有害甲藻Stoeckeria algicida在辽东湾的时空分布

Stoeckeria algicida为甲藻纲胸甲球藻科,有侵噬鱼类细胞杀鱼的能力,可导致鱼类成群死亡,同时也会杀死其他海洋微藻。由于该藻个体微小、形态学鉴定困难,研究较为迟缓,我国海域几乎没有该藻的研究报道。近几年,高通量测序技术的发展极大地推动了微型/微微型浮游植物的鉴定研究,为了解我国辽东湾海域是否存在Stoeckeria algicida及其分布情况,以18S rD NA V4区作为目标基因,结合高通量测序技术,专门设计了微型/微微型浮游植物鉴定引物对V4(F/R),随后对辽东湾2014年四季海水中微型和微微型浮游植物多样性进行了检测。结果发现,Stoeckeria algicida除了春季未检出外,其他季节均有检出,温度是影响该藻繁殖的主要因素。虽然Stoeckeria algicida在整个环境样品中优势度不太明显,但其夏季密度较高(最高达2.753×10~3个/L),高值区主要分布在辽东湾东西两岸,致灾风险较高,应引起有关方面足够重视。Stoeckeria algicida在我国海域首次报道,其危害后果严峻,必须加强监测监管。     

利用真核微藻表达外源蛋白的现状与展望

正近年来,随着基因工程技术的迅猛发展,利用真核微藻表达外源蛋白技术得到广泛关注。综述了真核微藻表达外源重组蛋白的研究现状,并分析了其中存在的关键科学问题。在此基础上,阐述了真核微藻作为新型表达系统所面临的机遇与挑战,最后展望了真核微藻表达重组蛋白的发展前景。     

真核微藻蓝光受体及其功能研究进展

真核微藻是一类能够进行光合作用的真核生物,起源于内共生事件,因种类繁多、分布广泛及进化历史复杂等特点,使其成为逆境生理研究的理想实验材料。光是环境中重要的信号因子之一,不仅为真核微藻提供能量,而且还为它们提供信息,调节其生长发育过程。为适应光强、光质及光周期等光信息,真核微藻在漫长的进化过程中形成了一套精细的光信号接收和转导系统。光受体在光信号通路中发挥重要的作用,起着接收和转化的桥梁作用。按照光信号波段的不同,目前已知光受体分为红光受体、蓝光受体、绿光受体及紫外光受体。蓝光受体能感受蓝光和近紫外光波段(320-400 nm),在调控植物体的多种生理过程中发挥重要的作用。以高等植物拟南芥中蓝光受体结构及功能研究进展为参照,总结了真核微藻蓝光受体研究进展,重点关注真核微藻蓝光受体基因克隆、蛋白结构、分子进化、光化学特性、生理功能及光信号转导等方面,并提出了今后真核微藻蓝光受体研究工作应注意的问题和关注的重点,以期为真核微藻蓝光受体的研究提供参考。     

长山群岛海域真核微藻粒级结构及扇贝摄食选择性

采用高通量测序-分子鉴定分级技术于2019年对长山群岛全海域真核微藻粒级结构进行了研究。结果发现,春季以中（47%）、小粒级（41%）为主,夏季以小（39%）、大粒级（38%）为主,秋季以大粒级（60%）为主,春、夏、秋季小、中、大粒级微藻比例为42：47：11、39：23：38、22：18：60。小粒级微藻优势种为细小微胞藻（Micromonas pusilla）、融合微胞藻（Micromonas commoda）和金牛微球藻（Ostreococcus tauri）,中粒级微藻优势种为剧毒卡尔藻（Karlodinium veneficum）、大粒级微藻优势种为柔弱几内亚藻（Guinardia delicatula）、平野亚历山大藻（Alexandrium hiranoi）、多纹膝沟藻（Gonyaulax polygramma）,综合整个真核微藻群落,春季由中粒径的剧毒卡尔藻占据优势（23.9%）,夏季由大粒径的平野亚历山大藻占据优势（29.4%）,秋季由大粒径的多纹膝沟藻占据优势（66.8%）,有毒甲藻在该海域中占有绝对优势,贝毒累积风险较高,小粒径微藻中金牛微球藻和抑食金球藻曾在渤海引发褐潮,潜在威胁该海域贝类养殖业。虾夷扇贝对小粒级和大粒级微藻的选择性较低,对中粒级微藻的选择性较高,尤其对水体中优势种剧毒卡尔藻一直表现出主动选择。光学需氧量、无机氮、溶解氧、石油类及部分重金属Cd、As、Hg影响着整个长山群岛海域真核微藻粒级结构时空演变。     

