翡翠贻贝精子的超微结构

利用透射电镜研究翡翠贻贝 (Pernaviridis)精子的超微结构。精子为典型的原生型 ,包括头部、中段与尾部三部分。头部由顶体和细胞核组成。顶体明显突出呈倒漏斗形。亚顶体腔呈锥形 ,其中的亚顶体物质呈伞状分布 ,中轴一直延伸至核的后端。细胞核近似球形 ,被管状的核前窝几乎分成相似的两部分。 4～ 5个椭圆形的线粒体围绕着中心粒复合体形成精子的中段。中心粒为中空的圆柱形 ,具有卫星体结构。尾部细长 ,轴丝为典型的“9 2”结构。本文讨论了双壳类精子形态的种属间的差异。     

翡翠贻贝外套膜转录组及贝壳珍珠质层和肌棱柱层蛋白质组分析

贝类贝壳在生物材料学及仿生学研究中占据着重要地位。贝壳基质蛋白质是贝壳中的主要有机质成分,对贝壳的形成以及贝壳的力学性能至关重要。翡翠贻贝(Perna viridis)贝壳主要由肌棱柱层和珍珠质层两种微观结构组成,其结构层次较简单,是研究贝壳基质蛋白质及其与贝壳形成关系的极好材料。为深入研究翡翠贻贝贝壳基质蛋白质的分子组成以及分布特点,首先采用扫描电子显微镜,观察翡翠贻贝贝壳内表面珍珠质层和肌棱柱层的微观结构;采用刮取法获得贝壳内表面珍珠质层和肌棱柱层的粉末;对不同层次的贝壳粉末,利用酸溶法去除碳酸钙成分,所获得的有机质组分通过离心将其分为酸可溶性组分和酸不溶性组分。采用Illumina深度测序技术对翡翠贻贝外套膜组织进行大规模测序和序列组装,在此基础上,采用LC-MS/MS质谱技术结合外套膜转录组数据库搜索,对翡翠贻贝肌棱柱层和珍珠质层贝壳基质蛋白质开展组学分析。扫描电镜观察结果表明,翡翠贻贝贝壳有两种不同形貌结构的层次,其中珍珠质层为片状堆叠结构,而肌棱柱层为柱状结构。翡翠贻贝外套膜转录组测序共计获得 69 859 条Unigene。蛋白质组学鉴定结果表明,翡翠贻贝贝壳中总计鉴定到蛋白质54种,其中38种为肌棱柱层所特有蛋白质,3种珍珠质层特有蛋白质,另有13种在珍珠质层和肌棱柱层均被鉴定到。肌棱柱层特有蛋白质的分子多样性明显强于珍珠质层。上述研究为进一步探讨贝壳不同微观层次的形成机制,以及贝壳基质蛋白质对贝壳不同结构层次的调控作用机制奠定了基础。     

厚壳贻贝一种新型贝壳胶原蛋白的重组表达与功能分析

贝壳是一种具有优异力学性能的生物硬组织,贝壳基质蛋白质对贝壳的形成具有重要意义。厚壳贻贝(Mytilus coruscus)贝壳中发现一种类似胶原蛋白质的新型贝壳基质蛋白质,命名为collagen-like protein 2（CLP-2）。然而,该蛋白质的结构与功能以及对贝壳形成的影响机制尚不清楚。为此,本研究对CLP 2开展了序列分析;进一步采取密码子优化结合原核重组表达策略,开展了CLP-2的重组表达;在此基础上分析了重组CLP-2对酸钙结晶的诱导、结晶速率抑制以及碳酸钙结合能力。对CLP-2的序列分析结果表明,该蛋白质序列中含有信号肽及两个Von Willebrand factor A（VWA）结构域。CLP-2在数据库中尚无高同源性蛋白质存在,表明这是一种较为新颖的贝壳基质蛋白。所获得的重组CLP-2对碳酸钙体外结晶表现出明显的诱导作用,扫描电镜以及傅里叶红外光谱结果表明,重组CLP-2可诱导碳酸钙晶体的形貌由立方体形转化为球形,并在高浓度下进一步转化为哑铃形;同时,重组CLP-2可促使碳酸钙晶体的晶型由方解石型向文石型转化;重组CLP-2在体外具有碳酸钙晶体结合作用;此外,重组CLP-2能显著抑制碳酸钙晶体的结晶速度（P<0.01）,并具有浓度依赖性。上述结果表明,厚壳贻贝贝壳CLP-2蛋白质在贝壳,特别是文石型肌棱柱层的生物矿化过程中具有重要作用。上述研究为深入了解贻贝贝壳的形成机制,以及胶原类蛋白质对生物矿化过程的影响奠定了基础。     

