三倍体Gong鳟性转换及性腺发育的研究

用高温刺激法获得三倍体Gong鳟幼体。从开始投饵起,分别投喂含有不同浓度17β－雌二醇（0－60μg/μg）的饵料,持续60天,获得了93％－100％的高三倍体雌性率（对照组的雌性率为52％）。三倍体雄鱼可观察到其精子形成,但性腺成熟度较相对的二倍体差;三倍体雌鱼的生殖细胞多停留在偶线期到粗线期,未发现有性成熟个体。实验结果推测,三倍体Gong鳟雌鱼的不育现象看来不是由于雌雄基因组的组成差异而致,更可能是由于三组染色体造成对Gong鳟卵巢发育更大的阻力。雌二醇处理对三倍体Gong鳟幼鱼的生长和成活没有明显的影响。     

虹鳟spindlin基因克隆及不同倍性的表达分析

spindlin基因是减数分裂纺锤体相关因子, 为了研究spindlin基因在二倍体和三倍体雌性虹鳟减数分裂过程中出现的差异, 通过cDNA末端快速扩增(RACE)技术获得spindlin基因cDNA 4529 bp(GenBank登录号: MN378564), 其中3′非编码区(UTR)和5′非编码区(UTR)分别长3662 bp和141 bp, 开放阅读框(ORF)长726 bp, 编码241个氨基酸, 该蛋白质序列的相对分子量为28.3 kD, 理论等电点值为5.94, 无跨膜结构。同源性分析表明, 虹鳟(Oncorhynchus mykiss)与银大马哈鱼(Oncorhynchus kisutch)同源最高, 高达99.59%。系统发育进化树显示, 虹鳟与大鳞大马哈鱼(Oncorhynchus tshawytscha)和红点鲑(Salvelinus alpinus), 聚为一支。实时荧光定量(RT-PCR)结果显示, spindlin基因在二倍体雌性虹鳟卵巢、肾、肝、脾、肌、鳃、心、眼、肠和鳍组织中均有表达, 其中, 在卵巢中的表达量极显著高于其他组织(P<0.01)。对于二倍体雌性虹鳟, 在受精后240—300d (days post fertilization, dpf)发育阶段, spindlin基因在卵巢组织中的相对表达量显著下降。对于三倍体雌性虹鳟, 该基因在240—330 dpf阶段的表达量显著上升。在同一发育阶段中, spindlin基因在二倍体雌性虹鳟卵巢中的表达量较三倍体雌性虹鳟相对较高, 且均存在极显著差异(P<0.01)。通过免疫组化结果发现, 二倍体雌性虹鳟在240 dpf阶段, Spin蛋白在初级卵母细胞核内信号最强, 在270—330 dpf阶段逐渐减弱; 三倍体虹鳟卵巢在240—330 dpf发育阶段, 在卵原细胞中信号逐渐增强。减数分裂异常是性腺败育的关键原因, 研究结果表明三倍体虹鳟在减数分裂过程中出现异常与spindlin基因的低表达有关, 这可能是卵巢发育阻滞的原因之一。     

三倍体雌性虹鳟性腺发育过程中lc3b基因的表达分析

旨在证明细胞自噬与三倍体雌性虹鳟性腺败育的关系。利用RT-PCR扩增虹鳟自噬基因lc3b(Microtubule associated protein 1 light chain 3 beta,lc3b)的编码区序列,通过透射电镜观察三倍体雌性虹鳟不同发育阶段性腺中自噬小体的形态,结合不同发育阶段性腺中lc3b基因和蛋白的表达,来证明细胞自噬的发生。结果表明,在200 dpf-270 dpf(Days post fertilization,dpf)这个发育阶段,lc3b基因以及自噬蛋白LC3B-II/LC3B-I的表达处于相对较高的水平,在这个时期内,都观察到了自噬体结构的存在。这些结果证明了性腺细胞自噬为三倍体雌性虹鳟性腺败育的主要原因之一。     

