栉孔扇贝、虾夷扇贝及其杂交子代的群体遗传多样性分析

采用扩增片段长度多态性(Amplified Fragment Length Polymorphism,AFLP)标记对栉孔扇贝(Chlamys farreri,♀)、虾夷扇贝(Patinopecten yessoensis,♂)及其Fl群体的遗传多样性进行分析.5对引物组合共扩增得到315个位点,其中311个为多态位点,总的多态位点比例为98.73%.栉孔扇贝、虾夷扇贝及其F1群体的Shannon信息指数分别为0.477 1±0.260 6、0.423 9±0.277 7、0.548 8±0.220 9;3个群体的Nei's基因多样性指数分别为0.329 1±0.185 0、0.289 7±0.195 9、0.382 8±0.161 2,表明杂交子代群体的遗传多样性水平比2个亲本群体都高.栉孔扇贝与虾夷扇贝群体间的遗传距离为0.411 6,栉孔扇贝与F1群体间的遗传距离是0.131 6,虾夷扇贝与F1群体间的遗传距离是0.278 0,表明杂种子代与母本栉孔扇贝遗传距离较近.聚类分析显示,虾夷扇贝群体、栉孔扇贝群体以及F1群体的所有个体分别聚到一起,且都形成独立群体;分子方差分析(AMOVA)结果显示,变异来源有30.47%来自群体间,有69.53%来自群体内,表明群体内的遗传多样性比较丰富.杂种优势的产生与群体遗传多态性有关.     

基于线粒体12S rDNA序列分析岩扇贝与国内主要经济扇贝的亲缘关系

应用PCR扩增和测序技术获得了引进种岩扇贝(Crassadoma gigantea)及我国主要经济扇贝:虾夷扇贝(Patinopecten yessoensis)、栉孔扇贝(Chlamys farreri)和海湾扇贝(Argopecten irradians)的12SrDNA全序列,并进行了相关分析。结果表明,四种扇贝12SrDNA的序列长度在906~964bp之间,其A+T含量在50.57%~56.77%之间;单倍型数、多态性位点数、单倍型多样性指数、核苷酸多样性指数和平均核苷酸差异数分别在2~3、2~3、0.264~0.623、0.000 75~0.002 07和0.427~1.882之间,均表现出较低的遗传多样性水平;种间遗传距离在0.079~0.306之间,其中虾夷扇贝与栉孔扇贝的遗传距离最近,其次是岩扇贝与虾夷扇贝,而岩扇贝与海湾扇贝的遗传距离最远,并且系统进化树分析结果显示虾夷扇贝先与栉孔扇贝聚为一支,再与岩扇贝进行聚类,而海湾扇贝则与其它扇贝聚为另一个单系群,进一步阐明了四种扇贝间的亲缘关系。     

两种扇贝杂交和自交子一代生长性状的比较

以虾夷扇贝和栉孔扇贝为材料,建立4个试验组合：虾夷扇贝♀×栉孔扇贝♂(正交组合)、栉孔扇贝♀×虾夷扇贝♂(反交组合)、虾夷扇贝♀×虾夷扇贝♂(虾夷扇贝自交组合)、栉孔扇贝♀×栉孔扇贝♂(栉孔扇贝自交组合),分析各试验组合受精率、孵化率、幼虫生长及存活、附着变态率、海区保苗率以及养成阶段生长与存活等性状。试验结果显示：4个组合的受精率、孵化率分别在97%、89%以上,组合间差异不显著(P>0.05);幼虫期,反交组合的日平均增长量、眼点幼虫规格较栉孔扇贝自交组合分别提高9.03%、3.74%,正交组合的日平均增长量、眼点幼虫规格较虾夷扇贝自交组合分别提高5.02%、4.32%,组合间的幼虫存活率无显著差异(P>0.05);4个组合的附着变态率差异不显著(P>0.05),正交组合的海区保苗率较虾夷扇贝自交组合提高5.13个百分点;在养成期,反交组合的生长速度、月增长量较栉孔扇贝自交组合分别提高7.41%、7.45%,正交组合的生长速度、月增长量较虾夷扇贝自交组合分别提高17.37%、17.36%,反交、正交组合的存活率较栉孔扇贝自交、虾夷扇贝自交组合分别提高5.85、18...     