邻苯二甲酸二丁酯对翡翠贻贝抗氧化酶及脂质过氧化水平的影响

实验室条件下,研究了不同浓度邻苯二甲酸二丁酯(DBP)长期胁迫(15 d)对翡翠贻贝内脏团和外套膜抗氧化酶(超氧化物歧化酶SOD、过氧化氢酶CAT)及脂质过氧化(LPO)水平(以MDA含量表示)的影响,以及受胁迫翡翠贻贝在清洁海水中恢复阶段上述生化指标的变化特征.结果表明:胁迫阶段,0.5和2.5 mg.L-1DBP下翡翠贻贝内脏团SOD活性表现为先抑制后逐渐恢复,12.5和62.5 mg.L-1下则持续受到显著抑制;不同浓度组CAT活性均明显被抑制.LPO水平明显升高.外套膜中,2.5 mg.L-1下SOD活性受到持续诱导,其他浓度组则先被抑制,后随曝露时间延长逐渐被诱导;各浓度组CAT的变化波动较大,没有明显规律;而LPO水平明显升高.净化恢复阶段,12.5和62.5 mg.L-1DBP胁迫下的内脏团SOD和CAT活性恢复较慢,其LPO水平随时间延长逐渐恢复至对照组水平;外套膜中SOD活性呈持续升高趋势,CAT活性和LPO水平则随时间延长恢复到对照组水平.     

邻苯二甲酸二乙基己酯对翡翠贻贝(Perna viridis)生化指标的影响

实验条件下,研究邻苯二甲酸二乙基己酯(DEHP)5个浓度组(0、0.38、1.92、9.60和48.00mg.L-1)长时间胁迫下翡翠贻贝(Perna viridis)内脏团和外套膜中抗氧化酶(SOD和CAT)活性和丙二醛(MDA)含量的变化,以及胁迫解除后这些指标的恢复情况。结果表明:在胁迫过程中,翡翠贻贝内脏团SOD活性表现为先显著升高,随后受抑制而逐渐降低(P<0.05),CAT活性则表现为先被抑制后受诱导,15d后恢复到对照组水平,MDA含量呈显著增加的趋势(P<0.05);外套膜中的SOD活性在胁迫初期在低浓度组被抑制,而在高浓度组则被诱导(P<0.05),4d后SOD活性逐渐恢复到对照组水平,各浓度组MDA含量均出现明显的增加(P<0.05);净化阶段,低浓度组(0.38mg.L-1)内脏团SOD活性和CAT活性逐渐恢复到对照组水平,但MDA含量升高;净化7d后,除高浓度组(48.00mg.L-1)外,其余浓度组外套膜中SOD活性均已经恢复到对照组水平,MDA含量也没有出现明显升高的现象。研究表明,DEHP对翡翠贻贝内脏团和外套膜抗氧化防御系统酶具有明显的影响,DEHP诱导引起2种组织内脂质过氧化损伤,并且短期内这种损伤无法消除。     

