养殖水体中氨氮对大菱鲆的急性毒性效应研究

为了评估大菱鲆(Scophthalmus maximus)耐氨氮性状的遗传参数和为循环水养殖大菱鲆提供科学指导,作者研究了养殖海水中氨氮对4月龄和8月龄大菱鲆的急性毒性效应,同时对比了氨氮对白化和正常个体、雌性和雄性个体的急性毒性效应差别。结果表明:非离子氨对4月龄大菱鲆的24、48、72、96 h LC50分别为2.19、1.94、1.80、1.72 mg/L,对8月龄大菱鲆的24、48、72、96 h LC50分别为3.64、3.02、2.93、2.86 mg/L,非离子氨对4月龄和8月龄大菱鲆的安全质量浓度分别为0.17和0.29 mg/L;氨氮对白化和正常大菱鲆以及雌性和雄性大菱鲆的急性毒性效应均没有显著性差异,研究结果为进一步选育大菱鲆的耐氨氮品系提供科学依据。     

Accumulation and effects of benzo(a)pyrene on cytochrome P450 1A in waterborne exposed and intraperitoneal injected juvenile turbot (Scophthalmus maximus)

Juvenile turbot (Scophthalmus maximus) were exposed to benzo(a)pyrene (BaP) for 14 d using a glass bead generator flow-through system. Exposure was followed by a recovery period of 16 d. The highest BaP concentration in the edible portion of the fish, 16.5 ± 4.3 μg BaP/kg, was observed on the first day. Then concentrations dropped following first-order kinetics. BaP was below detection level at the end of the experiment. A statistically significant increase in bile fluorescence was observed from day 9 until the end of the experiment, suggesting the elimination of BaP metabolites by this route. No significant differences between control and exposed fish in EROD activity and CYP1A concentration, measured by immunodetection method, were observed. Intraperitoneal injection of 2.5 mg BaP/kg in juvenile turbot induced EROD activity. Under waterborne experimental conditions, bile fluorescence was observed to be a more sensitive biomarker of BaP exposure than EROD activity and CYP1A measurement.     

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。     

Potassium channel in peripheral blood lymphocytes of turbot (Scophthalmus maximus)

In order to provide pertinent evidence of ion channel with immune response in the fish,whole cell patch-clamp technique was employed for potassium ion channel study in turbot(Scophthalmus maximus) . Lymphocytes were isolated by Percoll density gradient centrifugation from peripheral blood samples,and electrophysiological characters of the channel were analyzed. In the recorded cells,activated voltage of the channels was-42.5 ± 3.7 mV and the average peak current was 313.12 ± 28.2 pA. The channel was identified as voltage dependent,the current was outward and it could be inhibited by 10 mmol/dm 3 TEA or 5 mmol/dm 3 4-AP,a specific potassium channel inhibitor,identifying the existence of potassium channel in peripheral lymphocytes of the turbot.     

大菱鲆精子运动特征与精液pH的相关性研究

为探究大菱鲆(Scophthalmus maximus)精液质量与pH的关系及甘油对大菱鲆精子激活的影响,本研究通过计算机辅助精子分析系统分析检测了威海和烟台4家大菱鲆育苗场(A、B、C、D)的精子质量。研究发现,不同育苗场的大菱鲆精子活力存在显著的差异,A2A1DCB,精子预测寿命分别为11.7、7.1、6.8、4.6、3.3 min,所取精子的质量的存在显著差异。研究结果发现,大菱鲆精子运动特性与精子活化率相关性显著,其中精子活化率与平均曲线运动速度、平均直线运动速度、平均路径速度、精子头侧摆幅度、精子平均鞭打频率呈显著正相关(P0.05),与运动直线性、运动摆动性、运动前向性等参数无显著相关性;大菱鲆精子运动特性受pH影响显著,精液pH与平均曲线运动速度、平均直线运动速度、平均路径速度、精子头侧摆幅度、精子平均鞭打频率呈显著正相关(P0.05),而与运动直线性、运动摆动性、运动前向性等参数无显著相关性。同时还发现,精子活化率高、寿命长的大菱鲆精液pH偏弱碱性(pH=8.0),即大菱鲆精子适宜在弱碱性环境中活动。除此之外,向激活海水中加入梯度甘油(0.1、1、10、100、500、1 000 mmol/L),发现可以显著提高大菱鲆精子寿命,其中甘油浓度为100 mmol/L时,精子寿命延长至15.17 min±0.29 min,推测甘油在精子激活后可能被精子吸收并氧化分解提供能量。     

