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.     

Nutrient compositions of cultured Thalassiosira rotula and Skeletonema costatum from the Jiaozhou Bay in China

Carbon, nitrogen, phosphorus, silicon composition of cultured two different sized phytoplankton common species of Thalazsiosira rotula and Skeletonema costatum from the Jiaozhou Bay were measured. Carbon, nitrogen, phosphorus, silicon contents in cell were obvious higher in T. rotula than in S. costatum, but the percents of nitrogen, phosphorus, sihcon contents in cell dry mass in T. rotula were lower than those in S. costatum. The dry mass concentrations of nitrogen,phosphorus, silicon in S. costatum were much higher than those in T. rotula, particularly silicon, the former was 6. 4 times of the latter, showing that S. costatum could more assimilate these elements. Especially, S. costatum had competitive dominance for assimilating silicon, which is beneficial to its becoming a major dominant species in relative short silicon of the Jiaozhou Bay. There were some differences in numerical value of nutrient ratios both laboratory-cultured phytoplankton and different sized suspended particulates （ mainly phytoplankton） in the Jiaozhou Bay, which was caused by the changes of environment. High contents of carbon, nitrogon and relative low phosporus, sil- icon, high molar ratio of nitrogen to phosphorus （far higher than Redfield value） and low ratio of silicon to phosphorus and silicon to nitrogen （far lower than Redfield values） in the two diatoms and different sized suspended particulates were consistent with those in the seawater. Relative short silicon in the seawater and phytoplankton showed that silicon was possibly affectting phytoplankton growth in the Jiaozhou Bay.     

浒苔绿潮暴发期间海水溶解有机碳的生物可利用性及其碳汇作用

自2007年以来,浒苔绿潮已经连续15年在南黄海暴发。浒苔(Ulva prolifera)作为主要肇事藻种,在暴发过程中向海水释放大量的溶解有机碳(DOC)。然而,这些藻源DOC能否长期保存在海洋中,主要取决于它们的生物可利用性,目前关于此方面的研究甚少。本研究在浒苔绿潮大规模暴发时期(2019年6月),分别在浒苔暴发海区和无浒苔海区各选择3个站位富集表层海水,在实验室进行长期(300 d)的DOC降解实验。结果发现,在60 d内,不同站位富集海水中的DOC浓度随着微生物的利用快速下降,微生物丰度也在第60天达到峰值,表明这些被消耗的DOC是生物可利用性高的活性DOC(LDOC)。60 d后,剩余的DOC可抵抗微生物的降解,在60～300 d内保持稳定,表明这些DOC是具有强稳定性的惰性DOC(RDOC)。最终发现,浒苔暴发海水的RDOC占富集DOC的46%,明显高于无浒苔海水的(36%)。并且,荧光溶解有机物(FDOM)中活性的类蛋白组分随着微生物的利用被快速消耗,惰性的类腐殖质组分逐渐积累,暗示了在降解过程中LDOC逐渐向RDOC转化。可见,浒苔绿潮暴发除了在短时间内增加海水中的D...     

气相色谱法分析小球藻脂肪酸的组成

利用超声波破碎技术在1 mol/L HCl-CH3OH 溶液中提取小球藻脂肪酸并酯化,并用气相色谱和气质联用仪对其进行定量和定性分析,鉴定出 C14:0, C16:2, C16:1, C16:0, C18:2, C18:1, C18:3, C18:0, C20:0共9种脂肪酸,其中不饱和脂肪酸占总脂肪酸的65%.6份平行样的相对标准偏差为6.04%~9.84%,样品加标回收率范围为77.0%~122.6%,各种脂肪酸甲酯的仪器检测限<0.3 ng.初步探讨了脂肪酸甲酯在弱极性色谱柱上的保留时间规律     

Variations of dissolved organic matter during a plankton bloom: qualitative aspects,based on sugar and amino acid analyses

Water samples collected during a spring plankton bloom in the northern North Sea were investigated for dissolved organic carbon (DOC), total dissolved sugars (TDS) and dissolved amino acids (TDAA). Their concentrations varied in the range of 0.6 to >4 mg 1^?1 for DOC, 40—400 γg 1^?1 for TDS and 100—850γg 1^?1 for TDAA. TDS and TDAA contributed to less than 20% of the DOC, but showed variations independent of each other. Changes in the efficiency of heterotrophic utilization and its selectivity seem to have caused these variations. These, together with differences in the sources of dissolved organic matter — phytoplankton and detrital materials — were also reflected in the qualitative composition of TDS and TDAA in the water column. In the TDS fraction glucose and mannose showed maxima during the phytoplankton bloom and galactose, rhamnose, xylose, arabinose and fucose after it. Glutamic acid registered maximum in the early stages of the phytoplankton bloom, histidine towards its end and aspartic acid, glycine, alanine and lysine after it. Serine was present in large amounts throughout the investigated time interval. The results of the present study are discussed in the light of previously published data on the formation and utilization of dissolved organic matter in the sea.     