几种养殖扇贝微卫星标记的研究进展

我国扇贝养殖规模占世界首位,栉孔扇贝( Chlamys farreri ) 、华贵栉孔扇贝( Chlamys nobilis ) 、虾夷扇贝( Patinopecten yessoensis) 和海湾扇贝( Argopecten irradians ) 是4 种主要养殖扇贝种类。本文主要概述我国4种主要养殖扇贝微卫星标记开发现状以及在遗传多样性分析、遗传图谱构建、物种鉴定等方面取得的研究成果和应用进展,为微卫星标记在扇贝类遗传学研究提供基础资料。     

3种扇贝遗传结构的RAPD分析

用RAPD方法对扇贝科中的虾夷扇贝、海湾扇贝和栉孔扇贝的基因组DNA的特异性遗传标记进行分析,筛选了43个随机引物,有13个引物获得了片段长度在250～2000 bp之间且重复性良好的谱带,每个引物分别获得了4～17个大小不等的片段,共获得了170个扩增片段,其中12个引物得到特异性片段58个,可以将两种或者3种扇贝区别开来。通过Popgene 3.2分析软件包统计表明,虾夷扇贝与海湾扇贝的遗传距离最大,为0.2481;栉孔扇贝与海湾扇贝的遗传距离次之,为0.2161;虾夷扇贝和栉孔扇贝遗传距离最小,其值为0.2133。根据遗传距离指数,用MEGA3统计软件中的UPGMA方法进行聚类分析,构建系统树。     

墨西哥湾扇贝和扇贝“渤海红”及其杂交子代的遗传分析

利用SSR (Simple Sequence Repeats)分子标记技术,对扇贝“渤海红”、墨西哥湾扇贝(Argopecten irradians concentricus)及其杂交子代3个群体共90个个体的遗传多样性进行分析。结果显示,8个SSR位点共扩增出67个等位基因,各位点等位基因数范围为4~14个,平均等位基因数为8.5个。扇贝“渤海红”有效等位基因数(Ne)、平均观测杂合度(Ho)和多态信息含量(PIC)最高,分别为2.3947、0.504和0.462;3个群体的遗传分化指数(Fst)、基因流(Nm)和固定系数(Fis)分别为0.1398、1.5387和0.3698。对亲代与杂交子代间的遗传分化分析表明,杂交子代与扇贝“渤海红”的遗传距离最小(0.1188),遗传相似度最大(0.888),杂交子代的遗传结构更偏向亲本扇贝“渤海红”。研究结果可为扇贝“渤海红”和墨西哥湾扇贝群体种质资源评估和杂交新品种的选育提供理论参考。     

栉孔扇贝与虾夷扇贝杂交及子一代的遗传性状

20 0 0～ 2 0 0 3年在山东省长岛增殖站进行了以栉孔扇贝为母本与虾夷扇贝为父本 (正交 )、虾夷扇贝为母本与栉孔扇贝为父本 (反交 )、栉孔扇贝自交、虾夷扇贝自交的苗种培育 ,并在同一海区进行了养殖试验。结果表明 ,水温 15～ 18℃条件下 ,正、反交均可正常受精 ,受精率在 90 %以上 ,与对照组没有明显差别 ;成体的外部形态与母本基本相同 ,是偏向母本类型的异源二倍体 ;正交组在第 2年高水温季节栉孔扇贝出现大量死亡的情况下 ,成活率达 95 % ,生长速度提高 2 3% ,反交组在苗种中间暂养和养殖过程中的成活率比虾夷扇贝提高 16 % ,生长速度未见显著差别 ;正、反交子一代生殖腺发育正常 ,可排放精、卵。试验证明 ,栉孔扇贝♀×虾夷扇贝♂的杂交子一代 ,外部形态虽与栉孔扇贝基本相同 ,但显著提高了栉孔扇贝的生产性能尤其是抗逆能力 ,远缘杂交育种是解决栉孔扇贝大面积死亡的重要途径之一 ,其杂交子一代已经具有生产使用价值。     

