大银鱼人工授精技术研究

大银鱼Protosalanx hyalocranius Abbott系小型经济鱼,因具有繁殖力强,在水库和湖泊中能自行繁殖,繁殖温度低(2～6℃),富竞争性[1],种群形成快,经济价值高等特点而成为广泛移植的理想鱼.现在我国大部分地区都已移植,移植体主要是人工授精的受精卵.多年来,大银鱼人工授精方法很多,技术上许多关键性问题尚未完全解决.本文对亲鱼死亡时间、精子激活时间、受精时加水量(精子浓度)对受精率的影响及3种人工采精方法的比较进行了研究,以期进一步完善大银鱼人工采卵授精技术.     

养殖施氏鲟的人工繁殖

对6尾雌性养殖施氏鲟(AcipenserschrenckiiBrandt)进行人工催产,其中4尾雌鲟顺利排卵;采用输卵管切割手术取卵,获成熟卵7.7kg,共计42.77×104粒;以"半干法"授精,获受精卵28.82×104粒,受精率67.37%;孵出鱼苗12.69×104尾,孵化率43.63%。本研究对人工催情方法和活体取卵方法与手术技巧进行分析,结果发现,人工养殖亲鱼催产效应时间长于野生鱼(已报道);不同时间取出鱼卵的授精效果差异较大,即成熟卵均能受精,但受精率随产卵时间的延长而逐渐降低;经产雄鱼与初产雄鱼间精子活力差异显著,经产雄鱼精子平均快速泳动时间及精子寿命均长于初产雄鱼精子。     

大银鱼人工授精技术研究

大银鱼ＰｒｏｔｏｓａｌａｎｘｈｙａｌｏｃｒａｎｉｕｓＡｂｂｏｔｔ系小型经济鱼 ,因具有繁殖力强 ,在水库和湖泊中能自行繁殖 ,繁殖温度低 (2～ 6℃ ) ,富竞争性[1] ,种群形成快 ,经济价值高等特点而成为广泛移植的理想鱼。现在我国大部分地区都已移植 ,移植体主要是人工授精的受精卵。多年来 ,大银鱼人工授精方法很多 ,技术上许多关键性问题尚未完全解决。本文对亲鱼死亡时间、精子激活时间、受精时加水量 (精子浓度 )对受精率的影响及 3种人工采精方法的比较进行了研究 ,以期进一步完善大银鱼人工采卵授精技术。1　材料和方法1 1　材料…     

扁吻鱼胚胎发育及仔稚鱼形态与生长观察研究

文章采用干法受精获取受精卵的方法,对扁吻鱼(Aspiorhynchus laticeps)胚胎发育和仔稚鱼的形态变化进行观察.研究结果表明,扁吻鱼卵为沉性卵,卵细胞呈青灰色、微黏性.该次孵化共获取成熟卵21.45×104粒,受精率88.95％,孵化率93.27％.扁吻鱼成熟卵卵径(2.46±0.13)mm,受精卵吸水后...     

中国蛤蜊的诱导产卵和胚胎发育观察

用常规方法对成熟期的中国蛤蜊进行人工诱导产卵，可获得大量的受精卵，且受精卵能正常分裂发育。中国蛤蜊卵呈圆球形，粉红色，卵径50Ixm、卵核径约20um。精卵在水中相遇后立即受精，在24～24．5℃的水温条件下，24h后进入D型幼虫期，4.5d进入壳顶初期，8～10d进入壳顶后期。     

索氏六须鲶受精早期精子入卵的扫描电镜观察

用扫描电镜对索氏六须鲶受精早期精子入卵过程进行系统观察,结果显示,索氏六须鲶在受精后３０－６０ｓ内完成精子入卵过程,在受精后２ｍｉｎ精子开始逐渐解体,直至受精后１５ｍｉｎ左右,受精卵外精子全部解体。通过对精子和卵子的精孔管的测量判定索氏六须鲶精方式为单精受精。     

伊犁裂腹鱼胚胎及早期仔鱼发育的观察

2010年5月通过人工催产,人工授精方法获得伊犁裂腹鱼(Schizothorax pseudaksaiensis Herzenstein)受精卵,在人工培育的条件下对其胚胎和仔鱼发育进行观察.结果显示:伊犁裂腹鱼成熟卵为淡黄色,卵径为1.70～1.80 mm.受精40 min后,受精卵卵周隙接近最大化,此时卵径为2.9...     

