基于转录组的假微型海链藻硫代谢相关途径分析

为分析近海广布藻华原因种应对广盐环境的细胞关键生命代谢活动过程的调控策略,本研究对不同盐度以及不同处理时间的假微型海链藻(Thalassiosira pseudonana)进行转录组测序,获得T.pseudonana硫代谢途径酶编码基因的差异表达结果。研究结果显示:在不同盐度下,共发现13个硫代谢途径的差异表达基因;随培养时间延长,低盐度培养的实验组相较于对照组有更多的硫代谢编码基因发生显著差异表达,如在硫酸盐同化过程中,在培养的第二天ATP硫酸化酶的基因表达量相对于第一天显著降低,其原因可能是因为藻细胞对硫酸盐同化需求的减少,从而可能会限制亚硫酸盐的形成;在低盐胁迫下,与半胱氨酸合成相关的酶(如丝氨酸乙酰转移酶)基因表达量显著降低,说明藻细胞减少了含硫氨基酸(如半胱氨酸)的合成,这些都会影响其正常的生命代谢活动。本研究表明,沿海河口区的盐度波动可能会对T.pseudonana的硫代谢产生重要影响,这一发现有助于深入理解近海广布藻华原因种应对盐度压力时的分子响应机制。

Effects of Ocean Acidification on Nitrogen Metabolism of Skeletonema costatum

Ocean acidification(OA),caused by the rising concentration of atmospheric CO_(2),leads to changes in the marine carbonate system.This,in turn,affects the physiological processes of phytoplankton.In response to increased pCO_(2) levels,marine microalgae modulate their physiological responses to meet their energy and metabolic requirements.Nitrogen metabolism is a critical metabolic pathway,directly affecting the growth and reproductive capacity of marine microorganisms.Understanding the molecular mechanisms that regulate nitrogen metabolism in microalgae under OA conditions is therefore crucial.This study aimed to investi-gate how OA affects the expression profiles of key genes in the nitrogen metabolic pathway of the marine diatom Skeletonema costatum.Our findings indicate that OA upregulates key genes involved in the nitrogen metabolic pathway,specifically those related to nitrate assimilation and glutamate metabolism.Moreover,pCO_(2) has been identified as the predominant factor affecting the expression of these genes,with a more significant impact than pH variations in S.costatum.This research not only advances our understanding of the adaptive mechanisms of S.costatum in response to OA but also provides essential data for predicting the ecological consequences of OA on marine diatoms.

Asexual Reproduction of Aurelia coerulea Polyps in situ on Settling Plates in Jiaozhou Bay,China

Jellyfish blooms are increasingly prevalent coastal phenomena,with Aurelia coerulea being a significant contributor.The complex life history of jellyfish,featuring a sessile polyp stage,is a key factor in bloom formation,yet it is challenging to study in the field.This study aimed to describe the asexual reproduction of A.coerulea polyps in Jiaozhou Bay,China,and to assess their ability to form sedentary colonies and their relationship with environmental factors.Using settling plates,we observed polyps in situ and documented three asexual reproduction modes:budding,strobilation,and podocyst formation.Environmental parameters including temperature,salinity,dissolved oxygen(DO),chlorophyll a(Chl a),and the biomass of phytoplankton and zooplankton were monitored.Budding was the predominant reproductive method,featuring stolon budding in autumn and direct budding during the winter and spring seasons.Strobilation took place in the colder months,releasing ephyrae between April and May.Polyp abundance was positively correlated with seawater temperature and negatively correlated with DO concentration.Polyps at 1.0 m depth showed higher survivorship and proliferation over 8 months,whereas those at 2.0 m depth experienced a steady decline and died within 4 months.The study enhances our understanding of the life cycle and asexual reproduction strategies of A.coerulea polyps in the field,with implications for predicting and managing jellyfish blooms.The findings underscore the importance of temperature and DO in polyp survival and suggest that initial polyp population proliferation is crucial for long-term survival.

