The effect of PCBs on glycogen reserves in the eastern oyster Crassostrea virginica

Recent declines in Chesapeake Bay oyster populations have been attributed to disease, and reduced water quality from pollution. The stress associated with pollutant exposure may reduce energy available for growth and reproduction. Polychlorinated biphenyls (PCBs) are lipophilic contaminants that may potentially affect mobilization of lipid reserves, increasing reliance on glycogen stores, which could otherwise be utilized to supply energy for gametogenesis. Thus, PCBs may indirectly affect glycogen stores in oysters in a deleterious manner. To test for this effect, reproductively inactive oysters were exposed to PCBs by feeding individuals 0.7 g of algal paste containing 0, 0.35, or 3.5 μg PCBs daily for 8 weeks. Additionally, a group of oysters was exposed to PCBs (0, 0.35, and 3.5 μg) plus 0.3 g of non-toxic artificial sediment to examine interactive effects of sediment particles and PCBs. Adductor muscle, mantle, and gonadal tissues were analyzed for glycogen content. Results suggest that glycogen content is reduced in the adductor muscle with increasing PCB exposure, but there are no effects of PCBs in the mantle and gonadal tissues.     

Esterification of vertebrate-like steroids in the eastern oyster (Crassostrea virginica)

The esterification of two model vertebrate steroid hormones - estradiol (E2) and dehidroepiandrosterone (DHEA) - was studied in the oyster Crassostrea virginica. The activity of acyl-CoA:steroid acyltransferase was characterized in microsomal fractions isolated from oyster digestive glands. The apparent Km and Vmax values changed with the fatty acid acyl-CoA used (C20:4, C18:2, C18:1, C16:1, C18:0 or C16:0), and were in the range of 9-17 microM, and 35-74 pmol/min/mg protein for E2, and in the range of 45-120 microM, and 30-182 pmol/min/mg protein for DHEA. Kinetic parameters were also assessed in gonadal tissue. The enzyme saturated at similar concentrations, although conjugation rates were lower than in digestive gland. Preliminary data shows that tributyltin (TBT) in the low microM range (1-50) strongly inhibits E2 and DHEA esterification, the esterification of E2 being more sensitive to inhibition than that of DHEA. Overall, results indicate that apolar conjugation occurs in oysters, in both digestive gland and gonads, at a very similar rate to mammals, suggesting that this is a well conserved conjugation pathway during evolution. Esterification, together with other mechanisms, can modulate endogenous steroid levels in C. virginica, and might be a target for endocrine disrupters, such as TBT.     

Effects of PCBs sorbed to algal paste and sediments on the stress protein response (HSP70 family) in the eastern oyster, Crassostrea virginica.

This study examined the stress protein response (HSP70 family) of reproductively inactive oysters fed 0.7 g algal paste containing 0, 0.35 and 3.5 micrograms polychlorinated biphenyls (PCBs) daily. A second set of treatment groups investigated the combined effect of PCBs and sediments (0.3 g sediments daily per oyster) on HSP70 response. After 8 weeks of PCB exposure, oyster tissues (mantle and gill) were sampled and analyzed for HSP70. Preliminary results did not show a significant effect in HSP70 response in oysters fed PCB sorbed to algal paste, albeit PCBs accumulated up to 1342 ng/g dry weight in the mantle, and up to 180 ng/g dry weight in gill tissues. However, the addition of sediments caused a significant increase in HSP70 levels of gills and mantle, although the mantle was less sensitive to the sediments.     

Accumulation and cytosolic partitioning of metals in the American oyster Crassostrea virginica

The ability of the oyster to accumulate trace metals, particularly the transition metals (Cd, Cu, Zn, and Hg), from food and water has been well documented. There is little information, however, on how these metals interact to affect whole animal retention, partitioning, and binding to metallothionein (Mt). In this study oysters were exposed to Cd and Cu, both alone and in combination, and significant effects have been demonstrated on subcellular partitioning. These studies showed that Cu can displace Cd from Mt, but the Cd is not lost from the animal. Two metal-binding protein peaks have been separated by gel chromatography (<10 K and 24 K daltons). The lower molecular weight protein peak has been characterized as Mt, consisting of two isoproteins, but the higher molecular weight protein peak appears to be an aggregation of metal-binding proteins, some of which may be Mt or a Mt dimer. In environments that are contaminated with trace metals, Mt binds primarily Hg, Cu, and/or Cd and Zn, but in noncontaminated areas Mt binds primarily Zn on one of the two isoMts.     

