三疣梭子蟹野生群体同工酶的遗传多态性分析

采用聚丙烯酰胺不连续凝胶垂直电泳技术对三疣梭子蟹野生群体的同工酶进行检测.分析了48个样本的11种同工酶在三疣梭子蟹肌肉中的表达情况.11种同工酶(MDH、LDH、ME、SOD、 EST、SDH、IDH、ADH、POD、CAT、AAT)共检测出20个基因座位,其中Me-2、Sod-3、Cat-3、Ldh-2共4个座位呈多态(P0.99),多态座位百分数P为20%.平均每个座位的有效等位基因数目Ae为1.230,预期杂合度He为0.094,实际杂合度Ho为0.175.同时还分析了三疣梭子蟹4个多态座位的Hardy-Weinberg遗传偏离指数d.与日本绒螯蟹、中华绒螯蟹等相比,三疣梭子蟹野生群体的遗传多样性水平较高,种质资源还处于较好的状态.     

基于微卫星标记的中华绒螯蟹遗传多样性分析

为了解中华绒螯蟹不同群体的遗传结构，利用微卫星分子标记，分析中华绒螯蟹单年系F5代选育群体、“长江2号”和长江野生群体的遗传多样性。结果表明，6个微卫星位点的等位基因数为7～11，有效等位基因数为4.539 4～9.529 4，观测杂合度为0.638 9～0.861 1，期望杂合度为0.790 7～0.907 7，多态信息含量为0.779 7～0.895 1,6个微卫星位点均具有高度多态性。3个河蟹群体的期望杂合度为0.817 6～0.847 8，多态信息含量为0.784 1～0.812 5，表明所有群体均具有高度遗传多样性。遗传分化指数值为0.052 69～0.084 82，表明所有群体间均有不同程度的遗传分化。AMOVA分析显示，群体间变异占总变异的5.34%，群体内个体间的变异占总变异的94.66%。基于Nei氏遗传距离构建的UPGMA系统进化树显示，单年系F5代选育群体先与“长江2号”群体聚为一类，再与长江野生群体聚为一类。     

星突江鲽同工酶的遗传多样性研究

采用聚丙烯酰胺垂直板不连续凝胶电泳技术对星突江鲽Platichthys stellatus野生亲本繁殖子一代的同工酶进行了检测.分析了44个样本的9种同工酶在肌肉中的表达情况.9种同工酶共记录了20个基因座位,其中Amy-3、Gdh、Ldh、Me和Sdh-1共5个座位呈多态,多态座位百分数是25%.平均每个座位的等位基因有效数值A,为1.204 6,预期杂合度H_e 为0.107 8,实际杂合度H_o为0.182 9.同时还分析了星突江鲽各个多态座位的Hardy-Weinberg遗传偏离指数d.这些数据说明,星突江鲽野生亲本繁殖子一代的养殖群体遗传多样性水平较高,有利于对其资源进行开发以及对其遗传育种工作的开展.     

菲律宾蛤仔4个野生群体的同工酶分析

采用聚丙烯酰胺不连续凝胶垂直电泳技术对黄、渤海沿岸4个野生菲律宾蛤仔群体的同工酶进行了检测,8种同工酶共记录了18个基因座位,其中Est-1、Est-2、Sod-1、Sod-2、Ldh-1和Ldh-2共6个位点是多态的,多态座位百分数为33.33%。测得4群体平均等位基因数目为1.4412,平均有效等位基因数目1.1260,平均实际杂合度0.0570,平均期望杂合度0.0825,Hardy-Weinberg遗传偏离指数-0.2358,表明菲律宾蛤仔4个野生群体的遗传多样性属偏低水平。     

莱州湾三疣梭子蟹的生化遗传分析

采用聚丙烯酰胺不连续凝胶垂直板电泳技术对三疣梭子蟹的同工酶进行检测。分析了13种同工酶在莱州湾三疣梭子蟹肌肉中的表达情况,并对同工酶的表型进行了生化遗传分析。结果表明,13种同工酶有22个基因位点编码,其中Ldh-1,Me-3,Gdh-1,Sdh-1,G6pd-1和Sod-3共6个位点是多态的,多态位点百分数是27.3%。三疣梭子蟹的平均每个位点的等位基因的有效数目Ae为1.2270,平均每个位点的预期杂合度He为0.1170,实际杂合度Ho为0.2159。同时还分析了三疣梭子蟹的各个多态位点的Hardy-Weinberg偏离指数:Ldh-1、Me-3和Sod-3的d值都为1.0000,Gdh-1、Sdh-1和G6pd-1的d值分别为0.5047、0.8726和0.0597。这些数据表明,莱州湾三疣梭子蟹的种质资源还处于较好的状态。     