壳损伤及海水酸化对厚壳贻贝脲酶和碳酸酐酶的影响

海洋酸化是当前全球面临的最为紧迫的环境问题之一,已显现出对具生物矿化现象物种的严重影响。以往研究发现,贻贝表现出对海洋酸化较强的耐受性。为探究贻贝对海洋酸化耐受性的可能机制,选择两种对生物矿化具有重要影响的酶(碳酸酐酶和脲酶)为研究对象,分析其在壳损伤以及酸化海水条件下基因表达量和酶活力的变化;进一步对上述条件下的贻贝贝壳内表面开展了显微观察。研究结果表明,相比对照组,壳损伤或酸化海水处理诱导碳酸酐酶和脲酶的基因表达量产生不同程度的上调(P<0.05),酶活力测试与基因表达量分析结果具有类似特征,但存在时序性差异。而壳损伤叠加海水酸化处理则诱导碳酸酐酶和脲酶的基因表达量及酶活性在外套膜中均明显下调(P<0.05),但碳酸酐酶在血细胞中明显上调(P<0.05);在酸化海水中添加尿素则明显上调血细胞和外套膜中碳酸酐酶和脲酶的基因表达量以及酶活性(P<0.05)。贝壳内表面显微观察结果进一步表明,海水酸化及壳损伤导致损伤部位附近的贝壳内表面产生明显纹理质地改变,尿素可诱导海水酸化条件下壳损伤部位修复层的重新出现。上述结果表明,碳酸酐酶和脲酶可能参与了对壳损伤修复及海洋...     

厚壳贻贝外套膜对细菌肽聚糖胁迫的代谢响应

为了解贻贝(Mytilus)外套膜免疫相关机制, 对贻贝进行肽聚糖(Peptidoglycan, PGN)胁迫, 并利用超高压液相色谱−质谱联用技术, 对贻贝外套膜在PGN胁迫后48h的代谢物组成及含量进行组学鉴定, 同时对胁迫前后的外套膜组织开展了游离氨基酸组成, 外套膜黏液抑菌活性及抗氧化能力分析。结果表明, PGN胁迫导致贻贝外套膜部分代谢物含量发生显著变化(P<0.05), 从中共鉴定到486种差异代谢物, 包括232种上调和254种下调代谢物; 其中, 上调差异代谢物主要富集于细胞信号转导及氨基酸代谢相关途径, 而下调SDM主要富集到脂质代谢和维生素代谢相关途径。此外, PGN胁迫导致外套膜中过氧化氢酶活力上调(P<0.05), 并导致外套膜黏液对枯草芽孢杆菌的抑制率显著上升(P<0.05)。研究为深入了解贻贝应对免疫胁迫的分子策略, 以及贻贝健康养殖奠定了基础。     

厚壳贻贝McCaspase 3-4基因的克隆及其在幼虫变态中的作用

为进一步了解细胞凋亡及其效应基因caspase-3在海洋贝类厚壳贻贝(Mytilus coruscus)变态中的作用,研究通过RACE技术克隆并鉴定了一个厚壳贻贝caspase-3基因的cDNA全长,并对该基因在幼虫变态中的作用进行了研究。碱基序列和氨基酸序列特征分析显示,该基因编码的蛋白质具有典型的caspase P20和P10结构域,但核心半胱氨酸活性位点QACXG五肽保守序列发生了突变。系统进化分析结果显示该基因与无脊椎动物Caspase-3聚在一起,并与地中海贻贝(Mytilus galloprovincialis)caspase 3/7-4相似度最高,因此将其命名为McCaspase 3-4。随后,通过实时荧光定量PCR技术对该基因在成贝不同组织及幼虫变态不同时间点的表达水平进行了分析。结果显示, McCaspase 3-4在成贝外套膜和唇瓣中的表达量显著高于其他组织。肾上腺素诱导眼点幼虫变态12h后, McCaspase 3-4的表达水平出现显著上升,并在刺激24h后达到最高,表明该基因可能在幼虫变态早期发挥作用。利用RNA干扰技术敲降眼点幼虫McCaspase 3-4基因...     

