大菱鲆幼鱼蛋白质的生态营养需要量研究

采用单因素随机区组设计,选择平均体质量34.5 g±5.5 g的大菱鲆(Scophthalmus maximus L.)幼鱼225尾,平均分为5组(A～E),每组3重复。分别喂以蛋白质水平为450,48%,50%,52%和54%的5种膨化颗粒饲料,探寻饲料蛋白质水平对大菱鲆幼鱼生长性能和水生态因子的影响,试验期88d。结果表明:不同蛋白质水平膨化饲料对大菱鲆幼鱼的生长性能和饲料利用率均产生了显著影响(P<0.05),净增质量、质量增长率、特定生长率随饲料蛋白水平的提高而增加,饲料系数随蛋白水平增加而降低;水生态因子中除pH外, 对其它指标都有显著影响(P<0.05),化学耗氧量(COD)、氨氮(NH3-N)、亚硝氮(NO2—N)、磷酸盐(PO4-P)的浓度都随饲料蛋白水平的提高而增加。其中,C组质量增长率比A,B组显著提高21.6%和14.6%(P<0.05),而比D,E组低1.6%和5.1%;C组饲料系数比A,B 组低0.22和0.16(P<0.05),而仅比D,E组高0.02和0.03;C组NH3—N比A,B组提高13.0%和4.0%,而比D,E组降低达23.1%和48.1%(P<0.05)。本试验从幼鱼生长性能和水生态因子两方面表明,大菱鲆幼鱼膨化饲料中蛋白质的生态营养需要量为50%。     

大菱鲆不同家系生长性能的比较

为培育优良的大菱鲆养殖新品种,收集了4个不同地理群体的大菱鲆作亲本,根据巢式设计,采用人工受精技术,在群体间和群体内构建了全同胞和半同胞家系,对其不同生长时期(生长阶段)全同胞家系的生长性能进行了比较.综合不同生长时期家系的体质量差异分析和不同生长阶段家系的生长速度分析,结果表明:来自英国和法国的群体间和群体内家系的生...     

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。     

一种新的牙鲆病原菌——兔莫拉氏菌

目前国内外已报道的牙鲆(Paralichthys olivaceus)细菌病的病原有鳗弧菌(Vibrio anguillarum)以及弧菌(Vibrio spp.)属的其他一些种类、爱德华氏菌(Edwardsiella tarda)、屈挠杆菌(Flexibacter maritimus)、肠球菌(Enterococcus seriolicida)、链球菌(Streptococcus spp.)、卡帕奇诺卡氏菌(Nocardia kampachi)、变异微球菌(Micrococcus varians)[1].     

用显微注射方法分析牙鲆肌肉发育调节基因MyoD, Myf5的启动子序列

克隆了牙鲆(Paralichthys olivaceus)肌肉发育调节基因MyoD和Myf5的启动子序列,分别为608 bp和1 073 bp。对序列进行分析发现MyoD和Myf5的启动子含有参与肌肉发育调控的基因的结合位点。将MyoD和Myf5的启动子连接到GFP载体上,构建了重组质粒MyoDP-GFP和Myf5P-GFP,并注射到一细胞或两细胞的斑马鱼胚胎中,对这两个基因的启动子进行了瞬时表达研究。结果表明,牙鲆MyoD和Myf5的启动子可以驱动绿色荧光蛋白特异地在斑马鱼肌肉纤维中表达,因此牙鲆608 bpMyoD和1073 bpMyf5的启动子包含着可以驱动MyoD和Myf5正常表达的必需元件,它们是肌肉特异的,并且可以跨物种行使其功能。     

几种鲆鲽鱼类精子超微结构及生理特性的比较研究

借助透射电镜分别对石鲽(Kareius bicoloratus)、大菱鲆(Scophthalmus maximus L.)和牙鲆(Paralich- thys olivaceus)成熟精子的超微结构进行了比较研究,进一步测定了石鲽和牙鲆精子的基本生理特性,并研究了17℃和4℃保存精子的活力变化。结果显示3种鱼类精子均为有鞭毛精子,无顶体,尾部轴丝9+2型。不同之处包括:成熟精子的核致密度不同,以石鲽最高,大菱鲆和牙鲆则较低;在大菱鲆和牙鲆精子头部,核前端侧面都有一缺刻状结构,石鲽精子未发现此结构;头部核膜与质膜之间的细胞质含量以大菱鲆的最多;精子中段的线粒体数量不同,大菱鲆较多(10个左右),石鲽次之(8个左右),牙鲆最少,仅为5个左右;中段植入窝深度不同,以大菱鲆陷入核中最深,其他2种比较接近。石鲽和牙鲆精液的pH值比较接近。精子密度以牙鲆较高,为10.18×10~9个/mL±3.37×10~9个/mL。牙鲆和石鲽精子活力的保存时间在17℃下分别为48 h和36 h,4℃下分别为7 d和8 d。精子活力完全失去之前,随着保存时间的延长,精子的激活率和寿命逐渐下降,而涡动时间则相差不大。     

