条斑星鲽卵巢发育规律和性类固醇激素周年变化研究

采用组织学方法、形态测量法、放射免疫法(RIA)等技术方法,研究了人工养殖条件下条斑星鲽雌性亲鱼卵巢发育成熟和性类固醇激素的年周期变化规律.结果表明,条斑星鲽属于非同步分批产卵类型.卵母细胞按其组织学特点可分为6个时相,卵巢发育依据其形态特征可划分为相应的6个时期.卵巢年周期发育过程中,性腺指数(GSI)、肝脏指数(HSI)和肥满度(CF)发生变化,表现出与性腺发育协同一致的变化规律.统计分析表明,卵巢年周期发育过程中,GSI和HSI值与CF值都存在显著的正相关关系,GSI值的变化与水温调控呈显著的负相关关系.亲鱼血浆中雌二醇(E2)在6月份开始升高,9月份达到年周期中的第1个小高峰,翌年2月份,E2表达水平达到最高峰值,4月产卵结束后,E2表达水平降低并维持较低水平至下一个生殖季节.血浆中睾酮(T)含量在7月达到第1个峰值,并在翌年1月份达到第2个峰值,在4月份产卵后降至最低水平并维持至下一个生殖季节.统计分析表明,E2与T存在正相关关系,E2与光周期调控存在正相关关系,同时与GSI和CF呈弱的正相关关系.     

圆斑星鲽卵巢发育的组织学和数量形态特征研究

研究了圆斑星鲽卵巢年周期发育的组织学规律及其数量形态特征变化.结果表明,圆斑星鲽产卵属1年1次成熟分批产卵类型,卵巢发育属非同步分批成熟类型.性成熟亲鱼在年周期发育中经历5个发育时期,同时可见5个发育时相的卵母细胞,不同发育时期的卵巢中卵母细胞类型和组成比例不同.性腺指数(GSD、肝脏指数(HSI)和肥满度(CF)在亲鱼的年周期发育中呈现规律性的变化,与卵巢发育呈现密切的关联.亲鱼卵巢发育的有效积温为1050日度.     

半滑舌鳎卵巢发育的组织学和形态数量特征研究

采用组织学方法系统研究了人工养殖条件下半滑舌鳎亲鱼卵巢的组织发育周年变化特征.结果表明,卵母细胞发育可分为6个时相,卵巢发育分为6期.卵巢不同发育时期都由不同类型的卵母细胞组成,半滑舌鳎为非同步分批多次产卵类型.周年发育过程中,6月龄前的鱼卵巢发育处于第Ⅰ期;9～12月龄卵巢可发育至第Ⅱ期;12月龄半滑舌鳎性腺可发育达到Ⅲ期,并一直保持至24月龄.3龄雌鱼达到性成熟,卵巢可发育至第Ⅳ期;随着水温和光周期的调控,卵巢发育进入成熟期(Ⅴ期),发生水合现象是卵母细胞成熟并准备排卵的信号;产卵结束后卵巢退化至第Ⅵ期,排出的卵母细胞退化吸收,以Ⅲ期卵巢越冬后,卵巢退化至Ⅱ期并持续至下一个繁殖周期.人工养殖条件下,卵巢内存在卵母细胞闭锁现象.性腺周年发育过程中,性腺指数(GSI)在性腺达到成熟期时达到峰值,产卵结束后性腺快速退化,GSI值各月份差异显著(P<0.05),亲鱼性腺重量在卵巢发育成熟过程中一直处于生长状态.亲鱼肝脏指数(HSI)在产卵前7～8月份(性腺达到成熟期前30 d左右)达到最大,表明7～8月份是性腺进行卵子发育、卵黄能量储备的重要时期,亲鱼在产卵前通过积极摄食储备体内能量,肝脏在卵母细胞卵黄积累方面具有重要的生理作用.肥满度(CF)值在繁殖盛期10～11月份达到最大,表现出与GSI类似的变化趋势.     