不同浓度微塑料对厚壳贻贝生理和转录物组的影响

微塑料(<5 mm)广泛分布在海洋环境中,对双壳贝类构成严重威胁。本文主要对不同浓度聚苯乙烯微塑料(PS微塑料,直径90μm, 200μg/L MPs、2μg/L MPs)对厚壳贻贝(Mytilus coruscus)的酶活、能量代谢和转录物组的影响进行研究。结果表明,厚壳贻贝微塑料暴露后SOD、CAT活性上升,MDA含量升高,摄食率显著下降(P<0.05),并呈现剂量效应。转录物组分析结果表明,与对照组相比,高浓度微塑料暴露组共鉴定到3 087个差异基因,包括上调的差异基因1 989个,下调的差异基因1 098个;低浓度微塑料暴露组共鉴定到2 184个差异基因,上调的差异基因有1 514个,下调的差异基因有670个。与低浓度微塑料暴露组相比,高浓度微塑料暴露组共鉴定到81个差异基因,包括上调的差异基因41个,下调的差异基因40个。分别对不同比较组的上、下调DEGs进行富集分析,上调的差异基因主要富集到RNA转运(RNA transport)、甲状腺激素信号通路(thyroid hormone signaling pathway)和黑色素生成(melanogenesis)等...     

厚壳贻贝两种新型防御素的分子鉴定

贻贝是全球范围内具有重要经济价值和生态价值的双壳贝类。贻贝抗菌肽具有极强的分子多样性,也是当前抗菌肽研究的重要对象。防御素是贻贝抗菌肽的重要成员, 从厚壳贻贝中鉴定到2种新型防御素, 但其分子特性和免疫机制尚不清楚。为此, 对厚壳贻贝体内新发现的2种防御素开展研究。序列分析结果表明,2种新型防御素均具有节肢动物防御素结构特征,因而被命名为arthropod like defensin (ALD)。利用荧光定量PCR研究了2种防御素在贻贝不同组织及不同发育阶段的表达量差异。进一步分析了2种防御素在3种不同微生物诱导下的表达量时间曲线。利用固相化学合成技术对2种防御素的成熟肽区进行合成并开展了功能验证。研究结果表明, 2种ALD 主要表达部位在外套膜和消化腺, 且ALD-1具有雄性特异表达特征。此外, ALD-1和ALD-2在贻贝幼虫阶段均未表达; 在不同微生物刺激下, 2种ALD表现出不同的免疫反应模式, 显示出2种防御素具有不同的免疫调节机制。化学合成的2种ALD均具有抑菌活性, 其对不同微生物的抑制率在20%~80%之间。上述研究为深入了解贻贝免疫防御的分子机制,以及贻贝抗菌肽的免疫功能和后续的分子资源开发奠定了基础。     

锡林郭勒草原布氏田鼠体重和体长关系的研究

1991～ 1998年对内蒙古阿巴嘎旗那仁宝力格苏木布氏田鼠 (Microtusbrandti)体长、体重和胴体重资料进行了调查和分析 ,得到如下结论 :田鼠体长、体重、胴体重的均值在低密度回升期均逐年增高 ;雌性越冬鼠的胴体重均值低于雄性越冬鼠且有显著性差异 (P <0 0 5 ) ;秋季雄鼠与雌鼠的 3项指标的均值无显著差异(P >0 10 ) ,体长、体重和胴体重间相关极显著 (P =0 0 0 0 1)。分别给出了体重与体长 :胴体重与体长的模型W =aLb,其中W为体重或胴体重 ,L为体长 ;胴体重与体重的关系适合于模型NW =W / (a bW ) ,其中NW为胴体重 ,W为体重。     

Influence of salinity on the physiological conditions in mussels, Perna perna and Perna viridis (Bivalvia: Mytilidae)

