大菱鲆南方移养技术研究

利用秋末至次年夏初福建省海区自然水温一般处于大菱鲆(Scophthalmusmaximus)生长最适水温范围这一条件,采用陆上工厂化养殖和海上网箱养殖二种方式,试验其养殖效果。利用海上网箱养殖方式,可以在秋末夏初自然水温处于大菱鲆最适生长范围期间,将大菱鲆鱼苗饲养至平均体质量500g/尾的商品规格。从2003年11月26日至2004年4月2日,鱼苗从平均全长11.3cm、平均体质量113.6g生长至平均全长27.1cm、平均体质量500.4g,成活率86.7%。而同一阶段,陆上水泥池的水温一般要比自然海区低1～2℃,生长速度也较慢。另外,大菱鲆对高水温有相当的耐受能力,在平均水温达27.0℃、最低25.2℃、最高达29.2℃的6月份,成活率仍可达85.9%。结果表明,在南方开展大菱鲆阶段性养殖在技术上是可行的。     

褐藻低聚糖对提高大菱鲆免疫机能的作用

研究了在养殖过程中投喂酶解制备的褐藻低聚糖对大菱鲆(Scophthalmus maximus)鱼苗免疫机能的影响,对大菱鲆鱼苗血液中的酸性磷酸酶、过氧化物酶、超氧化物歧化酶和头肾淋巴细胞吞噬能力进行了测定。实验发现,褐藻低聚糖能够提高大菱鲆血液中酸性磷酸酶、过氧化物酶、超氧化物歧化酶活力;头肾淋巴细胞吞噬率较对照组提高15.4%,吞噬指数则无显著差异。结果表明在大菱鲆鱼苗的养殖过程中褐藻低聚糖可作为一种非特异免疫调节剂提高大菱鲆机体的免疫能力。     

急性热应激对大菱鲆血液生化指标的影响

为探讨大菱鲆(Scophthalmusmaximus)大规格幼鱼热耐受性和热应激过程中敏感指标的变化,本研究设定18℃、21℃、24℃和27℃4个温度梯度,采用1℃/h的升温速率,对驯化温度为18±1℃的苗种进行96 h急性胁迫实验,统计各温度组96 h成活率和测定血浆部分生化指标的变化情况。结果表明,在pH 7.85、盐度29.5和溶解氧高于6.5 mg/L的环境条件下,平均体质量183.65±15.99 g的大菱鲆幼鱼96h高起始致死温度(UILT50)为28.05℃;温度和胁迫时间对血浆肾上腺素(EPI)、皮质醇(Cortisol)、血糖(GLU)、还原型谷胱甘肽(GSH)、超氧化物歧化酶(SOD)和碱性磷酸酶(AKP)含量/活性都存在显著影响,除GSH外,温度和胁迫时间存在显著的交互影响作用;热应激组血浆EPI含量随胁迫时间延长呈逐渐上升趋势,同一胁迫时间以21℃组含量最高;热应激组Cortisol含量的响应时间随温度升高相应提前、随胁迫时间呈先升后降的趋势,均在48 h达最高值; 24℃和27℃组GLU含量随胁迫时间呈先升后降趋势,胁迫3h～6h即达最高值,上升幅度与温度正相...     

大菱鲆配合饲料中植物蛋白替代鱼粉的可行性研究

研究了植物蛋白豆粕部分或全部替代鱼粉对大菱鲆生长和消化酶活性的影响。用含不同豆粕量的饲料对4个实验组的大菱鲆（Scophthalmus maximus L．）在18℃下进行50d的饲喂实验,结果表明不同饲料配方下的大菱鲆对蛋白质和碳水化合物的表观消化率随饲料中豆粕质量分数的增加呈现下降的趋势;生长实验表明,30％豆粕质量分数实验组和10％豆粕质量分数实验组（常规质量分数）的生长状态良好,50d的体质量增长率分别为82．25％,85．3％,特殊生长率为1．19,1．23,饲料系数分别为1．35,1．28,蛋白质效率分别为1．44,1．49,各生长参数差别不显著。50％豆粕质量分数实验组及70％豆粕质量分数实验组生长缓慢;各实验组大菱鲆消化器官比重随豆粕添加量的增多而呈增大的趋势;不同饲料配方组大菱鲆蛋白酶、淀粉酶活力随豆粕质量分数的增多而升高,脂肪酶无明显变化。以上各实验结果均表明,饲料中以豆粕替代鱼粉量20％对大菱鲆生长无负面影响。     

