大叶藻(Zostera marina L.)PSⅠ和PSⅡ复合物的分离鉴定

采用蔗糖密度梯度离心法分离纯化大叶藻类囊体膜 ,经 1 0 %SDS增溶后 ,用蔗糖密度梯度超速离心分离其色素蛋白质复合物。经稳态光谱分析、DCIP光还原活性测定及P680 、P70 0 差示光谱检测结果表明 ,2 0 %蔗糖层的CP3和 40 %蔗糖层 (上 )的CP4为PSⅡ复合物 ,具有光化学活性 ;40 %蔗糖层 (下 )的CP5为PSⅠ复合物 ,其P70 0 特征吸收峰位于 695nm处。CP3和CP4的DCIP光还原活性 :CP3为 34.2 7微电子当量 /(mgchl·h) ,CP4为 7.2 9微电子当量 /(mgchl·h)。

大叶藻(Zostera marina)的热值和元素含量(碳、氮、磷)及其相关性

选取辽宁獐子岛近岸海域大叶藻样品,对其须根、根状茎和不同生长阶段叶片的灰分含量、热值和元素含量(碳、氮、磷)及其相关性进行研究,结果表明:在不同生长阶段,叶片长度随着生长顺序先增大后减小;灰分含量随着叶片生长逐渐升高,且极显著小于根状茎和须根(P<0.01);不同部位干质量热值差异极显著(P<0.01),以初生叶最高,为(14.89±0.040)k J/g,后逐渐减小,在须根中最小,为(8.15±0.079)k J/g;不同生长阶段叶片的去灰分热值差异不大,但极显著大于须根和根状茎(P<0.01);碳、氮、磷含量均呈同一趋势:初生叶>幼叶>成熟叶1>成熟叶2>衰老叶>根状茎>须根;N/P比值为27.08,说明大叶藻存在P限制;N/P比值在根状茎中最大,须根中最小,这说明大叶藻存在能量的转移和存贮;大叶藻不同部位灰分含量与干质量热值、去灰分热值、碳含量、氮含量、磷含量之间呈极显著的负相关(P<0.01),干质量热值与碳含量、氮含量、磷含量之间呈极显著的正相关(P<0.01);大叶藻不同部位的热值与碳、氮、磷元素含量之间均呈显著的线性关系(P<0.05)。

Influence of Temperature and Salinity on Germination of Eelgrass (Zostera marina L.) Seeds

Seagrass restoration as part of ocean ecosystem protection has been launched for many years all over the world, but intensive research on this subject in China has just begun in recent years. Seed broadcasting has been widely accepted as the most potentially useful method for seagrass restoration over large areas. We examined the influence of key environmental factors on seed germination to help promote eelgrass bed restoration. Under anoxic conditions, the influence of temperature and salinity on the germination rate of eelgrass (Zostera marina L.) seeds was examined at different combinations of four temperatures (4, 9, 14, and 24℃) and nine salinities (5 to 45, increment of 5). The effect of significant interaction of temperature and salinity on germination rate was observed (ANOVA) (P<0.001). The highest germination rate (83.3 ± 3.5)% was reached in 8 weeks at 14℃ and salinity 5. Higher temperature significantly increased the germination rate at salinity 5 (P<0.001) during the whole observation period except for 24℃, while lower salinity significantly increased the germination rate at 14℃ (P<0.001). Although significant interaction was found between temperature and salinity (P<0.001), the influence of salinity was stronger than that of temperature for the germination of eelgrass seeds. These results provide useful information for the propagation of artificial seedlings for seagrass restoration in China.

