太平洋牡蛎(Crassostrea gigas)精液的超低温保存研究

筛选了六种抗冻保护剂(GLY[甘油],DMSO[二甲基亚砜],PG[丙二醇],EG[乙二醇],DMA[二甲基乙酰胺],MeOH[甲醇])及其五种不同浓度(6%,8%,10%,12%,14%,V/V)、三种稀释液(HBSS溶液,人工海水[ASW],过滤海水[FSW])、四种稀释比例(1︰1,1︰2,1︰4,1︰6)、三种添加剂(蔗糖,葡萄糖,海藻糖)、三个分步降温程序(程序A,程序B,程序C)对太平洋牡蛎(Crassostrea gigas)精液冷冻保存的影响,成功建立了太平洋牡蛎精液超低温保存方法:以10%DMSO为抗冻保护剂,HBSS溶液为稀释液,1:4的稀释比例,添加海藻糖,采用分布降温法冷冻保存太平洋牡蛎的精液,37°C水浴解冻后精液运动率达到71.27%±3.24%,显著高于其它实验组(P0.05),受精率和孵化率分别达到95.04%±1.99%、93.33%±1.33%,与鲜精(88.89%±15.16%,94.90%±0.95%)、程序冷冻前精液(95.96%±2.15%,92.67%±14.83%)差异不显著(P0.05),证明该太平洋牡蛎精液超低温保存方法可以应用于生产实践和科学研究中。     

太平洋鳕(Gadus macrocephalus)精液超低温冷冻方法的建立及精子超微结构分析

采用分步降温法冷冻保存太平洋鳕(Gadus macrocephalus)精液,并用扫描电镜和透射电镜技术研究了精子的超微结构损伤。分析了添加剂(蛋黄)及五种抗冻剂(PG(丙二醇)、DMSO(二甲基亚砜)、EG(乙二醇)、GLY(甘油)、MeOH(甲醇))不同浓度(8%、10%、12%、14%、16%、18%)对太平洋鳕精子冷冻保存效果的影响。实验结果表明,蛋黄能够显著提高太平洋鳕冷冻保存效果(P0.05);12%PG中添加10%蛋黄保存的冻融精子运动率最高(85.87%±1.6%),显著高于其他冻存组(P0.05)。添加蛋黄的12%浓度的DMSO组解冻后精子运动速率较高,平均直线速度、平均曲线速度、平均路径速度分别达到了(120.39±20.78)μm/s、(120.39±20.78)μm/s、(155.64±12.02)μm/s,与其他各实验组差异显著(P0.05)。通过扫描电镜和透射电镜观察新鲜和冻融后的鳕鱼精子发现,鲜精中75.5%精子形态结构正常、24.5%精子形态结构异常;运动率最高的冻精中67%精子形态结构正常、33%精子形态结构异常。超低温保存对太平洋鳕精子结构产生了显著影响,细胞膜破裂、线粒体肿胀变形,鞭毛脱落、断裂。     

浒苔腐烂液的生物毒性及其对太平洋牡蛎胚胎发育的影响

为防范浒苔绿潮次生灾害和降低海水养殖风险,文章采用人工授精的太平洋牡蛎胚胎、不同相对浓度的浒苔腐烂液以及基于发光细菌的生物毒性检测法,通过实验分析浒苔腐烂液的生物毒性及其对太平洋牡蛎胚胎发育的影响。研究结果表明:随着浒苔腐烂液相对浓度的提高,太平洋牡蛎D形幼虫相对孵化率逐渐降低至0,滞育和畸形等非正常胚胎增多,囊胚-原肠期和担轮幼虫期占比降至极低,易发生卵膜破裂和卵裂球分散,这些与太平洋牡蛎胚胎发育的阶段特点和浒苔腐烂液生物毒性的作用机制等有关;浒苔腐烂液相对浓度(0.25～10g/L)与生物毒性呈显著正相关,生物毒性(10%～97%)与相对孵化率呈显著负相关,且生物毒性达重毒水平时(由56%至70%)相对孵化率大幅下降(由66%至18%);建议在太平洋牡蛎胚胎发育的相对孵化率为50%时发出浒苔绿潮的渔业高风险预警,对应的浒苔腐烂液相对浓度为4.6g/L、生物毒性为55%。     

