双酚A暴露对不同黑色素含量长牡蛎的影响研究

长牡蛎的壳色有很多种,其中壳色较深个体(黑壳)比较浅个体(白壳)的外套膜中含有更多的黑色素。本研究采用高效液相色谱-质谱法(HPLC-MS/MS)测定天然海水中白壳与黑壳长牡蛎外套膜双酚A (BPA)含量,发现黑壳长牡蛎外套膜中的BPA含量较白壳长牡蛎低(P0.05)。由于黑色素合成过程中的关键——酶酪氨酸酶是一种酚氧化酶,可以通过对氧化酚类或芳胺类等多种底物的氧化起催化作用,推测其在BPA等酚类毒物的降解中也发挥重要作用。为了进一步研究不同黑色素含量牡蛎对BPA降解能力和对BPA损伤抵抗能力的差异,本实验进行了浓度为1 mg/L,1.5 mg/L和2 mg/L的BPA亚慢性暴露实验:通过对暴露后长牡蛎外套膜组织中BPA含量分析发现,白壳与黑壳长牡蛎外套膜中BPA含量均随着暴露浓度的升高而升高,且3种浓度BPA亚慢性暴露后,黑壳长牡蛎外套膜BPA含量均低于白壳长牡蛎,但是差异不显著(P0.05)。采用3种浓度BPA亚慢性暴露后,黑壳长牡蛎外套膜组织中活性氧(ROS)荧光值均显著低于白壳长牡蛎(P0.01)。通过对外套膜组织学观察发现,2 mg/L的高浓度BPA暴露使白壳长牡蛎外套膜组织发生了明显的病理变化,黑壳长牡蛎外套膜未发生明显的病理变化。可见,黑壳长牡蛎对BPA暴露可能具有更强的抵抗能力。     

Seasonal variation in the reproductive activity and biochemical composition of the Pacific oyster (Crassostrea gigas) from the Marlborough Sounds,New Zealand

Reproductive cycles of the Pacific oyster Crassostrea gigas (Thunberg) from the Marlborough Sounds, New Zealand, were followed between June 1998 and January 2000. Histological examination of the gonad confirms an annual cycle with a winter inactive period followed by rapid gonad development and a single short spawning period. The population gonad index correlated with seawater temperature and changes in tissue dry weight, condition index, and biochemical components. In winter, few individuals with early gametogenic stages were present and rapid development of primary oocytes (diam. 11 μm) occurred during spring (September‐November). The developmental rate and the diameter of mature oocytes (37 μm) was similar for the 1998 and 1999 seasons. For a standard 110‐mm‐length oyster, maximal tissue body weight and condition index were recorded in December. Rapid weight loss in January was length dependent and was attributed to spawning. Temperature was the environmental variable which best correlated with the timing of gametogenesis. Food availability (phytoplankton biomass) may have been responsible for inter‐annual variations. The biochemical composition (% glycogen, lipid, protein) of separated gonad and somatic tissues were variable seasonally and annually. Gametogenesis (oocyte diameter) was associated with increased gonad protein and glycogen and a decrease in lipid concentrations. These changes are similar to those in Pacific oyster populations from other parts of the world.     

利用四倍体与二倍体杂交规模化培育全三倍体太平洋牡蛎苗种

利用引进美国的四倍体太平洋牡蛎(Crassostrea gigas)雄贝与我国的二倍体雌贝杂交，规模化培育全三倍体牡蛎苗种。杂交组的平均受精率为48．27％，对照组为80．53％，杂交组较低的受精率可能与四倍体雄贝的精子活力及精子数量有关。杂交组的孵化率、幼虫成活率和变态率分别为81．10％，42．3％和1．82％，对照组分别为79．96％，37．0％和1.80％。杂交组共培育出稚贝1200万粒。不同时期取样的流式细胞仪分析结果表明，杂交组的三倍体率为100％。     

Multi-technique approach to characterise the effects of cryopreservation on larval development of the Pacific oyster (Crassostrea gigas)

Cryopreservation experiments were conducted on D-stage larvae of the Pacific oyster (Crassostrea gigas) to investigate the effects of two cryoprotectant solutions and three cooling rates on larval development from 1 to 22 days post-fertilisation. Cryoprotectant solutions were made up to final concentrations (after 1:1 dilution with larvae) of 10% ethylene glycol, 1% polyvinylpyrrolidone and either 0.2 or 0.4 M trehalose. Three cooling rates (0.5, 1 and 2 °C min^?1 between ?10 and ?35 °C post-holding) were tested in an orthogonal design with the two cryoprotectants. Results indicate that control larvae out-performed all cryopreservation treatments for survival, feeding consumption and shell length parameters. However, larvae exposed to 0.4 M trehalose did considerably better than those exposed to 0.2 M trehalose, regardless of cooling rate conditions. Scanning electron and light microscopy observations were used to assess larval morphology and organogenesis, indicating that treatments with surviving larvae were morphologically and developmentally similar to control larvae.     