微卫星标记在大菱鲆(Scophthalmus maximus L.)家系系谱印证中的应用研究

用8个微卫星标记对大菱鲆7个人工选育家系进行了系谱鉴别和遗传多样性研究。8个微卫星位点的平均多态信息含量为0.6721,平均杂合度为0.7206。8个微卫星位点在7个家系分析中,共检测到40个等位基因,每个位点的等位基因数在38个之间,每个位点平均有5个等位基因。根据已知亲本及子代基因型,成功的推断出了7个家系中缺失亲本的基因型。在双亲未知的情况下8个微卫星位点累积排除概率是97.07%,已知单亲时累积排除概率达99.63%。用UPG-MA法对7个家系的137个子代个体进行了聚类分析,90.5%同一家系的子代个体能完全聚类到一起,分类结果与系谱来源基本一致。微卫星标记可以为大菱鲆混养家系进行亲权鉴定和系谱构建提供技术支持。     

两株养殖大菱鲆体表出血病原菌的分离鉴定

从秦皇岛和胶南患出血病的大菱鲆(Scophthalmus maximus)肾脏中分别分离得到优势菌株MHK_2和JN,人工感染实验证实这两株菌对大菱鲆有较强的致病性,半数致死量(LD_(50))分别为6.30×10~3cfu/尾和2.51×10~4cfu/尾。对病原菌16s rDNA序列对比分析及进化树分析表明,MHK_2和JN与迟缓爱德华氏菌(Edwardsiella tarda)高度一致。API ID32E快速鉴定结果表明,MHK_2和JN生理生化特征与非致病性迟缓爱德华氏菌标准菌株LSE_6一致,属于迟缓爱德华氏菌。经API ZYM酶活检测,MHK_2和JN的碱性磷酸盐酶和酸性磷酸酶活性明显高于LSE_6,这两项酶活特征有望作为区别致病性和非致病性迟缓爱德华氏菌的一个标准。     

牙鲆性腺分化的组织学研究

运用组织学方法对牙鲆(Paralichthys olivaceus)的性腺分化和发育过程进行了观察.结果表明,牙鲆的性腺发育可分为3个阶段,即原始性腺阶段、性腺分化阶段和性腺分化后阶段.培育水温为18℃～20℃下,孵化后第45天、平均全长为22.0 mm±2.8 mm的牙鲆,其性腺分化尚未开始,属于原始性腺.70日龄、平均全长为38.0 mm±1.7 mm的牙鲆,部分个体中观察到卵巢的雏形,其余个体中性腺变化并不大;到了第110天、平均全长达到86.5 mm±5.9 mm时,雌性个体卵巢的卵巢腔进一步增大,并出现卵原细胞向卵母细胞的转变.精巢的分化开始于90日龄、平均全长为63.5 mm±3.4 mm的幼鱼,此期精原细胞快速增殖,体细胞进行有丝分裂并逐渐形成输精管原基;进一步的细胞学分化则出现在100日龄、平均全长为76.0 mm±8.6 mm的个体中,此时可以看到精小叶的形成;在平均全长为140.0 mm±15.2 mm时,精巢中出现初级精母细胞,标志着性腺分化的基本完成.     