扇贝血细胞胞内酶活性的细胞学观察

分别对海湾扇贝、虾夷扇贝、栉孔扇贝的3种血细胞胞内酶：碱性磷酸酶(ALP)、过氧化物酶(POD)、酚氧化酶(PO)进行了免疫细胞化学研究,并对血细胞酶反应阳性率作了统计。结果表明,3种扇贝血细胞ALP阳性率相差不大,海湾扇贝为46．6％,虾夷扇贝为42．4％,栉孔扇贝为36．8％;3种扇贝血细胞POD阳性率相差较大,栉孔扇贝阳性率高达50．2％,海湾扇贝为44．2％,虾夷扇贝仅为17．2％。3种扇贝血细胞PO孵育反应后,由于海湾扇贝、虾夷扇贝PO反应活性很低,瑞氏染液复染后,无法区分阴性、阳性细胞。实验中仅对栉孔扇贝PO阳性率进行了统计,栉孔扇贝血细胞PO阳性率为39．3％。     

栉孔扇贝♀×虾夷扇贝♂杂交子一代与其双亲同工酶的比较研究

采用不连续垂直聚丙烯酰胺凝胶电泳法,对栉孔扇贝(Chlamys farreri)♀、虾夷扇贝(Pa-tinopecten yessoensis)♂及其杂交子一代的ADH、MDH、SDH、IDH、ME和SOD共6种同工酶的酶谱进行分析,以探讨双亲遗传物质在杂交子代中的表达情况及杂交子代基因的表达特征。结果表明,杂交子代的同工酶酶谱与母本栉孔扇贝的相似,而与父本虾夷扇贝的明显不同,说明父本虾夷扇贝的同工酶在杂交子代中没有表达。幼虫阶段的同工酶酶谱亦表明,父本虾夷扇贝的同工酶在杂交子代的幼虫阶段仍然没有表达。另外,在对母本栉孔扇贝群体和杂交子代群体的同工酶酶谱进行比较研究时发现,在杂交子代中出现了一定比例的杂合子,因此不能确定杂交子代是由雌核发育而来。     

栉孔扇贝和虾夷扇贝杂交子代的GlSH鉴定及其免疫学特性

以栉孔扇贝[Chlamys farreri(Jones etPreston)](♀)和虾夷扇贝(Patinopecten yessoensis)(♂)杂交子代担轮幼虫为材料，分别用栉孔扇贝和虾夷扇贝基因组作探针，采用基因组荧光原位杂交(GISH)的方法，对杂交后代杂交子的确切身份进行初步鉴定。结果表明，子代分别继承了双亲各     

Association between the interannual variation in the oceanic environment and catch rates of bigeye tuna (Thunnus obesus) in the Atlantic Ocean

The environmental processes associated with variability in the catch rates of bigeye tuna in the Atlantic Ocean are largely unexplored. This study used generalized additive models (GAMs) fitted to Taiwanese longline fishery data from 1990 to 2009 and investigated the association between environmental variables and catch rates to identify the processes influencing bigeye tuna distribution in the Atlantic Ocean. The present findings reveal that the year (temporal factor), latitude and longitude (spatial factors), and major regular longline target species of albacore catches are significant for the standardization of bigeye tuna catch rates in the Atlantic Ocean. The standardized catch rates and distribution of bigeye tuna were found to be related to environmental and climatic variation. The model selection processes showed that the selected GAMs explained 70% of the cumulative deviance in the entire Atlantic Ocean. Regarding environmental factors, the depth of the 20 degree isotherm (D20) substantially contributed to the explained deviance; other important factors were sea surface temperature (SST) and sea surface height deviation (SSHD). The potential fishing grounds were observed with SSTs of 22–28°C, a D20 shallower than 150 m and negative SSHDs in the Atlantic Ocean. The higher predicted catch rates were increased in the positive northern tropical Atlantic and negative North Atlantic Oscillation events with a higher SST and shallow D20, suggesting that climatic oscillations affect the population abundance and distribution of bigeye tuna.     

