虹鳟生长性状的随机回归分析

生长和抗逆是水产动物遗传育种工作中最重要的农艺性状,虹鳟的生长性状关乎虹鳟规模化养殖的生产经济效益,为了从遗传上精细解析虹鳟的生长性状,我们从渤海、丹麦、挪威、唐纳森和加利福尼亚5个虹鳟(Oncorhynchus mykiss)种系间的双列杂交开始,进行了连续4代的继代选育。本研究测量了第4代总共4368个实验个体在516日龄、608日龄、668日龄、883日龄和1036日龄5个时间点的生长数据。采用随机回归测定日模型,对虹鳟生长性状进行了动态遗传分析。根据贝叶斯信息准则,确定3阶勒让德多项式为拟合体重和体长的加性遗传效应和永久环境效应变化的最优子模型。利用双变量随机回归模型同时分析体重和体长两个生长性状。它们的遗传力在400～1000日龄之间呈现递减趋势,分别从0.288下降到0.164和从0.469下降到0.186,并且在该生长区间内体长的遗传力始终高于体重的遗传力。无论体重还是体长性状,在不同日龄之间的遗传相关都随着生长间隔的增大而降低,但是两个性状在生长初期和后期之间的遗传相关较高(遗传相关系数0.75以上),尤其是体重(遗传相关系数0.85以上),该研究结果为虹鳟早期的遗传选育提供了理论支撑。两个性状之间在相同日龄之间的遗传相关均在0.75以上,在不同日龄之间的遗传相关随着生长间隔的增大由0.83下降到0.63。以上的研究结论为虹鳟生长性状(主要是体长和体重)的遗传选育提供了理论基础,同时也为虹鳟的体长和体重两个性状的联合选育提供了精确的遗传分析结果,由于两性状在前期有较高的遗传相关,因此建议在虹鳟生长前期(400日龄)进行联合选择。     

Isolation and partial characterization of rainbow trout (Oncorhynchus mykiss) gill mucin

Gill mucin from rainbow trout was isolated utilizing two rounds of cesium chloride density ultracentrifugation followed by gel filtration on Sepharose CL-2B. Neither density ultracentrifugation nor gel filtration alone was sufficient for purification of the mucin. Isolated gill mucin had a density of 1.5 g/ml and eluted at the void volume of the Sepharose CL-2B column. Silver-stained reducing 6% polyacrylamide gel electrophoresis of gill mucin produced a band at the origin with a smear entering the separating gel. There was no evidence of a link protein in gill mucin on reducing 12% polyacrylamide gel electrophoresis. Gill mucin had an amino acid profile similar to that of mucins in other species. Specifically, 35.1% of the total amino acids were represented by threonine and serine, while another 27.5% were alanine and proline. Gill mucin contained galactose (26.7 ± 3.2%), galactosamine (22.5 ± 4.4%), glucose (16.6 ± 8.7%), fucose (16.1 ± 1.5%), glucosamine (12.0 ± 1.9%) and mannose (5.1 ± 4.4%). Uronic acid levels from purified mucin were very low (0.7 ± 0.1%). Sialic acid was also present (0.06 g/g of mucin protein). The periodic acid-Schiff assay routinely utilized for detection of mucins was relatively insensitive for detection of gill mucin (6 × less sensitive than for pig gastric mucin) so a rabbit antiserum was raised. The antiserum produced profiles similar to the periodic acid-Schiff assay of fractions following gel filtration. Immunofluorescence of formalin-fixed rainbow trout gill tissue sections showed that the antiserum detected mucin within branchial goblet cells.     