福建太平洋牡蛎种群微卫星DNA分析

本文采用Ucdcg153、157、202微卫星引物对福建两个太平洋牡蛎(学名为长牡蛎Crassostrea gigas)养殖种群进行扩增分析和序列测定.与采自漳浦旧镇的近江牡蛎(C. ariakensis)比较,前者对以上三种引物全部呈阳性反应,后者只对Ucdcg157显阳性.与Genebank提供的相关序列比较,福建太平洋牡蛎与Genebank样品应属同源,其中漳浦霞美和厦门白礁的样品则可能代表同一种群的两个衍生品系.本项调查结果同时显示,近亲繁殖的育苗方式已经导致福建太平洋牡蛎种群裂化;任其发展不利于维护良种优势.本次采集的样品中未发现显示葡萄牙牡蛎、熊本牡蛎和美洲牡蛎微卫星特征的个体.     

长牡蛎Wnt5b基因的鉴定及在黑色素生成中的作用

Wnt基因家族编码一类保守的分泌型糖蛋白,广泛参与细胞分化与增殖、细胞运动、凋亡、免疫调控以及色素生成等生物学过程。本研究鉴定了一个长牡蛎Wnt家族基因——CgWnt5b,比较了黑、白壳色长牡蛎中CgWnt5b转录水平的表达情况,并对其在调控黑色素生成方面的功能进行初步研究。研究结果显示,CgWnt5b编码序列全长1116 bp,编码371个氨基酸。生物信息学分析预测CgWnt5b蛋白相对分子量为42 KDa,等电点为9.28。对CgWnt5b与哺乳动物Wnt基因家族19个成员进行系统发育分析,确定了CgWnt5b为哺乳动物Wnt5a和Wnt5b的同源基因。荧光实时定量PCR检测长牡蛎外套膜、鳃、消化腺、闭壳肌、性腺、唇瓣以及血淋巴组织中CgWnt5b的mRNA分布,结果显示,CgWnt5b在外套膜、闭壳肌和唇瓣组织中的相对表达水平较高,在消化腺、鳃、血淋巴和性腺组织中相对表达量较低。黑壳长牡蛎中CgWnt5b mRNA表达量显著低于白壳长牡蛎,且siRNA敲降CgWnt5b的表达后,多个黑色素生成相关酪氨酸酶基因的表达显著上调。综上,本研究在长牡蛎中鉴定了一个Wnt家族成员CgWnt5b,可能通过抑制黑色素生成关键基因TYR的表达抑制长牡蛎中黑色素的生成。     

太平洋牡蛎配子和精子入卵的扫描电镜观察及胚胎发育的初步研究

通过扫描电镜观察 ,太平洋牡蛎精子由头部、中段和尾段组成。精子全长约 2 1 .6μm ,尾段长约 1 8.2 μm ,末端观察到“针眼”样结构 ;头部最大处的直径约 2 .6μm ;中段直径约 1 .1 μm ;未成熟精子尾段中部出现膨大 ,该处直径约 0 .3μm。卵呈梨形 ;卵的直径约为 41 .9μm。卵表面有微绒毛组成的皱折结构 ,可见卵黄膜孔。卵质收回时 ,在卵表面卵柄处可见明显的树枝状结构。受精后 ,受精卵表面的卵黄膜剥离处可见较多数量的皮质小泡。在水温2 1 .2～ 2 3.5℃ ,精卵混合 34min时 ,释时出第 1极体 ;1h2 2min时 ,放出第 2极体 ;2 1h55min时 ,胚胎发育至初期D形幼虫。     

太平洋牡蛎配子和精子人卵的扫描电镜观察及胚胎发育的初步研究

通过扫描电镜观察,太平洋牡蛎精子由头部、中段和尾段组成.精子全长约21.6μm,尾段长约18.2μm,末端观察到"针眼"样结构;头部最大处的直径约2.6μm;中段直径约1.1μm;未成熟精子尾段中部出现膨大,该处直径约0.3μm.卵呈梨形;卵的直径约为41.9μm.卵表面有微绒毛组成的皱折结构,可见卵黄膜孔.卵质收回时,在卵表面卵柄处可见明显的树枝状结构.受精后,受精卵表面的卵黄膜剥离处可见较多数量的皮质小泡.在水温21.2～23.5℃,精卵混合34min时,释时出第1极体;1h22min时,放出第2极体;21h55min时,胚胎发育至初期D形幼虫.     