黄颡鱼XX伪雄鱼诱导与全雌种群规模化繁育

研究利用3种雄性化因素, 包括17α-甲基睾丸酮(MT, 5 mg/kg)、来曲唑(LZ, 300 mg/kg)和高温(33.5℃) 联合处理12—65日龄黄颡鱼幼鱼, 并将性成熟的XX伪雄鱼与正常XX雌鱼进行人工繁殖, 开展了全雌黄颡鱼(Tachysurus fulvidraco)规模化繁殖与苗种培育工作。研究发现, MT、LZ和高温共同作用可诱导XX黄颡鱼逆转为生理型雄性, 完全性逆转个体运动型精子比例与XY雄鱼无显著性差异, 组织学切片也显示其精巢中存在大量精子细胞, 推测XX伪雄鱼具有正常的繁殖功能。随后, 以XX伪雄鱼为父本, 正常XX雌鱼为母本开展了规模化人工繁殖, 获得了57万尾基因型全部为XX的黄颡鱼苗种, 并将其成功培育成大规格鱼种。在幼鱼60日龄和120日龄时取样发现, 分别有2.8%和12.0%的个体发生了不同程度的雄性化, 推测其可能受到池塘自然高温的影响而发生了性逆转。其余XX雌鱼卵巢发育良好, 来年繁殖季节可作为规模化人工繁殖的雌性亲本。研究成功开展了全雌黄颡鱼规模化繁育工作, 为全雌黄颡鱼规模化繁育体系的建立提供了基础, 也为黄颡鱼新品种选育中雌性选育提供了保障。     

低鱼粉饲料中添加苏氨酸对三倍体虹鳟抗氧化能力、消化生理及肠道炎症因子基因表达的影响

研究低鱼粉饲料中不同水平苏氨酸对三倍体虹鳟(Oncorhynchus mykiss)生长、抗氧化能力、消化生理及肠道炎症因子基因表达的影响, 以初始体质量(18.42±0.20) g的三倍体虹鳟为研究对象, 在室内200 L流水水族箱中进行摄食生长试验56d。在基础饲料中添加L-苏氨酸, 配制苏氨酸水平分别为0.45%(对照)、0.76%、1.09%、1.29%和1.64%的5种等氮等能饲料。将450尾三倍体虹鳟随机分为5组, 每组处理3个重复, 每个重复30尾鱼。结果表明, 当添加量为1.09%、1.29%和1.64%时, 血清超氧化物歧化酶(SOD)活力显著提高, 同时显著降低了丙二醛(MDA)的含量(P<0.05), 过氧化氢酶(CAT)在1.29%组含量最高, 相比对照组差异显著(P<0.05)。在饲料中添加苏氨酸(0.76%—1.64%)时, 肠道脂肪酶(LPS)和胰蛋白酶的消化酶活性显著高于对照组(P<0.05), 但随着饲料苏氨酸水平达到1.64%时, 消化酶活性有所降低。1.09%和1.65%组的肠道组织形态最佳, 绒毛发达, 排列整齐, 无融合和脱落现象。苏氨酸水平对肠道炎症因子IL-2、IL-8、IL-10、IgM、TNF-α和PepT1都有显著影响(P<0.05), 随着苏氨酸水平的增加, IL-2、IgM和PepT1呈现先升高后降低的变化趋势, 在1.29%和1.09%组的表达量达到最高; IL-8呈逐渐升高趋势; 而IL-10和TNF-α呈下降的变化趋势。研究结果表明, 在低鱼粉饲料(15%)中添加适量水平的苏氨酸, 对提高三倍体虹鳟幼鱼生长、消化酶活性、促进肠道形态结构发育和增强免疫具有积极的作用。     

黄海中南部细纹狮子鱼的生物学特征及资源分布的季节变化

根据2009年7—8月、10月和2010年1月、5月黄海中南部渔业生物底拖网调查数据,对该海域细纹狮子鱼的生物学特征及其分布的季节变化作了分析。结果表明,细纹狮子鱼平均体长和平均体重从春季(4.7 cm、3.3 g)到冬季(34.2 cm、764.9 g)呈显著增加,并且雄性个体平均体长显著大于雌性个体(P<0.05,春季除外)。性比(♀∶♂)随体长组和季节变化,体长越大趋向于雄性,反之趋向于雌性;夏季雄鱼居多(0.70∶1,P<0.05),秋季则为雌鱼居多(1.35∶1,P<0.05),而冬季(产卵群体)和春季性比接近于1∶1。细纹狮子鱼各季节摄食等级均在2.5以上,冬季雄性个体摄食等级显著大于雌性个体(P<0.05),但雄性个体肥满度为全年最低(1.52)。细纹狮子鱼相对资源量和贡献率从春季(0.17 kg/h,1.54%)到秋季(15.36 kg/h,33.05%)呈上升状态,而冬季(2.37 kg/h,5.60%)有所下降。相比2000年,夏秋季相对渔获量和贡献率提高明显。全年集中分布于7.8—13.6℃,3.20%—3.38%的水域,平均体重和水深有显著的相关性(秋季除外)。另外,根据性成熟个体分布区和稚幼鱼分布的相关历史资料分析发现,除海州湾外,黄海中部深水区可能是细纹狮子鱼的产卵场。     