Glucose-stimulated lipolysis in rainbow trout,Oncorhynchus mykiss,liver

Rainbow trout, Oncorhynchus mykiss, were used to characterize further the influence of glucose on hepatic lipolysis. Liver was removed from fed fish, cut into 1 mm³ pieces and incubated for up to 5 h in Hanks medium containing either 2 mM, 5.5 mM, 10 mM, or 25 mM glucose. Glucose-stimulated lipolysis was indicated by tissue triacylglycerol (TG) lipase activity and by medium concentrations of glycerol and fatty acids (FA). Triacylglycerol lipase activity in liver pieces incubated in the presence of higher concentrations (25 mM) of glucose was significantly higher than that in liver pieces incubated in lower concentrations (2 mM) of glucose, rising from 0.075 ± 0.002 (mean ± SEM) nmol FA released/h/mg protein to 0.092 ± 0.004 units. Similarly, higher concentrations of glucose stimulated significantly more FA release and glycerol release from liver pieces than that stimulated by lower concentrations of glucose. Glycerol release from liver pieces incubated in the presence of 10 mM glucose and 25 mM glucose was ca. 2-fold to 2.8-fold, respectively, higher than that from liver pieces incubated in the presence of either 2 mM or 5.5 mM glucose. Fatty acid release from liver pieces incubated in the presence of 10 mM or 25 mM glucose was ca. 1.8-fold higher than that from liver pieces incubated in the presence of either 2 mM or 5.5 mM glucose. Notably, increased glycerol release was not accompanied by a parallel increase in FA. Fatty acid reesterification was more pronounced in liver pieces exposed to higher glucose (10 mM and 25 mM) than in liver pieces exposed to lower glucose (2 mM and 5.5 mM). ¹⁴C-incorporation studies indicated that glucose serves as a carbon source for reesterified FA in trout liver. The route of reesterification appears to be from glucose to glycerophosphate to phosphatidic acid to diacylglycerol to TG. Increasing concentrations of glucose did not affect glycerol kinase activity, indicating that glucose-stimulated lipolysis was not accompanied by increased glycerol recycling within the liver. These results suggest that glucose stimulates fatty acid reesterification and directly enhances net lipolysis in trout liver incubated in vitro.A part of this study was presented at the Annual Meeting of the American Society of Zoologists, December 26–30, 1991, Atlanta, GA.     

Lipid-stimulated somatostatin secretion in rainbow trout,Oncorhynchus mykiss

Previous work has shown that somatostatins (SS) affect teleost lipid metabolism indirectly by inhibition of insulin (INS) and directly by stimulation of hepatic lipolysis. In the present study, rainbow trout (Oncorhynchus mykiss) were used to characterize further the lipid-SS relationship by evaluating how lipid, contributes to SS secretion bothin vivo andin vitro. In vivo hyperlipidemia was induced for up to 3 h by short-term (2 min) infusion of a triacylglycerol (TG)-rich lipid emulsion (20% Intralipid^®). Plasma total lipid concentration increased 118 and 155% over control levels 1 h and 3 h, respectively, after infusion; much of this increase was due to elevated plasma fatty acids (FA), which increased 39 and 520%, respectively, over the same time-frame. The hyperlipidemic pattern was attended by a significant increase in the plasma concentration of SS. The specific effects of fatty acids were evaluated on isolated Brockmann bodies. Palmitic acid and oleic acid stimulated SS release 378 and 82%, respectively, over baseline levels. These results indicate that lipids, and in particular fatty acids, modulate SS secretion in rainbow trout.     

国内虹鳟代表性养殖群体的高通量SNP芯片检测及遗传分析

本研究旨在对国内虹鳟(Oncorhynchus mykiss)代表性养殖群体开展全基因组水平的遗传评估。利用57K单核苷酸多态性(single nucleotide polymorphism,SNP)芯片,检测了来自不同地域的6个虹鳟养殖群体样本共计48尾,包括黑龙江虹鳟、黑龙江金鳟、四川虹鳟、四川金鳟、北京虹鳟和北京金鳟,共获得有效SNP位点50201个,在中国虹鳟中的多态比例达到97.7%,表明该芯片虽然基于美国和挪威虹鳟群体设计,但对中国群体同样具有良好的适用性。各群体最小等位基因频率均值为0.240~0.267,与国外主流养殖群体相近,黑龙江虹鳟、四川虹鳟和北京虹鳟群体内遗传多样性丰富,多态位点比例为83.6%~84.9%,与国外主流养殖群体相近,而黑龙江金鳟、四川金鳟和北京金鳟,多态位点比例相对较低,在60.2%~76.9%范围内。应用6个中国虹鳟群体和2个美国虹鳟群体数据开展系统发育分析、主成分分析和群体遗传结构STRUCTURE分析,结果显示8个群体可分为3个祖源类群,其中3个金鳟群体为遗传联系较紧密的一个类群,黑龙江虹鳟和北京虹鳟为一个类群,而四川虹鳟与2个美国虹鳟群体为一个类群,部分中国养殖群体中有显著离群个体存在,表明群体遗传背景不均一。本研究表明,高密度SNP芯片在我国虹鳟养殖群体遗传分析中具有广泛的应用前景,能够为种质资源评估、本土化良种培育、制种和引种工作提供基因组水平的参考信息。     