几种益生菌对大菱鲆幼鱼生长及消化酶活性的影响

作者以初体质量为(4.80±0.11)g 的大菱鲆(Scophthalmus maximus)为研究对象,在循环水水族箱中进行60 d 饲喂实验,研究饵料中添加单一益生菌和复合益生菌对大菱鲆生长及消化道酶活性的影响.以基础饲料为对照组,在基础饲料中分别添加枯草芽孢杆菌(Bacillus subtilis)、地衣芽孢杆菌...     

饵料中添加益生菌对大菱鲆幼鱼肠道菌群及部分免疫指标的影响

作者以初体质量为(4.80±0.11)g的大菱鲆(Scophthalmus maximus)为研究对象,在循环水水族箱中进行60 d饲喂实验,研究饲料中添加单一益生菌和复合益生菌对大菱鲆肠道菌群及部分血液免疫指标的影响。以基础饲料为对照组,在基础饲料中分别添加枯草芽孢杆菌(Bacillus subtilis)、地衣芽孢杆菌(Bacillus licheniformis)、枯草芽孢杆菌+地衣芽孢杆菌(1:1)、嗜酸乳酸菌(Lactobacillus acidophi-lus)、双歧杆菌(Bifidobacterium inopinatum)、嗜酸乳酸菌+双歧杆菌(1:1)、枯草芽孢杆菌+地衣芽孢杆菌+嗜酸乳酸菌+双歧杆菌(1:1:1:1),每克饲料约含菌3.0×109个/g干质量,共配制7种实验饲料。实验结果显示,饲料中添加益生菌对肠道中总菌数无显著影响,但显著降低了肠道弧菌数(P<0.05),显著提高了大菱鲆酚氧化酶活力、溶菌酶活性和总抗氧化力。添加不同的益生菌对血细胞数、血清蛋白浓度及超氧化物歧化酶的影响不同,例如添加的益生菌中含有乳酸菌能显著提高大菱鲆血清蛋白浓度(P<0.05),而添加芽孢杆菌无此效果。上述实验结果表明,与添加单一益生菌相比,在饲料中添加复合益生菌对降低大菱鲆肠道弧菌总数,提高机体免疫力具有更好的促进作用。     

军曹鱼生物学特性及苗种规模化繁育技术

简要介绍了军曹鱼(Rachycentron canadum)的形态、分布、食性、生殖、胚胎和仔稚鱼发育过程等生物学特性以及军曹鱼苗种繁育的技术工艺。选择2~3龄鱼作为亲鱼,在海区网箱中采用营养强化结合激素诱导的方法,使其在整个繁殖季节多次成熟产卵。在大亚湾地区其繁殖季节为4~10月,亲鱼繁殖的适宜水温为25~31℃、适宜盐度为30~34,大批量人工催产的受精率为30%~60%、孵化率为55%~82%。采用室外池塘进行种苗大规模培育,初孵仔鱼在水温26~32℃、盐度28~33条件下培育35~40 d,全长8~11 cm,达到商品苗规格。     

大菱鲆卵子排放后在卵巢腔内老化特征初探

为了探明大菱鲆(Scophthalmus maximus)卵子在卵巢腔内老化的原因及特征,采用连续跟踪采卵的方式,通过荧光染色、生理生化指标测定,对不同采卵时间(0, 3, 6, 9, 12 h)的卵子活性氧和脂质过氧化情况(氧化产物丙二醛MDA)、抗氧化系统(超氧化物歧化酶SOD,还原型谷胱甘肽GSH)、卵巢液pH值及受精率和孵化率进行对比研究。结果表明,随着卵子在体内老化时间的延长,其活性氧含量及脂质过氧化程度逐渐升高,活性氧含量及过氧化程度与排卵后卵子老化呈正相关;SOD酶活力,GSH酶活力均显著下降,MDA含量显著提高;受精率和孵化率均显著下降。排卵后6h,孵化率降到2.87%～10.60%,此时的卵质已经极度恶化,不能再继续用于后续的生产中。综上,在卵巢腔中,卵子排放后,随着时间的延长,活性氧含量显著升高,抗氧化能力逐渐下降,受精率和孵化率显著降低,老化逐渐加剧。     