低盐胁迫下假微型海链藻脂肪酸代谢的转录组学分析

文章以假微型海链藻(Thalassiosira pseudonana)转录组测序数据为基础,对差异表达的功能基因进行GO(gene ontology)和KEGG(kyoto encyclopedia of genes and genomes)富集分析,以研究低盐胁迫条件对其脂肪酸代谢通路相关功能基因表达的影响。结果发现,不同盐度培养下的假微型海链藻在生长期的第2天与第4天时细胞中与脂肪酸生物合成、脂肪酸延伸的相关基因(如:ACC1、arm)的表达量相对于生长第1天均显著升高,与脂肪酸降解相关基因(如:ACADM、ECI1)的表达量无显著变化;与脂肪酸的生物合成、延长相关基因的表达量在试验组和对照组间存在差异(如:起重要作用的KASⅠ、ACAA2等基因表达量显著增加,但也有少数处于下降或者微变动的状态),与脂肪酸降解过程相关基因的表达量显著上升。

Amplicon-Based Illumina Sequencing and Quantitative PCR Reveals Nanoplankton Diversity and Biomass in Surface Water of Qinhuangdao Coastal Area, China

Aureococcus anophagefferens caused brown tides for three consecutive years from 2009 to 2011 in the coastal waters of Qinhuangdao, China, with numerous, widespread ecological and economic impact on ecosystems. To understand the population dy- namics of nanoplankton during the brown tides, sequences of the V9 region of the 18S rDNA gene, used as a marker, were analyzed by Illumina sequencing to assess nanoplankton biomass, and real-time fluorescence quantitative PCR was performed to analyze spa- tial variation in the 18S rDNA copy concentrations of nanoplankton off the Qinhuangdao coast in July, 2011. The results showed that A. anophagefferens and Minutocellus polymorphus were the dominant species in the local phytoplankton community during the brown tide in July 2011. The highest 18S rDNA copy concentrations of A. anophagefferens and M. polymorphus were detected at stations SHG and FN, respectively. The central area most strongly affected by the brown tide migrated southward from 2011 to 2013. Redundancy analysis (RDA) showed that the decreasing NOx concentration might provide suitable nutrient conditions for the A. anophagefferens outbreak. During the brown tide caused by A. anophagefferens, other phytoplankton, such as diatoms, cryptophytes, chlorophytes, dinoflagellates and other flagellates, could co-occur with it. For zooplankton, due to less selective feeding behavior, Amoebozoa was the most abundant zooplankton at station SHG, while Ciliophora was the most abundant zooplankton at other sta- tions for its more selective feeding.

Seasonal and spatial distribution of ammonia-oxidizing microorganism communities in surface sediments from the East China Sea

Ammonia oxidation plays a significant role in the nitrogen cycle in marine sediments. Seasonal and spatial distribution of ammonia-oxidizing archaea （AOA） and betaproteobacteria （13-AOB） in surface sediments from the East China Sea （ECS） were investigated using ammonia monooxygenase ct subunit （amoA） gene. In order to characterize the community of AOA and 13-AOB, real-time quantitative polymerase chain reaction （qPCR） was carried out in this study, along with environmental parameters. The abundance of 13-AOB amoA gene （2.17x 10^6-4.54x10^7 copy numbers per gram wet weight sediment） was always greater than that ofAOA amoA gene （2.18x 105-9.89x 10^6 copy numbers per gram wet weight sediment） in all sampling stations. The qPCR results were correlated with environmental parameters. AOA amoA gene copy numbers in April were positively related to temperature and nitrite concentration （p〈0.05）. 13-AOB amoA gene copy numbers in August correlated negatively with salinity （p〈0.01）, and correlated positively with ammonium concentration （p〈0.05）. With the increase of salinity, the amoA gene copy ratio of AOB to AOA had a tendency to decrease, which suggested 13-AOB dominated in the area of high level ammonium and AOA preferred high salinity area.

Microbial Community Composition and Function in Sediments from the Pearl River Mouth Basin

This study was conducted to characterize the diversity and function of microbial communities in marine sediments of the Pearl River Mouth Basin(PRMB)in the South China Sea.The results showed that the bacterial and archaeal communities varied greatly with depth.Proteobacteria in bacterial communities and Nitrososphaeria and Woesearchaeota in archaeal communities were dominant in the shallow sediments(1-40 cm),while Chloroflexi in bacterial communities and Bathyarchaeia in archaeal communities were dominant in the deep sediments(50-200 cm).Regarding ecological functions based on the metatranscriptomic data,genes involved in various pathways of nitrogen metabolism and sulfur metabolism were observed in the tested sediment samples.Metagenomic analysis revealed that Proteobacteria contribute the most to nearly all genes involved in nitrogen and sulfur metabolism.Moreover,Thaumarchaeota contribute the most to certain genes involved in nitrification,denitrification and assimilatory sulfate reduction pathways.The most abundant bacterial genus,Candidatus Scalindua,is crucial for nitrification,dissimilatory nitrate reduction,denitrification and assimilatory sulfate reduction pathways.