虹鳟源嗜冷黄杆菌的分离鉴定及致病性研究

为确定患病虹鳟的病原,本实验从患病鱼溃烂肌肉中分离到2株细菌,分别命名为CH06和CH07,经回归感染证实分离菌株为导致此次虹鳟患病的病原菌,并进一步对其形态特征、理化特性、分子特征、血清型及耐药性进行分析。结果显示,CH06和CH07株在TYES琼脂平板上呈煎蛋状外观,产黄色素,氧化酶和过氧化氢酶呈阳性,能水解明胶和酪蛋白,不能水解淀粉,不能利用果糖、半乳糖和七叶苷等。16S rRNA比对结果显示,CH06和CH07株与嗜冷黄杆菌模式株NBRC 15942的同源性分别为99.35%和99.42%。综合菌株理化和分子特性确定CH06和CH07株为嗜冷黄杆菌。利用多重PCR方法鉴定CH06和CH07株的血清型均为1型(Fd型);多位点序列分型(MLST)分析表明,CH06和CH07株的基因型分别为ST-12和ST-78型,且均属于CC-ST10克隆型。人工感染结果显示,CH06和CH07株对虹鳟幼鱼具有较高致病性,其半致死浓度(LD₅₀)分别为7.1×10⁵和1.1×10⁵ CFU/mL,攻毒剂量与临床病症出现时间呈反比,从人工感染实验鱼的肌肉、脾脏等组织中可重新分离到嗜冷黄杆菌。组织病理变化显示,病鱼肝细胞肿胀,空泡变性,部分肝细胞溶解坏死,细胞核溶解消失;脾脏充血、出血,淋巴细胞减少,红细胞和含铁血黄素增多;肌纤维间隙增宽、断裂、弯曲不齐,部分肌细胞肌浆溶解呈蜂窝状。CH06和CH07株对10种抗菌药物的耐药谱略有不同,均对氨苄西林和甲氧苄啶-磺胺甲噁唑敏感;CH06株对恩诺沙星、氟苯尼考等耐药,而CH07株对恩诺沙星和氟苯尼考中度敏感。本研究首次报道了我国虹鳟源嗜冷黄杆菌的分离鉴定及生物学特性,以期为虹鳟细菌性冷水病的防控提供科学依据。     

虾青素和角黄素对虹鳟肌肉着色和肝脏总抗氧化能力的影响

分别在基础饲料(对照组)中添加100 mg/kg的虾青素、角黄素,混合色素(50 mg/kg虾青素+50 mg/kg角黄素)饲喂初始体重为(56.60±0.63) g的虹鳟60 d,考察虾青素和角黄素对虹鳟肌肉着色和肝脏总抗氧化能力的影响。结果显示,饲料中添加了虾青素、角黄素和混合色素后对虹鳟增重率、饲料系数及肌肉常规成分、肌肉失水率、含肉率均无显著影响(P> 0.05)。虾青素组、角黄素组和混合色素组虹鳟肌肉的比色卡得分、红度、虾青素含量和血清总类胡萝卜素含量均比对照组有显著提高(P< 0.05);虾青素组虹鳟肌肉比色卡得分(26.25)和红度值(18.40)显著高于角黄素组(22.38, 14.13)和混合色素组(24.00, 15.70)(P< 0.05);虾青素组虹鳟肌肉虾青素含量为4.75 mg/kg (30 d)和6.45 mg/kg (60 d),均显著高于混合色素组的3.87 mg/kg (30 d)和5.48 mg/kg (60 d)(P< 0.05);在虹鳟血清总类胡萝卜素含量方面,虾青素组 > 混合色素组 > 角黄素组;虾青素组、角黄素组、混合色素组虹鳟肝脏的总抗氧化能力之间无显著差异(P> 0.05),分别为2.39 U/mg,2.25 U/mg,2.39 U/mg,均较对照组(2.03 U/mg)显著提高(P< 0.05)。上述结果表明：饲料中添加100 mg/kg虾青素、角黄素及虾青素+角黄素混合(1∶1)均能有效改善虹鳟肌肉颜色,提高肝脏总抗氧化能力,虾青素、虾青素+角黄素混合(1∶1)对虹鳟肌肉的着色效果优于角黄素。     