条斑星鲽精巢年周期发育规律和血液性类固醇激素含量变化

采用组织学方法和放射免疫法(RIA)等技术方法,研究了人工养殖条件下条斑星鲽雄性亲鱼精巢发育规律和性类固醇激素的年周期变化规律。实验结果表明,条斑星鲽精巢中可见5个时相的生殖细胞类型,精巢发育可分为5期。条斑星鲽雄鱼GSI值自10月开始升高,12月达峰值(P≤0.05),之后显著下降并保持较低值至下一次生殖周期开始。HSI值在11～12月保持较高表达值,其他各月份保持相对稳定水平。CF值在5～8月保持较高水平,其他月份保持平稳状态。雄鱼血浆中睾酮(T)水平自9月开始升高并在12月(Ⅴ期精巢)达到峰值,其后显著下降并在其后的月份保持较低水平,而雌二醇(E2)在2月出现峰值,其他月份保持相对稳定水平。     

养殖银鲳性腺发育规律和性类固醇激素变化

为掌握养殖银鲳的性腺发育规律及其与性类固醇激素水平的关系,本研究采用组织切片技术和酶联免疫吸附法(Elisa),观察养殖条件下银鲳的精巢与卵巢发育特征和血清中性类固醇激素的周年变化,并分析性腺成熟指数(GSI)、肝重指数(HSI)和肥满度(CF)与性腺发育的关系。结果显示,养殖银鲳雌鱼1龄即可发育成熟,成熟卵巢呈一对""形的囊状结构。11月份有近一半个体发育至III期,33.3%已发育至IV期,12月—翌年1月越冬期间约2/3退化至II期,3月又迅速发育至IV期,4月份达到V期并产卵,5月份产卵结束卵巢进入恢复期IV期。银鲳精巢属小叶型,III期精巢出现小叶腔,IV期精巢出现精子,精巢发育速率快于卵巢,即"雄性早熟",7月龄约20%个体可达到性成熟,越冬时退化至III期,3月重新发育,精子分批成熟排精。雌鱼GSI值为0.19%~12.89%,HSI为0.97%~2.30%,CF为2.30~3.08 g/cm~3,雄鱼的GSI为0.08%~2.62%,HSI值为0.73%~1.83%,CF为2.11~2.80 g/cm~3,GSI值在V期时达到最大,HSI和CF值则于IV期达到峰值。雌鱼血清中雌二醇(E2)周年表达水平为23.27~59.13 pg/m L,雄鱼为15.90~36.20 pg/m L,雌鱼的睾酮(T)为14.57~68.67 nmol/L,雄鱼的T为18.62~66.49 nmol/L,E2水平与HSI值呈现显著正相关关系,T表达水平与GSI值呈现显著正相关关系。研究表明,银鲳血清中性类固醇激素含量与精巢和卵巢的发育密切相关,可作为了解养殖银鲳性腺发育的重要指标。     

不同激素对雌性花鳗鲡性体指数和发育过程中促性腺激素基因(GtHα,FSHβ,LHβ)表达的影响

为评估不同激素诱导花鳗鲡卵巢发育的效果并探究促性腺激素在花鳗鲡发育过程中的作用,实验比较了不同激素组合处理后花鳗鲡性体指数(gonadosomatic index,GSI)的变化,并采用实时荧光定量PCR(qRT-PCR)对处于不同发育阶段雌鱼脑垂体中促性腺激素基因GtHα(促性腺激素亚基),LHβ(促黄体生成素亚基)及FSHβ(卵泡刺激素亚基)的表达进行了定量研究.结果发现,鲤脑垂体萃取液(CPE)与人类绒毛膜促性腺激素(hCG)组合能够有效促使雌性花鳗鲡性腺发育成熟,GSI平均达22.9％,与对照组相比差异显著.单独注射雄烯二酮(ADSD)和注射ADSD、CPE、hCG混合制剂诱导效果不明显,GSI仅为1％ ～3％.荧光定量发现,经CPE与hCG处理的亲鱼LHβ和GtHα的表达量不断上调,其中发育早期GtHα的表达量极显著升高,而FSHβ的表达量呈现下调趋势.研究表明,CPE与hCG组合注射能够成功诱导花鳗鲡卵巢发育成熟,而ADSD对花鳗鲡卵巢发育无明显促进作用.LH在花鳗鲡卵巢发育整个过程中扮演重要角色;FSH在花鳗鲡性腺发育早期作用显著,可能是诱导鳗鲡性腺早期发育的关键因素.     