Generation and analysis of expressed sequence tags from the salt-tolerant eelgrass species, Zostera marina

Zostera marina, a monocotyledonous angiosperm, is one of the most important seagrass species. To inves- tigate the salt-tolerance mechanism and discover salt-tolerant genes in Z. marina, a cDNA library was con- structed. Single-pass sequencing of the 5＇ ends of 4 081 clones yielded 4 002 high quality expressed sequence tags （ESTs）, which were assembled into 241 contigs and 1 673 singletons, representing 1 914 unigenes. The average length of the ESTs was 582 bp, with sizes ranging from 100-1 500 bp. Basic Local Alignment Search Tool （BLASTX） analysis revealed that 1 664 unigenes had significant homology to known genes in the Na- tional Center for Biotechnology Information （NCBI） non-redundant （nr） database （E-value≤5-10）. Among them, the two most abundant genes encoded metallothionein （157 ESTs） and chlorophyll a/b-binding pro- tein （38 ESTs）, accounting for 7.1% and 1.7% of the total ESTs, respectively. Using Kyoto Encyclopedia of Genes and Genomes （KEGG）, 1 462 unigenes were assigned to 1 161 pathways （E-value≤5-10）. A total of 938 unigenes were assigned Gene Ontology （GO） terms based on the GO hierarchy analysis, and InterProScan searches recognized 1 003 InterPro families. Three genes for metallothionein in Z. marina that belonged to Class II was identified. Results of this study will improve understanding of the molecular mechanisms of saline tolerance in Z. marina.

Identification of Light-Harvesting Chlorophyll a/b-Binding Protein Genes of Zostera marina L.and Their Expression Under Different Environmental Conditions

Photosynthesis includes the collection of light and a/b-binding （LHC） proteins. In high plants, the LHC gene family constituting the light-harvesting complex ofphotosystems I and II. the transfer of solar energy using light-harvesting chlorophyll includes LHCA and LHCB sub-families, which encode proteins Zostera marina L. is a monocotyledonous angiosperm and inhab- its submerged marine environments rather than land environments. We characterized the Lhca and Lhcb gene families of Z. marina from the expressed sequence tags （EST） database. In total, 13 unigenes were annotated as ZmLhc, 6 in Lhca family and 7 in ZmLhcb family. ZmLHCA and ZmLHCB contained the conservative LHC motifs and amino acid residues binding chlorophyll. The average similarity among mature ZmLHCA and ZmLHCB was 48.91% and 48.66%, respectively, which indicated a high degree of diver- gence within ZmLHChc gene family. The reconstructed phylogenetic tree showed that the tree topology and phylogenetic relation- ship were similar to those reported in other high plants, suggesting that the Lhc genes were highly conservative and the classification of ZmLhc genes was consistent with the evolutionary position of Z. marina. Real-time reverse transcription （RT） PCR analysis showed that different members of ZmLhca and ZmLhcb responded to a stress in different expression patterns. Salinity, temperature, light intensity and light quality may affect the expression of most ZmLhca and ZmLhcb genes. Inorganic carbon concentration and acidity had no obvious effect on ZmLhca and ZmLhcb gene expression, except for ZmLhca6.

Habitat suitability analysis of eelgrass Zostera marina L. in the subtidal zone of Xiaoheishan Island

We present a GIS-based habitat suitability index(HSI) model to identify suitable areas for Zostera marina L. restoration in the subtidal zone of Xiaoheishan Island. The controlling factors in the model,in order of importance,are Secchi depth,sediment composition,water temperature,salinity,current velocity,water depth and nutrient quality. Specific factor piecewise functions have been used to transform parameter values into normalized quality indexes. The weight of each factor was defined using expert knowledge and the analytic hierarchy process(AHP) method. All of the data thus obtained were interpolated using the inverse distance weighted(IDW) interpolation method to create maps for the entire region. In this study,the analysis of habitat suitability in the subtidal zone of Xiaoheishan Island was conducted for four seasons. According to the GIS-based HSI model,the optimal habitat of Z ostera marina L. appears in spring,although habitat remains suitable all year round. On the whole,the optimum site for eelgrass restoration is located in the eastern region,followed by the western and southern regions. We believe that the GIS-based HSI model could be a promising tool to select sites for Z ostera marina L. restoration and could also be applicable in other types of habitat evaluation.