Effects of constant light on haematological parameters of cultured rainbow trout (Oncorhynchus mykiss) in the Southern Hemisphere

The effects of artificial photoperiod regimes on reproductive patterns have been studied in several species, as have haematological parameters. However, information on how artificial photoperiods may affect blood components is scarce, especially under field conditions. We have assessed the effects of constant light [long day (LD) photoperiod: 24 h (light):0 h (dark)] on haematological parameters of cultured rainbow trout in Chile (Southern Hemisphere). In the initial stage (March up to June), two groups of fish were maintained under similar conditions and under the ambient (natural) photoperiod (NP). One group was then placed under the LD photoperiod regime for 2 months (June/July), following which it was returned to␣the NP (August–January); the control group remained under the NP throughout the experiment (March–January). All fish were assessed for haematological parameters and growth performance at various times during the experiment. During the initial stage, no differences were found between groups. However, at the end of the LD 24:0 period, the LD fish had increased haematocrit values and erythrocyte numbers and diminished mean corpuscular haemoglobin. After the LD fish had been returned to the NP, they developed secondary sexual characteristics and had a 40–44% decrease in the number of leukocytes.     

Triiodothyronine is necessary for the action of growth hormone in acclimation to seawater of brown (Salmo trutta) and rainbow trout (Oncorhynchus mykiss)

Brown (BT) and rainbow trout (RT) in freshwater (FW) were treated with ovine growth hormone (GH), GH + iopanoic acid (IOP), and GH + IOP plus triiodothyronine (T₃) for RT only. After 1 week of treatment, trout were transferred to 30 o/oo SW and treatment continued. In FW, GH treatment increased significantly plasma T₃ level (BT) and T₃/T₄ ratio (BT and RT) by stimulating T₄ to T₃ deiodination. In the GH + IOP group, the plasma T₃ levels and T₃/T₄ ratio fell significantly as T₄ to T₃ deiodination was inhibited. In GH + IOP + T₃-treated RT, plasma T₃ and T₃/T₄ ratios increased significantly relative to other groups. No mortality occurred and plasma osmolarity (PO) was not altered by any treatment in FW. After transfer to SW, all IOP + GH trout died within 2 (BT) or 3 days (RT). All GH-treated or control BT survived to the end of the experiment (6 days). RT survival rates tended to be improved in GH and GH + IOP + T₃ groups relative to controls. Correlatively on day 1 the PO increase was significantly higher in IOP + GH groups (BT and RT) than in the other groups and significantly lower in GH and GH + IOP + T₃ treated RT than in controls from days 1 to 6. These data confirm the requirement of T₃ and deiodination of T₄ to T₃ for the development of hypoosmoregulatory mechanisms in SW as previously shown (Lebel and Leloup 1992). Furthermore, the suppression of the hypoosmoregulatory effect of GH, when conversion of T₄ to T₃ was inhibited by IOP and the reversal when T₃ was added to IOP + GH treatment suggests that GH osmoregulatory action in SW acts via the simulation of T₄-5′ monodeiodination which increases T₃ production.     

Influence of ionic composition on acid-base regulation in rainbow trout (Oncorhynchus mykiss) exposed to environmental hypercapnia