环境样本中微型和微微型浮游植物高通量测序的引物优化

分别以18Sr DNA的V4区和V9区为目标基因,采用高通量测序平台和生物信息学方法,分析海水样品中微型和微微型浮游植物多样性。利用在线分析软件对V4(F/R)、V9(F/R)和C4(F/R)3对引物的敏感性、特异性进行了评估和比较,发现自行设计的引物V4(F/R)对真核藻类的扩增特异性高于V9(F/R)和C4(F/R)。高通量测序结果显示,检测样品平均获得68834条原始序列,高质量数据占99%以上,获得基因注释的序列数达94%以上。3对引物V4(F/R)、V9(F/R)、C4(F/R)鉴定的平均微型/微微型浮游植物OTUs数分别为78、42、58,引物V4(F/R)鉴定效率相对较高,同时对细小微胞藻(Micromonas pusilla)、(金牛微球藻Ostreococcus tauri)、密球藻(Pycnococcus provasolii)、抑食金球藻(Aureococcus anophagefferens)、赤潮异弯藻(Heterosigma akashiwo)等优势种检出频率高于引物V9(F/R)。相对已发表的2对引物,设计的引物V4(F/R)对海洋微型和微微型藻类多样性检测更为高效。     

水东湾海域浮游植物潮汐分布特征及其与环境因子的关系

2014年秋、冬两季,每个季节在大潮期和小潮期对水东湾海域浮游植物群落结构和环境因子进行了调查,共鉴定出4门57属134种。其中硅藻门42属106种,占浮游植物种类数的79.1%;甲藻门13属26种,占浮游植物种类数的19.4%;蓝藻门1属1种,占浮游植物种类数的0.8%;针胞藻纲1属1种,占浮游植物种类数的0.8%。优势种15种,主要为尖布纹藻Gyrosigma aluminatum、圆海链藻Thalassiosira rotula、中肋骨条藻Skeletonema costatum、丹麦细柱藻Leptocylindrus danicus和舟形鞍链藻Campylosira cymbelliformis等。4个航次共有种类数在18—40种,Jaccard种类相似性指数范围在0.200—0.404,多样性指数和均匀度平均值分别为2.60和0.60。秋季大、小潮期多样性指数差异较显著(P0.05),冬季细胞丰度、多样性指数和均匀度大、小潮期均无明显差异。浮游植物细胞丰度变化范围为0.95×10~4个/L—28.0×10~4个/L,平均为6.86×10~4个/L,平均丰度冬季小潮期(9.46×10~4个/L)秋季小潮期(7.56×10~4个/L)冬季大潮期(5.97×10~4个/L)秋季大潮期(4.44×10~4个/L)。主成分分析(PCA)表明:盐度和营养盐可能是影响水东湾海域生态环境的主导因子。对水东湾海域浮游植物群落结构与主要环境因子进行Spearman相关性分析,细胞丰度与盐度在秋季大、小潮期为负相关,在冬季大、小潮期呈显著正相关;与无机氮和磷酸盐在冬季大、小潮期呈极显著负相关,在秋季大、小潮期均无相关性。冬季小潮期水温与多样性指数、均匀度和细胞丰度均呈正相关;从测定结果来看浮游植物细胞丰度、多样性指数和均匀度与叶绿素a含量均无统计学意义上的相关性。     