菲律宾蛤仔受精及早期胚胎发育过程的细胞学观察

利用HOECHST 33258荧光染料对已固定的菲律宾蛤仔受精卵及早期胚胎样品进行染色的方法,连续观察记录了菲律宾蛤仔受精及早期胚胎发育的细胞生物学过程。结果表明,菲律宾蛤仔的成熟未受精卵子处于第一次成熟分裂中期;精卵混合后精子迅速结合于卵子周围并开始进行顶体反应;精子人卵后去致密并且激发卵子恢复两次成熟分裂,排放出第一及第二极体,完成成熟分裂;受精卵完全排放出第二极体的同时早期的精卵原核形成,随后二者相互靠拢,体积扩散膨大,在细胞中央位置形成各自的染色体组后完成联合;菲律宾蛤仔早期胚胎发育速度进程较快;在受精过程中存在极少数的多精受精现象。     

管角螺受精过程的荧光显微观察及保育卵形成分析

为了研究管角螺受精卵在受精和早期卵裂过程中的核相变化以及保育卵的形成,实验通过Hoechst 33258染色、荧光显微观察方法,对管角螺受精和早期卵裂过程中核行为的细胞学变化进行了详细观察,并对其胚胎发育过程中的数量变化进行统计分析。结果显示,管角螺的成熟未受精卵呈卵圆形,平均卵径(0.33±0.03)mm,核相处于第一次成熟分裂中期,在水温为(26±2)°C条件下,1 h精子入卵,2~3 h受精卵先后排出第1、第2极体,完成第一次和第二次成熟分裂,6 h雌、雄性原核形成,靠近融合,第一次卵裂形成2细胞,12~24 h时2细胞继续分裂生成4细胞。研究表明,管角螺受精卵在成熟分裂和卵裂过程中表现出发育的不同步性,同个卵荚内多个发育阶段共存,受精率为43.14%,而受精卵正常发育概率仅5.70%。卵荚内多数卵子不受精或者受精卵停止发育,成为营养卵(保育卵),为正常可育卵消化利用,这两种来源的保育卵比率约为3∶2。管角螺胚胎发育过程中,1~6 d内胚胎数减少明显,担轮幼虫发育率为0.48%,之后数量稳定,直至稚螺出膜。     

南美白对虾的性腺发育、交配、交卵和受精

以池养南美白对虾（Penaeus vannamei）亲虾进行人工促熟，并使之交尾、产卵、受精。实验在上海金山区抗州湾河口低盐地区一套自行设计的水循环系统中进行，育苗用水通过兑配使盐度为27-28。育苗期间基本不换水和移动亲虾。亲虾205尾，雌雄比约1：1。雌虾切除眼柄后3-4d，性腺即开始发育，6-7d产卵，但未受精。手术后约30d，亲虾出现追尾现象、随之发生交配并出现受精卵。实验进行了约60d，共收集虾卵456.7万粒，其中受精卵出现后所产卵为137万粒，通过设置在水循环系统内的集卵网袋，可收集到95％以上的卵，共获受精卵61万粒，平均日受精率为44.5％，无节幼体25.1万尾，受精卵的平均孵化率为41.2％，最高达87.5％。文中还重点讨论了雌雄亲虾性腺发育的特点，产卵交配规律，以及亲虾规格大小、饵料种类、水质状况对南美白对虾交配和受精的影响。     

低渗溶液浓度对黄颡鱼精子活力和受精率的影响

为提高黄颡鱼(Pelteobagrus fulvidraco)人工授精的稳定性,分别用卵子和精子同步激活法以及预激活精子法,在6个梯度浓度(0%～0.5%)范围的低渗溶液中进行人工授精试验,比较精子活力、受精率和孵化率。结果显示:低渗溶液浓度与精子活力、精子激活率和受精率密切相关,在相同浓度的低渗溶液组中采用预激活精子法的受精率显著高于卵子和精子同步激活法(P<0.05),受精率相对应提高了7.7%～14.5%;低渗溶液的浓度和人工授精方式对孵化率无显著影响。结果表明在浓度0.3%的低渗溶液中采用预激活精子法的效果最好。     

Production of androgenetic amago salmon<Emphasis Type="Italic"> Oncorhynchus masou ishikawae</Emphasis> with dispermy fertilization