大菱鲆体重性状遗传参数及选育遗传进展研究

本研究估计了大菱鲆收获体重性状的遗传参数和选育遗传进展。数据共包括3个世代（G0,G1,G2）的508个全同胞家系10952尾个体。G0,G1,G2的体重估计遗传力分别为0.11±0.08,0.18±0.09,0.17±0.07;世代间估计遗传力为0.19±0.04。每一世代母本和共同环境效应分别为0.10±0.04,0.14±0.04,0.13±0.03;世代间为0.12±0.01。G0和G1世代选择差分别为,18.24g和21.19g。对应的G1和G2世代的遗传进展为22.06g,11.93g;百分比表示分别为6.36%,3.52%。连续两代选择之后总遗传进展为10.10%。以上结果说明针对大菱鲆体重性状的选育项目是成功的。     

大菱鲆周期蛋白依赖激酶基因cdk1、cdk6克隆及表达分析

通过同源克隆以及5′/3′RACE的方法扩增得到大菱鲆(Scophthalmus maximus)细胞周期蛋白依赖激酶基因cdk1和cdk6的全长c DNA序列,在m RNA水平上分析了它们组织表达特征,并对比分析了静水压处理对其胚胎期表达的影响。结果显示,cdk1基因全长c DNA序列为1281 bp(Gen Bank登录号:KP339306),编码区为912 bp;cdk6基因c DNA全长1400 bp(Gen Bank登录号:KT186374),编码区长度为978 bp。组织RT-PCR分析表明,cdk1基因表达具有广泛性,在性腺、肾脏等生长发育较快的组织中表达量较高;cdk6基因也在多个组织中表达,在性腺中表达量最高。通过Real time RT-PCR检测基因在胚胎中的表达水平发现,静水压处理组cdk1、cdk6在胚胎发育中,总体变化趋势与对照组类似。大菱鲆胚胎对照组中cdk1在原肠期以前相对神经胚期及以后时期表达量较高,静水压处理组基因表达变化与对照组趋势相同;对照组中cdk6在原肠期出现表达高峰,但静水压处理组在原肠期表达量相对较低。静水压处理对cdk1和cdk6的表达量的影响不同,这可能与基因还有除调控细胞周期的其他功能有关。为解析这两个基因在大菱鲆胚胎发育中的作用及其机制提供了参考。     

Morphological and histological changes in the brains of turbot (Scophthalmus maximus) with gonadal development

The brain plays a critical role in controlling reproduction through the hypothalamus-pituitary-gonadal (HPG) axis in vertebrates. Turbot (Scophthalmus maximus) has become an economically important marine fish in Europe and North China. Previous research investigating turbot reproduction has focused on the role of the HPG axis in regulating egg and sperm production. However, the morphology and histology of the organs in the HPG axis have not been studied. In this study, we investigated the morphology and histology of brains in female and male turbot at different stages of gonadal development. The results showed that the brains of both female and male turbot were composed of seven parts that are typical of advanced teleosts: the telencephalon, diencephalon, cerebellum, hypothalamus, pituitary gland, myelencephalon, and olfactory bulbs. The telencephalon was well-developed and contained five distinct lobes, with the contiguous diencephalon at the caudal portion. The torus longitudinales and rostral torus semicircularis of the mesencephalon flattened along the dorsal surface, and the rostral corpus cerebellum was located in the dorsal portion. The actual total brain volume in mature males was significantly greater (p<0.05) than that of females with gonadal development. Notably, the pituitary volume in male turbot significantly increased (p<0.05) from immature to mature stage, but this difference did not occur in females. The data together illustrate a distinct sex difference in the turbot brain during gonadal development, providing insight into their HPG axes.     