湛江港湾潜在赤潮生物的时空分布及其影响因素

2009年2月(冬)、5月(春)、8月(夏)和11月(秋)分别对湛江港湾的浮游植物和环境因子进行了调查,并对该海域潜在赤潮生物的时空分布及其影响因素进行了分析。结果表明,湛江港湾海域潜在赤潮生物共有126种,其中硅藻门26属85种,占潜在赤潮生物种类数的67.5%,甲藻门14属36种,占潜在赤潮生物种类数的28.6%,蓝藻门1属2种,占潜在赤潮生物种类数的1.6%,金藻门2属2种,占潜在赤潮生物种类数的1.6%,针胞藻纲1属1种,占潜在赤潮生物种类数的0.8%。种类数以春季最多,达89种,秋季次之,为71种,夏季61种,冬季最少,仅有58种。细胞丰度在12.36×104～43.55×104cells·L-1,春季最高,夏季次之,冬季最低,从各主要优势种的细胞丰度和细胞大小判断,该海域潜在赤潮生物的丰度均未达到赤潮发生的阈值。4季均出现的种类共有31种,季节相似性指数在0.43～0.50。优势种共有19种,全为硅藻,没有观测到甲藻。优势度最大的种类主要有浮动弯角藻(Eucampia zoodiacus)、旋链角毛藻(Chaetoceros curvisetus)和中肋骨条藻(Skeletonema costatum),没有全年优势种。中肋骨条藻、旋链角毛藻和冰河拟星杆藻(Asteri-onellopsis glacialis)为3季优势种。分析表明,潜在赤潮生物细胞丰度与叶绿素a、水温、盐度和pH值存在着极显著的正相关,与DIN和SiO32-存在着极显著的负相关,与PO43-不存在显著相关关系。     

莱州湾金城海域网采浮游植物年际变化及与环境因子的关系

为了解莱州湾金城海域浮游植物群落年际变化特征,在该海域设置了15个监测站位,于2009年和2010年采用浅水Ⅲ型浮游生物网逐月连续进行24个航次浮游植物调查,同步监测其他环境因子。浮游植物样品经鲁格氏溶液固定后在OLYMPUS BX_(51)显微镜下进行分类、鉴定、计数,共鉴定出浮游植物81种,隶属于硅藻门、甲藻门、黄藻门和金藻门4门25科43属。其中硅藻16科33属67种,占种类数的82.7%;甲藻6科7属11种,占种类数的13.6%;黄藻2科2属2种,占种类数的2.5%;金藻1科1属1种,占种类数的1.2%。2009年和2010年金城海域浮游植物种类组成和比例基本一致,浮游植物丰度年际变化和种类数年际变化均呈现明显的双峰模型,每年2月和9月为高峰期,5月为全年最低值。2009年和2010年细胞丰度均值分别为465×10~4(38.4×10~4—1351×10~4)个/m~3和457×10~4(41.8×10~4—1380×10~4)个/m~3,均低于1982—1983年平均水平。浮游植物群落年际相似性在60%水平聚类分为四个类群,其中Ⅰ类为5月份样品,优势种为夜光藻和洛氏角毛藻;Ⅱ类为11—4月份样品,优势种有中肋骨条藻、旋链角毛藻和夜光藻;Ⅲ类为8、9、10月份样品,优势种为洛氏角毛藻和丹麦细柱藻;Ⅳ类为6、7月份样品,优势种为洛氏角毛藻和尖刺拟菱形藻。浮游植物群落Shannon-Winner多样性指数平均2.715 (1.446—3.807),Margalef丰富度指数平均2.418 (1.545—3.153),Pielou均匀度指数平均0.523 (0.263—0.706)。2009年和2010年多样性指数和均匀度高峰值出现在3月和10月,低值出现在7月和9月;均匀度指数年际变化低值出现在6—9月。浮游植物群落结构主要受水温和盐度环境因素的影响,其中水温与洛氏角毛藻丰度呈极显著正相关(R=0.613,P0.01),与旋链角毛藻、中肋骨条藻和夜光藻丰度呈极显著负相关(R=-0.662,-0.649,-0.649,P0.01);化学需氧量与旋链角毛藻和中肋骨条藻均呈显著负相关(R=-0.589,-0.480,P0.05);盐度与中肋骨条藻呈极显著负相关(R=-0.585,P0.01);其他环境因子与各优势种丰度相关性不明显。     