圆斑星鲽sox9基因的克隆与表达

本研究筛选到圆斑星鲽(Verasper variegatus)的性别相关基因sox9，并通过RACE技术获得了全长序列，基因全长为3287 bp，包括1431 bp的ORF，编码477个氨基酸，368 bp的5¢ UTR和1488 bp的3¢ UTR。在3¢ UTR中有多聚腺苷酸尾和加尾信号AATAAA。通过荧光定量PCR测定了sox9基因在圆斑星鲽成鱼不同组织中的表达水平，发现sox9基因在圆斑星鲽的脑、眼、鳃、心、肝、胆、肠、精巢、卵巢、肾和肌肉等各个组织中都有不同程度的表达。在鳃、脑和精巢组织中检测到较高水平的sox9转录，其中精巢中的转录水平显著高于其他组织，sox9基因在性腺中的表达显示出性别两相性差异。其在精巢中的表达水平要显著高于卵巢，说明sox9基因与雄性性腺发育相关。通过测定sox9基因在圆斑星鲽幼鱼不同发育时期(20、30、40、50、60、70和80日龄)的表达水平，发现其在20~50日龄表达量逐渐下降，在60日龄时表达量上升，推测表达量上升可能与幼鱼性腺分化相关。     

提早“家鱼”产卵的主要技术措施

实践证明:从提早亲鱼的培育时间着手,如能使亲鱼性腺在四月上、中旬发育成熟,四月中、下旬催产获得鱼苗,为当年培育大规格鱼种,提高越冬成活率和成鱼高产提供了可靠的保证。利用工厂温热水和地下温泉水培育亲鱼,促使家鱼提早产卵获得成功,但推广受到限制。目前鱼苗生产上可以采用控制水温、亲鱼培育及注射外源激素等技术措施。一、控制水温。温度对性腺的发育、成熟具有显著的影响,它通过改变鱼体代谢强度,加速或抑制性腺发育成熟的过程。为了提高亲鱼培育池的水温,冬季应     

雄性黄颡鱼芳香化酶基因mRNA在熟期组织中表达分析

在水温分别为20(常温对照组)、25、30℃条件下,采用RT-PCR方法,研究性腺成熟期雄性黄颡鱼组织中P450芳香化酶(P450aromA和P450aromB)的mRNA表达水平,同时测定性腺成熟系数(GSI)、肝重指数(HSI)、肥满度(CF)和血浆睾酮(T),雌二醇(E2)含量.结果表明,P450aromA在性腺...     

池塘养殖黄斑篮子鱼初次性成熟性腺发育研究

研究池养黄斑篮子鱼(Siganus oramin)初次性成熟的性腺发育及季节性变化,可为其全人工繁殖提供基础数据。2019年1—7月,每月取1~+龄黄斑篮子鱼进行解剖取性腺切片,计算性腺成熟系数(GSI)和肝体指数(HSI)。结果显示,实验期间性腺发育可分为Ⅱ、Ⅲ、Ⅳ、Ⅴ、Ⅵ5个时期,未发现Ⅰ期性腺。1月全部为Ⅱ期;2月卵巢仍为Ⅱ期,部分精巢达Ⅲ期;3月卵巢约一半发育至Ⅲ期,个别Ⅳ期,精巢主要为Ⅳ期,部分Ⅴ期,能挤出精液;4月GSI最大,精巢基本都为Ⅴ期,卵巢少部分发育成熟,绝大部分停留在Ⅲ、Ⅳ期,少量为Ⅱ期;5月繁殖个体性腺短暂进入Ⅵ期后迅速退化,未成熟个体性腺停止发育并退化;6月性腺退化至Ⅱ期;7月保持Ⅱ期,GSI进一步降低。池养黄斑篮子鱼初次性成熟雌性GSI为1.74‰～133.44‰,雄性GSI为1.06‰～138.51‰,均在3—5月出现一次高峰,HSI与GSI的变化趋势一致,但提前1个月左右达到峰值。黄斑篮子鱼卵巢发育是完全同步型,为一次产卵鱼类,繁殖期为3月底至5月初。     