The influence of ambient calcium, bicarbonate and chloride levels on acid-base regulation was investigated in rainbow trout acclimated and exposed to hypercapnia in five different water types. In soft water (low [Ca⁺⁺] and [HCO₃ ⁻]), compensation of the respiratory acidosis was slow and incomplete within 72h. High ambient [HCO₃ ⁻] clearly improved extracellular HCO₃ ⁻ accumulation, and pH recovery was accomplished within 24h. This may result from stimulation of branchial HCO₃ ⁻ (influx)/Cl⁻ (outflux) exchange. Elevation of ambient [Cl⁻] had a small, positive effect on pH compensation. High ambient [Ca⁺⁺] improved the degree of pH compensation. Plasma [HCO₃ ⁻] and [Cl⁻] showed an inverse 1:1 relationship in all acclimation groups, revealing an ubiquitous chloride-mediated acid-base regulation. Ventilation activity was increased by hypercapnia and only returned to control values in hard water (high [HCO₃ ⁻]and [Ca⁺⁺]). During progressive hypercapnia (up to 3% CO₂), hard water acclimated fish obtained significantly higher plasma [HC0₃ ⁻] (51.2 mM) than fish acclimated to low [Ca⁺⁺]/high [HCO₃ ⁻] (44.7 mM). This suggests an additive effect of ambient Ca⁺⁺ on plasma HCO₃ ⁻ accumulation. At levels of CO₂ above 1%, some mortality was induced in low [Ca⁺⁺]/high [HCO₃ ⁻] water. Dying fish could be distinguished from surviving fish by an excessive Cl⁻loss and increasing extracellular anion gap.     

Biochemical characterization of growth hormone receptor in rainbow trout (Oncorhynchus mykiss) before and after purification

The aim of this work was to verify if, compared to mammals, the lower molecular weight of GH-R previously reported in salmonid is real or due to the experimental process. For this purpose, we compared the apparent molecular weight of GH-R, obtained by SDS-PAGE after cross-linking with ⁵⁰ I-rtGH, obtained from rainbow trout crude liver membrane preparation, incubated in different buffers with those obtained after purification with affinity chromatography.Using crude liver membrane preparation, two specific bands of ⁵⁰ I-rtGH-protein complex were observed: the major one corresponds to a MW of 70 kDa and the minor one to 45 kDa. However, the pattern of electrophoresis varied according to the different incubation buffers tested. Digestion of the cross-linked complex with -galactosidase and phospholipase did not significantly modify the position of bands, whilst N-glycosidase F induced a large smear including 4 more pronounced bands (50, 65, 97 and > 130 kDa), the heavier band corresponding to the most intensive signal.GH receptors were purified using solubilisation and affinity chromatography. The yield of the liver GH-R from crude liver membrane preparation by the solubilization technique was optimized (48%) using Triton 1% for 1 h (12°C ). Specific binding sites in the solubilized membrane proteins were saturable when incubated with increasing ⁵⁰ I-rtGH concentrations, and revealed a high affinity constant (Ka=0.7×10⁹ M^–1). After affinity chromatography, specific binding activity was increased 64,000 fold. However, the purity of the preparation was partial and the purification yield was very low (about 0.3%). This enriched fraction, analysed by SDS-PAGE after cross-linking, showed a very intense band (about 63 kDa) which disappeared with an excess of cold rtGH.These results suggest that the lower molecular weight observed in salmonid (41 kDa), compared to mamals, is not due to the experimental process. The significance of GH-R size difference between salmonids and mammals is discussed.     

Purification and characterization of hepatic triacylglycerol lipase isolated from rainbow trout,Oncorhynchus mykiss

Trialcylglycerol (TG) lipase was isolated and partially purified from rainbow trout liver. Triacylglycerol lipase activity was assayed by measuring¹⁴C-oleic acid release from¹⁴C-triolein.¹⁴C-oleic acid release was linear for up to two hours. Optimal activity occurred at pH 7.0 and 15°C. Most of the lipase activity was recovered in the cytosolic fraction. A 27,000-fold purification was achieved after Sepharose (Bio-gel A 0.5 M, 200–400 mesh) chromatography of a resuspended 20% ammonium sulfate fraction. The molecular weight of the trout hepatic lipase as determined by size-exclusion chromatography and by SDS-polyacrylamide gel electrophoresis was 40–43 kD. Lipase-mediated hydrolysis of TG resulted in the production of diacylglycerols, monoacylglycerols, and fatty acids. Kinetic analysis indicated that V_max=0.016 nmol/h/mg protein and that K_m=0.28 mM triolein. Lipolytic activity was enhanced in the presence of cAMP/ATP-Mg²⁺. These results suggest that the liver of trout possesses a neutral TG lipase that is responsible for mobilizing stored TG and is catalytically activated by phosphorylation.A part of this work was presented at the Annual Meeting of the American Society of Zoologists, December 26–30, 1990, San Antonio, TX.     