日本蟳精子获取和超低温冷冻保存技术研究

本文比较了胰酶消化法、网搓法和机械研磨法3种方法对获得游离日本蟳精子的效果。对获得的精子进行超低温冷冻保存,用曙红Y进行染色,检测精子的存活率。使用7种不同稀释液、不同浓度的二甲基亚砜(DMSO)、甘油(GLY)、1,2-丙二醇(PG)为抗冻保护剂,在6种不同的降温程序下,对日本蟳精子进行超低温冷冻保存,后在不同的温度下解冻复苏精子。实验结果表明:3种精子获取方法中以搓洗法的实验效果最好,胰酶消化法的效果次之,最差的是机械研磨法。其中搓洗法中以500目的筛绢效果最好,精子密度达2.56×10~(13)个/mL。日本蟳精子冷冻保存实验结果表明,稀释液以去钙人工海水、抗冻保护剂以5%DMSO、平衡时间20 min、降温程序P-1(0℃平衡20 min;-5℃/min降至-80℃;-80℃平衡5 min;-20℃/min降至-180℃;然后置于液氮中保存),水浴解冻37.5℃的效果最好,精子存活率最高。     

底物中钙赋存形态对太平洋牡蛎(Crassostrea gigas)幼虫附着的诱导效应

为探讨底物中钙赋存形态对牡蛎幼虫附着的诱导效应,本研究设计3因子随机区组实验,检验实验底物钙含量、钙赋存形态及海水中牡蛎幼虫丰度对实验底物上太平洋牡蛎(Crassostrea gigas)稚贝附着效果(密度和壳高)的影响。结果表明,在3个因子中,实验底物钙含量对附着稚贝密度和壳高的影响不显著(P0.05),而其它2个因子(钙赋存形态和海水中牡蛎幼虫丰度)均显著影响着实验底物上附着牡蛎稚贝的密度和壳高(P0.05)。在相同的牡蛎浮游幼虫丰度下,3种钙形态底物对牡蛎幼虫附着的诱导能力大小顺序为:有机钙碳酸钙=硫酸钙(P0.05);壳高的结果却恰恰相反。在同样的钙形态底物中,高牡蛎幼虫丰度下牡蛎稚贝附着量均显著高于低幼虫丰度的处理组(P0.05)。     

太平洋牡蛎精液的超低温保存

利用解剖法采集太平洋牡蛎精液 ,用自然海水做基础溶液 ,配制不同浓度的 DMSO(二甲亚砜 )作为超低温保护剂 ,在液氮 (L N2 )面上不同高度进行预处理 ,投入 LN2 内保存 ,升温解冻后 ,检测精子成活率。结果表明 ,DMSO处理浓度、预处理温度及时间和升温方式对超低温保存成活率均具有显著影响。其中 ,用 10 % DMSO在 LN2 面上 17cm预处理 6 min,采用 16～ 17℃流水不完全解冻 ,效果最好 ,超低温保存精子成活率可达 70 %。与新鲜精子相比 ,在 L N2 内保存一天的精子 ,授精能力没有明显变化。     