长牡蛎nacrein 基因的克隆、结构及进化分析

首次在长牡蛎(Crassostrea gigas)中克隆得到一种新的贝壳基质蛋白nacrein-like protein F3的全长c DNA序列。nacrein-like protein F3基因c DNA全长1499bp,其中编码区长度为1242bp,编码一条含413个氨基酸残基的多肽链。氨基酸序列比对和结构域分析均表明其为合浦珠母贝(Pinctada fucata)nacrein的同源蛋白,含有1个保守的α-碳酸酐酶结构域,但由于重复结构域的插入,α-碳酸酐酶结构域被间隔成2个亚结构域。系统进化分析显示nacrein-like protein F3与贻贝(Mytilus californianus)nacrein-like protein进化关系最近。此外,在软体动物中,双壳纲nacrein-like proteins进化速度相对较快,推测与寒武纪时期剧烈的环境变化有关,如影响贝壳形成的海水化学变化。     

Effects of sea urchin (Evechinus chloroticus) grazing in Dusky Sound,Fiordland, New Zealand

Evechinus chloroticus (Val.) is known to be an important grazer of subtidal barren habitats in northern New Zealand but its role in structuring rocky reef communities at colder southern localities is poorly understood. The present study was prompted by a proposal for the establishment of an E. chloroticus fishery in Dusky Sound, Fiordland, New Zealand. To determine the ecological importance of E. chloroticus in Dusky Sound, sea urchins were experimentally removed from shallow fucoid fringe (upper zone), mid‐depth barrens (middle zone), and deeper algal meadow (lower zone), at eight sites and compared with a paired set of eight control sites. Two years of continuous removal of E. chloroticus resulted in conspicuous changes to the algal community at all depth zones examined. The density of fucoid algae (Carpophyllum spp., Sargassum spp., Cystophora spp., and Landsburgia quercifolia) increased mostly in the upper and middle zones, although smaller increases were also observed in the lower zone. Changes in cover of herbaceous, turfing and crustose algae observed in the middle and lower zones were larger than in the upper zone. Density increases of the kelp Ecklonia radiata were observed mainly in the lower zone, although still important in the middle zone. These results show that E. chloroticus can have a strong influence on the structure of algal assemblages in Dusky Sound hence the development of an E. chloroticus fishery in this area should be treated with caution.     

Spatial variability in growth and reproduction of the Pacific oyster Crassostrea gigas (Thunberg, 1793) along the west European coast

The Pacific oyster Crassostrea gigas was introduced in Europe for commercial purposes in the mid 1960s. It was initially thought that low winter temperatures would restrain this species' reproduction and settlement; however, its present distribution in areas where no introduction has taken place suggests that natural invasion and expansion has occurred. Along the European coast, wild populations of Pacific oysters are already found from northern Germany to southern Portugal. Whether C. gigas will continue to further expand through northern waters will depend on its physiological performance. In this study, the performance of wild oyster populations has been studied in terms of growth and reproduction at three stations: La Rochelle (France; 46°N), Yerseke (Oosterschelde estuary, The Netherlands, 51°N), and Texel (Wadden Sea estuary, The Netherlands, 53°N). The French population had the lowest somatic-shell mass ratio and an increase in maximum shell length, somatic and gonadal mass was observed from France to the Netherlands. In addition, mean oocyte diameter decreased significantly from south to north. The combination of increasing gonadal mass and decreasing oocyte volume suggests an increasing reproductive output in terms of egg numbers from France to The Netherlands. Differences in temperature between locations will at least be partly responsible for the observed patterns; however, other environmental factors (such as food availability, predation pressure, sediment type and/or seston concentration) cannot be excluded. Since smaller eggs (oocytes) are thought to have a longer development time, the environmental conditions along the Dutch coast may result in increased larval dispersal and possibly in further population expansion.     

Catastrophic reduction of the oyster,Tiostrea chilensis (Bivalvia: Ostreidae), in Foveaux Strait,New Zealand,due to infestation by the protistan Bonamia sp.

Between 1985 and 1992, mortality due to infestation by the protistan Bonamia sp. severely reduced the numbers of the oyster, Tiostrea chilensis in Foveaux Strait, New Zealand. In 1990, two dredge surveys gave relative estimates, and a dive survey an absolute estimate, of the size of this oyster population; this allowed us to estimate dredge efficiency precisely. This estimate was used to re‐evaluate the dredge survey of 1975–76 and relate the size of the population in 1990 to that in 1975. By 1990 the population density had declined by 67% while a dredge survey in 1992 showed that the population density had declined by 91% from 1975. A dredge survey in 1993 showed that the population had increased slightly (but not significantly) over the 1992 level. By 1992 the population had probably been reduced below 10% of the virgin level. The fishery was closed in 1993 to allow the population to rebuild. The population surveys have followed stratified random and grid pattern sampling designs and the merits of the designs and methodology are discussed.     