The effect of acid water on oxygen consumption,circulating catecholamines and blood ionic and acid-base status in rainbow trout (Salmo gairdneri,Richardson)

Chronically cannulated rainbow trout were exposed in acid water (pH 4.0) for 72h. The gill potential was strongly dependent on water pH, being blood side negative in neutral water, but positive in acid water. Catecholamine levels increased irregularly during acid exposure, and the Bohr and Root effects were not completely erased by the effect of catecholamines during acid exposure. Long term exposure to low water pH, although causing an acidosis in the fish, did not suppress resting oxygen consumption. Prolonged exposure to acid conditions, however, resulted in an increase in ammonia excretion. Changes in plasma sodium and chloride were similar to that reported previously for trout exposed to low calcium, acid water. We conclude that exposure of trout to pH 4 soft water, although impairing oxygen transport, does not limit resting oxygen consumption but does reduce the scope for activity. More extreme acid conditions do impair resting oxygen uptake.     

Changes in plasma prolactin and growth hormone concentrations during freshwater adaptation of juvenile chum salmon (Oncorhynchus keta) reared in seawater for a prolonged period

Freshwater adaptability of chum salmon was examined in juvenile fish reared in seawater for 4 months. The fish, weighing about 40g, were transferred directly to fresh water in October, when their cohorts are migrating in the North Pacific Ocean. Plasma sodium concentration decreased from 167 mM in seawater to about 130 mM during the first 24h, and increased gradually during 2–7 days after the transfer. No immunoreactive prolactin (PRL) was detected in the plasma of the seawater-adapted fish nor during the first 24h in fresh water. Significant levels of PRL were detected after 2–3 days. The maximal level (2.6 ng/ml) was observed after 5 days and became undetectable again after 7 days; no significant correlation was seen between the changes in plasma sodium and PRL levels during the transfer. Plasma growth hormone levels were relatively constant, except for a significant decrease 12h after the transfer. Although plasma thyroxine levels were highly variable during the experiment, a significant decrease and an increase were observed 12h and 5 days after the transfer, respectively. The present study indicates that juvenile chum salmon retain hyperosmoregulatory ability even after prolonged rearing in seawater. Examination of turnover rates, rather than changes in plasma levels, seems to be essential to clarify the osmoregulatory roles of the hormones.     

三倍体雌性虹鳟性腺发育阶段细胞色素相关基因的表达

Cyp19a1b、Cyp11a1、Cyp11b2、Cyp17a1、Hsd3b1等细胞色素相关基因能够调节硬骨鱼类性类固醇的合成,对性腺发育和性别决定产生影响。本研究以全雌三倍体虹鳟(Oncorhynchus mykiss)为研究对象,正常雌性二倍体虹鳟为对照,选取31~68 dpf(days post fertilization)时间段的虹鳟仔鱼脑组织,采用q RT-PCR和酶联免疫的方法研究以上几种基因的表达状况和脑芳香化酶的活性变化,以期探明导致三倍体雌性虹鳟性腺发育异常的关键原因。q RTPCR结果显示,二倍体中Cyp19a1b在30~50 dpf时表达量上调并且维持在较稳定水平,但50~56 dpf时表达量逐渐下调,之后56~68 dpf表达量持续上调;三倍体中Cyp19a1b表达量在30~35 dpf开始上调,35~47 dpf逐渐下调,47~55 dpf开始第二次上调,之后维持在较稳定水平直至68 dpf,但三倍体Cyp19a1b的表达量显著(P0.05)低于同期二倍体的。二倍体Cyp11a1表达量在34 dpf出现峰值,三倍体Cyp11a1在38 dpf时出现峰值。二倍体Hsd3b1表达量在33~42 dpf时维持在较高水平,在38 dpf时出现高峰;三倍体Hsd3b1表达量在47~59 dpf时较高,在49 dpf出现高峰。二倍体中Cyp11b2在37 dpf出现峰值,之后开始下调;三倍体在40 dpf出现峰值,之后逐渐下调,但三倍体Cyp11b2表达量显著低于同期二倍体。二倍体Cyp17a1的表达量在35~46 dpf时逐渐上升,在45 dpf时达到高峰之后直至69 dpf逐渐下降,并且维持在较为平稳的水平上;但是在相同的实验条件下未检测到同一时期三倍体Cyp17a1的表达量。酶联免疫结果显示,在40 dpf时二者的脑芳香化酶活性到达高峰,但在40~60 dpf时期,二倍体虹鳟脑芳香化酶活性显著(P0.05)高于三倍体虹鳟,尤其在45~50 dpf时,该酶活性分别较三倍体的高1.15倍和1.12倍。以上结果表明三倍体虹鳟早期性腺发育迟缓的原因之一是Cyp19a1b、Cyp11a1、Cyp11b2、Cyp17a1、Hsd3b1等基因的表达晚于二倍体,且表达量低于二倍体,造成雌二醇不能正常合成,最终导致性腺发育迟缓。     