Colorimetric determination of nanomolar concentrations of silicate in natural waters after liquid-liquid extraction using methyl isobutyl ketone

A sensitive solvent extraction method for the determination of nonamolar concentrations of silicate in natural waters is developed. According to the traditional aqueous silicate method, silicomolybdenum blue formed by the reaction between silicate and ammoni- um molydate and reduced by metol-sulfite reagent is extracted by methyl isobutyl ketone. The absorbance can be enhanced substantially up to 10-folds. The detection limit of silicate is 8 nmol/dm^3 , which is one tenth smaller than the traditional method, with the precision of 4.0% at a silicate level of 50 nmol/dm^3 and 3.2% at a silicate level of 6 μmol/dm^3. Comparing the calibration curves in the distilled water and seawater, it can be seen that the salt effect also exists in the extraction method. However, the salt effect is a linear function of the salinity and can be corrected by simple calibration. The proposed method is successfully applied to the determination of silicate in natural waters. Natural concentrations of arsenate, arsenite and phosphate cause negligible interference.

渤海和南黄海沉积物中氨氧化微生物对硝化潜势的相对贡献

硝化作用是海洋氮循环的核心过程。作为硝化过程关键步骤的氨氧化过程的主要参与者,氨氧化古菌和氨氧化细菌对硝化作用的相对贡献是海洋氮循环关注的热点问题之一。本文选取渤海和南黄海20个站位的表层沉积物,通过微宇宙培养实验研究了沉积物中氨氧化古菌和氨氧化细菌对硝化潜势的相对贡献。结果表明,渤海和南黄海海域表层沉积物中潜在硝化速率(以氮计,下同)为0.0046~0.2831μmol/(g·d),其中氨氧化古菌潜在硝化速率为0.0043~0.2743μmol/(g·d),氨氧化细菌潜在硝化速率为0.0004~0.0560μmol/(g·d)。氨氧化古菌是硝化潜势的主要贡献者,在渤海海域的贡献率为59.79%~97.95%,在南黄海海域的贡献率为18.47%~94.26%。渤海海域潜在硝化速率显著高于南黄海海域。此外,本研究海域中盐度是影响潜在硝化速率的关键环境因子,对渤海海域的分析则表明越高的NO_(3)^(-)浓度可能指示着越高的硝化潜势。在河口及近海沉积物中,氨氧化古菌在硝化过程中起着更加重要的作用;河口和近岸沉积物硝化潜势总体高于远海。本研究为进一步认识近海海洋氮循环过程提供了参考依据。

基于qPCR技术的黄东海沙海蜇时空分布特征研究

沙海蜇(Nemopilema nomurai)是东亚海域常见的大型水母,近年来其频繁大规模暴发导致了严重灾害。本文利用荧光定量PCR方法分析了2012年10月、2013年5月、8月和9月黄海、东海海区沙海蜇线粒体16S rDNA(mt-16S rDNA)的浓度和分布特征。研究表明,沙海蜇mt-16S rDNA在4个调查月份中均有检出,最早于5月上旬在长江口近岸出现较高检出值,检出浓度的高值区与长江口近岸温度高值区和盐度低值区位置一致;2013年8月的检出浓度在4个调查月中最高,检出浓度的高值区与温度高值区位置一致;2012年10月的检出浓度最低,检出浓度的高值区与温度低值区位置一致。从8月到10月沙海蜇mt-16S rDNA的浓度高值区呈由南向北转移的趋势,这与已有研究中沙海蜇水母体丰度分布趋势相吻合。2013年8月和9月的分层次采样结果显示,在大部分站位的底层水体中沙海蜇mt-16S rDNA浓度较高,这可能与沙海蜇成体或微小个体的沉降或垂直移动有关。研究结果表明,荧光定量PCR检测技术可实现水体中水母DNA定量检测,可以弥补传统拖网调查方法的不足,为全面了解水母的时空分布及水母栖息地溯源提供方法。

Characterization of Bacterial Communities in Aerosols over Northern Chinese Marginal Seas and the Northwestern Pacific Ocean in Autumn