Genetic parameter estimation for juvenile growth and upper thermal tolerance in turbot (Scophthalmus maximus Linnaeus)

Twenty-six half-sib groups(53 full-sib families) of turbot, Scophthalmus maximus Linnaeus, were obtained by artificial insemination. We measured growth in the offspring(40–50 individuals/family) and subjected them to a thermal tolerance challenge over a period of 34 d. There was no significant difference in daily mortality(range: 0.580%–1.391%) between Days 1–13 during the thermal tolerance challenge. However, daily cumulative mortality increased rapidly between Days 14 and 29, especially on Days 15 and 16(20.232% and 34.377%, respectively). Mortality was highest on Day 16(14.145%). We estimated the genetic parameters using the average information restricted maximum likelihood method. We used a likelihood ratio test to evaluate the significance of effects in models with and without identity as an effect, and compared the final log-likelihoods(maximum log L). Lastly, we estimated phenotypic and genetic correlation between the upper thermal tolerance limit(UTT) and body weight(BW). In this study, the positive phenotypic correlation was low between UTT and BW(0.093±0.029). The genetic correlation between UTT and BW was negative(-0.044±0.239). The heritability for upper thermal tolerance was low(0.087±0.032), which is of approximately moderate heritability. The heritability for body weight was high(0.303±0.074). Our results suggest there is significant potential for improvement in the culture of turbot by selective breeding.     

Changes of melatonin and its receptors in synchronizing turbot (Scophthalmus maximus) seasonal reproduction and maturation rhythm

In most fish, reproduction is seasonal or periodic under the suitable conditions. In turbot(Scophthalmus maximus) farms, one of the most economically important marine flatfish species, changes in daylength could cause changes in the spawning time. In this study, to characterize the regulation of reproductive physiology following light signals, three melatonin receptors(Mtnr) investigated in turbot were named sm Mtnr1, sm Mtnr2,and sm Mtnr1 c. Distinct expression profiles demonstrated that Mtnr m...     

水位深度对工厂化流水养殖大菱鲆生长的影响

为探讨水深对工厂化流水养殖水环境的影响,本实验将9 000尾初始体质量为141.62±24.47 g的大菱鲆(Scophthalmus maximus)按照低水深(20 cm)、中水深(40 cm)、高水深(60 cm)条件分为3个不同养殖水深组,实验周期为80 d。实验期间,跟踪检测长期和特定时期(投喂后8 h内)不同养殖水深水体中总氨氮(TAN)、亚硝酸盐(NO₂^--N)、固体悬浮物(SS)、化学需氧量(COD)等参数,并在实验结束时对大菱鲆成活率、体质量、饲料系数水平进行测量。研究表明,水池内水流速度与水深呈负相关,但各组间无显著性差异。高水深组的固体悬浮物含量显著(P<0.05)低于其他两组,低水深组的化学需氧量显著(P<0.05)低于其他两组,各水深组中氨氮、亚硝酸盐都在大菱鲆幼鱼安全浓度范围内,且无显著性差异。在投喂后,固体悬浮物含量在各水深组中呈先升高后降低趋势,其中低水深组波动最大。氨氮含量在投喂后3 h开始上升,其中低水深组涨幅最大。各水深组中化学需氧量随着投喂时间延长而逐渐积累,而亚硝酸盐含量基本保持不变。实验结束,低水深组中大菱鲆增质量率、特定生长率、体质量变异系数均显著(P<0.05)高于高水深组,而存活率、肥满度、饲料系数在各组之间没有显著差异。研究结果显示,增加水深有利于提高养殖水环境水质恶化的缓冲能力。在保证养殖系统水质指标安全的前提下,低水深在大菱鲆工厂化流水养殖中是一个可行的方案。     

Rearing in intermediate salinity enhances immunity and disease-resistance of turbot (Scophthalmus maximus L.)