Anesthesia induces stress in Atlantic salmon (<Emphasis Type="Italic">Salmo salar</Emphasis>), Atlantic cod (<Emphasis Type="Italic">Gadus morhua</Emphasis>) and Atlantic halibut (<Emphasis Type="Italic">Hippoglossus hippoglossus</Emphasis>)

Stress in response to anesthesia with benzocaine, MS-222, metomidate and isoeugenol was studied in Atlantic salmon (Salmo salar), Atlantic halibut (Hippoglossus hippoglossus), and Atlantic cod (Gadus morhua) with no concomitant stress from handling or confinement in association with anesthesia or sampling. All of the anesthetics tested induced a stress response in all species, displayed by a release of cortisol to the water. MS-222 anesthesia elicited the highest cortisol release rates, reaching maximum levels 0.5 h post-exposure and returning to basal levels after 3–4 h. Benzocaine anesthesia caused a bimodal response where the initial peak in cortisol release rate was followed by a second increase lasting towards the end of the trial (6 h). This bimodality was more profound in Atlantic salmon than in Atlantic halibut and Atlantic cod. Metomidate anesthesia induced the lowest release of cortisol of the agents tested in both Atlantic halibut and Atlantic cod, but resulted in a bimodal response in Atlantic salmon where the initial increase in cortisol release was followed by a larger increase peaking at 2–2.5 h post exposure before returning to basal levels after 5 h. The stress induced in Atlantic salmon by isoeugenol anesthesia resembled that of MS-222, but did not reach the same elevated level. Overall, the cortisol release was most profound in Atlantic salmon followed by Atlantic halibut and Atlantic cod.     

Assessment of sex and evaluation of ovarian maturation of fish using ultrasonography

Ultrasonography is a non‐invasive method that can be used to assess sex and maturity of fish. This study was conducted to examine the feasibility of using ultrasound images to sex juvenile and mature halibut Hippoglossus hippoglossus (Linnaeus), mature winter Pleuronectes americanus (Walbaum) and yellowtail flounder Pleuronectes ferruginea (Storer) and mature haddock Melanogrammus aeglefinus (Linnaeus). In females, both immature and mature ovaries were relatively easy to distinguish, but in males only mature testes were consistently distinguishable. Ovarian maturation of haddock and ovulatory cycles of halibut may also be assessed using ultrasound.     

Fillet shelf-life of Atlantic halibut Hippoglossus hippoglossus L. fed elevated levels of α-tocopheryl acetate

Fish fillet quality may be influenced by the antioxidant level in preslaughter diet. Thus, the effects of dietary α‐tocopheryl acetate supplementation and feeding time on the flesh quality of farmed Atlantic halibut Hippoglossus hippoglossus L. were investigated. Halibut of mean initial weight of 312 ± 12.3 g were divided into two groups and fed commercial diets, supplemented with different levels of α‐tocopheryl acetate at the dietary inclusion levels of 189 and 613 mg kg^?1 diet. Fish were sampled after 6,9,12 and 24 weeks. Over the experimental period, they reached a final mean weight of 1320 ± 108.4 g. Tissue α‐tocopherol of fillet and liver was significantly affected by the levels of α‐tocopheryl acetate given with the diets (P < 0.001). In storage on ice, fillets of fish fed the diets high in α‐tocopheryl acetate exhibited significantly lower (P < 0.001) levels of lipid oxidation. The colour of fillets in all groups deteriorated slightly, but diet did not affect this process. Halibut fed the supplemented diets for longer periods were better protected against lipid oxidation (P < 0.001) and colour deterioration (P < 0.01) than those fed for shorter periods. However, after 9 days of storage, lipid oxidation levels were still extremely low [< 0.6 µg malondialdehyde (MDA) g^?1 fillet], even in fillets of fish fed the low α‐tocopheryl acetate diet for a short period preslaughter. Different slaughtering methods tested at the end of the trial showed that percussive stunning can delay the onset of rigor mortis by 8–12 h compared with bleeding of the fish. These results suggest that halibut fillets have enhanced shelf‐life stability even at low doses of dietary α‐tocopheryl acetate, and that other factors in the antioxidant defence mechanisms of the species might play a major role in the prevention of lipid oxidation.     