Growth performance,digestive enzyme activity and immune response of Japanese sea bass,Lateolabrax japonicus fed with fructooligosaccharide

In this experiment, a feeding trial was performed to determine the effects of fructooligosaccharide (FOS) on growth performance, digestive enzyme activity and immune response of Japanese sea bass, Lateolabrax japonicus juveniles (initial weight 38.3 ± 0.5 g), and the fish were examined following feeding with six levels of FOS (0, 0.5, 1, 2, 4 and 6 g/kg) for 28 days. Significant enhancement of weight gain (WG) and specific growth rate (SGR) was found in fish fed 1 g/kg FOS incorporated diets (p < .05), while the feed conversion ratio (FCR) in the 1, 2 g/kg FOS groups reduced significantly compared with the control (p < .05). Besides, the crude lipid in the 4, 6 g/kg FOS groups increased significantly compared with the control (p < .05). On the other hand, the erepsin and lipase activities significantly elevated in intestine of fish fed 2 g/kg FOS (p < .05) and the lysozyme activity in serum of fish fed 2 g/kg FOS were significantly higher than that in the control (p < .05). Moreover, the alkaline phosphatase activities in serum of fish fed 0.5, 1, 2 g/kg FOS were significantly higher than in control (p < .05). Regression analysis showed that the relationships between dietary FOS levels and either SGR, FCR, erepsin or lysozyme activities were best expressed by regression equations, and the optimal inclusion levels are 1.37, 1.80, 3.06, 3.11, 1.93 and 1.80 g/kg for SGR, FCR, erepsin, lipase, lysozyme and total superoxide dismutase activities, respectively. Overall, this study revealed that FOS incorporated diets could beneficial for L. japonicus culture in terms of increasing the growth, digestion and immune activities. Under the present experimental condition, the optimal supplementary level of FOS in the diet of L. japonicus is 1–3 g/kg.     

Rare serotype occurrence and PFGE genotypic diversity of Streptococcus agalactiae isolated from tilapia in China

Previously, we reported 10 PEGE types of 85 tilapia Streptococcus agalactiae(GBS), which shifted from Streptococcus iniae in China, by using PEGE method. Presently, larger and more representative tilapia GBS were isolated, for the ?rst time in China, to characterize their serotypes and genetic diversities more precisely than had done before. 168 GBS strains were distributed in ?ve provinces of China, in which Guangdong, Guangxi and Hainan were the major ones, holding36.9%(62/168), 37.5%(63/168) and 19.6%(33/168), respectively. Serotypes, Ia, Ib and III, were observed in these strains and the most predominant one was Ia(95.2%), which mainly distributed in Guangdong, Guangxi and Hainan. Ia initially occurred in 2009, it shoot up to 32.1% in 2010,but decreased to 16.1% in 2011 before went up to 45.2% in 2012. Ib sporadically occurred during2007–2011, III onlyoccurred in 2012. 14 different PFGE types, including 4 new types(N, O,P and Q), were observed, in which B, D, F and G were the predominant types, holding 83.9%(141/168) of the total GBS strains. Ia corresponded to 11 PFGE types(A–H, N–P), in which type D predominated(51%). Ib represented 3 genotypes(I, J and Q) and III harbored only 2genotypes(N and F). Type N and Fsynchronously presented in Ia and III. In summary, the genetic diversity of tilapia GBS varied by serotypes and changed with geographical locations and years.Although Iastillpredominated, new rareserotypeIII alreadyoccurred in China.     