Ultrastructural characteristics of two distinct gonadotropes (GTH I- and GTH II-cells) in the pituitary of rainbow trout Oncorhynchus mykiss

The salmonid pituitary produces two chemically distinct gonadotropins (GTHI and GTHII). Ultrastructural characteristics of GTHI- and GTHII-producing cells were studied in the trout pituitary with electronmicroscopic immunocytochemistry using antisera against salmon GTHIβ- and IIβ-subunits. In pituitaries from vitellogenic fish, GTHI-cells were characterized by numerous dilated cisternae of the granular endoplasmic reticulum (GER) and a small number of Iβ-positive granules (diameter, 100–300 nm), whereas GTHIIβ-immunoreactivity was found on granules (diameter, 200–400 nm) and large globules (diameter, 500–4000 nm) in apparently different cells (GTHII-cells). Distinct cellular distributions of GTHI and GTHII were maintained during gametogenesis, although morphological characteristics of GTHI- and GTHII-cells overlapped each other due to changes in number and size of the granules, globules and cisternae of the GER. Interestingly, the globules in the GTHI-cells were immunonegative for GTHIβ, although in the GTHII-cells they were always stained with GTHIIβ-antiserum. These results confirm that GTHIβ and GTHIIβ are synthesized in distinctly different cell-types in the salmonid pituitary and indicate that morphological characteristics cannot be used to distinguish these two cell-types.

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Résumé L'hypophyse des salmonidés produit deux gonadotropines (GTHI et GTHII) chimiquement distinctes. Les caractéristiques ultrastructurales des cellules produisant la GTHI et la GTHII ont été étudiées par immunocytochimie en microscopie électronique en utilisant des anticorps dirigés contre les sousunités GTHIβ et GTHIIβ de saumon. Chez les poissons en vitellogenèse, les cellules à GTHI se caractèrisent par un réticulum endoplasmique granulaire (REG) contenant de nombreuses citernes dilatées et un petit nombre de granules positives à la GTHIβ (diamètre 100–300 nm) tandis qu'une immunoréactivité à la GTHII était trouvée sur des granules (diamètre 200–4000 nm) et de grands globules (diamètre 500–4000 nm) dans des cellules apparemment différentes (cellules à GTHII). Des distributions cellulaires distinctes de la GTHI et la GTHII se sont maintenues à des stades plus avancées de la gamétogenèse, bien que les caractéristiques morphologiques des cellules à GTHI et GTHII se recoupent suite à des changements dans le nombre et la taiile des granules, des globules et des citernes du REG. II est à remarquer que les globules prśents dans les cellules à GTHI étaient immunonégatifs à la GTHIβ alors que dans les cellules à GTHII ils étaient toujours marqués par l'anticorps contre la GTHIIβ. Ces résultats confirment que, dans l'hypophyse des salmonidés, les GTHIβ et GTHIIβ sont synthetisées dans des types cellulaires différents et indiquent que des caractéristiques morphologiques ne peuvent être utilisées pour distinguer ces deux types cellulaires.

     

The effect of dietary coconut oil on reproductive traits and egg fatty acid composition in rainbow trout (Oncorhynchus mykiss)

Two hundred and twenty rainbow trouts (IBW: 700 g) were randomly allotted to four tanks, with a male/female ratio of 0.56. Fish were fed for 168 d with four experimental diets containing herring oil, cod liver oil and coconut oil with the following inclusion rates: diet A: 12-1-0% respectively; diet B: 6-1-6%; diet C: 0-1-12%; diet D: 0-0-13%. Irrespective of the dietary treatment, weight gains of broodstocks were high (> 3 g/d) and FCR below 2. No significant difference was observed concerning the total amount of eggs spawn, egg average weight (82.5 mg/egg) and lipid content (5.4 mg/egg). However, the fatty acid profile of eggs was significantly affected by the dietary treatments. The content of unsaturated fatty acids, particularly the n-3 fatty acid series (EPA and DHA) significantly decreased with increasing levels of coconut oil in the diet.     