养马岛附近海域藻华期间有色溶解有机质的生物可利用性研究

本研究利用吸收光谱和荧光激发-发射矩阵光谱-平行因子分析(EEMs-PARAFAC),研究了养马岛附近海域海水中有色溶解有机质(CDOM)的浓度、组成、来源和生物可利用性,并估算了浮游植物生长繁殖对CDOM及具有生物可利用性CDOM的贡献。结果表明,表、底层海水中CDOM浓度(以吸收系数a₃₅₀计)平均值分别为1.62±0.42 m^-1和1.30±0.47 m^-1,光谱斜率(S_275-295)平均值分别为0.022±0.003 nm^-1和0.023±0.003 nm^-1。利用PARAFAC模型识别出4种荧光组分,分别为陆源类腐殖酸C1、类色氨酸C2、类酪氨酸C3和微生物源类腐殖酸C4。荧光指数(FIX)、腐殖化指数(HIX)和生物指数(BIX)显示,CDOM受陆源输入和海洋自生源的综合影响。降解实验结果显示,表、底层海水中生物可利用性CDOM百分比(%△a₃₅₀)平均值分别为(23.36%±17.94%)和(8.93%±20.30%)。C1、C2和C4组分的荧光强度在培养之后降低,而C3组分的荧光强度上升。各荧光组分生物可利用性依次递减的顺序为:%△C1(23.75%±8.96%)＞%△C4(20.83%±11.71%)＞%△C2(11.67%±38.87%)＞%△C3(-29.61%±39.90%),显示培养之后CDOM的平均分子量和腐殖化程度降低。表层海水中a₃₅₀、%△a350与Chl a之间存在显著线性相关关系,据此可以估算出浮游植物生长繁殖对CDOM的贡献为36.9%,对具有生物可利用性CDOM的贡献为85.0%。     

Development of lipids during a spring plankton bloom in the northern North Sea: I. Particulate fatty acids

The plankton spring bloom in the northern North Sea was extensively investigated during a period of three months in 1976 at a fixed station occupied by the R.V. “Meteor”. Samples of different depth-profiles, representative of the phytoplankton development, were collected eleven times to analyze the concentration of fatty acids of the particulate matter. The water column was divided into an upper and lower layer according to the thermocline depths, because different processes take place in these layers. During the exponential growth phase the fatty acid concentration rose only slightly due to increases in polyunsaturated fatty acids (18:4, 20:5, 22:6), which are typical for marine plankton. With the exhaustion of nutrients the biochemical composition changed and the fatty acid concentration increased sharply from about 3 to 20 μmol C dm^? finally to about 30% of the particulate carbon. The main proportion consisted of oleic acid (28.3%) and palmitic acid (24.2%). The first phytoplankton bloom, dominated by diatoms (Chaetoceros species), was characterized by the increase in fatty acids with 16 carbon atoms, whereas during the second smaller bloom, with dinoflagellates as the main species, more fatty acids with 18 carbon atoms occurred. After the stationary growth phase the phytoplankton biomass strongly decreased, resulting in an increase of particulate matter below the thermocline. The fatty acid pattern there was similar to that during the stationary phase of the phytoplankton bloom in the upper layer.     

秋季东海溶解态和颗粒态氨基酸的分布与组成*

本文对2012年秋季中国东海31个站位的海水样品中溶解态氨基酸（THAA）和颗粒态氨基酸（PAA）的分布与组成进行了研究。结果表明：表层海水中溶解游离氨基酸(DFAA)的平均浓度为0.12±0.04 μmol/L (0.06~0.19 μmol/L),溶解结合氨基酸(DCAA)的平均浓度为0.61±0.51 μmol/L (0.15~1.79 μmol/L),PAA的平均浓度为0.11±0.06 μmol/L (0.02~0.27 μmol/L)。THAA的水平分布特点大致为近岸高、远岸低;PAA的水平分布特点是近岸海域向远海海域分布呈现逐渐减小的趋势。THAA的垂直分布特点是由表层向底层逐渐降低。DCAA、PAA与Chl-a有很好的相关性,而DFAA与Chl-a的相关性不明显。东海表层海水中THAA的主要组分是天门冬氨酸、谷氨酸、丝氨酸、甘氨酸、苏氨酸及丙氨酸,PAA的主要组分是天门冬氨酸、谷氨酸、丝氨酸、甘氨酸、丙氨酸及亮氨酸。在表层海水中氨基酸是作为一个整体而对海洋生物地球化学过程产生影响的。     