中国东部陆架边缘海网采浮游植物种类组成和季节变化

为揭示中国东部陆架边缘海浮游植物群落季节变化规律,根据2006年6–7月、2007年1–2月、2007年11月和2009年4–5月在中国东部陆架边缘海域(25.00°–39.00°N,118.00°–129.00°E)进行的综合采样调查,对调查海域网采浮游植物(网孔直径77μm)的物种多样性和分布特征进行了研究。4个航次共鉴定出浮游植物4门70属257种(不包括未定名种),其中硅藻是主要功能群,其次是甲藻,主要的优势种为骨条藻(Skeletonema spp.)、细弱海链藻(Thalassiosira subtilis)、囊状海链藻(T.scrotiformis)、伏氏海线藻(Thalassionema frauenfeldii)、菱形海线藻(T.nitzschioides)、具槽帕拉藻(Paralia sulcata)、洛氏角毛藻(Chaetoceros lorenzianus)、旋链角毛藻(C.curvisetus)、尖刺伪菱形藻(Pseudo-nitzschia pungens)和夜光藻(Noctiluca scintillans)。浮游植物细胞丰度为0.02×104–31,350.21×104cells/m3,最低值出现在冬季黄海海域,最高值出现在春季长江口邻近海域。4个季节的浮游植物细胞密度呈现春季夏季秋季冬季的趋势,浮游植物各生物多样性指数的等值线均呈现西北–东南走向。     

2012年夏季挪威海和格陵兰海浮游植物群落结构的色素表征

对2012年中国第5次北极科学考察期间的挪威海和格陵兰海两个断面的光合色素进行了高效液相色谱（HPLC）分级分析,通过藻类色素化学分类分析软件（CHEMTAX）获得了不同浮游植物类群对叶绿素a的贡献,进而得到该海域表层和次表层（30 m）的浮游植物群落结构。结果表明：表层总叶绿素a的浓度为23.59 ng/L,低于次表层的30.38 ng/L,其中浮游植物根据粒径划分对总叶绿素a的贡献由高到低依次是微型浮游植物、小型浮游植物和微微型浮游植物。该海域同时存在葱绿叶绿素（Prasino）、墨角藻黄素（Fuco）、别藻黄素（Allo）、多甲藻素（Perid）、玉米黄素（Zea）、19-丁墨甲藻黄素（19’BF）和19-六已墨甲藻黄素（19’HF）等色素,其浓度和分布与温盐和营养盐等环境因子存在一定的相关性。不同粒径浮游植物色素组成显示,微微型浮游植物群落中以S型定鞭藻（28%）、N型定鞭藻（21%）、硅藻（18%）和青绿藻（12%）占优;微型浮游植物群落的优势类群为S型定鞭藻（53%）、N型定鞭藻（20%）和硅藻（12%）;而小型浮游植物群落主要为硅藻（63%）和甲藻（17%）。     

我国近海赤潮多发区域及其生态学特征

根据有关的研究资料,分析我国近海赤潮多发区分布、主要赤潮生物种类、环境状况及赤潮发生生态学特点.结果表明,我国辽东湾、渤海湾、莱州湾、大连湾、长江口、舟山海域、杭州湾、厦门湾、柘林湾、大鹏湾、珠江口等为赤潮多发区.引发赤潮的因素较多,它与气象、水动力、营养盐及生物环境的变化密切相关,人类活动(如海水养殖、陆源污水排放等)影响加剧近海富营养化是引发赤潮的重要因素;但富营养化并非发生赤潮的唯一条件,低营养海区也可能发生赤潮.目前对诱发赤潮的关键因子及赤潮发生机理,应加强定点长期监测和开展深入研究.     

Resolving phytoplankton taxa based on high-throughput sequencing during brown tides in the Bohai Sea,China

Large-scale blooms formed by pico-sized phytoplankton, which strongly inhibited feeding activity and growth of cultured scallops, have been recorded along the coast of Qinhuangdao in the Bohai Sea since 2009. Based on pigment profiles and clone library analysis of phytoplankton samples during the blooms, the major bloom-forming species was primarily identified as pelagophyte Aureococcus anophagefferens Hargraves et Sieburth, the causative species of intensive brown tides in the United States and South Africa. Due to the indistinct morphological features of the bloom-forming microalgae and limited knowledge on the composition of phytoplankton communities, there were still disputes concerning the causative species of the blooms. In this study, the method of high-throughput sequencing targeted 18S rDNA V4 region was used to study the composition of pico- and nano-sized phytoplankton communities in 2013 and 2014. A total of 18 groups of eukaryotic microalgae at the class level and more than 2000 operational taxonomic units (OTUs) were identified in phytoplankton samples collected from the brown-tide zone in the Qinhuangdao coastal waters. For both years, A. anophagefferens was the most dominant species during the bloom period and its maximum relative abundance exceeded 60 percent. Along with other evidence, the results further confirm that A. anophagefferens is the major causative species of the pico-sized phytoplankton blooms in the Bohai Sea. The outbreak of brown tide exhibited a strong inter-annual variation between 2013 and 2014, and an increasing dominance of dinoflagellates could be observed in the Qinhuangdao coastal waters.     