长江口中华鲟幼鱼对底质的选择

采用单因子实验方法,研究了中华鲟幼鱼对沙(直径＜0.2 cm)、小砾石(直径1～2 cm)、中砾石(直径4～5 cm)、大砾石(直径13～15 cm)4种底质类型的选择.结果表明:单尾鱼实验时,中华鲟幼鱼(体长283.0 mm±18.6 mm,体质量86.2 g±17.8 g,n=30)在4种底质中的时间百分比分别为沙51.50%±21.20%、小砾石20.27%±9.70%、中砾石18.03%±6.17%、大砾石10.20%±11.20%,幼鱼在沙底质中时间明显长于其它3种底质,且差异极显著(P＜0.01);多尾鱼实验时,中华鲟幼鱼(体长278.0 mm±14.3 mm,体质量79.2 g±19.6 g,n=30)在4种底质中的数量百分比分别为沙46.25%±8.06%、小砾石19.38%±7.72%、中砾石18.75%±7.19%、大砾石15.63%±8.92%,幼鱼在沙质底质中的数量明显高于其它3种底质,且差异极显著(P＜0.01);多尾鱼实验中,活动鱼(在水层中游动,不贴底)在4种底质中的数量无显著差异(P＞0.05);非活动鱼(贴底游动或静止)在沙底质中的数量显著高于其它3种底质(P＜0.01).单尾鱼(从时间角度)和多尾鱼(从数量角度)实验均表明,中华鲟幼鱼明显选择沙底质.     

甲胺磷和17β－雌二醇对萼花臂尾轮虫实验种群动态的影响

对暴露于不同浓度的甲胺磷(0.01、0.1、1.0、10.0、100.0、1 000.0和10 000.0 μg·L-1)和17β-雌二醇(0.001、0.01、0.1、1.0、10.0、100.0、和1 000.0 μg·L-1)溶液中的萼花臂尾轮虫的种群动态进行了研究.结果表明:甲胺磷和17β-雌二醇对轮虫种群动态的影响存在差异.甲胺磷和17β-雌二醇对实验期间轮虫的平均种群增长率均有显著影响,但对轮虫种群中的混交雌体受精率的平均值无显著影响;甲胺磷对实验期间轮虫种群中的携卵雌体数/非携卵雌体数和混交雌体百分率的平均值均有显著影响,而17β-雌二醇则无显著影响;甲胺磷对实验期间轮虫的休眠卵总产量无显著影响,而17β-雌二醇的影响较显著.与对照相比,10.0～10 000.0 μg·L-1的甲胺磷和100.0 μg·L-1的17β-雌二醇均使轮虫的平均种群增长率显著提高;0.1～10 000.0和10.0 μg·L-1的甲胺磷分别显著地降低了实验期间轮虫种群中的携卵雌体数/非携卵雌体数和混交雌体百分率的平均值;1 000.0 μg·L-1的17β-雌二醇显著降低了轮虫的休眠卵总产量.在实验浓度范围内,轮虫平均种群增长率(Y,d-1)与甲胺磷浓度(X,μg·L-1)之间的关系为Y=-2×10-8X2 0.0002X 0.3474.种群增长率可用于监测和评价甲胺磷对轮虫种群动态的影响.     

红鳍东方鲀转录因子GATA4基因的表达

利用荧光实时定量PCR(q RT-PCR)技术对红鳍东方鲀(Takifugu rubripes)成鱼的肾脏、肌肉、脑、脾脏、肝脏、精巢和卵巢等组织进行了GATA4基因的表达分析,并对幼鱼不同发育时间的个体水平上的GATA4基因的表达进行分析。结果显示,红鳍东方鲀成鱼的肌肉、脾脏、肾脏和脑等4个组织中的GATA4的表达量差异不显著,精巢、卵巢和肝脏的GATA4的表达量显著高于其他组织(P0.05),GATA4在精巢、肝脏中的表达量显著高于卵巢中(P0.05)。在雄性幼鱼5个发育时期中(23、30、40、60和80日龄),GATA4的表达量从23日龄到30日龄逐渐升高,30日龄最高且明显高于同时期雌鱼GATA4的表达量(P0.01),随后到40日龄之间表达量逐渐降低,从60日龄到80日龄又有上升的趋势。在雌性幼鱼5个发育时期中,GATA4表达量从23日龄到40日龄逐渐升高,40日龄最高且明显高于同时期雄鱼GATA4的表达量(P0.01),到60日龄期间表达量降低,从80日龄后又有上升趋势。各时期雌雄幼鱼,除40日龄外,其他各个时期在雄性中GATA4的表达量均高于在雌性中的表达量。     

