藻蓝蛋白(F666)的荧光，我们为此提出在该藻胆体中光能从C-藻蓝蛋白传给别藻蓝蛋白-B有两条途径：一是直接传给别藻蓝蛋白-B，另一是传递给别藻蓝蛋白和别藻蓝蛋白-B的复合物，此复合物在0.lM到0,6M磷酸缓冲液中比较稳定．即： c-藻红蛋白→c—藻蓝蛋白—①别藻蓝蛋白-别藻蓝蛋白-B复合物、②别藻蓝蛋白-B

内蒙古草原地耳和发菜的固氮特性及在氮素循环中的作用

以内蒙古干旱半干旱草原上的固氮蓝藻－普通念珠藻和发状念珠藻为研究对象，对其分布、生态特点、形态结构、生物量及不同条件下的固氮活性进行了研究，并根据降水与气温状况对其年固氮量进行了估算。结果表明：同发状念珠藻相比，普通念珠藻的分布更为广泛，其形态也有较大的差异。普通念珠藻的生物量在以禾本科为主的退化草场和沙地上较大，在pH值偏碱、含水量300％和温度30℃的情况下，其固氮活性较高，达2500nmolC2H2/g•h（干重）以上，年固氮量也高达4.19kg/h.m2，占草原氮素总输出的10％左右。由此可见，在维持干旱半干旱草原与荒漠草原生态系统的氮素平衡中，固氮蓝藻这类低等固氮植物可能起着重要作用。

发菜类囊体膜的组成及其光合特性的研究

发菜（Nostoc flagelliforme Born. et Flah.）的细胞壁由纤维素、半纤维素、糖脂和蛋白质组成。未经破碎的细胞难以进行各种光合特性的研究。由于纯度较高的发菜类囊体膜制备比较困难，对它的光合机理的研究一直是停留在整体水平上进行。我们采用French Press低温下高压破碎细胞，建立了一种快速简便的制备方法。在提取液中加入一定浓度的Ca2+ （Ca2+既有助于维持类囊体膜的放氧活性又可以使类囊体膜在较低的离心速度下使类囊体膜得到凝集沉淀），从而在较短时间内、在高速离心的情况下得到了纯度较高并具有较高放氧活性的发菜类囊体膜。在此基础上，我们采用改进的Allen（1991）的温和绿胶系统，首次对陆生蓝藻发菜类囊体膜色素蛋白复合体进行了分离，共分离出了11条绿色的色素蛋白复合物条带和两条浅黄色的条带。7条绿色的色素蛋白复合物条带属于PSI组分，4条绿色的蛋白复合物条带属于PSII组分，其中一条浅黄色条带系未被报道过的新的色素蛋白复合物条带，经其光谱性质的分析初步鉴定为类胡萝卜素蛋白复合物，此复合物的分离有助于解释发菜独特的适应荒漠、半荒漠地带高光辐射的特性。 本文还对干燥状态、复水30分钟后和复水生长24小时后的野生发菜及人工培养的发菜藻丝体膜脂及其脂肪酸组成进行了分析。发菜的膜脂由MGDG、MGDG、SQDG和PG组成，其酯酰基部位连接有16:0、16:1、18:0、18:1、18:2和18:3六种脂肪酸。野生发菜中具有高含量的不饱和脂肪酸，其含量可达总脂的73%，其中16:1和18:3分别达到28.9mol%和34.3mol%，远远高于已报道的其它蓝藻，所以我们推测发菜具有极强的抗逆性和其膜脂不饱和程度密切相关。分析不同处理的发菜的膜脂和脂肪酸组成表明，复水对野生发菜的膜脂及其脂肪酸组成没有显著影响，说明发菜的膜脂和脂肪酸组成在干燥状态下能保持很高的稳定性。从野生发菜分离出的藻丝体在25 ℃条件下培养，其膜脂脂肪酸组成发生了显著变化，主要表现为脂肪酸的不饱和程度的大幅度降低，18:3从34.9mol%降低到8.6mol%，16:1从28.9mol%降低到13.9mol%。上述结果表明了发菜具有极强的通过改变其膜脂的脂肪酸组成而适应生存环境的能力。