大型水母沙海蜇暴发的温带贫营养海洋生态系统中细菌和浮游植物的关系

Bacterial abundance, phytoplankton community structure and environmental parameters were investigated to study the relationships between bacteria and phytoplankton during giant jellyfish Nemopilema nomurai blooms in the central Yellow Sea during 2013. N. nomurai appeared in June, increased in August, reached a peak and began to degrade in September 2013. Results showed that phosphate was possible a key nutrient for both phytoplankton and bacteria in June, but it changed to nitrate in August and September. Phytoplankton composition significantly changed that pico-phytoplankton relative biomass significantly increased, whereas other size phytoplankton significantly decreased during jellyfish bloom. In June, a significantly positive correlation was observed between chlorophyll a concentration and bacterial abundance(r=0.67, P0.001, n=34).During jellyfish outbreak in August, there was no significant correlation between phytoplankton and bacteria(r=0.11, P0.05, n=25), but the relationship(r=0.71, P0.001, n=31) was rebuilt with jellyfish degradation in September. In August, small size phytoplankton occupied the mixed layer in offshore stations, while bacteria almost distributed evenly in vertical. Chlorophyll a concentration significantly increased from(0.42±0.056) μg/L in June to(0.74±0.174) μg/L in August, while bacterial abundance just slightly increased. Additionally, the negative net community production indicated that community respiration was not entirely determined by the local primary productivity in August. These results indicated that jellyfish blooms potentially affect coupling of phytoplankton and bacteria in marine ecosystems.     

蛋氨酸和裂壶藻强化对卤虫体内游离氨基酸含量的影响

以卤虫(Artemia parthenogenetica)无节幼体作为试验材料,研究蛋氨酸或裂壶藻(Schizochytrium sp.)强化对卤虫体内游离氨基酸(FAA)含量的影响。试验分为4组,分别为蛋氨酸(Met)组、裂壶藻组、蛋氨酸加裂壶藻组以及对照组,试验进行16 h,每4 h取样1次。结果显示：试验结束时,蛋氨酸组的卤虫体内游离Met含量显著高于裂壶藻组(P<0.05)。除对照组的丝氨酸(Ser)、谷氨酸(Glu)、甘氨酸(Gly)含量显著高于蛋氨酸+裂壶藻组外(P<0.05),其他所有的必需氨基酸及非必需氨基酸含量在16h时都没有显著差异。试验过程中FAA水平随时间的变化趋势为：对照组和蛋氨酸组基本上呈现先减少再升高的趋势,且16 h和0 h的含量差别不大;裂壶藻组和蛋氨酸加裂壶藻组的卤虫,其体内FAA含量基本上呈下降趋势,或小幅升高后下降,16 h时的含量大多低于0 h时。试验结果表明：给卤虫无节幼体强化Met可以提高其体内Met含量,对其他种类的FAA影响不大。     

Development of taxonomic rRNA-targeted probes of two harmful algae: Prorocentrum minimum and Karenia mikimotoi by fluorescence in situ hybridization

Harmful algal blooms recently have been under the spotlight throughout the world, because of their negative impact on the marine environment, aquaculture, fisheries as well as public health. The development of methods for rapid and precise identification and quantification of causative species is essential for the warning and monitoring of blooms, among which the techniques based on taxonomic probes are the most favored. In this study, two harmful algae, i.e., Prorocentrum minimum and Karenia mikimotoi were taken into consideration. The partial large subunit rDNA (D1-D2) of both species were firstly PCR-amplified, cloned and sequenced. The obtained sequences were then introduced to carry out alignment analysis for gene specific regions. Three respective candidate probes for each species were designed and used to screen the optimal probe by performing fluorescence in situ hybridization (FISH) tests. The results showed that the probes Pmin0443 and Kmik0602 displayed the best hybridization for P. minimum and K. mikimotoi, respectively. Both the specific (taxonomic) (Pmin0443 and Kmik0602) and the control probes (UniC0512 and UniR0499) were used for cross-reactivity tests with other microalgae in our laboratory. The probes Pmin0443 and Kmik0602 are specific and could be served as taxonomic probes introduced into the techniques targeting rRNA, such as FISH, sandwich hybridization, and DNA-microarray assay of P. minimum and K. mikimotoi in the future. Finally, FISH analyses with both probes were performed on the simulated field samples. The probes could hybridize exclusively with the target cells well, and no significant difference (p >0.05) was observed in the cell densities of the samples determined by FISH and light microscopy (LM). All suggest that the probes are specific and could be introduced into FISH for the monitoring of both harmful algae.     