Growth hormone (GH) and reproduction: a review

Interaction between growth and reproduction occurs in many vertebrates and is particularly obvious at certain stages of the life cycle in fish. Endocrine interactions between the gonadotropic axis and the somatotropic axis are described, the potential role of GH being emphasised. A comparative analysis of these phenomena in mammals, amphibians and fish, suggests a specific role of GH in the physiology of puberty, gametogenesis and fertility. It also shows the original contribution made by studies on the fish model in this field of investigations.     

Purification and partial characterization of GtHs (cfLH and cfFSH) from Indian walking catfish (Clarias batrachus) (L.) and development of a homologous ELISA for cfLH

Two gonadotropins (GtH; Qa and Qb) were purified by gel filtration and ion exchange chromatography from the pituitaries of Indian walking catfish (Clarias batrachus). The presence of GtH during purification was assessed by in vitro oocyte maturation and in vivo steroidogenic activity, and their identities were determined by elution profiles, molecular weight, biological activities and yield. The molecular weights of Qa and Qb were 37 and 42 kDa, respectively, and composed of distinct subunits (Qa: 20 and 14 kDa and Qb: 26 and 18 kDa). Polyclonal antibodies raised against Qa immunostained Qa, Qb and pituitary GtH cells. A competitive Qa‐ELISA was developed whose sensitivity was 6.25 ng mL^?1 (1.25 ng well^?1) with intra‐ (3.5%) and inter‐ (12.4%) assay coefficients of variation. Displacement curves parallel to the standard were obtained with plasma and pituitary extracts of catfish, Qb and carp GtHII. The assay was validated by measuring the plasma Qa levels after LHRH treatment and in relation to ovarian growth in the female catfish during different reproductive phases. Based on the results, Qa and Qb corresponded to fish LH and FSH respectively. The findings will increase the knowledge of the mechanisms controlling fish reproduction and identification of sensitive phases in fish in captivity for hormonal manipulation.     

Controlled infection of Poecilia reticulata Peters (guppy) with Tetrahymena by immersion and intraperitoneal injection

Tetrahymena is a protozoan parasite, which infects guppy, Poecilia reticulata Peters, and causes substantial economical losses in commercial farms worldwide. Studies of guppy infected by Tetrahymena require standardized infection protocols. The LD50 for Tetrahymena infection of guppies by intraperitoneal (IP) injection was calibrated, and the level obtained was 946 parasites per fish. Guppy infection with Tetrahymena by immersion, imitating the natural route of infection via the integument, was studied under normal or stress conditions. Exposure to cold and netting (CNI) and to cold only (CI) followed by immersion exposure to 10 000 Tetrahymena per mL resulted in 22.5% and 19.2% mortality, respectively, as compared to 14.2% and 10% in groups that were netted only (NI) or non‐stressed (I). Histopathology revealed that immersion infection resulted in a systemic infection. Lysozyme levels, measured 3 weeks after infection, were significantly higher in the CNI group (288 μg per mg protein) compared with CI‐, NI‐ and I‐treated groups (94.5, 64 and 62.3 μg mg^?1, respectively). There was no evident parasite immobilization activity in body homogenates, suggesting no development of acquired immunity. Re‐infection by IP injection revealed no increase in protection in any of the treatment groups, mortality range of 56.3–75%, higher than in the non‐exposed control (40.6% mortality).     

Involvement of gonadal steroids in final oocyte maturation of white perch (Morone americana) and white bass (M. chrysops): in vivo and in vitro studies