With the aim of improving artificial androgenesis in teleost fishes, we tested two methods for producing androgenetic diploids of amago salmon (Oncorhynchus masou ishikawae), namely, fertilization of gamma-ray irradiated eggs with fused spermatozoa (sperm-fusion method) and the fertilization of irradiated eggs with untreated sperm followed by the blocking of cell division (mitosis-inhibition method). Our results showed that the optimal condition for sperm fusion was to treat the sperm with 50% polyethylene glycol (molecular weight 7500) for 100 s. The efficiency of the two methods of androgenesis was compared in terms of fertilization rate, hatching rate, and larval survival after hatching. The rate of fertilization was lower with the sperm-fusion method than with the mitosis-inhibition method, but the reverse was true for the hatching rate. The survival rate of hatched larvae was the same with the two methods. Androgenesis was confirmed with a recessive albino color marker, and all viable offspring were found to be heterozygous based on analysis of the microsatellite markers. Our results suggest that androgenesis with the sperm-fusion method is a promising approach with potential applications in both aquaculture breeding programs and the preservation of endangered freshwater fishes.     

两种颜色栉孔扇贝卵及其早期胚胎磷脂和核酸含量的比较

用钼蓝比色法和紫外分光光度法分别测定栉孔扇贝（Chlamys farrPri）两种颜色的卵子及其早期胚胎中磷脂含量和核酸含量，并进行比较。结果表明，橘红色卵子的RNA含量较浅黄色的略高，而两种颜色卵子的DNA含量却非常相似；在2细胞期，高受精率的受精卵其RNA量迅速增加，DNA和磷脂含量变化不大，但低受精率的受精卵相对未受精卵而言，其DNA、RNA和磷脂含量略有降低；在32～64细胞期，受精率高的胚胎中DNA和磷脂含量增加幅度大，RNA基本保持恒定。磷脂含量与卵子或胚胎的颜色可能有关，核酸、磷脂含量的变化与受精率高低有关。本实验旨为鉴定扇贝卵子的质量提供了科学依据。     

Development of an in vitro culture system for producing eel larvae from immature ovarian follicles in Japanese eel <Emphasis Type="Italic">Anguilla japonica</Emphasis>

To standardize conditions during the final maturation and ovulation of ovarian follicles from Japanese eel, we have developed a culture system for the production of fertilizable eggs from post-vitellogenic ovarian follicles in vitro. Post-vitellogenic ovarian follicles were incubated in culture medium supplemented with 17α,20β-dihydroxy-4-pregnen-3-one (DHP) with or without bovine serum albumin (BSA) to assess the effects of protein concentration. Eggs that ovulated during incubation were fertilized, and the remaining follicles were incubated in prostaglandin F_2α (PGF_2α) for a further 3 or 6 h before fertilization. Male eels were injected repeatedly with human chorionic gonadotropin. The quality of eggs obtained under the different culture conditions was evaluated after artificial fertilization in terms of hatching success. Hatching rates tended to decrease with increasing concentrations of BSA in the incubation medium in a dose-dependent manner. The addition of PGF_2α drastically increased the number of eggs that ovulated, but the rate of hatching was greatly decreased compared with eggs obtained earlier by DHP incubation alone. The larvae obtained from artificially fertilized eggs produced in vitro survived for 14 days without feeding. We conclude that in vitro culture systems thus have a great potential for the acquisition of good quality eggs under tightly controlled artificial conditions, culminating in the production of eel larvae.     

中国对虾受精过程的细胞学研究

通过光镜和电镜切片技术,对中国对虾的受挤过程进行了观察。结果表明,中国对虾成熟卵子处于第1次成熟分裂前期,卵子激活后产生胶质层,精子在卵子的诱导下产生顶体反应后进入卵子。中国对虾为多精入卵、单精受精。精子入卵后,卵子完成成熟分裂并举起围卵膜,但精子的存在并非此过程所必需。在卵子进行第1次成熟分裂时,其纺锤体旋转90°。卵子完成成熟分裂后,精卵原核原核化并向卵子中央移动,最后,精卵原核结合形成合子核,卵子完成受挤过程。     

Vitamin contents of eggs that produce larvae showing a high survival rate in the Japanese eel Anguilla japonica

This paper describes the relationship between the egg vitamin concentrations and the egg quality in the Japanese eel Anguilla japonica . No notable relation was found between any vitamin and the fertilization rate. Hatching and survival rates of larvae, however, significantly increased with an elevated level of egg vitamin C (VC). In contrast to VC, the relation between vitamins E (VE) and A (VA) concentrations and survival rate showed a clear peak, with a reduced survival rate at both higher and lower vitamin concentrations. The ratio of VE to lipid or highly unsaturated fatty acid (HUFA) in eggs positively correlated with hatching and survival rates of larvae. High-quality (HQ) eggs were determined as eggs that produced larvae having a survival rate higher than 80% at 8 days post hatch, and low-quality (LQ) eggs were determined as eggs that did not hatch. The level of VC of HQ was significantly higher than LQ. The results of this study suggest that HQ eggs, which produce larvae having a high survival rate, must have high levels of VC and VE/HUFA ratio and contain optimum levels of VA and VE in Japanese eel.     