发状念珠藻光合特征及其在复水－失水过程中能量传递变化的研究

摘要 "发状念珠藻（Nostoc flagelliforme Born. Et Flah.），俗名发菜，是生长于干旱、半干旱土壤表面的陆生蓝藻，具有极强的抗旱能力。发菜光合作用方面的研究大多处于整体细胞水平，且研究手段非常有限。本实验对发菜光合特征进行深入研究，并探讨了发菜在干湿交替过程中能量传递的变化情况。 叶绿素和藻胆素是发菜细胞中两种主要的光合色素。发菜复水后光合活性完全恢复时，在室温（20℃）或低温（77K）下，其绝大部分的荧光是由于藻胆素被激发而产生。在室温下，大部分荧光来自藻胆体;当叶绿素被激发后，产生的荧光非常微弱。在低温下，藻胆素被激发后，荧光发射光谱中可分辨出藻胆蛋白、光系统Ⅰ和光系统Ⅱ的发射峰;叶绿素被激发后，荧光发射光谱包括光系统Ⅰ和光系统Ⅱ的荧光。相比之下，室温荧光发射光谱不适于用做发菜细胞光合作用方面的研究。 我们设计了一种新方法，从野生发菜细胞中分离得到类囊体膜及细胞质膜，并对其性质进行分析。发菜细胞外复杂的胶质结构使得现有破碎其它蓝藻细胞的方法无法破碎发菜细胞。通过实验发现，联合使用细胞破碎仪和毛地黄皂甙（0.3%）可有效破碎发菜细胞;并且毛地黄皂甙在低浓度下（≦0.5%），对色素与蛋白的结合不会造成破坏作用。随后，通过蔗糖密度梯度离心可将细胞质膜与类囊体膜分离。发菜类囊体膜的光谱性质与其它蓝藻相似。细胞质膜除结合有类胡萝卜素外，还结合有少量叶绿素前体。类囊体膜和细胞质膜膜脂及脂肪酸组成相似。其中，十六碳烯酸[16：1(9)]和亚麻酸[18：3(9,12,15)]是含量最高的两种脂肪酸，分别占总脂肪酸含量的三分之一左右。高含量的多不饱和脂肪酸可能和发菜极强的抗旱能力有关。 我们首次对发菜捕光色素蛋白复合物－藻胆体的组成和结构进行分析。发菜藻胆体为“3核＋6杆”的半圆盘结构。组成藻胆体的藻胆蛋白包括藻蓝蛋白和别藻蓝蛋白。两个藻蓝蛋白六聚体通过连接肽组成藻胆体的“杆”结构。在“杆”结构中等量分布着两条连接肽（分子量分别为29kDa和34kDa）为杆连接肽和核杆连接肽。而“核”结构中核膜连接肽的分子量为103kDa。 发菜在无霜期，几乎每天经历一次复水－干燥过程：夜间的结露使发菜在黑暗中复水，而清晨太阳升起后，在光照下迅速失水进入休眠状态。我们研究了发菜在黑暗中的复水过程及在光照下失水过程中藻胆体与光系统能量传递的变化情况。发菜在黑暗中复水后，光系统Ⅱ活性无法恢复。藻胆体内及藻胆体与光系统Ⅰ的能量传递在5分钟内恢复;而藻胆体与光系统Ⅱ的能量传递只能部分恢复。我们设想，发菜在复水过程中通过双扳机－水和光－控制光合活性的恢复，以及在黑暗中部分恢复藻胆体与光系统Ⅱ的能量传递，将减少不必要的能量消耗与通过光合作用储备尽可能多的化学能－这两个生存策略有机的结合起来。发菜在光照下的失水过程中，光合活性在含水量降至90％前基本保持稳定，随后迅速下降。而在含水量达到150％后，藻胆体内的能量传递便开始受到抑制，并且随着含水量的降低，该抑制现象逐步加剧。这样，发菜在干燥过程中，通过抑制藻胆体内的能量传递，减少了传递到光系统Ⅱ反应中心的能量，从而避免了在光合活性下降过程中过剩光能对光系统Ⅱ产生的破坏作用。"

阳光紫外辐射对室内水培发状念珠藻生理特性的影响

发状念珠藻(Nostoc flagelli forme Bornet & Flahault)是一种重要的陆生经济蓝藻,室内培育出的原植体如何适应阳光辐射的问题尚需探讨。为此,作者将室内水培发菜置于阳光下培养,测定了其生长、有效光化学效率(ΔF/F′m)和色素的变化。结果表明,较高的可见光(PAR,395—700nm)和紫外辐射(UVR,280—395nm)均导致水培发菜的ΔF/F′m下降。第1天中午,PAR和UVR分别使ΔF/F′m下降了54%和13%;傍晚,ΔF/F′m有部分恢复。UVR对发菜适应阳光2

不同念珠藻的提取物对酪氨酸酶活性的抑制作用

研究了Nostoc sp.FACHB87、Nostoc paludosum FACHB89等10株念珠藻的甲醇提取物对酪氨酸酶的抑制作用。结果表明,随着提取物浓度的升高,对酪氨酸酶的抑制作用也增强。其中Nostoc sp.FACHB892的甲醇提取物对酪氨酸酶抑制作用最强,在33.3μg/mL的最高测试浓度下,抑制率可达到60.54%;Nostoc sp.FACHB95、Nostoc sp.FACHB106、N.punctiforme FACHB252、N.calcicola FACHB389和N.par

葛仙米对丁草胺胁迫的响应

研究了丁草胺对可食用蓝藻葛仙米(Nostoc sphaeroides)生理和代谢活性的影响．用不同浓度的丁草胺处理葛仙米,结果显示低浓度(5 mg·L-1)丁草胺使其光合作用、呼吸作用和光合系统Ⅱ活性增强,高浓度丁草胺(>5 mg·L-1)限制其光合作用、呼吸作用和光合系统Ⅱ活性．同时丁草胺对葛仙米膜结构和功能具有破坏作用,随着丁草胺处理浓度增大,质膜透性不断增大,丙二醛和超氧自由基阴离子含量升高；在低浓度丁草胺处理时,类胡萝卜素含量增加,超氧化物歧化酶(SOD)活性增强,高浓度丁草胺处理时,类胡萝卜素含

3种念珠藻多糖对自由基的清除作用

研究了地木耳(Nostoc commune)、葛仙米(N.sphaeroides)和发菜(N.flagelliform e)3种念珠藻多糖对自由基的清除作用。结果表明,3种念珠藻多糖对超氧阴离子自由基和羟自由基具有很强的清除作用,且呈量效关系,地木耳清除超氧自由基作用最强,最高清除率达72.3%,葛仙米和发菜分别为46.7%、35.5%;发菜清除羟自由基效果最强,最高清除率达74.3%,葛仙米和地木耳清除率分别为49.0%、46.7%;3种念珠藻多糖对DPPH自由基的清除作用不显著。