魁蚶母源大防御素在子代发育早期的动态变化

母源免疫因子能够通过卵细胞、母乳等方式转移给子代,在免疫系统发育成熟之前发挥重要的免疫保护作用,但贝类中的相关报道较少。为探索贝类非特异性免疫因子在子代发育早期的表达规律,本研究以魁蚶(Scapharca broughtonii)为研究对象,通过qRT-PCR和ELISA分析了大防御素在魁蚶胚胎及幼虫早期发育过程中mRNA和蛋白质的动态变化特征。结果显示,魁蚶卵子受精后发育至壳顶幼虫期,大防御素mRNA和蛋白质的表达变化趋势基本相同,即从受精卵期开始下降,至囊胚期最低,自担轮幼虫期表达量开始上升;与对照组相比,幼虫发育至担轮幼虫期,鳗弧菌(Vibrio anguillarum)处理组的大防御素升高更为显著,mRNA和蛋白的表达水平都基本高于对照组,说明鳗弧菌刺激能够上调担轮幼虫期后魁蚶幼体体内大防御素的表达量。总的来说,研究结果表明魁蚶母源性大防御素能够通过卵子传递给后代,从卵细胞受精后开始被消耗,当幼虫发育至担轮幼虫可能开始自身合成。     

不同固着基及2种金属离子(K~+、Cu~(2+))对香港巨牡蛎眼点幼虫固着变态的影响

用不同种类固着基及2种金属离子(K+、Cu2+)对香港巨牡蛎Crassostrea hongkongensis眼点幼虫固着变态的效果进行了研究。结果表明:黑色聚乙烯膜更有利于香港巨牡蛎眼点幼虫的固着,固着率达到49.85%,其次为黑色塑料板,固着率为19.33%,红色塑料板最低,固着率为5.14%;添加K+对牡蛎眼点幼虫的固着变态有诱导作用,当添加K+浓度为13.42mmol/L作用24h时,与对照组相比幼虫的固着提高率、变态提高率分别为42.39%、25.20%;而添加Cu2+对香港巨牡蛎眼点幼虫的固着变态没有诱导作用,随着Cu2+处理时间及处理浓度的增加,香港巨牡蛎眼点幼虫的死亡率明显提高,推测浓度过高的Cu2+对香港巨牡蛎眼点幼虫有毒性作用。     

吡啶硫酮类防污剂对华美盘管虫早期不同发育阶段的毒性效应研究

以中国南海常见的多毛类——华美盘管虫(Hydroides elegans)作为受试生物,分别研究了吡啶硫酮铜和吡啶硫酮锌对早期不同发育阶段的华美盘管虫的急性毒理效应。发现暴露于吡啶硫酮铜的华美盘管虫精子和卵子的受精半数抑制浓度(IC50)分别为19.49μg/L和88.44μg/L,担轮幼虫24 h半数致死浓度(24h-LC50)和48 h半数致死浓度(48h-LC50)分别为7.35μg/L和5.00μg/L,后担轮幼虫24h-LC50和48h-LC50分别为8.57μg/L和5.87μg/L;暴露于吡啶硫酮锌的华美盘管虫精子和卵子的受精IC50分别为36.74μg/L和159.59μg/L,担轮幼虫24 h-LC50和48 h-LC50分别为8.57μg/L和6.67μg/L,后担轮幼虫24 h-LC50和48 h-LC50分别为12.03μg/L和8.07μg/L。表明华美盘管虫早期发育的不同阶段对吡啶硫酮铜和吡啶硫酮锌的敏感性表现出一定差异,与配子受精率和后担轮幼虫的毒性反应相比,担轮幼虫对吡啶硫酮铜和吡啶硫酮锌的毒性表现出较高的敏感性;而且,对吡啶硫酮铜的敏感性普遍高于吡啶硫酮锌。     