EFFECTS OF NUTRIENT ENRICHMENT ON NATURAL POPULATIONS OF THE BROWN TIDE PHYTOPLANKTON AUREOCOCCUS ANOPHAGEFFERENS (CHRYSOPHYCEAE)1

The brown tide picoalga Aureococcus anophagefferens Hargraves et Sieburth was present in approximately equal numbers in 12 large scale (13,000 L) mesocosms at the start of a nutrient addition experiment in June 1985. Increases in abundance in untreated systems mimicked the pattern of bloom development in Narragansett Bay, Rhode Island, the seawater source for the experiment. Aureococcus increased to maximal values of 2.6 × 10⁹ cells. L^?1 and persisted at high numbers (10⁸ cells·L^?1) for 7–8 weeks. In nutrient addition tanks, the picoalgae bloomed briefly (1–3 weeks) but rapidly declined to the usual level (～10⁷ cells·L^?1) for eukaryotic algae in Narragansett Bay. The decline in picoalgae abundance was followed by an increase in total diatoms in all nutrient treated tanks. Mean picoalgae abundance in the mesocosms and the bay was significantly (P < 0.05) and inversely correlated (r =–0.93) with mean concentration of dissolved inorganic nitrogen. The persistence of the brown tide species in control mesocosms and Narragansett Bay appears related to its ability to grow at very low concentrations of dissolved inorganic nitrogen, levels previously shown to limit diatom growth.     

VARIATION IN DCMU-ENHANCED FLUORESCENCE RELATIVE TO CHLOROPHYLL A: CORRELATION WITH THE BROWN TIDE BLOOM1

DCMU–enhanced fluorescence and extracted chlorophyll a were simultaneously measured in Narragansett Bay, Rhode Island and the MERL (Marine Ecosystems Research Laboratory) mesocosms during the 1985 brown tide bloom. Marked differences in the relationship between these variables were observed as the phytoplankton community shifted from dominance by picoalgae to diatoms. The fluorescence to chlorophyll a ratio was significantly (P< 0.05) higher in the mesocosms and the bay when the brown tide species (Aureococcus anophagefferens Hargraves et Sieburth) dominated the phytoplankton community compared with other taxa. Although several factors could have affected the relationship we believe the high ratios are related to the pigment composition and/or small size of the brown tide organism.     

Bait worm packaging as a potential vector of invasive species

Invasive species have become an increasingly greater concern for the ecological health of coastal ecosystems, yet vectors of these introductions often are unclear. This project evaluated the potential for the brown seaweed Ascophyllum nodosum ecad scorpiodes (Hauck) Reinke, packaged with bait worms (Nereis virens) harvested from the coast of Maine (USA), as a vector of invasive marine fauna and flora. Often, the seaweed and contents of the bait boxes are discarded into the water by recreational fishermen after using the bait worms, and any included non-native species may then be introduced. Bait boxes were purchased from several commercial vendors in Connecticut and New York over a two-year period. Subsamples of the seaweed were placed in laboratory culture and the growth of associated macro- and microalgae was monitored. Marine invertebrate species present in the samples were also identified and quantified. Results indicated 13 species of macroalgae and 23 species of invertebrates were associated with baitboxes. Among the highly diverse microbial assemblage detected, two species of potentially toxic marine microalgae, Alexandrium fundyense Balech and Pseudonitzschia multiseries (Hasle) Hasle, were found both prior to and after incubation at various temperatures, indicating these harmful algae are brought to and can survive in receiving waters. These findings highlight the need to consider alternative choices of bait box packaging materials or appropriate disposal methods of the seaweed in order to minimize the transport of species which are not native to the receiving coastal waters.     