Artificial gynogenesis and assessment of homozygosity in meiotic gynogens of spotted halibut (<Emphasis Type="Italic">Verasper variegatus</Emphasis>)

In the present study, methodology of gynogenetic induction in spotted halibut were developed and optimized; the sex ratio of putative meiotic gynogenetic diploids was determined using AFLP-based molecular sexing technique; the homozygosity of gynogenetic population was assessed as opposed to cultivated population. The results showed that high percentage of meiotic gynogenetic diploids were generated when the eggs fertilized with irradiated heterologous sea perch frozen sperm (30–50 mJ cm⁻²) were cold shocked in sea water of −1°C for 40–75 min at 5 min after fertilization. About 15,200 diploid gynogenetic larvae were achieved and they exhibited normal morphology similar to diploid control. The gynogenetic diploids were 100% female, which first confirmed the female homogamete (XX/XY) sex determination in spotted halibut. The genetic analysis showed that the average H _O was, respectively, 0.404 and 0.724 in gynogenetic population and cultivated population, indicating an increase of homozygosity in gynogenetic population.     

Change in nutrient composition of Artemia grown for 3–4 days and effects of feeding on‐grown Artemia on performance of Atlantic halibut (Hippoglossus hippoglossus,L.) larvae

Major challenges in culture of Atlantic halibut larvae have been slow growth during the late larval stages and inferior juvenile quality due to pigmentation errors and incomplete eye migration during metamorphosis. The hypothesis of this study was that feeding on‐grown Artemia would alleviate these problems. Artemia were grown for 3–4 days on Origreen or Origo. The growth and nutrient composition of Artemia nauplii and on‐grown Artemia were analysed, and both Artemia types were fed to Atlantic halibut larvae, on‐grown Artemia from 15 days post‐first feeding (dpff). The body length of Artemia increased with 20%–70% in response to on‐growing. In all experiments, protein, free amino acids and the ratio of phospholipid to total lipid increased, while lipid and glycogen decreased. The fatty acid composition improved in some cases and not in others. The micronutrient profiles were not negatively affected in on‐grown Artemia. All these changes are thought to be beneficial for marine fish larvae. The final weight of Atlantic halibut postlarvae was similar, and 90% of the juveniles had complete eye migration in both groups. It is concluded that the present version of Artemia nauplii probably covers the nutrient requirements of Atlantic halibut larvae.     

Distribution of α-tocopherol in fillets of turbot (Scophthalmus maximus) and Atlantic halibut (Hippoglossus hippoglossus), following dietary α-tocopheryl acetate supplementation

The present study investigated the distribution of α‐tocopherol (vitamin E) in fillets of turbot (Scophthalmus maximus) and Atlantic halibut (Hippoglossus hippoglossus). Turbot and Atlantic halibut were fed commercial diets, supplemented with different levels of α‐tocopheryl acetate at the dietary target levels of 100, 500 and 1000 mg α‐tocopheryl acetate kg^?1 diet. The actual levels were 72, 547 and 969 for turbot, while halibut received 189, 613 and 875 mg α‐tocopheryl acetate kg^?1 diet. Turbot were fed the diets for 24 weeks, while Atlantic halibut were fed for 20 weeks prior to slaughter. At the end of the feeding periods fish had reached a final weight of around 1 kg. Fish were slaughtered and filleted. From the four fillets obtained per fish, 22 samples were taken from designated areas and analysed for their α‐tocopherol content. The average concentrations of α‐tocopherol incorporated in turbot and Atlantic halibut increased with increasing levels of α‐tocopheryl acetate in the diet. Atlantic halibut had significantly (P < 0.05) more α‐tocopherol in positions 2/II and 1/I than in position 9/IX. Turbot had significantly (P < 0.05) more α‐tocopherol in position 2/II than in positions 1/I, 4/IV and 11/XI. By mapping α‐tocopherol concentrations in fish fillets, a high degree of quality prediction may be established. Moreover, this study may help scientists in their choice of sampling position, when investigating if α‐tocopheryl acetate supplementation resulted in successful α‐tocopherol incorporation.     