The development of a physiologically-based pharmacokinetic model using the distribution of 2,3,7,8-tetrachlorodibenzo-p-dioxin in the tissues of the eastern oyster (Crassostrea virginica)

A physiologically-based pharmacokinetic model (PBPK) was developed to describe the kinetics of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the eastern oyster (Crassostrea virginica). The estimated t(1/2) of elimination for a bolus dose of TCDD in C. virginica is approximately 14-24 days based on both the experimental data and the PBPK model. The highest dioxin concentration reached during 28-days was in the digestive gland followed by the mantle, gonad, hemolymph, gill, adductor muscle, and the kidney/heart. A binding protein for 2,3,7,8-TCDD had been reported in the literature for both the digestive gland and gonad. Incorporating a binding component in the model resulted in a better fit for the data. The PBPK model predicted the distribution and the elimination concentrations for 2,3,7,8-TCDD within each of the tissue compartments. This model will serve as a useful tool for predicting the kinetics of other persistent organic pollutants as well as, allow for a more refined ecological risk assessment by estimating dioxin concentrations in sensitive tissues such as the gonad.     

The effects of glutathione depletion on reproductive success in oysters, Crassostrea virginica.

Glutathione (GSH) is a ubiquitous tripeptide that functions as a very important modulator of cellular homeostasis, including detoxification of metals and oxyradicals. Therefore, depletion of GSH may predispose organisms to pollutant stress. Reproductively active oysters (Crassostrea virginica) were exposed to buthionine sulfoximine in the laboratory to deplete gonadal GSH. The effects of metal exposures (Cd and Cu) on fertilization and developmental assays were evaluated using gametes from control and GSH-depleted adults. Fertilization success was not affected by GSH status, i.e. the fertilization rates of gametes derived from GSH-depleted adults were the same or slightly higher. However, GSH depletion did increase the susceptibility of developing embryos to metal toxicity, i.e. adverse effects on embryonic development were observed at lower metal concentrations with gametes derived from GSH-depleted adults. These effects may be related to diminished removal of free radicals or increased availability of metals. Whereas sperm penetration of embryonic membranes and fertilization success may be facilitated by free radicals, the persistence of free radicals during subsequent developmental periods may adversely affect differentiation and normal development. GSH probably also plays an important role in scavenging toxic metals and reducing metal interactions with essential developmental processes. These results suggest that parental depletion of GSH may increase the susceptibility of embryos to metal toxicity.     

Expression of P-glycoprotein in southeastern oysters, Crassostrea virginica

These studies provide important fundamental information regarding the expression of P-glycoprotein (p-gp) in southeastern oysters (Crassostrea virginica). Using rhodamine transport studies, p-gp activity was detected in newly fertilized embryos. A monoclonal antibody (C219) was used to evaluate p-gp expression in oyster tissues. On the basis of laboratory studies, p-gp expression tended to be higher in gill tissues than mantle tissues, and was generally not related to salinity differences. Seasonal studies were conducted with oysters collected monthly for 1 year from Lighthouse Creek, an unpolluted site. There was a general pattern of higher p-gp expression in the warmer months and lower expression in the colder months. In contrast, total gill protein concentrations decreased during the warmer months and increased during the colder months. These studies indicate that there are seasonal patterns in p-gp expression which may represent an adaptive response to natural stressors associated with summer conditions.     

Factors that influence the accumulation of copper and cadmium by transplanted eastern oysters (Crassostrea virginica) in the Patuxent River, Maryland

To investigate the continued accumulation of copper and cadmium by oysters in the Patuxent River, MD, which have been at high levels since at least the mid 1960s, hatchery-raised Eastern oysters were transplanted into trays at four sites in the upper estuary. At each site two groups of oysters were used to determine growth and mortality, and another group was sampled for meat condition, metal concentration and body burden. Copper in oysters in the discharge of a coal-fired power plant was significantly greater than at all other sites, but maximum concentration of 310 micrograms g-1 was well below the maximum of 1880 micrograms g-1 detected in 1982. Cadmium levels were also significantly greater in the discharge than at the other sites, but probably because of higher discharge temperatures. Trace metal loadings, the local salinity gradient and the higher temperatures caused by power plant operation all appear to contribute to metal accumulation by oysters in the upper Patuxent estuary.     