Perna genus was introduced to Venezuela, but nowadays, Perna perna and Perna viridis coexist and are commercially exploited from their natural beds. The aim of this work was to determine the effect of salinity on the physiological conditions of these species by studying RNA/DNA and Protein/DNA ratios. The organisms were collected from natural beds at La Esmeralda, Sucre State, Venezuela, and acclimatized for 15 days under laboratory conditions at 25 degrees C, 36 per thousand salinity, pH between 7 and 8 and more than 90% of oxygen saturation. Later, they were divided in two groups: for one group, the salinity concentration was increased (36 to 45 per thousand), and for the other, the salinity was decreased (36 to 15 per thousand). The rate of change was 1 per thousand every day. Ten organisms per group of both species were taken at each of 15, 20, 25, 30, 36, 40 y 45 per thousand salinity concentrations. Protein (colorimetric method) and nucleic acids (RNA and DNA by fluorometric method) concentrations were measured in the digestive gland, gills and adductor muscle tissues. Results indicate that P. perna can physiologically compensate the increase in salinity, but not when the salinity decreased, when proteins are the most needed macromolecules. The Protein/DNA index is directly related to salinity changes in both cases. P. viridis shows physiological compensation to salinity increases and decreases. The RNA/DNA index value in both cases supports this. Digestive gland and muscle tissues are the regulating tissues in both species. These results show that P. viridis has a higher degree of adaptability to salinity changes and, therefore, a greater potential for aquaculture than P. perna.     

Foot glands in Perna indica and Perna viridis (Pelecypoda: mytilidae) histology and histochemistry

The foot of Perna viridis is found to contain three main types of glands, the white gland, phenol gland and the enzyme gland. But in Perna indica there are only two glands, the phenol gland and the enzyme gland. Besides these, mucous glands are found in both of the species. The shape and size of the cells of these glands vary from species to species. Glycogen and 1 : 2 glycol groups are found in these gland cells. Proteins rich in disulfides and sulfhydryls are present in the phenol glands of both the species and in the white gland of p. viridis but they vary in the intensity of staining. The presence of phenols is confirmed in the phenol gland cells. Phospholipids and lipoproteins are intense in the white and phenol gland cells. They are absent in the enzyme gland. Alkaline and acid phosphatases activity in the enzyme gland cells could be demonstrated. The secretions of these glands help in the formation of the byssus threads. The mucous gland cells are subepithelial localised they secrete acid and neutral mucopolysaccharides together with glycoproteins. Which participate in the attachment of the byssus disc.     

Greening of the coasts: a review of the Perna viridis success story

The green mussel Perna viridis has been receiving a lot of attention from workers working in the research areas of intertidal ecology, aquaculture, pollution monitoring, biofouling, zoogeography and invasion biology. P. viridis is a remarkable species in terms of its ability to reach very high biomass levels, to withstand environmental fluctuations, to concentrate a variety of organic and inorganic environmental pollutants, to colonise artificial marine habitats and to invade new geographic territories. This review collates data available on salient aspects of the distribution, biology and ecology of P. viridis. It is argued that the remarkable success of P. viridis as an invasive species basically stems from its long larval duration, fast growth rate, high fecundity, early maturity, high productivity and ability to withstand fluctuating environmental conditions (temperature, salinity, water turbidity and pollutants). Relevant aspects of the data are compared with the data available for a similar species Perna perna, which too is an invasive species, but to a more limited extent.     

Characterization of subpopulations and immune-related parameters of hemocytes in the green-lipped mussel Perna viridis

The green-lipped mussel Perna viridis is distributed widely in the estuarine and coastal areas of the Indo-Pacific region and extensively cultured as an inexpensive protein source. Morphology and immunological activities of hemocytes of P. viridis were investigated using flow cytometry and light and electron microscopy. Three major types of hemocytes were identified in the hemolymph, including dense-granulocyte, semi-granulocyte (small and large size) and hyalinocyte. Other hemocytes, which occurred in low numbers, included granulocytes with different electron-dense/lucent granules and hemoblast-like cells. Based on flow cytometry, two subpopulations were identified. Granulocytes were larger cells, and the more abundant, containing numerous granules in the cytoplasm, and hyalinocytes were the smaller and less abundant with the fewest granules. Flow cytometry revealed that the granulocytes were more active in cell phagocytosis, contained the higher lysosomal content, and showed higher esterase activity and reactive oxygen species (ROS) generation compared with hyalinocytes. Immune functions assessed by the flow cytometry indicated that the granulocytes were the main hemocytes involved in the cellular defence in P. viridis.     