Maturation and artificial fertilization of eggs of captive common wolf fish, Anarhichas lupus L. from the White Sea

Abstract. A stock of mature common wolffish, Anarhichas lupus L., were kept in tanks at the surface temperature in the sea and at a lowered temperature. At temperatures of 5–8C in August and September all females matured. The fertilization rate of the eggs was studied in relation to various aspects of the fertilization procedure including sperm concentration, the duration of the contact of eggs with the sperm and the time of exposure of the gametes outside of the body of spawners. The maximum fertilization rate was attained at sperm concentrations not less than 1.0 times 10⁶ per ml and when eggs were exposed to sperm for 1–2h. Fertilization of eggs in a narrow cylindrical vessel improved the efficiency of fertilization, with rates of up to 80–95%. Features of the fertilization of eggs of the wolffish are discussed in relation to the reproductive biology of the species.     

Changes in fertility of rainbow trout eggs retained in coelom

ABSTRACT:    Effects of prolonged retention time of ovulated eggs in the parental coelom on fertilization success were studied in rainbow trout Oncorhynchus mykiss using cryopreserved sperm with a uniform fertilizing ability. Proportions of successful fertilization, eyed eggs and hatched alevins were examined at different time periods up to a retention time of 14 days beyond the ordinary stripping time, and were compared with eggs incubated in artificial coelomic fluids (ACF). Eggs that were retained longer in the coelom showed gradual decreases in all the proportions of successful fertilization, eyed eggs and hatched alevins. The progress of cleavage after fertilization slowed with prolonged retention times. Eggs incubated in ACF lost their fertilizing ability much sooner than those retained in the coelom. The hatching rate of eggs retained for 2 weeks in coelom was 36%, while it was 1% in those eggs incubated for 4 days in ACF. Thus, eggs retained in the coelom showed higher fertilization success than those incubated in ACF.     

Evaluation of different procedures for fertilization and larvae production in Hediste diversicolor (O.F. Müller, 1776) (Nereididae,Polychaeta)

Different experimental trials were performed to clarify some aspects of the biology of the polychaete Hediste diversicolor (O.F. Müller, 1776) as a further step towards the development of appropriate breeding protocols for indoor farming systems. In particular, the trials were addressed to evaluate the effectiveness of two fertilization conditions (in vitro and “natural‐like”); induce gamete spawning by exposing mature individuals to thermal shock or to tissue homogenates; estimate the density effects on larval growth and survival; and evaluate the most suitable parameters to be used as proxy for biomass assessment. The highest percentages of fertilized eggs and larvae were obtained by the in vitro fertilization condition. Mature organisms were induced to spawn by exposure to thermal shock although the spawned eggs revealed low rates of fertilization and hatching. The treatment with male tissue homogenates induced females to successful spawning, and the resulting eggs showed high fertilization and hatching rates. The density of larvae in the rearing phase had no effect on growth or on survival rates of juveniles. Finally, allometric evaluations showed that fresh weight and L3 length are the most reliable parameters to be used as proxy for biomass assessment of this species.     

Artificial fertilization in turbot, Scophthalmus maximus (L.): different methods and determination of the optimal sperm–egg ratio

With the aim of improving artificial fertilization (AF) in turbot, Scophthalmus maximus (L.), a series of fertilization experiments was carried out under dry conditions and different wet conditions (eggs/sea water: 2V/V and V/V). Another series of fertilization experiments was carried out with different quantities of sperm pool to determine the optimal ratio of spermatozoa to eggs for each AF method. Sperm pool from two males and eggs from spawns with a viability rate of > 70% were used. The sperm pool’s density (0.4–5.18 × 10⁹ sperm mL^–1) and motility (1–5) had been assessed previously. Significantly different fertilization rates were found when comparing 2V/V and V/V wet conditions. Significantly higher fertilization rates were found in dry fertilization when the sperm–egg ratio was > 9000 spermatozoa per egg and, under wet condition V/V, at 3000–4000 spermatozoa per egg.