Plasma estradiol-17 (E₂), testosterone (T), 17,20-dihydroxy-4-pregnen-3-one (DHP) and 17,20,21-tri-hydroxy-4-pregnen-3-one (20-S) levels were measured by radioimmunoassay (RIA) in white perch (Morone americana) and white bass (M. chrysops) that were induced to undergo final oocyte maturation (FOM) with human chorionic gonadotropin (hCG). Plasma DHP levels increased in females of both species in association with oocyte germinal vesicle migration (GVM) and germinal vesicle breakdown (GVBD) and decreased thereafter. Plasma 20-S levels also increased with oocyte GVM in white bass, but were several-fold lower than DHP levels. Circulating E₂ and T levels were greatest during GVM and GVBD in both species and decreased to low levels during oocyte hydration and ovulation. Follicles from white perch and white bass which received a priming injection of hCG in vivo, produced both DHP and 20-S in vitro after exposure to hCG and their oocytes underwent GVBD. Ovarian incubates from unprimed fish of either species produced only E₂ and T and their oocytes did not complete GVBD. Oocytes from unprimed bass, but not perch, matured when follicles were exposed to hCG in vitro. Both trilostane and cycloheximide blocked in vitro production of DHP and 20-S and oocyte GVBD by white perch follices. DHP and 20-S were equipotent inducers of FOM in the GVBD bioassay. None of several other structurally-related steroids tested were effective within a physiological range of concentrations. These results indicate a role for DHP and 20-S in the control of FOM in white perch and white bass.     

Cardiac,ventilatory and metabolic responses of two ecologically dissimilar species of fish to waterborne cyanide

Changes in heart rate, ventilatory activity and oxygen consumption were determined in trout (Salmo gairdneri) and brown bullhead catfish (Ictalurus nebulosus) during exposure to a steadily increasing concentration of waterborne cyanide selected to produce death in 8–9 hours for each species. The lethal cyanide concentration for the bullheads was an order of magnitude higher than for trout. Trout developed an immediate and gradually increasing bradycardia throughout the exposure period. Cyanide produced tachycardia in the bullhead followed by a gradual onset of bradycardia as the concentration of cyanide was raised. Pericardial injection of atropine (a muscarinic cholinergic antagonist) indicated that bradycardia in the trout was due initially to increased vagal tone but later due to the direct effect of cyanide on the heart. Hyperventilation in the trout persisted throughout the exposure period, although the rate and amplitude fluctuated and was variable between individual fish. During the last hour of exposure (highest cyanide concentration), ventilation was characterized by rapid, shallow breaths followed by a sudden respiratory arrest. The bullheads exhibited hyperventilation during the first 3 hours of exposure followed by a gradual, linear drop in ventilation rate and amplitude until death occurred. Cardiac and ventilatory responses in both species were attributed to stimulation of central and peripheral chemoreceptors by cyanide. Evidence is presented which suggests the initial response in the bullheads was due, at least in part, to gustatory stimulation by the cyanide. Oxygen consumption of the trout remained above pre-exposure levels for the majority of the test period. Oxygen consumption in the bullhead paralleled the changes in heart and ventilatory rates. Whole-body lactate levels of fingerlings of both species during cyanide exposure were measured to estimate the extent of anaerobiosis. Whole-body lactate levels were much greater in the bullheads than the trout, indicating a higher capacity for anaerobiosis, possibly due to a greater fuel supply. Overall, the trout responded to cyanide in a manner similar to that produced by environmental hypoxia whereas the bullheads experienced a gustatory stimulus which masked the hypoxia-like response.     

An overview of stress physiology of fish transport: changes in water quality as a function of transport duration

This study brings an integrated analysis about the relationship between water deterioration and its physiological consequences in live fish transport. The analysis was focused on the transport water and its deterioration, and physiological challenges imposed on the fish. Usual commercial handling procedures employed to mitigate fish stress during transport were discussed. Future topics of research for the establishment of safer fish transport protocols were proposed. Transport was classified into short (≤8 h) or long transport (>8 h). The main issue in short transports should be the prevention of water pH reduction, while in long transports it is the increase in ammonia. Plasma cortisol is the most employed marker for stress and is acutely elevated upon short episodes of transport, but remains elevated even in long‐transport events. Plasma glucose is perhaps a better marker for handling stress. Plasma lactate, pH, osmolality CO₂ and ions should be more often evaluated. Plasma Na⁺ and Cl^‐ are very useful markers of acidosis, due to their respective exchange for H⁺ and , for acid–base regulation. The establishment of species‐specific transport protocols should be preceded by such combined analyses of water and physiological parameters.