注射LHRHa对泥鳅血清性类固醇激素变化研究

研究了注射促黄体激素类似物(LHRHa)后,泥鳅(Misgurnus anguillicaudatus)血清性类同醇激素的变化规律,并探讨在泥鳅繁殖季节时.孕酮(P)、睾酮(T)和雌二醇(E2)对性腺发育的作用及调节机制.实验共分两组,对照组和实验组;对照组只注射生理盐水;实验组注射LHRHa,雌鱼0.2 μg/g,雄鱼减半.注射前尾静脉采血,作为血液样本分析基础水平(Oh),注射药物后分别在7、24、48、72和96h尾静脉采血.测定雌鱼睾酮、雌二醇和孕酮,雄鱼孕酮和睾酬血清浓度.实验结果表明:注射LHRHa,雌鱼血清睾酮和雌二醇浓度显著高于对照组,雄鱼血清睾酮和孕酮显著高于对照组(P<0.05);24h浓度较高.雌鱼孕酬、睾酮和雌二醇分别为(0.710±0.082)ng/mL、(9.00±0.57)ng/mL和(696.4±26.2)pg/mL,雄鱼孕酮和睾酮分别为(0.527±0.121)ng/mL和(9.62±0.62)ng/mL.实验组雌鱼孕酮变化基本规律为,基础水平(0-7h)-逐渐升至最高(7-24h)-逐渐降至基础水平(24-48h)-维持基础水平(48-96h).实验组雌鱼睾酮和雌二醇与雄鱼孕酮和睾酮变化规律基本相似,其规律为,逐渐上升至最高(0-24h)-逐渐降至基础水平(24-72h)-维持基础水(72-96h).24h对照组雌鱼睾酮和雌二醇显著升高,浓度分别为:睾酮(2.20±0.18)ng/mL,雌二醇(269.1±36.6)pg/mL.对照组雄鱼血清孕酮和睾酮浓度实验期间均无显著变化.研究认为:LHRHa能够刺激泥鳅性类同醇激素分泌,特别是睾酮的分泌,显著提高雌鱼性腺指数(GSI).但刺激P的分泌调控能力有限,实验期间处于较低水平,诱导排卵效果差,泥鳅的性类固醇激素可能有特殊的调节机制.雌二醇和睾酮对性腺成熟有重要作用,孕酮对介导卵细胞最终成熟和排卵可能起重要作用,而雌二醇和睾酮无明显效果.     

Gonadal sex reversal in triploid rainbow trout (Oncorhynchus mykiss).

Sex determination in salmonids is primarily governed by sex chromosomes; however, phenotypic expression and successful development of the gonads may be influenced by additional factors. Exposure to exogenous steroids during the critical period of gonadal differentiation will reverse the expected phenotypic sex of both female and male trout. Triploidy, a viable condition in rainbow trout (RBT), alters the degree of gonadal development in a gender-specific manner. Males produce testes with similar morphology and function as diploid fish, but females produce underdeveloped ovaries devoid of growing oocytes. One possible explanation for this observed gender difference is that the timing of meiotic initiation may influence ovarian/testicular development in triploid RBT. To determine whether the early entrance of germ cells into meiosis results in the lack of ovarian development in triploid females, the objective of this study was to sex-reverse genotypic triploid female RBT (XXX) into phenotypic males and genotypic triploid male RBT (XXY) into phenotypic females. Male fish were exposed to estradiol-17beta (E(2)) and females were exposed to the non-aromatizable androgen 17alpha-methyldihydrotestosterone (MDHT). Over 90% of the male fish treated with exogenous E(2) developed gonadal structures indistinguishable from the gonads of triploid females. Triploid female RBT treated with MDHT developed testes; however, not all fish treated with this androgen were completely sex reversed. The results of this investigation are consistent with the hypothesis that the failure of ovarian development in triploid RBT is due to the early onset of meiosis and does not appear to be due to genotypic sex. J. Exp. Zool. 284:466-472, 1999.     