Effects of feeding time,water temperature,feeding frequency and dietary composition on apparent nutrient digestibility in rainbow trout Oncorhynchus mykiss and common carp Cyprinus carpio

A series of experiments was conducted to evaluate the effects of feeding time (daytime vs nighttime feeding), dietary fat content (8 and 20%), feeding frequency (trout 1–3 times/day, carp 2–7 times/day), water temperature (trout 18 and 11°C, carp 25 and 17°C), on the apparent nutrient digestibility in rainbow trout and common carp. The feeding time had little effect on the macronutrient digestibility in both species. In trout, starch digestibility decreased with the decrease of water temperature and with increase of feeding frequency, but in carp, increase of the feeding frequency markedly decreased the macronutrient digestibility and phosphrrus absorption of the high fat diet. Fat digestibility of the beef tallow diet decreased relative to the pollock oil diet in carp, without affecting the phosphorus absorption. Inclusion of raw starch, the digestibility of which was lower than that of gelatinized starch, increased the phosphorus absorption in carp. These results suggest that reduction of water temperature and increase of feeding frequency notably decreased starch digestibility in trout while in carp, the effects of feeding frequency and water temperature on macronutrient digestibility and phosphorus absorption are notable for a high fat diet.     

Survival of salmonids in seawater and the time-frame of growth hormone action

In salmonids, growth hormone (GH) effectively promotes adaptation of freshwater (FW) fish to seawater (SW), but it has been unclear whether GH has osmoregulatory actions apart from those consequent to an increase in body size. Our objectives were first, to examine the minimum time and dose required for GH to enhance SW adaptation; and second, to optimize the conditions for the acute GH response in developing a convenient GH bioassay based on its plasma ion lowering effect. Trout showed markedly improved SW survival when transferred from fresh water 6, 24, or 48h after a single chum salmon GH injection (0.25 μg/g). Preadapting trout to 1/3 SW enhanced the plasma ion lowering effect of ovine GH (oGH) injected 48h before transfer of the fish to 80% SW. Endogenous plasma GH levels were elevated in control trout switched from low salinities to 80% SW but were depressed in oGH-injected fish after transfer. Under optimal test conditions (1/3 SW preadaptation, 48h pre-transfer injection, and 100% SW final challenge), the reduction in plasma Na⁺, Ca⁺⁺, and Mg⁺⁺ levels of oGH-injected fish was dose-dependent. The oGH doses giving minimum and maximum responses were 50 and 200 ng/g, respectively. In short, GH exerts acute osmoregulatory actions that promote SW adaptation in the absence of changes in body size. Compared with growth GH bioassays, the osmoregulatory assay is superior in economy of time, animal costs, and hormone quantity required and potentially in specificity.     

Intraperitoneal injection of sulfur amino acids enhance the hepatic cysteine dioxygenase activity in rainbow trout (Oncorhynchus mykiss)

As part of the investigation into cysteine metabolism in fish, sulfur amino acids and their derivatives were injected intraperitoneally to fingerling rainbow trout (Oncorhynchus mykiss) to examine how the doses of these compounds affect the hepatic cysteine dioxygenase [EC 1.12.11.20] in this species. A dose of 0.25 mmol L-cysteine per 100 g body weight induced the enzyme activity as much as 2.5 times that of the control fish within 4h after the injection. The activity increased proportionally to the increasing dose of cysteine up to the dose of 0.15 mmol per 100 g body weight. The induction was observed to be rather specific to L-cysteine. These findings suggested that the cysteine sulfinate pathway might play an important role in the metabolism of excess cysteine in rainbow trout. The dosage of L-cysteine larger than 0.50 mmol per 100g body weight led to mortality of the fish. The pathway of cysteine catabolism was considered to function to prevent toxic accumulation of cysteine in rainbow trout, as in the case of mammals.