Studies were conducted to investigate the non-specific immune responses and disease-resistance of juvenile turbot Scophthalmus maximus,cultured at four different salinities(8,20,32 and 40) . Three concentrations(3.75 × 10 7,3.75 × 10 8 and 3.75 × 10 9 CFU/ml) of Vibrio anguillarum suspension were employed at each salinity to determine the 4-day LD 50 . The serum lysozyme activity,the alternative complement pathway activity(ACH50) and the phagocytosis percentage of head kidney in turbot were tested at 24,48 and 72 h post-challenge of V. anguillarum(1.1 × 10 8 CFU/ml,0.1 ml) ,respectively,to evaluate the non-specific immune responses at the selected rearing salinities. Fish reared at salinity 20 had the lowest mortality,namely,the highest 4-day LD 50 value(8.88 ± 0.17) . Besides,the lysozyme activity,ACH50 and the phagocytosis of turbot were the highest at the salinity 20,but with the lowest at the salinity 40 treatment regardless of sampling time. In addition,the non-specific immune activities kept increasing within 72 h post-challenge of V. anguillarum,except that the lysozyme activity increased from 24 to 48 h,and then decreased from 48 to 72 h at 40 significantly. These results together indicate that rearing in intermediate salinity(20) was able to enhance the immunity and disease-resistance of turbot.     

减数分裂期雄性大菱鲆孕激素及受体基因的表达分析

为研究孕激素在大菱鲆(Scophthalmus maximus)精原细胞增殖到减数分裂过程中的作用,以6~22月龄的大菱鲆精巢为材料,通过组织学方法和定量amh、sycp3基因确定精原细胞增殖期和减数分裂期的大菱鲆精巢发育过程。通过酶联免疫吸附技术(ELISA)检测6~22月龄雄性大菱鲆血清中的孕酮(P4)和17α,20β,双羟孕酮(DHP)含量变化,利用q RT-PCR技术和原位杂交技术检测孕酮核受体(pgr)和膜受体(m PRα)在不同组织和不同发育时期的表达情况。结果表明孕激素在精原细胞增殖期高表达,开始出现初级精母细胞时降低,在精子细胞数量增加时表达量再次升高,在精子细胞变态形成精子时降低。pgr在减数分裂初期定位于精巢中的sertoli细胞,在精原细胞增殖至减数分裂期表达量逐渐增加,出现精子后显著降低;m PRα在精原细胞增殖和初级精母细胞增加时表达量都很低,在精子细胞不断增加时显著增加。推测在精原细胞增殖和减数分裂阶段孕激素可能主要通过调节pgr的表达量来促进精巢发育,而在减数分裂II期pgr和m PRα都发挥作用,在精子细胞变态成熟时,主要是m PRα发挥作用。     

大菱鲆CD55负调控补体激活及其广谱细菌结合能力

CD55蛋白在哺乳动物中是一种补体调控因子,但其在鱼类中的功能未知。本研究探索了大菱鲆(Scophthalmus maximus) CD55(SmCD55)蛋白的功能。结果表明,SmCD55蛋白由344个氨基酸残基构成,含有一个N端信号肽和3个补体调控蛋白(CCP)功能域。SmCD55基因在大菱鲆多种组织中均有表达,其中,在肌肉中表达量最低,在脑组织的表达量最高。通过原核表达获得了重组的SmCD55(rSmCD55)蛋白,发现rSmCD55蛋白能够抑制大菱鲆血清的溶血活性和杀菌活性,表明其负调控补体系统的激活。此外,本研究还发现rSmCD55蛋白可以结合包括重要水产病原菌迟缓爱德华氏菌(Edwardsiella tarda)、荧光假单胞杆菌(Pseudomonas fluorescens)、鳗弧菌(Vibrio anguilla­rum)、哈维氏弧菌(Vibrio harveyi)和海豚链球菌(Strep­tococcus iniae)在内的多种细菌,其中与哈维氏弧菌的结合能力最强。以上这些研究结果丰富了鱼类补体系统的理论知识,加深了对鱼类补体激活调控的了解。     