Does triploidization produce functional sterility of triploid males of tench <Emphasis Type="Italic">Tinca tinca</Emphasis> (L.)?

Triploidy interferes with gametogenesis in all fish species tested so far. In fish it results in complete female sterility however, males are still able to develop testis. The reason why sterility levels in triploid fishes differ among species and between sexes is unclear. In the present study the reproductive capacity of triploid males of tench was studied. Flow cytometry revealed sperm cells of triploids to be largely aneuploid with high mosaic DNA, oscillating from haploid DNA to diploid DNA content. Analysis of variance showed an insignificant influence of ploidy level on the percentage of motile spermatozoa, as well as on spermatozoa velocity. Experimental crosses between normal diploid female and triploid males resulted in the appearance of triploid progeny, which exhibited genotypes composed of microsatellite alleles inherited from the founder female and additional allele derived from the donor male. We can conclude that the triploid males analysed in the present study were capable to fertilize eggs derived from diploid females.     

Effect of paired encounters on plasma androgens and behaviour in males and females of the spiny damselfish Acanthochromis polyacanthus

Sexually mature male and female spiny damselfish, Acanthochromis polyacanthus (Pomacentridae) were introduced to resident fish of both sexes in paired encounter trials and the effect on activity, agonistic behaviour and plasma levels of testosterone (T) and 11-ketotestosterone (11-KT) in males, and T and 17β-estradiol (E₂) in females, and cortisol in both sexes was determined. Introduced males showed suppressed levels of T in the presence of both resident males and females, whereas resident males showed elevated T in response to introduced males but not females. Plasma 11-KT levels were unchanged in either resident or introduced males. Resident males showed increased activity in the presence of males but not females. Resident females showed an increase in activity when paired with introduced females and greater aggressive behaviour towards females than males. There were no differences in plasma hormone levels in either resident or introduced female fish. Plasma cortisol levels were low in all fish and stress effects did not appear to account for differences in T levels in males. The results of the present study show that elevations in plasma T levels in resident male spiny damselfish are associated only with male presence but that suppression of T in introduced fish occurs irrespective of the sex of the resident. In contrast resident females, which show stronger evidence of aggressive behaviour do not show changes in plasma androgen levels. This suggests that aggressive behaviour in male and female fishes may be mediated by different endocrine pathways.     

Comparison of the gonadal development and plasma levels of sex steroid hormones in diploid and triploid sea bass, Dicentrarchus labrax L

The goal of this study was to compare the reproductive physiology of triploid and diploid European sea bass (Dicentrarchus labrax L.). Gonads of diploid and triploid fish (males and females) were examined both microscopically and macroscopically, together with the plasma levels of the major sex steroids produced (testosterone and estradiol-17beta) when fish were adults. Prior to sexual maturation, the gonadosomatic index (GSI) of triploid males was similar to that of diploids. However, the GSI in 4-year-old adult triploid males was 1.8 times lower than that of diploids (P < 0.05). All diploid males exhibited normal gonadal development. In contrast, in triploid males spermatogenesis was impaired during late meiosis, affecting severely spermiogenesis. This was achieved by an increasing imbalance in the amount of DNA present in daughter cells of the same type as spermatogenesis progressed, as demonstrated by abnormal cell sizes, culminating in inviable spermatids. Thus, no spermiating triploid fish were observed during 4 years, which included three full consecutive maturation cycles. Furthermore, the germ cells from triploids were significantly larger than those from diploids (P < 0.001). Seasonal profiles of plasma levels of testosterone in 4-year-old males were essentially similar in both ploidies. On the other hand, triploid females had rudimentary ovaries containing oogonia and primary oocytes that were arrested during meiotic prophase I, while diploid females exhibited all stages of ovarian development. Diploid females showed levels of testosterone and estradiol-17beta significantly higher than those of triploids (P < 0.05), in which no endocrine signs of maturation were observed at all. Regarding sex ratios, triploids had 10% more females than diploids (P < 0.05) but in both ploidies males predominated, as is usually found in this species under culture conditions. These results show that triploidy blocked the initial phases of meiosis in females and the latter ones in males, resulting in the absence of or reduced gonadal development, respectively. In conclusion, we provide an explanation for the lack of gonadal development in triploid male fish, and, to the best of our knowledge, we report for the first time a case in which induced triploidy completely blocks meiosis in both sexes, thus conferring functional sterility in the sea bass.     