广西茅尾海牡蛎天然种苗种类鉴定和群体组成的初步分析

了解广西茅尾海牡蛎种质资源现状,有助于提高牡蛎采苗效果。通过在茅尾海采集常见5种牡蛎成体,并在牡蛎不同繁殖高峰期分别采集牡蛎D型幼虫DⅠ、DⅡ,在不同附着期分别采集牡蛎幼苗MⅠ、MⅡ、MⅢ,采用多重PCR技术对牡蛎成体、稚贝及幼虫进行种类鉴定。香港巨牡蛎(Crassostrea hongkongensis)在茅尾海牡蛎稚贝和幼虫高峰期均属于优势种;稚贝中香港巨牡蛎、有明巨牡蛎(Crassostrea ariakensis)和熊本牡蛎(Crassostrea sikamea)3个种类的平均比例分别为88.3%、6.7%和5.0%,香港巨牡蛎与熊本牡蛎、有名巨牡蛎相比差异显著(P0.05),熊本牡蛎与有明巨牡蛎差异不显著(P0.05)。香港巨牡蛎是茅尾海牡蛎优势种,7月12日之前投放采苗器可采集到数量多、种类单一的香港巨牡蛎。7月12日之后投放采苗器,附着的稚贝中有明巨牡蛎和熊本牡蛎比例增加,影响采苗效果及养殖生产。     

Microbial Biomass and Organic Nutrients in the Deep-Sea Sediments of the Central Indian Ocean Basin

In order to assess the impact of deep-sea mining on the in situ benthic life, we measured the microbial standing stock and concentration of organic nutrients in the deep-sea sediments of the Central Indian Ocean Basin in the Indian pioneer area. Sediments were collected using box core and grab samples during September 1996. The total bacterial numbers ranged from 10 10 -10 11 cells per g -1 dry weight sediment. There was a marginal decrease in the number of bacteria from surface to 30 cm depth, though the subsurface section registered a higher number than did the surface. The highest numbers were encountered at depths of 4-8 cm. The retrievable number of bacteria were two orders less in comparison with the direct total counts of bacteria. An almost homogeneous distribution of bacteria, total organic carbon, living biomass, and lipids throughout the depth of cores indicates active microbial and benthic processes in the deep sea sediments. On the other hand, a uniform distribution of total counts of bacteria, carbohydrates, and total organic carbon in all the cores indicates their stable nature and suggests that they can serve as useful parameters for long-term monitoring of the area after the benthic disturbance. Further studies on temporal variability in this region would not only verify the observed norms of distribution of these variables but would also help to understand restabilization processes after the simulated benthic disturbance.     

海上大直径钢管桩打桩过程中桩周土强度弱化研究

海上大直径钢管桩打桩过程中,桩周土体受到强烈扰动而发生强度弱化,掌握桩周土体强度弱化规律对于准确预测打桩过程、保证工程安全具有重要意义。为研究土体强度弱化规律,开展了环剪试验模拟打桩对桩周土体的扰动,测试土体强度随剪切速率的变化规律,建立了描述土体强度弱化规律的拟合公式,引入到打桩分析软件中。研究结果表明：土体的强度折减程度不仅与土体本身的性质有关还受到土体的埋深和剪切速率的影响,埋深越深土体强度折减程度越低,剪切速率越高土体强度折减越高,在打桩分析中可采用这里推荐的线性折减方法来模拟不同深度处土体强度的折减规律。     

Geoacoustic Inversion via Genetic Algorithm and Its Application to Manganese Sediment Identification

An acoustic inversion method using a wide-band signal and two near field receivers is proposed and applied to multiple layered seabed models including a manganese sediment. The inversion problem can be formulated into a probabilistic model comprised of signals, a forward model, and additive noise. The forward model simulates wide-band signals, such as chirp signals, and is chosen to be the source-waveletconvolution plane wave modeling method. The wavelet matching technique, using weighted least-squares fitting, estimates the sediment sound-speed and thickness on which determination of the possible numerical ranges for a priori uniform distribution is based. The genetic algorithm is applied to a global optimization problem to find a maximum a posteriori solution for determined a priori search space. Here the object function is defined by an L 2 norm of the difference between measured and modeled signals. Not only the marginal pdf but also its statistics are calculated by numerical evaluation of integrals using the samples selected during importance sampling process of the genetic algorithm.     