台湾海峡牙鲆养殖技术研究

报道了台湾海峡牙鲆(Paralichthysolivaceus)的养殖技术。经过在室内水泥池189d的饲养,共获得平均全长32.20cm,平均体质量402.10g规格的牙鲆11.5万尾,养殖成活率达96%。养殖结果表明,台湾海峡牙鲆的生长速度快于黄、渤海牙鲆;水温与牙鲆的生长关系密切,台湾海峡牙鲆适宜的温度范围为18～27℃左右;饲料中添加适量中草药对提高牙鲆的成活率有较明显作用。     

牙鲆对海水中铅积累排放规律研究

研究了海水中铅在牙鲆内脏、肌肉、鳃组织中吸收、积累和排放规律及海水中总有机碳浓度对铅吸收的影响.结果表明,铅浓度为0.5mg/dm3时,各组织中铅蓄积量随暴露时间增加而增大,第9天均达到吸收平衡,此时铅蓄积量(mg/kg)内脏中的649.8大于鳃中的237.4大于肌肉中的59.71.染毒13d后将牙鲆移入清洁海水中排放的结果表明,随排放时间增加各组织中铅蓄积量明显下降,第10天时各组织中铅排出率为内脏团的84.85%大于鳃的61.26%大于肌肉的57.03%.海水中总有机碳浓度对铅蓄积量有明显影响,随着总有机碳浓度升高,各组织中铅蓄积量明显下降,表明海水总有机碳能降低铅的生物有效性.     

First isolation and characterization of a new species of Vibrio from diseased flounder (Paralichthys olivaceus Temminck et Schlegel)

Appropriate pathogenic bacteria were isolated from diseased (or dead) flounder (Paralichthys olivaceus L) occurring in a mariculture farms of Qinhuangdao, which all diseased fishes expressed bacterial septicaemia. The phenotypic information of the 12 pure cultures was studied extensively, including morphological characteristics, colony characteristics in different media, physiological and biochemical characteristics, and the mole fraction G C ratio of the DNA for representative strain. The results show that the isolates belong to a new species of Vibrio, and are designated as Vibrio qinhuangdaora sp.nov, by the area from which the diseased flounders are collected (Qinhuangdao) based on its biological properties following Rules of International Code of Nomenclature of Bacteria. At the same time,studies on the serum homology ofisolates, and pathogenicity of isolates were conducted by experimental infection. The results show that all the isolates are of serologic similarity, and the isolates have strong pathogenicity to flounder. The studies can extensively reflect the main biological properties of this new species of Vibrio.     

牙鲆(Paralichthys olivaceus)细菌败血感染症及病原检验与分析

采用调查统计、剖检病变、病原分离鉴定与人工感染健康鱼致病作用试验的方法，对自然发生的养殖牙鲆细菌败血感染症病例，进行了发病情况、临床表现及病理变化等方面的检验，同时取病(死)牙鲆的肝、腹水、腐烂肌肉组织为材料进行细菌分离，对分离于10尾鱼20株纯培养菌的形态特征、理化特性等进行鉴定。结果表明，分离出来的两种细菌，一种是弧菌属(Vibrio Pacini 1854)细菌的一个新种(sp．nov．)并定名为牙鲆弧菌(Vibrio olivaceus sp．nov．)；另一种是气单胞菌属(Aeromonas Kluyver and van Niel 1936)的嗜水气单胞菌(A．hydrophila)。通过人工感染试验等表明该两种分离菌为相应感染症的致病菌。     

大菱鲆体重和形态性状的遗传动态分析

In order to elucidate the genetic mechanism of growth traits in turbot during ontogeny, developmental genetic analysis of the body weights, total lengths, standard lengths and body heights of turbots was conducted by mixed genetic models with additive-dominance effects, based on complete diallel crosses with four different strains of Scophthalmus maximus from Denmark, Norway, Britain, and France. Unconditional genetic analysis revealed that the unconditional additive effects for the four traits were more significant than unconditional dominance effects, meanwhile, the alternative expressions were also observed between the additive and dominant effects for body weights, total lengths and standard lengths. Conditional analysis showed that the developmental periods with active gene expression for body weights, total lengths, standard lengths and body heights were 15–18, 15 and 21–24, 15 and 24, and 21 and 27 months of age, respectively. The proportions of unconditional/conditional variances indicated that the narrow-sense heritabilities of body weights, total lengths and standard lengths were all increased systematically. The accumulative effects of genes controlling the four quantitative traits were mainly additive effects, suggesting that the selection is more efficient for the genetic improvement of turbots. The conditional genetic procedure is a useful tool to understand the expression of genes controlling developmental quantitative traits at a specific developmental period(t-1→t) during ontogeny. It is also important to determine the appropriate developmental period(t-1→t) for trait measurement in developmental quantitative genetic analysis in fish.     

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.     