Organ responses to 14C-glucose injection in Atlantic halibut, Hippoglossus hippoglossus (L.), acclimated to diet of varying carbohydrate content

After adapting Atlantic halibut, Hippoglossus hippoglossus (L.), to three different levels of available dietary carbohydrate (22, 47 and 82 g kg^-1), the uptake of intraperitoneally injected glucose, both labelled and unlabelled, by the liver, white muscle, plasma, gills, heart, kidney and gastrointestinal wall, as well as concentrations of glucose in plasma and glycogen in organs, were studied. Even 16 h after glucose injection, halibut showed hyperglycaemia. The response, however, was moderate when adapted to the two higher than to the lowest starch diet, indicating some adaptative capacity. A negative correlation was found between plasma glucose and liver glycogen concentrations, hepatosomatic index and liver weight. The amount of glucose restored, relative to the amount injected, demonstrated that adaptation to dietary starch had significant effects on liver glycogen levels, with the highest incorporation of labelled glucose being observed in halibut adapted to the lowest dietary starch level. The amount of labelled glucose restored was highest in liver, followed by muscle > gastrointestinal wall > plasma > gills > kidney and heart, when evaluating the amount in the total organ. An indication of which organ was the most metabolically active in glucose utilization was found by expressing the results on a concentration basis, and it decreased in order, heart > gills > intestinal wall > kidney and muscle.     

Changes in Plasma Concentrations of Sex Steroids in Adult Pacific Halibut, Hippoglossus stenolepis

Blood samples were collected from captive Pacific halibut, Hippoglossus stenolepis , at intervals of about six weeks from early December 1986 to late November 1987. Concentrations of plasma androgen and estradiol-17β were determined by radioimmunoassay. The plasma concentrations of steroid were highest during autumn and winter in halibut that matured during late winter. The concentrations of steroids in samples collected in December were above 2 ng/mL (estradiol) or 1 ng/mL (androgen) in maturing females and below 0.5 ng/mL for both steroids in non-maturing females. The levels of steroids decreased rapidly about one month before spawning. In a mature male, androgen began to rise in August and November, and reached a peak of 7 ng/mL in early December. One month before spawning, the androgen concentration fell to 0.16 ng/mL. Estradiol concentrations were detectable in the male and varied little during the year. In immature fish, neither androgen nor estradiol changed significantly throughout the year. These results suggest that the concentrations of estradiol or androgen measured in blood samples taken during December may be used to determine the sex and state of maturation of Pacific halibut.     

Habitat utilization,feeding, and growth of wild spotted halibut <Emphasis Type="Italic">Verasper variegatus</Emphasis> in a shallow brackish lagoon: Matsukawa-ura,northeastern Japan

Habitat utilization, feeding, and growth of a rare pleuronectid flatfish, spotted halibut Verasper variegatus, were examined in a brackish lagoon in northeastern Japan: Matsukawa-ura. The distribution and date–length data of spotted halibut collected mainly from beam-trawl samplings during 1983–2008 indicated that age-0 juveniles [n = 25, 6.0–18.0 cm total length (TL)] and older spotted halibut (n = 71, 13.8–43.0 cm TL) inhabited almost the entire northern part of Matsukawa-ura. Comparative distribution surveys of spotted halibut, stone flounder, and marbled flounder during 1985–1989 revealed similar distribution patterns of these flatfishes; the highest densities were found around the central part of Matsukawa-ura, although significantly lower abundance was detected for spotted halibut. Linear growth equations of age-0 juveniles demonstrated that spotted halibut were able to achieve high growth after June, probably because abundant prey (e.g., mysids and gammarids) and suitable physical conditions (warmer water temperature and lower salinity) were present. The main prey items shifted from various crustaceans including mysids, caridean shrimps, and anomurans to brachyura Hemigrapsus spp. (≥20 cm TL). Our study shows that spotted halibut use the shallow brackish lagoon as an important nursery for juveniles, and also as a feeding ground for young and adults.