Phylogenetic diversity and phenotypic characterization of cultivable bacterioplankton isolated from polar oceans

A set of 27 marine planktonic bacteria isolated from the polar regions was characterized by 16S rDNA sequencing and physiological and biochemical testing. More than half of these bacteria were positive for caseinase, gelatinase and 13-glucosidase, and could utilize glucose, maltose or malic acid as carbon source for cell growth. Twelve isolates expressed nitrate reduction activities. Except for one antarctic isolate BSwlO175 belonging to Actinobacteria phylum, these isolates were classified as γ-Proteobacteria, suggesting that γ-Proteobacteria dominated in cultivable marine bacterioplankton at both poles. Genus Pseudoalteromonas was the predominant group in the Chukchi Sea and the Bering Sea, and genus ShewaneUa dominated in cultivable bacterioplankton in the Prydz Bay. With sequence similarities above 97%, genus Psychrobacter was found at both poles. These 27 isolates were psychrotolerant, and significant 16S rDNA sequence similarities were found not only between arctic and antarctic marine bacteria （ 〉 99% ）, but also between polar marine bacteria and bacteria from other aquatic environments （ ≥ 98.8% ）, including temperate ocean, deep sea, pond and lake, suggesting that in the polar oceans less temperature-sensitive bacteria may be cosmopolitan and have a bipolar, even global, distribution at the species level.     

Deep-Sea Sediment Compression Curves: Some Controlling Factors, Spurious Overconsolidation, Predictions, and Geophysical Reproduction

Oedometer tests have been carried out on 70 undisturbed surficial clays (at approximately 250 mm below the mudline), mostly collected by free-fall corers from sites widely scattered throughout the deep-sea North Atlantic. Acoustic measurements were also made, initially on contiguous samples and ultimately on the same sample using a geophysically instrumented oedometer which also collected electrical resistivity data. Apart from those quiescent areas below the carbonate compensation depth, such as north of the West Indies where very fine clays exist, most of the samples are silty clays whose geotechnical-geophysical properties are dependent on the type of clay minerals present (and their ability to take in moisture), the sand-size fraction, and the quantity of carbonate present. Thus the pure clays have high compressibilities which decrease on the addition of coarse particles, while the converse is true for the acoustic parameters, these increasing with the sand fraction. Using the notion of the intrinsic compression line for all samples, and comparison to it of the measured compression curves, it is clear that, contrary to some previously held ideas, most deep-sea clays are normally consolidated; the addition of carbonate has the effect of creating an open, stronger sediment skeleton. Interestingly, where information is available, the variation with depth of a sample's acoustic velocity follows the void ratio pressure relationship of the compression curve. This allows the construction of an in-situ sediment compression curve using the in-situ geophysical observations.     

INFORMATION FOR AUTHORS

<正>Acta Oceanologica Sinica(AOS)is a comprehensive academic journal edited by the Editorial Committee of Acta Oceanologica Sinica and is designed to provide a forum for important research papers of the marine scientific community which reflect the information on a worldwide basis.The journal publishes scholarly papers on marine science and technology,including physics,chemistry,biology,     

Estimation of Sediment Properties during Benthic Impact Experiments

Sediment properties, such as water content and density, have been used to estimate the dry and wet weights, as well as the volume of sediment recovered and discharged, during benthic impact experiments conducted in the Pacific and Indian Oceans. The estimates show that the weights of dry (355-1332 t) and wet sediment (1651-4888 t) recovered during the experiments, are not only a function of the total duration and distance covered during the experiment, but also depend on water content and density of the sediment. Estimation of dry sediment and its volumetric ratio in the discharge, are key parameters for calculating the discharged volume. The estimated volume of wet sediment recovered (1427-4049 m 3 ) as well as discharged (2693-6951 m 3 ) during the experiments, provide important inputs to evaluate the sediment resettlement and migration. Using these estimates, the average depth of excavation on the seafloor, can also been calculated. However, the stages of sediment recovery and discharge are expressed differently in some of the experiments. In order to standardize these, different stages have been identified, and definitions of certain terms have been suggested, for use in the future. The methods of calculating different properties, as well as weight and volume of discharged sediment are described in the paper for use in other applications concerning deep-sea discharges.