The characteristics of the bacterial community structure in aerosols of different particle sizes over northern Chinese marginal seas and the northwestern Pacific Ocean in autumn were analyzed using high-throughput sequencing and the quantitative real-time PCR method based on 16S rRNA genes.Combined with environmental factors and air mass sources,the bacterial abundance,community diversity,composition and structural characteristics in aerosols were studied,which might provide a scientific evaluation for a comprehensive understanding of the characteristics,long-distance transmission and ecological effects of aerosols from coastal waters to the distant ocean(northwestern Pacific Ocean).At the junction of the South Yellow Sea and the East China Sea,the bacterial abundance in the aerosols was substantially affected by transmission of continental air masses and was higher than those in other samples,and the bacterial abundance in coarse particle(>2.1μm)samples was significantly higher than that in fine particle(<2.1μm)samples(P<0.05).In contrast,aerosols collected at the northwestern Pacific Ocean(NWP5 site)were less affected by transmissions of continental air masses,and their bacterial abundance in coarse particle samples was significantly lower than that in fine particle samples.Significant differences in the richness and diversity of bacterial communities were observed among all samples(P<0.05).The differences in the bacterial community in different sea areas were greater than those in the same sample with different particle sizes(P>0.05).Among the environmental factors examined,a significant negative correlation was observed between bacterial community richness and temperature(P<0.05),and the bacterial community diversity was significantly positively correlated with the concentration of K^(+)(P<0.05).Canonical correspondence analysis showed that temperature,NH_(4)^(+) concentration and SO_(4)^(2−) concentration exerted significant effects on bacterial community structures(P<0.01)in aerosols of different particle sizes over northern Chinese marginal seas and the northwestern Pacific Ocean.

大气颗粒物中细菌群落结构的区域差异性研究

为分析大气细菌群落结构区域差异性,本研究采集了2014—2015年黄海、青岛和兰州大气生物气溶胶样品,对其进行了16S rRNA基因高通量测序分析,探讨环境因子(温度、风速和相对湿度)对大气细菌群落结构可能的影响。结果表明,3月黄海、青岛和兰州以及青岛不同采样时间的生物气溶胶中细菌群落结构在区域(p<0.01)和时间(p<0.05)分布上具有差异性,即黄海气溶胶中细菌群落多样性均高于陆地(p<0.01),3月青岛细菌群落的多样性高于兰州(p<0.01),且青岛春季大气生物气溶胶中细菌群落的丰富度和多样性高于冬季和夏季(p<0.05)。物种注释结果表明,变形菌(Proteobacteria)、厚壁菌(Firmicutes)、放线菌(Actinobacteria)和拟杆菌(Bacteroidetes)为这3个区域大气中的优势菌门。由于粗、细颗粒耐受环境变化的能力不同,不同区域粗粒径和细粒径样品的细菌群落的组成发生变化。例如,Bacteroides在黄海粗粒径样品中的相对丰度高于细粒径;Psychrobacter和Paenibacillus分别是3月青岛粗粒径和细粒径样品中的优势菌;沙尘发生时,兰州细粒径样品中的优势菌由Bifidobacterium转为Acinetobacter,粗粒径样品中的优势菌仍是Bifidobacterium但相对丰度值增大。典范对应分析表明,风速对细菌群落结构具有显著影响(p<0.01)。

实时荧光定量PCR分析中不同铁浓度处理下玛氏骨条藻内参基因的筛选

实时荧光定量PCR(qRT-PCR)是定量分析基因表达的常用方法,选择合适的内参基因对准确分析目的基因表达水平至关重要。本研究以不同铁浓度培养条件下的玛氏骨条藻为材料,定量分析Cytb、EF-1α、HPRT、UBC、GAPDH、β-actin以及β-tubulin 7个内参基因的表达情况,并利用GeNorm、NormFinder和BestKeeper软件对这些内参基因的稳定性进行综合评价。结果表明,Cytb和EF-1α的表达稳定性较好,EF-1α+Cytb组合的稳定性最佳,是玛氏骨条藻基因表达研究的理想内参基因,而其他基因的表达稳定性较差,不适合作为内参基因。本研究为玛氏骨条藻基因表达研究过程中内参基因的选择提供了方法学上的依据。

渤海沉积物中产甲烷途径及产甲烷菌群落特征

全球甲烷排放主要来源于厌氧环境中产甲烷菌的代谢活动。采集渤海典型站位沉积物样品,测定了沉积物中甲烷和硫酸盐含量。通过提供不同类型产甲烷底物进行培养,分析了各样品中甲烷产量;通过高通量测序,分析了产甲烷菌群落特征。结果表明,渤海沉积物中产甲烷途径以甲基营养型为主,同时具有H_(2)/CO_(2)还原型途径。同一站点随着深度的增加,甲烷产生量逐渐减少,产甲烷速率相应降低。古菌群落以Crenarchaeota、Asgardaeota和Nanoarchaeota为主,产甲烷菌Ca.Methanofastidiosales占据优势地位。本研究为全面了解产甲烷菌在海洋生境中产甲烷过程提供了重要参考。