人工诱导香港巨牡蛎雌核发育单倍体的研究

研究利用紫外线诱导香港巨牡蛎Crassostrea hongkongensis雌核发育的条件.结果表明,在强度为1820μW · (cm2·s)-1的紫外线(254nm)下分别照射精子0,5,10,15,20,25,30,35,40,45,50,55,60,70,80和90s后,照射30s的实验组能够保持较高的受精率(61.6%),该组的D形幼虫发生率为0,出现"Hertwig效应".研究表明,在强度为1 820 μW·(cm2·s)-1的紫外线下照射精子30s是获得香港巨牡蛎雌核发育单倍体的适宜条件.研究还表明受精率和D形幼虫发生率随照射时间的增加而下降,遗传失活的精子与正常卵子受精后其胚胎发育至D形幼虫前期停止.经紫外线照射的精子授精后所产生的单倍体胚胎发育速度慢于正常受精所产生的二倍体胚胎,各照射组均出现非整倍体.     

Zinc concentration effect at the organismal,cellular and subcellular levels in the eastern oyster

The purpose of this research was to demonstrate a concentration effect of zinc exposure at organismal, cellular and sub-cellular levels in the eastern oyster and to find associated protein expression signatures (PES) for each concentration of zinc. Oysters were exposed to six concentrations of zinc for 48 h in a controlled environment. At the organismal level, fecal material was observed as a measure of physiological health during metal exposures. At the cellular level, lysosomal destabilization was measured using hemolymph. This cellular response was significant only at the highest concentration, when the fecal index was lowest. Protein responses were monitored in the oyster following exposure to zinc. Gill tissue was excised and homogenized, and then analyzed using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and digital image analysis. Protein expression signatures were found to be specific to each concentration. The protein responses were linked to the other biological parameters measured, each of which followed a concentration gradient of zinc.     

The importance of dissolved organic compounds in the accumulation of copper by the american oyster,crassostrea virginica

The influence of various dissolved organic compounds upon copper accumulation by the American oyster, Crassostrea virginica was examined. Oysters were exposed to a range of copper concentrations (^64Cu) in the presence of filtered estuarine water (FEW), UV-photooxidized FEW and FEW amended with the additions of natural dissolved organic compounds—chitin and cellular orgaanic compounds (derived from the diatom, Thalassiosira pseudonana). Copper accumulation rates were significantly greater in the UV-treated medium and decreased with the addition of dissolved organics. When normalized on a per weight DOC basis, the reductions in copper accumulation rates were greater in the presence of chitin than cellular organic compounds. The results suggest that natural dissolved organics differ markedly with respect to their influence upon copper bioavailability.     

基于特定序列的长牡蛎视黄酸反应元件预测

视黄酸受体和核受体超家族中的大部分成员对细胞整个分化、增殖过程都具有调控功能。视黄酸受体结合配体后激活,通过结合靶基因启动子区特定的核苷酸序列调控靶基因表达。视黄酸受体结合序列是由核心序列[A/G]G[T/G]TCA间隔不同碱基构成的重复序列,称为视黄酸反应元件。为了实现对长牡蛎基因组中含有的视黄酸反应元件的快速筛选预测,本研究利用Perl工具编写了一个可以批量筛选视黄酸反应元件的脚本,并对长牡蛎基因组中启动子区域序列进行筛选预测,共筛选到412个启动子区含有视黄酸反应元件的基因。随后,将这些基因在各种数据库中比对分析,预测其参与的生物学过程及可能的生物学功能。结果显示,大部分基因与蛋白质结合、核苷酸结合、水解酶活性、蛋白激酶活性等功能有关。     

Reproductive cycle and gonadial changes in the New Zealand rock oyster Crassostrea glomerata

The annual reproductive cycle and the cyclical changes in the gonad of the New Zealand rock oyster, Crassostrea glomerata (Gould, 1850), during the breeding period of 1970–71 and 1971–72 have been described. The gonad passes through an intermediate phase, after a post‐spawning period, when sex is indeterminate. Gametogenesis begins in July and August, but follicles ripen mostly during the spring months of October and November. Maximum development is seen in November, and nearly all oysters are in spawning condition in December and January. Major spawning takes place in January or February depending upon water temperature, and spawning continues until the end of March. Larvae may be found in the‐ plankton until late in the season, to the end of April or even May. Following spawning, gonadial regression sets in, with leucocyte infiltration and phagocytosis of residual gonial cells. A greater percentage of oysters more than 1 y old are females. The majority of oysters of age 1 y and below are males.     