Eleven novel polymorphic microsatellite DNA markers from the green-lipped mussel Perna viridis

We report on the characterization of 11 polymorphic microsatellite loci in P. viridis, the first set of such markers developed and characterized for this species. The number of alleles per locus ranged from 2 to 7, whereas the observed heterozygosity ranged from 0.0447 to 0.4837. These markers should prove useful as powerful genetic markers for this species.     

Tissue-Specific Lipoprotein Lipase: Relationships to Body Composition and Body Fat Distribution in Normal Weight Humans

Twenty-six normal weight subjects (22 female, 4 male) were studied to determine the relationships of fasting levels of lipoprotein lipase in gluteal adipose tissue (ATLPL) and skeletal muscle (SMLPL) to body composition and body fat distribution. No relationship was found between fasting gluteal ATLPL and percent (%) body fat There was, however, an inverse relationship between fasting SMLPL (from the vastus lateralis) and %body fat (p=0.005). A strong inverse correlation was also seen between fasting ATLPL and waist/hip ratio (p=0.0006), a measurement of body fat distribution. These relationships existed with or without the male subjects included. The tissue-specific relationships of lipoprotein lipase to body composition and body fat distribution could relate to the development of obesity or the maintenance of normal body weight by the effects of the lipase on the partitioning of lipoprotein triglyceride fatty acids.     

Relationships Between Grain Weight and the Size of Floret Cavity in the Wheat Spike

The volume of the floret cavity at different floret positionsalong the spike and within a single spikelet was estimated in10 Triticum aestivum and three T. durum lines by injecting floretcavities with liquid silicone rubber which solidified thereafter.Highly significant correlation coefficients, ranging from 0.40to 0.76 were found between floret volume and grain weight; inmost lines the basal florets had a higher correlation than theterminal ones. The relationships between floret volume and grainweight were characterized by an intercept of about 30–60per cent of the mean grain weight (heavy-grained lines havinga larger intercept) and a slope of about 1 mg µI^–1.Differences in grain size and shape, both within spike as wellas among lines, closely reflected the variation in the sizeof floret cavity. The data support the hypothesis that grainweight is partly determined by the volume of the floret cavity. Grain weight, grain volume, floret cavity     

Biochemical markers of oxidative stress in Perna viridis exposed to mercury and temperature

Oxidative damage and antioxidant properties have been studied in Perna viridis subjected to short-term exposure to Hg along with temperature (72h) and long-term temperature exposures (14 days) as pollution biomarkers. The elevated thiobarbituric acid reactive substances (TBA-RS) levels observed in gills and digestive gland under exposure to Hg, individually and combined with temperature, as also long-term temperature stress have been assigned to the oxidative damage resulting in lipid peroxidation (LPX). Increased activities of antioxidants such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX), glutathione reductase (GR) and glutathione-S-transferase (GST) both in gills and digestive glands under long-term exposures to temperatures are more prominent to heat rather than cold stress suggesting activation of physiological mechanism to scavenge the ROS produced during heat stress. Also decreased values of reduced glutathione (GSH) on long exposures to temperature stress indicate utilisation of this antioxidant, either to scavenge oxiradicals or act in combination with other enzymes, was more than its production capacity under heat stress. The results suggest that temperature variation does alter the active oxygen metabolism by modulating antioxidant enzyme activities, which can be used as biomarker to detect sublethal effects of pollution.