基于响应面法分析温度和盐度及其交互作用对大菱鲆幼鱼抗氧化酶活性的影响

为了分析温度和盐度及其交互作用对大菱鲆(Scophthalmus maximus)抗氧化酶活性的影响,作者采用中心组合设计和响应曲面法研究温度(17~23℃)和盐度(20~40)对大菱鲆幼鱼血清超氧化物歧化酶(SOD)活力的效应。结果表明:温度对大菱鲆幼鱼血清SOD活力的线性效应极显著(P0.01),累积效应不显著(P0.05);盐度对大菱鲆幼鱼肠道SOD活力线性效应显著(P0.05),累积效应极显著(P0.01);温度和盐度对SOD活力的互作效应达到显著水平(P0.05)。响应曲面法分析表明,随着温度和盐度的增大,SOD的活力基本上均呈先降后升的趋势。建立SOD活力与温度、盐度及其温度-盐度互作效应的模型方程Y=74.76–9.53 X1+5.31 X2+1.8 X1 X2+5.31 X12+10.53 X22,并据此预测大菱鲆幼鱼血清SOD的活力。对所建模型进行优化,得到SOD活性最低的理论最佳条件为:温度21.84℃,盐度27.97,在此条件下SOD活性值为70.0463 U/mg。     

大菱鲆幼鱼视网膜结构及不同光谱下视蛋白基因表达特征的研究

大菱鲆(Scophthalmus maximus)是中国重要的养殖鱼类之一, 其生活史经历不同的栖息地和光环境, 视觉器官结构及视觉功能也具有适应性发育特征和可塑性。本研究以不同发育阶段的大菱鲆幼鱼为对象, 探究了其视网膜结构变化、视蛋白基因的表达特征及其与光谱之间的关系。结果表明, 随着大菱鲆幼鱼的生长发育, 其视网膜的外核层逐渐变厚; 视锥视杆层的厚度变化不明显; 而内核层与神经节细胞层逐渐变薄。视紫红质基因rh1与视蛋白基因总表达量的比例上升, 由2月龄的57.35%上升到9月龄77.19%; 视锥蛋白基因与视蛋白基因总表达量的比例下降; 其中红视蛋白基因lws 由2月龄的4.49%下降至9月龄的0.13%。将7月龄的大菱鲆幼鱼用不同光谱处理75 d后, 其视蛋白基因的表达会随光谱环境的变化而发生改变。与全光谱相比, rh2b1、sws2、sws1在红、黄光下的表达量显著下降, 而rh2b1在蓝、绿光下表达显著上升、sws2在绿光下表达显著上升, 其他则变化不显著(P<0.05)。黄、绿、蓝及全光谱下rh2基因家族与视蛋白基因总表达量的比例最高, 而红光下基因rh1的表达量占比最高。大菱鲆幼鱼在不同阶段以及不同光谱处理下表现出了视蛋白基因表达的可塑性以适应不同水层的光谱环境。本研究为探究大菱鲆对光环境的适应机制及工厂化养殖光照调控技术的建立提供理论参考。     

Impact assessment of modified clay on embryo-larval stages of turbot Scophthalmus maximus L.

Using modified clay is one of the most promising methods for the mitigation of harmful algal blooms(HABs).However,the environmental impact of modified clay has become a subject of global concern.In this study,turbot(Scophthalmus maximus L.)embryos were used as a model to assess the effect of modified clay on this sensitive stage of fish development.Results show that the 24 and 48h LC 50(median lethal concentrations)of the modified clay were 1.70 and 1.65 g/L,and the safe concentration was 0.47 g/L,which is much higher than the concentration used to treat HAB.The modified clay did not affect significantly the hatchability of turbot embryos but when the concentration exceeded 0.50 g/L,the deformity rate of newly hatched larvae increased significantly.The total length,specific growth rate(SGR)and yolk sac absorption rate of larvae reached their peaks at 0.50 g/L and then gradually decreased as the concentration of modified clay increased.Therefore,a moderate amount of modified clay does not harm the survival and hatching of turbot embryos,or the growth and development of newly hatched larvae.     