Development of lipids during a spring plankton bloom in the northern North Sea: II. Dissolved lipids and fatty acids

During the spring plankton bloom (1976) in the northern North Sea the fatty acid and lipid distribution of the chloroform extractable fraction of seawater was analyzed by gas chromatography and thin-layer chromatography. The fatty acids in the seawater made up about 3% of the total dissolved organic matter. Maximum concentrations during the first phytoplankton bloom were in the range of 5 μmol C dm^?3. Palmitic acid (30.4%) and oleic acid (21.4%) as well as myristic, stearic and palmitoleic acids were the main fatty acids. The concentrations of fatty acids were higher below the thermocline in the deeper layers of the water column. The oleic acid showed large fluctuations, especially below the thermocline and seemed to be bound in a polar fraction. Different lipid classes containing fatty acids were determined by thin-layer chromatography. An estimation showed that the free fatty acid fraction made up the main portion (47.9%), followed by the triacylglycerol fraction (22.8%) and a more polar fraction with 25.1%. The triacylglycerol fraction appears to be more stable than the other fraction in the water column.     

Biological control of short-term variations in the concentration of DMSP and DMS during a Phaeocystis spring bloom

In the spring of 1995, short-term variations in the concentration of particulate and dissolved dimethylsulfoniopropionate (DMSP) and dimethylsulfide (DMS) were monitored in the western Wadden Sea, a shallow coastal region in open connection with the North Sea. Significant correlations were found between abundance of Phaeocystis globosa and particulate DMSP; concentrations increased rapidly from 100 to 1650 nM in the middle of April. Highest DMS concentrations were found during the initial phase of the exponential growth of the bloom. DMS production and loss rates of DMSP and DMS were estimated experimentally during various phases of the bloom. DMS production and consumption were roughly in balance, with production only slightly exceeding consumption at the start of the bloom. Rates of production and consumption were highest during the exponential growth phase of Phaeocystis and declined in the course of the bloom (from 300–375 to less than 5 nmol dm⁻³ d⁻¹). Demethylation of DMSP increased during the bloom (from 11 to 1300 nmol dm⁻³ d⁻¹); it accounted for up to 100% of the DMSP loss at the end of the bloom. The shift from DMSP cleavage to demethylation in the course of a Phaeocystis bloom implies that DMS concentrations are not necessarily highest at the peak or towards the end of blooms.     

2005年春初浙江近海的低温特征及其对大规模东海原甲藻赤潮发生的影响

在长江口附近和浙江近海海域,近些年来每年春季几乎都发生大规模的东海原甲藻(Prorocentrum donghaiense)赤潮,时间通常是在4月底或5月初,但2005年却一直到5月底才发生。本文根据近几年在浙江近海进行的多个春季航次的调查资料和有关站位的气象资料,分析了该海区2005年春初的温度特征及其对大规模东海原甲藻赤潮发生的影响,阐明了2005年春季大规模东海原甲藻赤潮迟发的主要物理环境原因。结果表明,2005年3月浙江近海水温比多年同期平均值偏低,且这种情况在近岸区更为明显;与2004年同期相比,2005年春初(4月初)浙江近海水温偏低,2005年和2004年的这种水温差异在近岸和中上层的低盐水里最为明显,而在远岸和下层的高盐水水温差异则相对要小;近岸低盐水最低温度2005年比2004年低约2.6℃;2005年1—3月浙江近海特殊的气象条件(10 a中最低的气温和强劲的偏北大风)和2004/2005年冬季较弱的台湾暖流是形成4月初浙江近海低温特征的主要原因;2004年冬季和2005年春初浙江近水温偏低很可能是2005年春季东海大规模原甲藻赤潮迟发的主要原因。     

Mesoscale and microscale spatial variability of bacteria and viruses during a Phaeocystis globosa bloom in the Eastern English Channel