Why are triploid zebrafish all male?

Adult triploid zebrafish Danio rerio has previously been reported to be all male. This phenomenon has only been reported in one other gonochoristic fish species, the rosy bitterling Rhodeus ocellatus, despite the fact that triploidy is induced in numerous species. To investigate the mechanism responsible, we first produced triploid zebrafish and observed gonad development. Histological sections of juvenile triploid gonads showed that primary growth oocytes were able to develop in the juvenile ovary, but no cortical alveolus or more advanced oocytes were found. All adult triploids examined were male (n = 160). Male triploids were able to induce oviposition by diploid females during natural spawning trials, but fertilization rates were low (1.0 ± 3.1%) compared with diploid male siblings (67.4 ± 16.6%). The embryos produced by triploid sires were aneuploid with a mean ploidy of 2.4 ± 0.1n, demonstrating that triploid males produce aneuploid spermatozoa. After confirming that adult triploids are all male, we produced an additional batch of triploid zebrafish and exposed them (and a group of diploid siblings) to 100 ng/L estradiol (E2) from 5 to 28 dpf. The E2 treated triploids and nontreated triploids were all male. The nontreated diploids were also all male, but the E2 treated diploids were 89% female. This demonstrates that triploidy acts downstream of estrogen synthesis in the sex differentiation pathway to induce male development. Based on this and the observations of juvenile gonad development in triploids, we suggest that triploidy inhibits development of oocytes past the primary growth stage, and this causes female to male sex reversal.     

Gonadal development and an indication of functional protogyny in the Indian damselfish (Dascyllus carneus)

The objective of this study was to determine the sexual pattern of the Indian dascyllus Dascyllus carneus . After an initially undifferentiated state, gonads of D. carneus developed an ovarian lumen and primary growth stage oocytes, and subsequently cortical-alveolus stage oocytes. From ovaries with cortical-alveolus stage oocytes and from more developed ovaries, some gonads redifferentiated into testes. From a sample of 163 individuals, two had a gonad containing degenerating vitellogenic oocytes and proliferating spermatogenic tissue, nine had a gonad containing degenerating cortical-alveolus stage oocytes and spermatogenic tissue, and five had a gonad with degenerating primary growth stage oocytes and spermatogenic tissue. The size of these individuals overlapped greatly with the size range of mature females, suggesting that at least in some individuals, redifferentiation toward a testis occurred after spawning as females. This indicates that D. carneus is a functional, diandric protogynous hermaphrodite. Removal of a dominant male(s) did not induce a sex change in any of the ranking females in the laboratory and field groups. There was no difference in the number of chases and signal jumps performed by the ranking female between control and experimental field groups, or before and after removal of the male. However, the sizes of the ranking females were at or beyond the size range of individuals with a mixed-stage gonad, suggesting that the developmental window for female-to-male sex change may not be open ended. In 41 of 43 field groups, in which sex of fish was determined histologically or by the shape of the urogenital papilla, one to several highest size ranks were occupied by males, followed by one to several females. Mature males, however, were not limited to the highest ranks and occurred at various lower size ranks within groups. Individuals with a mixed-stage gonad also occupied various size ranks within groups.     

Massive production of all-female diploids and triploids in the crucian carp

In many species of aquaculture importance, all-female and sterile populations possess superior productivity due to faster growth and a relatively homogenous size of individuals. However, the production of all-female and sterile fish in a large scale for aquaculture is a challenge in practice, because treatments necessary for gynogenesis induction usually cause massive embryonic and larval mortality, and the number of induced gynogens is too small for their direct use in aquaculture. Here we report the massive production of all-female triploid crucian carp by combining artificial gynogenesis, sex reversal and diploid-tetraploid hybridization. Previously, we have obtained an allotetraploid carp population (4n = 200) by hybridization between red crucian carp (Carassius auratus red var; ♀) and common carp (Cyprinus carpio; ♂). We induced all-female diploid gynogens of the Japanese crucian carp (Carassius cuvieri; 2n = 100). We also generated male diploid gynogens of the same species treated gynogenetic fry with 17-α-methyltestosterone, leading to the production of sex-revered gynogenetic males. Finally, these males were used to cross with the female diploid Japanese crucian carp gynogens and the allotetraploid females, resulting in the production of fertile all-female diploid Japanese crucian carp (2n=100) and sterile all-female triploid hybrids (3n = 150), respectively. Therefore, diploid crucian carp gynogenetic females and sex-reversed male together with an allotetraploid line provide an opportunity to produce all-female triploid populations in a large scale to meet demands in aquaculture industry.     