Antibacterial Activity of Extracts from Zostera marina Against Pathogens of Apostichopus japonicus Skin Ulceration Disease

The purpose of this study was to investigate the antibacterial activity of extracts from Zostera marina against the pathogens of Apostichopus japonicus skin ulceration disease.When 95% ethanol (v/v) solvent was used to extract Zostera marina at 50℃,aqueous extract (ZA) showed obvious bacteriostatic effects on the tested bacterial strains (inhibition halo diameters between 8.23 mm and 13.62 mm),whereas the ethyl acetate extract (ZE) was almost inactive.The minimal inhibitory concentration (MIC) of ZA against four pathogens were homogeneous at 12.8 g L-1.ZA components were analyzed by thin layer chromatography (TLC) assay and six fractions were obtained.In another study,the six fractions showed inhibitory effects against the tested bacteria while their functions seemed to counteract the ZA activity.

Influences of diel and tidal cycles on fish assemblage in eelgrass(Zostera marina) bed of southern Korea during autumn

Short-term tidal and diel variations of autumn fish assemblage in a Zostera marina bed were investigated using 3 h interval samplings for 24 h in both spring and neap tide using a small beam trawl.A total of 1 346 fishes belonging to 19 species were collected at spring tide,whereas 1 115 fishes belonging to 17 species were at neap tide.The common fish species were Nuchequula nuchalis,Acanthogobius flavimanus,Takifugu niphobles,Acentrogobius pflaumii,and Pholis nebulosa with the former three species dominating at spring tide,while the latter two species being abundant at neap tide.Diel variation in abundance was significant with higher abundance at night than day,but there were no significant differences between spring and neap tides,and between ebb and flood tides（three-way ANOVAs）.Diel variation in the abundance of fishes may be influenced by tidal range and cycle,and day-night differences of food availability and behaviors of fishes indirectly.Non-metric multidimensional scaling（n MDS） ordination and analysis of similarity（ANOSIM） results revealed significant differences in species compositions both between day and night,and between spring and neap tide.Eelgrass beds are highly productive marine ecosystem,and thus,our results will contribute to conservation of seagrass ecosystem in the study area.

荣成天鹅湖大叶藻(Zostera marina)间2 种小型腹足类的分类学研究

小型腹足类生物在我国种类繁多,但其相关研究在国内基本属于空白。本文对荣成天鹅湖大叶藻间的2种小型腹足类生物进行了形态学及分子生物学(16S和28SrRNA基因片段)研究。结果表明,根据形态学所鉴定的短剑螺属未定种Machae roplax sp.,应为畦螺属未定种Lirularia sp.;分子生物学研究结果支持形态学对刺绣翼螺Alabapicta的鉴定。同时,与A.picta有一定形态学差异的其它4种相近形态的小型腹足类,在基因序列及遗传距离上与刺绣翼螺非常接近,亦应为刺绣翼螺Alaba picta。

A genome-wide analysis of the chloroplast NADH dehydrogenase - like genes in Zostera marina

The chloroplast NADH dehydrogenase-like(NDH)complex,homologous to respiratory complex I,participates in photosystem I cyclic electron flow(PSI-CEF)and chlororespiration in photosynthesis.Phylogenetic analyses indicated that Zostera marina,a widely distributed seagrass,has a complete NDH complex,which is rarely observed in marine macrophytes.We identified all 31 ndh genes necessary for the functional NDH complex,of which ndhB and pnsb3 occurred as duplication events.Secondary structural analyses of antiporter-like subunits showed that the long amphipathic helix of NdhF was lost in Z.marina,which could exhibit an alternative mode in the generation of trans-thylakoid proton gradient.The splicing pattern of ndh exhibited tissue-specific patterns and responded to light stress.RNA editing in Z.marina presented the ancestral pattern with many of the primitive editing sites and types.The partial editing in ndhF reflected the link between light stress and RNA editing.Moreover,the predominant expression in leaves of most ndh genes suggested that their major function is in photosynthesis.The quantitative real time-PCR results show that the expression of ndh was signifi cantly upregulated in response to light stress.Nevertheless,there were two diverse responsive mechanisms of the NDH complex in PSI-CEF and chlororespiration.Overall,the presence of a complete structure,upregulated gene expression level,and multiple post-transcriptional regulations could provide a molecular basis for the powerful NDH complex and enable Z.marina to maintain eff ective photosynthetic performance.