大菱鲆家系选育二代7种免疫因子的分析

为选育高成活率的大菱鲆(Scophthalmus maximus)抗鳗弧菌(Vibrio anguillarum)群体,对30个大菱鲆选育二代家系(F2)和1个普通养殖群体进行鳗弧菌攻毒实验(周期为14 d),统计分析死亡率。选取死亡率不同的6个选育家系和普通养殖群体构成7个实验组,采用半定量-聚合酶链反应(semi-quantitative polymerase Chain reaction,RT-PCR)技术对它们肝脏、脾脏、头肾的相关免疫因子——溶菌酶(Lysozyme)、抗菌肽(Hepcidin)、热激蛋白70(HSP70)、热激蛋白90(HSP90)、免疫球蛋白(Ig M)、C-型凝集素(C-type lectin)、Lily-型凝集素(Lily-type lectin)的表达量开展研究,并应用SPSS 16.0软件对攻毒前后各免疫因子的表达量与攻毒后存活率之间的相关性进行分析。研究结果表明:攻毒前后选育家系幼鱼肝脏、脾脏、头肾中7个免疫因子的表达量普遍高于普通养殖幼鱼,且在7个实验组中,攻毒后的表达量相较攻毒前均呈现下降趋势;攻毒前,肝脏中lysozyme、Ig M的表达量与存活率为极显著性正相关(P0.01),HSP70、HSP90、C-type lectin、Lily-type lectin的表达量与存活率为显著性正相关(P0.05);脾脏中,Hepcidin、HSP70和HSP90的表达量与存活率为极显著性正相关(P0.01);头肾中,HSP70的表达量与存活率为显著正相关(P0.05)。综上,可以推断选育家系相对于普通养殖群体大菱鲆抗鳗弧菌性能更强,且7种免疫因子在鳗弧菌感染鱼体的过程中发挥重要的抗感染作用;从F2中获得一个抗鳗弧菌性能较强的家系(23号),可用于今后大菱鲆抗鳗弧菌品系的选育指导工作,为大菱鲆高成活率群体的选育提供参考资料和理论依据。     

养殖密度对循环水系统中大菱鲆生长和蛋白质代谢的影响

在工厂化循环水养殖系统中,将初始体质量为186 g±2.0 g的大菱鲆(Scophthalmus maximus)放养于低(9.4 kg/m~2±0.2 kg/m~2)、中(13.6 kg/m~2±0.8 kg/m~2)、高(19.1 kg/m~2±1.3 kg/m~2)3个养殖密度,以研究不同养殖密度对大菱鲆生长、消化酶和蛋白质代谢的影响。养殖120 d后,低、中、高试验组养殖密度分别增长至26.1 kg/m~2±1.2 kg/m~2、38.2 kg/m~2±2.5 kg/m~2、52.3 kg/m~2±3.6 kg/m~2。结果表明:低密度组和中密度组中大菱鲆增质量率、特定生长率、肥满度和蛋白质效率均显著(P0.05)高于高密度组;而饲料系数显著低于高密度组(P0.05)。低密度组和中密度组大菱鲆总蛋白酶和淀粉酶活力显著低于高密度组(P0.05);但脂肪酶活力在3个密度组之间无显著性差异。与低密度组相比,高密度组显著提高了谷氨酸脱氢酶活力,同时降低了谷草转氨酶和亮氨酸氨肽酶活力(P0.05),而对谷丙转氨酶活力无任何影响。综上所述,在工厂化循环水系统中,增加养殖密度能提高养殖的产量,但过高的养殖密度会对大菱鲆生长、消化酶活力以及蛋白质代谢产生不利的影响。     

寄生养殖大菱鲆的海洋盾纤类纤毛虫（弗州拟尾丝虫相似种）的形态学和18S rDNA 序列分析

从养殖大菱鲆（Scophthalmus maximus）患病鱼体中分离出一种寄生纤毛虫, 通过活体观察、碳酸银法染色对其形态学特征进行分析, 初步鉴定为弗州拟尾丝虫相似种（Parauronema cf. virginianum Thompson, 1967）。同时对其18S rDNA 序列进行了测定和分子系统发育学分析...     