Sampling was conducted within inshore and offshore sites, characterized by highly dissimilar hydrodynamic and hydrobiological conditions, in the Eastern English Channel. The eutrophic inshore site was dominated by the influence of a dense bloom of the Prymnesiophyceae phytoplankton species Phaeocystis globosa, while the offshore site was characterized by more oceanic conditions. Within each site the microscale distributions of chlorophyll a and several flow cytometrically-defined subpopulations of heterotrophic bacteria and viruses were measured at a spatial resolution of 5 cm. The inshore site was characterized by comparatively high levels of microscale spatial variability, with concentrations of chlorophyll a, heterotrophic bacteria, and viruses varying by 8, 11 and 3.5-fold respectively across distances of several centimeters. Within the offshore site, microscale distributions of chlorophyll a and bacteria were markedly less variable than within the inshore site, although viruses exhibited slightly higher levels of heterogeneity. Significant mesoscale variability was also observed when mean microbial parameters were compared between the inshore and offshore sites. However, when the extent of change (max/min and coefficient of variation) was compared between meso- and microscales, the variability observed at the microscale, particularly in the inshore site, was substantially greater. This pattern suggests that microscale processes associated with Phaeocystis globosa bloom dynamics can generate heterogeneity amongst microbial communities to a greater degree than large scale oceanographic discontinuities.     

On an extremely dense bloom of the dinoflagellate Alexandrium tamarense in lagoons of the PO river delta: Impact on the environment

An extremely dense bloom of the potentially toxic dinoflagellate Alexandrium tamarense was observed in the lagoons of Cà Pisani (Veneto, Italy) in summer 1993. The lagoons were experiencing a significant eutrophication impact, receiving waste waters from intensive fish culture plants. During their bloom dinoflagellates in the lagoons reached densities of 2 to 4 × 10⁶ cells·dm⁻³ and a biomass of over 100 g·m⁻³. The bloom produced drastic ecological changes in the lagoons. It caused nocturnal anoxia, mortality of macrophytes and the build-up of labile organic matter in the water column. Grazing by the tintinnid Favella sp. contributed to the termination of the bloom of the flagellates. The results show that coastal aqua culture probably stimulates dinoflagellate blooms in shallow brackish lagoons.     

海蜇不同部位的氨基酸组成和含量分析

利用氨基酸自动分析仪对新鲜海蜇(Rhopilema esculentum Kishinouye)3个部位(伞部、口腕部和生殖腺)的氨基酸成分和含量进行了分析。结果表明,3个部位均含有所测定的17种氨基酸,总氨基酸质量比分别为146.6,150.0和245.6 mg/g,人体必需氨基酸质量分数分别为29.33%,29.46%和37.17%。海蜇的氨基酸分析结果显示,海蜇具有较高的营养保健价值。     

Studies on the absorption and utilization of amino acids in the test diets by the prawn Penaeus orientalis

This experiment was conducted to study the characteristics of absorption of amino acids by different regions of the alimentary tract and the utilization of free amino acids in the test diets of the prawn, Penaeus oricntolis.The midgut gland is the principal site of nutrient absorption. In the foregut, the digestibility coefficient of 15 amino acids tested accounted for 52.5% of the total digestibility coefficient in the entire alimentary tract. In the midgut, the digestibility coefficient was 47.5 % of the total. There was no evidence of the hindgut assimilating amino acids.The free amino acid in the test diet, methionine, was almost absorbed before entering the midgut. Methionine could not be absorbed synchronously with other amino acids bound in the dietary protein, but it did affect the synchronization of other essential amino acids. [3H]-lysine, incorporated into the test diet, was almost totally assimilated within the midgut gland. Five and a half hours after feeding, the labelled amino acid had be     

条斑星鲽幼鱼变态期间核酸及蛋白的变化

本试验测定了条斑星鲽(Verasper moseri)幼鱼变态过程中DNA、RNA、蛋白及其比值的变化,作为评价幼鱼变态阶段生长潜能的指标。结果表明,试验期间培育水温为16.5～18.0°C,DNA、RNA及蛋白的变化都带有发育阶段特异性。DNA含量在34～42日龄增加缓慢,然后快速增加直至44日龄,在45日龄和46日龄分别保持下降和上升趋势。蛋白含量和RNA含量的变化趋势相似,在试验期间保持快速增长的趋势。RNA/DNA比值从试验开始至35日龄呈上升状态,36日龄时下降,38日龄时达到最高值5.01,然后显著下降(P0.05),在试验结束时达到最低值1.76。蛋白/DNA在40日龄时达到最高值58.64,在46日龄时达到最低值21.28。RNA、DNA及蛋白含量跟全长和体质量有明显的线性关系(P0.05)。RNA/DNA与全长和体质量的关系比蛋白/DNA与全长和体质量的关系密切。蛋白/DNA比值的变化趋势跟RNA/DNA比值类似,但前者的变化滞后于后者,表明RNA/DNA比值是评价条斑星鲽生理状况的更有效的生理指标。