染料木黄酮对大菱鲆生长、消化酶活力和肠道组织结构的影响

以鱼粉为主要蛋白源配制了5种不同剂量的染料木黄酮(0、5、10、20、100mg·kg-1)的实验饲料,来研究其对大菱鲆生长、消化酶活力和肠道组织结构的影响。结果表明:饲料中添加不同剂量的染料木黄酮对大菱鲆幼鱼的存活、终末体重、特定生长率、饲料效率、摄食率以及肥满度、脏体比和肝体比均未产生显著影响(P0.05);饲料中添加染料木黄酮显著降低了鱼体粗蛋白和粗脂肪含量并显著提高了鱼体水分含量(P0.05),但对鱼体灰分含量无显著影响(P0.05);饲料中添加染料木黄酮显著降低了胃蛋白酶、胰蛋白酶和肠淀粉酶的活性(P0.05),但对肠蛋白酶和胃淀粉酶的活性未产生显著影响(P0.05);饲料中染料木黄酮的添加量达100mg/kg时会对鱼的肠道组织结构产生一定的破坏作用,而其他处理组间肠道组织结构没有明显的差异。实验结果表明,饲料中添加高剂量的染料木黄酮对大菱鲆的生长性能和消化吸收会造成一定的负面作用,使用大豆蛋白做蛋白源时,其含有的染料木黄酮的抗营养作用不可忽视。     

Estimating genetic parameters with molecular relatedness and pedigree reconstruction for growth traits in early mixed breeding of juvenile turbot

An introduced turbot population was used to establish families and to estimate genetic parameters of the offspring. However, there is a lack of pedigree information, and common environmental effects can be introduced when each full-sib family is raised in a single tank. Therefore, in the genetic evaluation, SSRs(simple sequence repeats) were used to reconstruct the pedigree and to calculate molecular relatedness between individuals, and the early mixed-family culture model was used to remove the impact of the common environmental effects. After100 d of early mixed culture, twenty SSRs were used to cluster 20 families and to calculate paired molecular relationships(n=880). Additive genetic matrices were constructed using molecular relatedness(MR) and pedigree reconstruction(PR) and were then applied to the same animal model to estimate genetic parameters. Based on PR, the heritabilities for body weight and body length were 0.214±0.124 and 0.117±0.141, and based on MR they were 0.101±0.031 and 0.102±0.034, respectively. Cross validation showed that the accuracies of the estimated breeding values based on MR(body weight and body length of 0.717±0.045 and 0.629±0.141, respectively) were higher than those of PR(body weight and body length of 0.692±0.052 and 0.615±0.060, respectively). The MR method ensure availability of all genotyped selection candidates, thereby improving the accuracy of the breeding value estimation.     

Egg production of turbot,Scophthalmus maximus,in the Baltic Sea

In the brackish water Baltic Sea turbot spawn at ~ 6–9 psu along the coast and on offshore banks in ICES SD 24–29, with salinity influencing the reproductive success. The potential fecundity (the stock of vitellogenic oocytes in the pre-spawning ovary), egg size (diameter and dry weight of artificially fertilized 1-day-old eggs) and gonad dry weight were assessed for fish sampled in SD 25 and SD 28. Multiple regression analysis identified somatic weight, or total length in combination with Fulton's condition factor, as main predictors of fecundity and gonad dry weight with stage of maturity (oocyte packing density or leading cohort) as an additional predictor. For egg size, somatic weight was identified as main predictor while otolith weight (proxy for age) was an additional predictor. Univariate analysis using GLM revealed significantly higher fecundity and gonad dry weight for turbot from SD 28 (3378–3474 oocytes/g somatic weight) compared to those from SD 25 (2343 oocytes/g somatic weight), with no difference in egg size (1.05 ± 0.03 mm diameter and 46.8 ± 6.5 μg dry weight; mean ± sd). The difference in egg production matched egg survival probabilities in relation to salinity conditions suggesting selection for higher fecundity as a consequence of poorer reproductive success at lower salinities. This supports the hypothesis of higher size-specific fecundity towards the limit of the distribution of a species as an adaptation to harsher environmental conditions and lower offspring survival probabilities. Within SD 28 comparisons were made between two major fishing areas targeting spawning aggregations and a marine protected area without fishing. The outcome was inconclusive and is discussed with respect to potential fishery induced effects, effects of the salinity gradient, effects of specific year-classes, and effects of maturation status of sampled fish.     