长牡蛎贝壳损伤后的再固着能力初探

牡蛎是重要的养殖贝类,营固着生活,能聚集形成牡蛎礁,在海洋生态系统中发挥着重要作用。本研究以成体长牡蛎为研究对象,将牡蛎从附着基上剥离下来,去掉其左壳腹缘端的表面鳞片并打磨平整,露出棱柱层,将其紧密粘合于水泥砖、橡胶片、磨砂玻璃三种固着基表面,背缘端用水泥固定,观察并记录牡蛎贝壳的修复及再固着情况。结果发现：左壳损伤后,在贝壳边缘会生长出新的透明壳膜,此时壳膜虽然会紧靠固着基表面生长,但却不能再次固着;将牡蛎从固着基上剥离下来,在三种固着基表面均未发现有“附着斑”形成。因此,为了进一步研究左壳损伤后贝壳修复的分子机制以及不能再次固着的原因,对长牡蛎左壳损伤后的外套膜转录组进行了分析,发现在损伤后的第1 d、第5 d、第9 d、第13 d、第17 d和第21 d,左侧外套膜中差异上调表达的基因显著富集在TGF-β等信号通路,而下调表达的基因显著富集在对温度以及对非生物刺激反应相关通路。通过基因表达数据,还发现壳损伤后Cgfmp、CgTyr、EGF-P1等与黏附相关的基因呈下调表达,这可能是导致成体长牡蛎无法再次固着的原因。长牡蛎贝壳损伤后再固着能力的研究,为贝类生物矿化、附着等机制的研究...     

Expression and activity of a multixenobiotic resistance system in the Pacific oyster Crassostrea gigas

The multidrug resistance (MDR) mechanism corresponds to a defence system relying on the expression of high molecular membrane proteins that can actively lower the intracellular concentration of a wide variety of toxins, thus maintaining them below their toxic level. Using RT-PCR, expression levels of a gene belonging to the class I of mammalian mdr genes, has been assessed in different developmental stages of the oyster Crassostrea gigas. While no expression was found in the oocyte or the trocophore stage, a rise of mRNA content was observed from the veliger stage to the juvenile stage, thus indicating the induction of the system as the animal is developing in the environment. The incubation of gill fragments in the dye rhodamine B and subsequent measurements of intracellular fluorescence using a microplate reader indicates that the system can effectively decrease the accumulation of the test compound in a competitive manner with known inhibitors or environmental contaminants as observed in vertebrate cells. The oyster MXR system is thus becoming active in adult oyster and could be of importance in environmentally contaminated areas.     

Cellular responses and disease expression in oysters (Crassostrea virginica) exposed to suspended field contaminated sediments.

Exposure of oysters to water soluble fractions derived from field-contaminated sediments (FCS) containing predominantly lower molecular weight organic aromatic compounds, has been previously demonstrated to enhance pre-existing infections caused by the protozoan parasite, Perkinsus marinus (Dermo), and the prevalence of experimentally induced infections. To further explore the role of pollution on the onset and progression of disease, effects of suspended FCS from an estuarine creek in Virginia, USA, dominated by higher molecular weight polycyclic aromatic hydrocarbons (PAHs) on cellular responses and Dermo disease expression in oysters (Crassostrea virginica) were examined. Sediments were collected from a PAH polluted estuarine creek in Virginia, USA. To test effects on cellular response, oysters from Maine were exposed daily to 0, 1.0, 1.5, or 2.0 g suspended FCS (corresponding to 0, 70.2, 105, or 140 microg PAHs, respectively) for 5, 10, 20, and 40 days. Hemocyte activities and plasma lipid, protein and lactate dehydrogenase (LDH) levels were then measured. Exposure stimulated neutral red uptake, MTT reduction, and 3H-leucine incorporation in oyster hemocytes at various exposure times, but did not affect the plasma protein, lipid and LDH levels. To test effects on Dermo expression, oysters from a Dermo enzootic area, with an initial estimated infection prevalence of 39%, were exposed daily to 0, 1.0, 1.5, or 2.0 g suspended FCS (corresponding to 0, 75.0, 113, or 150 microg PAHs, respectively) for 30 days. Exposure enhanced disease expression in oysters. However, no significant change was noted in any measured cellular or humoral parameters.     

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.     

Effects of furadan in the brown mussel Perna perna and in the mangrove oyster Crassostrea rhizophorae

Furadan is a carbamate pesticide used widely to combat agricultural pests. However little information is available about the toxicity of furadan in aquatic macroinvertebrates. The in vivo effects of furadan were evaluated in mussels, Perna perna, and oysters, Crassostrea rhizophorae. Glutathione S-transferase (GST), catalase (CAT) and cholinesterase (ChE) activities were measured in the gills of both species exposed to furadan (100 microg/l) for 96 h. No changes were observed in GST activity in the exposed groups. CAT activity was higher (9%) in the oysters exposed to furadan. ChE activity was inhibited by 64 and 35%, respectively, in C. rhizophorae and P. perna exposed to furadan, suggesting that the former is more susceptible to the toxic effects of furadan.