Isolation and Identification of Flavonoids Found in Zostera marina Collected in Norwegian Coastal Waters

In extracts of the seagrass Zostera marina, collected in coastal waters of West-Norway, fourteen different flavones and high amounts of rosmarinic acid were identified. Five of the flavones were found to be sulphated, among these were luteolin 7,3'-disulphate and chrysoeriol 7-sulphate structures previously not published with complete NMR assignments. Luteolin 7-O-β-(6''-malonyl) glucoside, and two other malonylated flavone compounds occurring in trace amounts, were identified for the first time in Z. marina. The sulphated flavones were fairly stable in slightly acidified (0.1% trifluoroacetic acid) extracts stored for months, however, under more acidic conditions (0.5% trifluoroacetic acid in the extracts) they were susceptible to undergo hydrolyses. When the solvents of purified fractions were removed by rotary evaporation, the sulphated flavones quickly decomposed to their corresponding aglycones due to the increased acid concentrations.

Artificial seed germination and seedling production of Zostera marina L.by salinity manipulation

Vast declines in Zostera marina seagrass beds demand effective methods of rehabilitation. In this study, we developed a practical method by reducing salinity to induce seed germination followed with recovering salinity to facilitate seedling production of Z. marina. The results showed that Z. marina seeds collected from natural seawater（salinity 30） were induced to germinate at reduced salinities. Percent germination（GR） was higher and mean-time-to-germinate（MTG） was shorter at lower salinities. The highest GR and shortest MTG occurred at salinity 0（deionized freshwater）. After germination in freshwater, seeds could develop into seedlings at salinities5–30 and continue the growth. Viability or development of germinated seeds was not significantly different during the 40 d of post-germination incubation at salinities 5–15 after 1–20 d of germination in freshwater. However,during the process of translating germinated seeds from salinity 0 and 5 to salinity 30, reducing the gradients of post-germination acclimation facilitated more seeds forming seedlings in less time. On average, after 60 d of static incubation, including 20 d in freshwater for germination followed with immediate shift to salinity 5 and increasing to salinity 30 at increment of 5 every two days until cultivation at constant salinity 30, 33% of Z. marina seeds produced healthy seedlings. The results indicate that the salinity-manipulation based method of artificial germination and seedling production is practical and effective in supporting rehabilitation of Z. marina bed.

Recent Trends of Zostera marina (Eelgrass) in a Highly Eutrophic Coastal Lagoon in the Mid-Atlantic Region (USA)

Spatial and temporal characteristics of Zostera marina (eelgrass) in the Barnegat Bay-Little Egg Harbor Estuary are compared before (2004-2011) and after (2012-2013) major fertilizer legislation (Fertilizer Law A2290) was enacted in New Jersey (USA) to reduce nutrient inputs from fertilizers to water bodies in the state. A significant decrease of Z. marina biomass and areal cover occurred in this eutrophic estuary between 2004 and 2011 concomitantly with increasing nitrogen and phosphorus loading from the watershed. The rate of decline in aboveground and belowground biomass was significantly sharper during 2004-2006 than during 2008-2010. In 2010, Z. marina biomass dropped to a very low level (mean aboveground biomass = 7.7 g dry wt mDŽ;mean belowground biomass = 27.0 g dry wt mDŽ), persisting through the last sampling period (October-November) in 2013. Biomass and areal cover of Z. marina decreased even further after Fertilizer Law A2290 was enacted in January 2012, with the lowest values recorded from August to November each year. These low values are the result of ongoing eutrophication of the system. More seagrass monitoring and research are necessary in future years to determine if the fertilizer law will have a positive effect on Z. marina condition in the estuary over the long term.