赤潮藻毒素生物合成研究进展

合成毒素是赤潮藻类的一个常见特征,已知能够产生毒素的微藻有70多种。作为次级代谢产物,藻毒素的产生可能是一种压制或清除其它藻类竞争者的一种反应,在群落演替、种间竞争中发挥重要作用。目前,人们对藻毒素生物合成机理依然知之甚少,相关基因的研究仍无明显突破。利用环境因子诱导毒素生成变化进而分离差异表达基因或者比较不同产毒藻株间基因表达的差异,从中克隆藻毒素生物合成基因似乎是一种极具潜力的研究方向。     

Multiple microalgal partners in symbiosis with the acantharian Acanthochiasma sp. (Radiolaria)

Acantharia (Radiolaria) are widespread and abundant heterotrophic marine protists, some of which can host endosymbiotic eukaryotic microalgae. Although this photosymbiotic association was first described at the end of the 19th century, the diversity of the symbiotic microalgae remains poorly characterized. Here, we examined the identity of the microalgae associated with the acantharian species Acanthochiasma sp. by sequencing partial 18S and internal transcribed spacer (ITS) ribosomal DNA genes from cultured symbionts and directly from isolated holobiont specimens. Single Acanthochiasma cells contained multiple symbiotic partners, including distantly related dinoflagellates (Heterocapsa sp., Pelagodinium sp., Azadinium sp. and Scrippsiella sp.) as well as a haptophyte (Chrysochromulina sp.). This original association of multiple symbiotic microalgae within a single host cell raises questions about the specificity and functioning of the relationship. These microalgae exhibit the common ecological feature of being abundant and widely distributed in coastal and oceanic waters, some occasionally forming extensive blooms. Some of the microalgal genera found in association with Acanthochiasma (i.e. Pelagodinium and Chrysochromulina) are known to occur in symbiosis with other heterotrophic protists such as Foraminifera and other Radiolaria, whereas Heterocapsa, Scrippsiella and Azadinium have never previously been reported to be involved in putative symbiotic relationships. The unusual association unveiled in this study contributes to our understanding of the ecological and evolutionary significance of photosymbiosis in Acantharia and also provides new insights into the nature of such partnerships in the planktonic realm.     

Emergence of brown tides caused by Aureococcus anophagefferens Hargraves et Sieburth in China

Large-scale blooms suspected to be “brown tides” occurred in early summer for three consecutive years from 2009 to 2011 in the coastal waters of Qinhuangdao, China, and had significant negative impacts on the shellfish mariculture industry. To identify the causative species of the blooms, phytoplankton samples were collected from regions with and without bloom in the coastal waters of Qinhuangdao in 2011, and clone libraries were built using eukaryote-specific 18S ribosomal RNA gene (18S rDNA). Altogether 50 clones, including 17 clones from bloom area and 33 clones from nearby regions without bloom were amplified. Blasted in GenBank, 17 clones amplified from the bloom area were assigned to Pelagophyceae (8 clones), Mediophyceae (2 clones), Cryptophyta (2 clones), Dinophyceae (2 clones) and unidentified eukaryotic species (3 clones). Those from the non-bloom site were assigned to Cryptophyta, Eustigmatophyceae, Prasinophyceae, Coscinodiscophyceae, Mediophyceae, Raphidophyceae and Dinophyceae, but not Pelagophyceae. All 8 pelagophyte clones from the bloom area were 99.7–100% similar to a single species, Aureococcus anophagefferens Hargraves et Sieburth, the causative species of brown tides on the east coast of USA. For nearly the entire length of the 18S rDNA, there were 0–6 base pair differences between the 8 amplicons and those of A. anophagefferens from USA. Furthermore, all of the 8 clones were clustered into the same well-supported clade with A. anophagefferens (posterior probability = 0.99) in a phylogenetic tree established for pelagophytes and other related microalgae. In our previous studies, the causative species of the bloom was tentatively identified as a pelagophyte, haptophyte or silicoflagellate, based on the pigment profile of the size-fractioned phytoplankton samples. Based on this study, we conclude that blooms in the coastal waters of Qinhuangdao of the Bohai Sea were brown tides caused by A. anophagefferens. As far as we know, this is the first report of brown tide events caused by A. anophagefferens in China, which is the third country in the world reporting A. anophagefferens blooms in addition to USA and South Africa.