大菱鲆鳍细胞系的建立

建立大菱鲆(Scophthalus maximus)鳍细胞系,文中采用胰蛋白酶、透明质酸酶和Ⅱ型胶原酶消化法启动大菱鲆鳍组织的原代培养,并通过培养液配方和培养条件的优化成功进行大菱鲆鳍细胞的继代培养。研究结果显示,在pH=7.0~7.4、温度20~24℃的培养条件下,培养于含有表皮细胞生长因子、碱性成纤维细胞生长因子、羧甲基壳多糖、N-乙酰葡萄糖盐酸盐的Leibovitz-15培养液(20%胎牛血清)中的大菱鲆鳍细胞,其生长分裂状况最好,细胞形态为成纤维细胞样。经继代培养后,细胞生长分裂依然十分旺盛,第60代大菱鲆鳍细胞系的群体倍增时间为62.4 h,虽然出现了染色体的非整倍性,但其特征性染色体仍为44条。该细胞系细胞经液氮冻存后仍保持有原有形态和较高存活率。现已成功建立了连续性大菱鲆鳍细胞系,目前已传至第65代。该细胞系的建立对于查清病毒对大菱鲆细胞的感染途径与感染机理等具有重要的理论意义,对于病毒疫苗研制具有重要应用价值。     

Hepatic biomarkers of sediment-associated pollution in juvenile turbot, Scophthalmus maximus L

Hatchery-reared turbot (Scophthalmus maximus L.) were exposed for 3 weeks, under laboratory conditions, to sediment collected from polluted sites in Cork Harbour and a reference site at Ballymacoda, Co. Cork, Ireland. The potential of surficial sediment for inducing hepatic biomarkers was assessed at two levels of biological organisation: expression of cytochrome P450 [Western blotting analysis and 7-ethoxy-resorufin O-dealkylase (EROD), 7-benzoxy resorufin O-dealkylase (BROD), 7-methoxy resorufin O-dealkylase (MROD), 7-pentoxy-resorufin O-dealkylase (PROD) activities] and DNA integrity (Comet assay). Positive controls were generated, either by exposing turbot to cadmium chloride-spiked seawater (Comet assay) or to beta-naphthaflavone by intraperitoneal injection (cytochrome P450 induction). The induction of cytochrome P450 activity (EROD, MROD and PROD) in animals following a 7-day exposure to contaminated sediments was significantly higher than those exposed to reference site sediment and remained elevated thereafter; BROD was not induced. DNA single-strand breaks were also significantly higher following exposure to contaminated sediments throughout the experiment. Although no direct correlation between induction of alkoxyresorufin O-dealkylase activities and a particular chemical class was established, the induction of MROD and PROD activities in fish exposed to sediments containing complex contaminant mixtures, appeared to be more sensitive than conventional EROD activity assays. We conclude from the present laboratory study that S. maximus is a suitable sentinel species for the assessment of moderately contaminated sediments and therefore allows for the further development of this model for future, ecologically relevant, field studies.     