远缘杂交在水稻遗传育种中的应用

水稻种内基因资源的充分利用为我国粮食增产作出了巨大贡献。为进一步挖掘种间、属间及亚远缘物种的有利基因,维持栽培稻遗传多样性,实现水稻超高产、优质和多抗育种,本文综述了远缘杂交所涉及的分子机理及水稻远缘杂交育种的思路,概述了水稻种间和属间远缘基因资源的研究成果、远缘杂交障碍及克服途径、多种技术在水稻远缘杂交方面的发展概况等,为加快远缘物种基因资源在水稻育种上的应用提供参考。

Nutrient concentrations and fluxes in the Changjiang Estuary during summer

In June 2003 and 2006 concentrations of nutrient were determined in the Changjiang Estuary. The data indicated that phosphate and nitrate did not behave conservatively in the estuary, but silicate behaved conservatively. An important mobilization of phosphate and nitrate was observed from the river up to halfway in the estuary. Both input flux （from river to estuary） and output flux （from estuary to coastal zone） of phosphate, silicate and nitrate were calculated from statistical interpretations of the salinity profiles. There was a large discrepancy between input and output fluxes of phosphate and nitrate. The river fluxes of silicate, phosphate and nitrate （fr） are augmented 5.3%, 28.9% and 36.6% in June 2003 and 1.0%, 62.5%, 31.7% in June 2006 by internal inputs （fi）.The phosphate and nitrate fluxes are enhanced through the estuarine process, while silicate flux is unaltered. The authors present some long-term data for nutrient concentrations and the ratios of silicon to nitrogen to phosphorus in the Changjiang Estuary. Silicate level falled in the last two decades, while concentration of nitrate increased. Phosphate concentration had no significant change.

Detection of Phaeocystis globosa using sandwich hybridization integrated with nuclease protection assay(NPA-SH)

Phaeocystis globosa Scherffel is one of the common harmful algae species in coastal waters of the southeastern China.In this study,sandwich hybridization integrated with nuclease protection assay(NPA-SH)was used to qualitatively and quantitatively detect P. globosa.Results showed that this method had good applicability and validity in analyzing the samples from laboratory cultures and from fields.The linear regression equation for P.globosa was obtained,and the lowest detection number of cells was 1.8×104 c...

Development of a real-time PCR method for Thalassiosira rotula rapid detection

Gene specific primers and DNA probe were designed based on the sequence of 18S rDNA cloned from the red tide alga Thalassiosira rotula. A real-time fluorescent quantitative PCR （RFQ - PCR） method was developed for quantitative detection of T. rotula. The RFQ - PCR assay data showed that the results obtained with the RFQ - PCR quite good agreement with those with the light microscope （LM） counting method, which suggested that the RFQ - PCR could be a useful method for red tide alga detection.

Effect of decreasing temperature on the strobilation of Aurelia sp.1

The worldwide proliferation of marine jellyfish has become a crucial ecological and social issue, and as a cosmopolitan species, Aurelia spp. have received increasing scientific attentions. In the present study, the responses of strobilation in Aurelia sp. 1 to decreasing temperature were illuminated through the expression levels of the retinoid x receptor （RxR） gene and the gene encoding a secreted protein, CL390. We observed that a higher final temperature decreased the strobilation prophase and strobilation interphase periods, and the growth rate of the strobilae ratio increased with increasing CL390 gene expression. The ratio of strobilae at 12~C was highest, and the strobilae showed the higher releasing ratios at both 12℃ and 16℃ compared with those at 4℃ and 8℃. Furthermore, more ephyrae were released at the higher final temperature. Additionally, up-regulation and down-regulation of the CL390 gene were observed in response to the four decreasing temperatures. Although the four CL390 gene transcript levels increased more significantly than the transcript levels of the RxR gene, similar trends were observed in both genes.

Effects of temperature and salinity on the asexual reproduction of Aurelia coerulea polyps

Aurelia coerulea polyp is an important stage in the outbreaks of this species.To test the combined effects of salinity and temperature on the survival and asexual reproduction of polyps,we maintained 864 polyps at various salinities(15,25,33,and 40)and temperatures(9,12,15,18,21,and 24℃).Polyps could mostly survive in all treatment combinations except in salinity 15 treatments with low temperatures(9-15℃).Budding occurred at all temperatures(9-24℃),while strobilation only occurred at the low temperatures(9-15℃).The range of 12-15℃was suitable for strobilation and ephyrae release.The optimal range of salinity for asexual reproduction was 25-33.Low(15)or high(40)salinity could significantly reduce the numbers of new buds or ephyrae,and low salinity of 15 retarded and even prevented strobilation at low temperatures.The optimal treatment for budding and strobilation was 21℃-salinity 25 and 12℃-salinity 33,respectively.Salinity had less of an impact than temperature on asexual reproduction,except for the polyps in high or low osmotic pressure conditions.