Biotic and Abiotic Conditions Can Change the Reproductive Allocation of Zostera marina Inhabiting the Coastal Areas of North China

The relationships between reproductive allocation and different factors have been widely recognized for terrestrial plants, but scarcely for marine angiosperms. Focusing on the seagrass Zostera marina, we examined the change in biomass when it was partitioned to growth, vegetative propagation and reproductive allocation of Z. marina. We also evaluated the relationships between reproductive allocation and biotic and abiotic conditions in the coastal areas of north China. The results indicated that the reproductive allocation of Z. marina at the levels of genet, shoot and spathe was positively correlated with shoot characteristics such as height and biomass(P < 0.01), but was negatively correlated with branch number and leaf number per genet(P < 0.01). In addition, the allocation was also affected by other abiotic factors such as water column phosphate concentration(P < 0.05) and sediment total petroleum(P < 0.01). These findings indicated that both biotic and abiotic conditions significantly influenced the reproductive allocation of Z. marina, which may affect further the population dynamics.

大叶藻(Zostera marina L.)胚的结构和种子萌发过程的研究

以采自山东荣成褚岛桑沟湾南岸成熟的大叶藻种子为材料,研究了种子的外部形态特征、胚乳成分及胚的解剖结构,并初步了解了大叶藻种子的萌发过程。结果表明,大叶藻种子是单子叶有胚乳的种子,呈暗褐色,椭圆形或长圆形,种皮革质且有17～20条纵肋,胚乳丰厚坚硬,胚乳细胞被淀粉粒充满;胚弯曲,由胚根、胚轴、胚芽和子叶组成,子叶U形棒状,其弯曲部有一裂缝,胚芽位于裂缝处,胚轴较短,其下是两个对生的胚根,有中央维管束贯穿胚根、胚轴、胚芽和子叶。在子叶出土萌发的植物类型中,大叶藻种子萌发过程是罕见的。萌发初期,子叶优先生长突破种皮而长出地面,此后胚根才开始生长并产生不定根。对大叶藻种子结构和萌发过程的研究,为探讨大叶藻种子萌发和幼苗更新的限制因素提供了一定的理论依据。

Cloning, characterization and expression analysis of a microsomal glutathione S-transferase gene from the seagrass Zostera marina

The response of glutathione S-transferase(GST)in Zostera marina to temperature variation was analyzed at molecular level by cloning the microsomal GST gene and texting the microsomal GST expression regularity under different temperature.Specific speaking,express ZmGST in Escherichia coli,then purify the recombinant protein and make the thermal stability analysis.Therefore,the experiments were carried out to provide a theoretical basis for the further elaboration to the population degradation mechanisms of Z.marina.In conclusion,the thermostability and the response of ZmGST gene to temperature changes can determine its temperature tolerance range,and affect its resilience in turn.

山东半岛典型海域大叶藻(Zostera marina)附着大型藻类群落结构

2012年3月、6月、9月和12月对青岛栈桥、威海双岛湾及荣成月湖3个典型海域大叶藻(Zostera marina)草场的附着大型藻类进行了调查,采用多样性指数、均匀度、聚类和排序等多元统计方法对各海域不同季节附着藻类的群落结构进行比较分析。结果表明:鉴定大型藻类17种,隶属3门14属,其中红藻门4属5种、褐藻门4属4种和绿藻门6属8种;三海域附着藻类种数最大值均出现在春季的栈桥和月湖(均为7种),最小值出现在秋季和冬季的月湖(均为1种);三海域附着藻类生物量最高值均出现在春季,而最低值分别为秋季(栈桥)、冬季(双岛)和夏季(月湖);栈桥的Shannon多样性和Pielou均匀度指数最低,双岛的物种多样性指数最高;一年中夏季的物种多样性和均匀度指数最高,冬季最低;聚类和排序结果表明,不同季节附着藻类群落结构在海域间和海域内不同采样点间均呈现显著性差异。季节变化和地域的差异对大叶藻附着藻类的群落结构特征有着重要影响。