大菱鲆仔稚鱼期消化酶及碱性磷酸酶活性的变化

针对海水鱼类仔稚鱼期高死亡率的现状,本实验采集不同发育阶段的大菱鲆,分析测定了酸、碱性蛋白酶、淀粉酶和脂肪酶以及消化道碱性磷酸酶的活性变化,以期为提高大菱鲆仔稚鱼期的成活率和生长率以及优化饵料配方提供理论依据。结果表明,大菱鲆初孵仔鱼体内就已具有碱性蛋白酶活性,而酸性蛋白酶活性在15日龄左右胃腺出现时才测得。蛋白酶活性的3个极低值分别出现于仔鱼开口期,卵黄囊完全消失期和仔鱼向稚鱼的转变期。早期仔鱼体内表现出较强的淀粉酶活性。5日龄时其活性急剧下降,并一直保持较低的比活力水平。脂肪酶活性在仔稚鱼期一直很弱,进入幼鱼期后比活力迅速升高。碱性磷酸酶活性从初孵到50日龄左右始终保持较小幅度的上升,直到60日龄变态完成时才出现大幅度的升高。大菱鲆仔稚鱼发育过程中,消化道中各种酶活性变化显著,仔稚鱼变态完成后,蛋白酶、脂肪酶和碱性磷酸酶活性显著增强,这标志着大菱鲆由稚鱼期向幼鱼期的转变及消化机能的完善。     

饲料中的维生素E对大菱鲆幼鱼生长、脂肪过氧化及抗氧化能力的影响

采用6种维生素E(VE)含量为26、52、66、132、191、339 mg/kg饲料干质量的等氮等能实验饲料,研究饲料中维生素E对大菱鲆(Scophthalmus maximus)幼鱼生长,肝脏和肌肉中的硫代巴必妥酸反应底物(TBARS)含量、超氧化物歧化酶(SOD)活性及总抗氧化力(T-AOC)的影响。结果表明大菱鲆的生长和体成分受饲料中VE含量的影响产生的差异不显著,然而肝脏和血清中的VE含量以及肝脏和肌肉中的硫代巴必妥酸底物(TBARS)含量、超氧化物歧化酶(SOD)活性和肝脏总抗氧化力(T-AOC)则受其影响显著(P<0.05)。结果显示大菱鲆幼鱼摄取不适宜量的VE时在表现出各种缺乏症之前会先发生脂肪过氧化现象,同时饲料中的VE能够有效提高大菱鲆幼鱼的抗氧化能力。     

大菱鲆Hepcidin基因的克隆和真核表达载体的构建

Hepcidin是鱼类的一种重要的抗菌肽.通过比较GenBank中不同Hepcidin基因的同源相似性,设计出特异性的引物,以我国海水养殖大菱鲆(Scophthalmus maximus L)肝脏为模板利用RT-PCR 技术克隆获得了一条基因.使用生物信息学手段对该基因序列进行了比对和分析,证实这条基因为Hepcidin1 precursor基因,属于Hepcidin基因家族.通过设计引物在Hepcidin基因的5′和3′端分别引入EcoR I和NotⅠ酶切位点.经过双酶切后获得具有EcoR I和Not I酶切位点的Hepcidin成熟肽片段,并与同样使用该酶线性化的表达载体pPIC3.5K连接.构建了表达载体pPIC3.5K/Hepc1,并将其导入到酵母基因组中,完成了真核表达载体的构建,为Hepcidin的真核表达作了基础性的准备工作.     

大菱鲆免疫球蛋白轻链IgL全长cDNA的分离、鉴定及表达分析

实验采用末端快速扩增(RACE)技术从大菱鲆脾脏cDNA文库中筛选得到了免疫球蛋白轻链IgL的全长cD-NA片段。该序列包含47 bp的5′末端非编码区(5′UTR),738 bp的开放阅读框(ORF)和202 bp 3′UTR,整个开放阅读框编码246个氨基酸。系统发生分析表明大菱鲆IgL基因与五条鰤的IgL基因起源关系最近。RT-PCR分析表明,大菱鲆IgL基因只在正常脾脏、肾脏和头肾组织中表达;IgL在大菱鲆胚胎发育细胞期就已开始表达,在大菱鲆胚胎尾芽期和体节期表达持续增强;在鳗弧菌感染大菱鲆脾脏和头肾早期均检测到IgL基因强烈表达,后期表达逐渐减弱;大菱鲆胚胎细胞(TEC)在用鳗弧菌感染48h后,IgL开始表达。这些结果均表明IgL基因在大菱鲆免疫应答中起着重要作用。