Comparison and history of Polydora websteri and P. haswelli (Polychaeta: Spionidae) as mud-blister worms in New Zealand shellfish

Shells of commercially valued bivalves in New Zealand, Crassostrea gigas, Perna canaliculus and Pecten novaezelandiae, are damaged by blister-causing Polydora polychaete species known to be close in morphology to the widely recorded oyster pest Polydora websteri Hartman. Recent New Zealand occurrences are here confirmed to relate to two species, P. websteri, and a second similar species, Polydora haswelli Blake & Kudenov, a new record for New Zealand, previously known only from Australia; the two species are described and compared. The worms have limited distributions, with P. websteri confirmed only for Pacific oysters (C. gigas) in northern New Zealand, although prior reports indicate it may also occur on scallops and have reached the northern South Island. Polydora haswelli has been found only in northern New Zealand, occurring on subtidal mussels and scallops and native oysters (Perna canaliculus, Pecten novaezelandiae, Ostrea chilensis), as well as co-existing with intertidal P. websteri on Pacific oysters. The worms are not present in Foveaux Strait O. chilensis beds, a major source of past oyster exports to Australia. The history of mud-blister worm outbreaks in Australasia is examined. While trans-Tasman exports of live oysters from New Zealand were commonplace during the nineteenth century, there is no evidence that mud-blister worms were present in New Zealand then. The earliest reports only date from the early 1970s and only from northern New Zealand, whereas a century earlier in the 1870s at least one of these pest worms had become widespread along eastern Australian coasts.     

强光照和黑暗条件下刺参体壁的基因表达差异

参对光照变化非常敏感,研究刺参对光照的分子响应非常重要。本研究应用RNA测序获取了刺参暴露于强光（“强光”）、正常光照（“对照”）和完全黑暗（“黑暗”）环境下体壁的基因表达谱情况,通过“对照”与“黑暗”,“对照”与“强光”和“黑暗”与“强光”的比较,在|log₂ ratio|≥1和FDR≤0.001的标准下分别发现了1161、113和1705个差异表达基因（DEGs）。基因本体分析表明,“cellular process”和“binding”在“生物过程”和“分子功能”类别中的DEGs富集最多,而“cell”和“cell part”在“细胞组分”类别中的DEGs富集最多。将DEGs与Kyoto Encyclopedia基因和基因组数据库上的于214、41和229条通路进行比对,发现了51、2和57条通路分别显著富集。本研究发现的光特异性DEGs可作为深入研究刺参对光照变化的生化适应机制的重要目标基因。     

Recent (post-1930) changes in the extent of subtidal seagrass (Zostera muelleri) beds of the eastern Bay of Islands,New Zealand

The surface areas of 12 subtidal seagrass Zostera muelleri beds at the islands and adjacent mainland shores of the eastern Bay of Islands (35° 12′ S, 174° 10′ E), New Zealand were estimated from aerial images. It appears that little subtidal seagrass existed until after the early-1950s, so significant beds here may be a relatively recent biome. Possible explanations for recent emergence of subtidal seagrass include that debilitating periods of turbid water and pulses of sedimentation associated with the land clearances of the late 1800s had worked through the marine ecosystem. An overall peak in seagrass-cover in the 1960s to 1980s was followed by declines in several beds in the 1990s to 2000s, with recovery since. The temporal presence of seagrass was well-correlated among the three mainland beds, and moderately well among the south-facing beds on the islands, consistent with mechanisms driving seagrass establishment and persistence operating at reasonably broad scales.     

The genus Pseudo-nitzschia (Bacillariophyceae) in New Zealand: analysis of the last decade's monitoring data

The diatom genus Pseudo-nitzschia blooms throughout New Zealand's coastal waters. More than a decade of monitoring data were analysed in this study and seasonality was a significant factor in the appearance of Pseudo-nitzschia blooms. Austral spring and summer were the main bloom periods, although there were regional differences. Between January 2000 and August 2011, 8.4% of 29,000 seawater samples analysed for Pseudo-nitzschia exceeded the voluntary trigger level for biotoxin testing (10×10⁴ cells l^?1), and these were analysed by fluorescent in situ hybridisation (FISH) assays. Pseudo-nitzschia fraudulenta and P. pseudodelicatissima (low domoic acid [DA] producers) were the dominant bloom formers throughout New Zealand (each contributing 25%) followed by the highly toxic P. australis (10%). Shellfish flesh testing for DA was triggered on 8477 occasions between 2000 and 2011, but no samples exceeded the regulatory limit (20 mg/kg); the maximum concentration was 13 mg/kg (Marlborough Sounds, mid-winter 2010).