石油烃水溶液对鳗草(Zostera marina)几种主要抗氧化物的影响

鳗草(Zostera marina)是北半球温带海域海草床的代表种,具有重要的生态学价值,其对海上溢油的生理响应特征尚未可知。本研究聚焦鳗草用石油烃水溶液(water accomadated fraction,WAF)培养过程中的生理指标变化,探究鳗草的抗氧化系统对WAF的响应特征。结果显示,实验组WAF质量浓度为1 mg/L,2.5 mg/L以及5 mg/L(分别用20%,50%和100%表示)的鳗草均在2 d出现显著的氧化损伤,丙二醛(MDA)含量一直高于对照组;超氧化物歧化酶(SOD)酶活性呈现先升高后降低的趋势;过氧化物酶(POD)酶活性随WAF培养时间增加均出现显著降低;总蛋白质的含量先增加后减少,且含量与对照组之间存在显著性差异;抗坏血酸(VC)的含量要比对照组的含量低;还原型谷胱甘肽(GSH)的含量与对照组差异不显著;单脱氢抗坏血酸还原酶(MDHAR)酶活性与对照组间存在显著性降低;过氧化氢(H2O2)的含量一直高于对照组。不同浓度的实验组均发生剧烈的氧化应激反应,说明鳗草是对WAF反应灵敏的物种。本研究初步揭示了鳗草抗氧化系统中几种关键酶对WAF持续性暴露的响应特征,为进一步研究海上溢油导致的生态破坏提供参考依据。

曹妃甸海草床生态特征及限制因子研究

通过开展曹妃甸海草床的分布、生物学特征、水环境和底质环境要素等调查研究工作,划定海草床分布区,对覆盖度进行分级,分析研究海草床生态特征及关键环境影响因子特征,为查明曹妃甸海草床资源环境家底、保护区划定、海洋生态修复及蓝碳研究工作提供技术参考。结果显示,曹妃甸海草床主要优势种类为鳗草,平均株高75.00±6.15 cm/shoot,且不同站点之间有差异;总平均茎枝密度为261.26±48.50 shoots/m^(2),总平均地上和地下生物量分别为302.53±71.79 g DW/m^(2)和114.71±27.40 g DW/m^(2)。海草床分布区面积42.90 km^(2),其中密集区、较密集区、一般区、较稀疏区和稀疏区分别占比为13.09%、12.93%、18.82%、18.75%和36.40%,海草分布在北部和南部两个区域,其中密集区与较密集区多分布在北部区域;草床区和裸沙区水环境差异不明显,底质环境差异较大;水温、营养盐、透光度、水流及人类工程活动是影响曹妃甸海域海草生长的关键因素。

中国温带海域新发现较大面积(大于0.5 km^2)海草床:Ⅱ声呐探测技术在渤海唐山沿海海域发现中国面积最大的鳗草海草床

近些年来随着生境的破坏其分布面积急遽下降,较大面积连续分布的海草床已经非常少见。本研究首次将声呐探测技术应用到海草床时空分布特征的相关研究中,在河北唐山乐亭-曹妃甸沿海发现目前中国面积最大的鳗草(Zostera marina)海草床,海草床面积高达29.17 km2,绘制了鳗草分布图和水深分布图,并对鳗草生物量和茎枝高度等基本生物学指标进行了调查。该发现极大丰富了中国海草数据库,并为鳗草的深入研究及保护提供了得天独厚的实验基地,也为周边海洋牧场的构建打下良好的基础。