养殖密度对陆基工厂化循环水养殖系统中大菱鲆(Scophthalmus maximus)幼鱼生长和胁迫反应的影响研究

Stocking density is widely recognized as a critical factor in aquaculture and a potential source of long-term stress.The influence of stocking density on growth and stress response of juvenile turbot(Scophthalmus maximus, ~3–75g, initial to final weight) was examined in fish held under low(LD, ~0.21–5.31 kg/m~2, initial to final density),medium(MD, ~0.42–10.81 kg/m~2) and high stocking density(HD, ~0.63–14.27 kg/m~2) for 120 days in a recirculating aquaculture system(RAS). In this trial, the growth curve for weight of juvenile turbot in RAS, all fitted by the Schnute model. No significant difference was found in growth performance among the three densities until at the final sampling(Day 120). The final weight and body weight increase(BWI) in the HD group were significantly lower than in other groups(P0.05, weight:(75.83±2.49) g,(75.39±2.08) g,(65.72±2.86) g and BWI:(2 436.12±28.10)%,(2 421.29±4.64)%,(2 097.88±20.99)% in LD, MD and HD groups, respectively). Similarly, the specific growth rate(SGR), feed conversion ratio(FCR) and coefficient of variation for weight(CV_w) were adversely affected by high stocking density(P0.05). However, there was no difference in survival and Fulton's condition factor(K) of turbot among the different groups. Physiological analyses demonstrated a clear increase in the plasma cortisol level and an obvious decrease in growth hormone(GH) concentration in the HD group on Day120(P0.05). There was no significant effect of stocking density on plasma glucose, Cl– and protein levels. All these findings would provide a reference for selecting the optimal stocking density of juvenile turbot in RAS.     

Studies on the edwardsiellosis and characterization of pathogenic bacteria from diseased flounder (Paralichthys olivaceus L.) and turbot (Scophthalmus maximus L.)

1IntroductionEdwardsiellahas been recognized as a world-wide-occurring pathogen of numerous marine fishspecies,and edwardsiellosis has been recorded in adiverse array of fish species,including chinooksalmonOncorhynchus tshawytschaL.(Amandi etal.,1982),cha…     

大菱鲆低温生长遗传参数及相关的估计

The objectives of this present research were to assess the heritability of growth traits under low temperature conditions in turbot(Scophthalmus maximus L.), and to analyze the correlation between body weight(BW) and body length(BL). There were 536 individuals from 25 full- and half-sib families involved in this study. During the entire 90-day period, which was initiated at 233 dph(day old) and ended at 323 dph, the individuals' BW and BL were weighed consecutively six times every 18 days. The heritability of BW and BL and the correlation between these two traits were estimated based on an individual animal model with the derivative-free restricted maximum likelihood(DFREML) method. These results showed that the specific growth rates(SGR) of 25 families was from 0.75±0.11 to 1.05±0.14 under water temperature of 10.5–12°C. In addition, the heritability of BW and BL estimated under low-temperature were 0.32±0.04 and 0.47±0.06, respectively. The BW had a medium heritability(0.2–0.4), and the BL had a high heritability(0.45), which suggested that selection for increased weight and length was feasible. Moreover, there was potential for mass selection on growth. The genetic and phenotypic correlations between BW and BL were 0.95±0.01 and 0.91±0.01(P 0.01), respectively. A significant correlation between BW and BL showed that BL could be instead of BW for indirect selection, which could be effectively implemented in the breeding program.