Enhancing mating performance after juvenile hormone treatment in Anastrepha fraterculus: a differential response in males and females acts as a physiological sexing system

Methoprene (a mimic of juvenile hormone) treatment can reduce the time required for sexual maturation in Anastrepha fraterculus (Wiedemann) (Diptera: Tephritidae) males under laboratory conditions, supporting its use as a treatment for sterile males within the context of the sterile insect technique (SIT). We evaluated sexual behaviour, mating competitiveness of methoprene-treated males, and female readiness to mate after methoprene-treatment in field cages. The study involved two strains of A. fraterculus from Argentina and Peru, which show several polymorphisms in relation to their sexual behaviour. We also analyzed whether methoprene treatment affected male and/or female behaviour in the same way in these two strains. Methoprene-treated males were equally competitive with untreated mature males, and became sexually competitive 6 days after emergence (3–4 days earlier than untreated males). In contrast, methoprene did not induce sexual maturation in females or, at least, it did not induce a higher rate of mating in 7-day-old females. These results were observed both for the Argentina and the Peru strains. Altogether, our results indicate that methoprene treatment produces sexually competitive males in field cages. In the absence of a genetic sexing system, and when sterile males and females of A. fraterculus are released simultaneously, the fact that females do not respond as do males to the methoprene treatment acts as a physiological sexing effect. Therefore, in the presence of mainly sexually immature sterile females, released sexually mature sterile males would have to disperse in search of wild fertile females, thereby greatly reducing matings among the released sterile insects and thus enhancing sterile insect technique efficiency.     

Reproductive performance of male and female phenotypes in three sex chromosomal genotypes(XX, XY, YY) in the killifish, Oryzias latipes.

The present study investigates the juvenile and adult reproductive performance of Japanese killifish, Oryzias latipes, which were successfully sex-reversed by feeding them male or female sex hormones during the fry stage. Sexual maturation of these laboratory grown fish of three known genotypes (XX, SY, YY) occurred earliest in untreated genotypes, next in genotypes treated with male hormone (methyl testosterone) and female hormone (estrone), respectively. The delay in sexual maturation caused by early exogenous, sex-hormone treatment may represent a disturbance in the delicate neural-gonadal axis. However, the degree of response was also strongly influenced by the animal's sex chromosomal genotype. XX fish, normally phenotypic females, were affected more by the male hormone than by the female hormone. XY and YY fish, normally phenotypic males, were delayed more by the female hormone than by the male hormone. This specific sex hormone-sex genotype interaction also influenced adult reproductive performance (sperm or egg production). Differences in the metabolism of male and female sex hormones by the XX, XY, and YY fish are probably responsible for these interesting findings.     

Gonadal morphology of normal and sex-reversed triploid and gynogenetic diploid coho salmon (Oncorhynchus kisutch)

In this study, the gonadal morphology of untreated and sex-reversed juvenile triploid and gynogenetic diploid coho salmon was compared with that of diploids. Testes of triploids were of the same size as those of diploids. Spermatogonia, however, were significantly bigger than those of diptoids in both diameter (P<0·001) and volume (P<0·01), suggesting that this characteristic can be a useful indicator of ploidy in the early stages of gonadal development. In females, induction of triploidy did not affect the lamellar structure of the ovaries but reduced their size considerably. Further, these ovaries had no oocytes. Treatment of triploids with oestrogen resulted in the feminization of genotypic males, which had ovaries similar to those found in tripioid females. However, gonads of triploid males partially sex-re versed into females were identified by their enlargement, the presence of remnants of the male vascular system, and by the appearance of ovarian lacunae and germinal and somatic cells typical of triploid females, Induction of gynogenesis resulted in 100% females, of which 34% had ovaries of reduced size with areas devoid of oocytes. However, and contrary to what has been found in cyprinids, no male germ cells were observed in these ovaries. This discrepancy may reflect differences, in the mechanisms of sex determination between salmonids and cyprinids. Treatment of gynogenetics with androgen increased the number of fish with abnormal ovaries but also resulted in the production of phenotypic-male gynogenetic diploids, of which 11% had testes indistinguishable from those of untreated control diploids.