台湾海峡野生牙鲆人工育苗技术研究

报道了2004年3月对产自台湾海峡的野生牙鲆(Paralichthys olivaceus)进行人工育苗的研究结果。共获153万粒受精卵,孵出仔鱼140.8万尾,孵化率达92%,在育苗水温为20~24℃,海水盐度为26~31的条件下,经63 d培育,育出全长3.8~5.5 cm的幼鱼53.39万尾,成活率为37.9%。     

植酸对牙鲆生长和消化酶活性的影响

以鱼粉为主要蛋白源,在基础饲料中分别添加不同含量的植酸纳使植酸水平分别达到0.04%(对照组)、0.21%、0.44%和0.81%,研究不同含量植酸对牙鲆(Paralichthys olivaceus)(2.58 g±0.01 g)摄食、生长和消化酶活性的影响.实验在室内循环水养殖系统中进行,为期10周.结果表明,植酸处理组牙鲆摄食率(2.4%、2.5%和2.4%)显著高于对照组(2.1%)(P＜0.05),但植酸处理组之间差异不显著(P＞0.05).植酸处理组(59.47、61.27和64.82 U/mg蛋白)胃蛋白酶比活力显著低于对照组(79.34 U/mg蛋白),但植酸处理组之间差异不显著.各处理组之间牙鲆肝脏、肠和胃中淀粉酶比活力及肝脏和肠中蛋白酶比活力均无显著差异.随着饲料中植酸含量的增加,干物质和蛋白质表观消化率显著下降.同时,牙鲆特定生长率、饲料转化率、蛋白质效率和净蛋白质利用率随着饲料中植酸含量的升高均呈下降趋势,其中各处理组之间牙鲆特定生长率无显著差异,而植酸处理组饲料转化率(74.0%、73.7%和69.2%)则显著低于对照组(84.0%),0.81%植酸处理组蛋白质效率(1.37)和净蛋白质利用率(25.0%)显著低于对照组(1.64和30.6%).本实验结果表明,随着饲料中植酸含量的增加,牙鲆生长性能和胃蛋白酶比活力呈下降趋势.植酸对牙鲆生长的抑制作用将随着其含量的升高或实验周期的延长突现出来.     

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。     

利用父母本微卫星标记估计大菱鲆(Scophthalmus maximus L.)幼鱼生长性状的遗传参数

The estimation of genetic parameters has played an important role in animal selective breeding for growth traits.Recently studies show that molecular markers can be incorporated into genetic evaluations. In order to improve the performance of an incomplete pedigree(i.e, only parents are known) in the genetic evaluations, 12 microsatellite markers have been applied in the estimation of the genetic parameters for body weight in a farmed population(n=1 890) of juvenile turbot(Scophthalmus maximus L.). A new relatedness called parental molecular relatedness(PMR) is estimated based on results of genotyping of 48 parents(31 males, 17 females) with microsatellites markers. The feasibility of PMR in estimation of genetic parameters is verified by comparison with pedigree related(PR) which is obtained from a complete pedigree. The results demonstrate that a high correlation(0.872) between them is found. Heritabilities are estimated using the PMR(0.52±0.13) and PR(0.55±0.22) with the same animal model. A cross-validation shows that the predictive abilities of models using the PMR and the PR are identical(0.81). From that, a conclusion can be made that PMR and PR predicted genetic values equally well in a population of juvenile turbot. Therefore PMR can be applied as an alternative of the PR when only parents are known. However, for a better performance, more markers and more families should be included in a further study.     

养殖大菱鲆细菌性红体病病原菌的分离与鉴定

从患有红体病的养殖大菱鲆(Scophthalmus maximus)的肌肉中分离得到一株优势菌并记为H1.经人工注射感染证实H1即为引起养殖大菱鲆红体病的病原菌,其半致死量LD50为2.82×105CFU/mL,而低浓度(1.41×103 CFU/mL)没有造成死亡,但注射部位有脓肿现象,注射相同体积1.5%无菌生理盐水的对照组没有明显的症状,注射部位也无异常.革兰氏染色显示该菌为革兰氏阴性,菌体呈杆状,周生鞭毛.综合该菌的形态、常规生理生化特征和API32E鉴定结果,发现H1与迟钝爱德华氏菌的表性特征非常相似,相似率迭到99.9%.在分子水平上对其16SRNA基因序列的测定,同源性分析,结果表明H1与迟钝爱德华氏菌的亲缘关系最近,相似度达到99%.综合上述研究结果,将该菌株初步鉴定为迟钝爱德华氏茵(Edwardsiella tarda).