基于线粒体控制区序列的三疣梭子蟹增殖放流亲蟹遗传多样性研究

为研究山东沿海三疣梭子蟹增殖放流亲蟹群体的遗传多样性状况,实验采用536 bp的线粒体DNA控制区片段作为分子标记,对4个亲蟹群体的遗传多样性和遗传结构进行了分析。结果显示,301个三疣梭子蟹个体共检测到155个单倍型,4个群体的单倍型多样度为0.972 3~0.993 0,核苷酸多样度为0.021 2~0.023 6,表现出丰富的遗传多样性。AMOVA分析和Fst分析结果均显示,4个三疣梭子蟹亲蟹群体间遗传分化微弱,未形成明显的遗传结构,NJ系统树中未出现与各群体相对应的的谱系分支。研究表明,4个增殖放流亲蟹群体的遗传多样性丰富,且其遗传结构与放流海域的野生群体间没有明显分化,种质资源质量较好。此外,群体历史动态分析显示,渤海南部和黄海北部的三疣梭子蟹历史上曾经历过群体扩张事件。     

日本蟳4个地理群体遗传变异的同工酶分析

采用聚丙烯酰胺不连续凝胶垂直电泳技术对中国沿海的大连黑石礁、山东莱州湾、江苏海州湾和浙江象山4个日本蟳地理群体的10种同工酶进行检测,并分析了群体间的生化遗传差异。实验共记录了28个座位,其中m-Mdh-1、Sod-3、Cat-3、Est-1、α-Amy-1、α-Amy-4、Ldh-2、Alp-1和Alp-3共9个座位表现为多态,4个群体的多态座位百分数（P0.99）均为32.14%,平均每个座位等位基因有效值Ae为1.283 0～1.297 9,平均预期杂合度He为0.146 2～0.155 6,平均观察杂合度Ho为0.273 8～0.278 3,遗传距离D为0.000 4～0.001 8。4个群体的聚类分析表明,莱州湾和大连群体亲缘关系最近,而象山群体与其他3个群体遗传距离较远。实验结果表明,日本蟳遗传多样性处于较高水平,种质资源维持良好,有利于其种质资源的开发及遗传改良等工作的开展。     

中华绒螯蟹野生群体和不同水系人工选育群体的遗传多样性分析

本研究利用10个微卫星分子标记分析了中华绒螯蟹(Eriocheir sinensis) 3个不同水系人工选育群体(“长江1号”、“光合1号”和七里海河蟹)和1个海河流域自然群体的遗传多样性和遗传分化水平。结果显示,10个位点在4个群体中的等位基因数(N)为3~17,平均等位基因数为8.5~9.7,平均期望杂合度为0.720~0.745,平均观测杂合度为0.566~0.661,平均多态信息含量为0.687~0.716,近交系数(Fis)范围为–0.080~0.827。在40个群体–位点组合中,有13个群体–位点组合显著偏离哈迪–温伯格平衡(P<0.05)。遗传多样性分析结果显示,与海河自然群体相比,3个人工选育群体遗传多样性水平略有降低,但仍保持在较高水平,具有较大的选育潜力。遗传分化分析结果显示,群体间遗传分化指数(Fst)范围为0.015~0.075,遗传相似度为0.7702~0.9401,遗传距离为0.0617~0.2611。基于Nei’s遗传距离构建了群体UPGMA系统进化树,自然群体和“光合1号”聚为一支,而七里海河蟹群体单独聚为一支。综上所述,4个中华绒螯蟹群体间的遗传分化水平较低,群体遗传多样性较高。本研究将为中华绒螯蟹选育繁育和种质资源利用与管理等提供理论基础。     

中国大陆沿海六水系绒螯蟹_中华绒_省略__群体亲缘关系_生化遗传差异分析

采用聚丙烯酰胺凝胶电泳分析比较了分布于我国大陆沿海 6个主要水系的两种绒螯蟹 (中华绒螯蟹、日本绒螯蟹 )群体间的生化遗传差异。结果发现 ,(1)辽河、黄河、长江和瓯江中华绒螯蟹群体间的生化遗传差异不显著 ,四个群体间的遗传距离D =0 .0 0 0 6～ 0 .0 0 53,属种内群体间差异。 (2 )珠江和南流江日本绒螯蟹群体间的生化遗传差异也不显著 ,两个群体间遗传距离D=0 .0 0 0 7,尚未达到亚种分化水平。 (3)辽河、黄河、长江和瓯江中华绒螯蟹与珠江和南流江日本绒螯蟹在SOD同工酶表型上有明显差异 ,SOD - 2位点仅在辽河、黄河、长江和瓯江中华绒螯蟹中表达 ,而SOD - 4位点仅在珠江和南流江日本绒螯蟹中表达 ,这些可作为中华绒螯蟹与日本绒螯蟹群体区分的生化遗传标志。辽河、黄河、长江和瓯江中华绒螯蟹群体同珠江和南流江日本绒螯蟹群体间的遗传距离较大 ,D =0 .0 72 1～ 0 .0 74 7。 (4 )聚类分析表明 ,在我国大陆沿海水系分布的绒螯蟹可分为两大类群 ,辽河、黄河、长江和瓯江水系的绒螯蟹属北方类群 ,即中华绒螯蟹 ;珠江和南流江的绒螯蟹属南方类群 ,即日本绒螯蟹。     

网箱养殖大黄鱼遗传多样性的同工酶和RAPD分析

采用垂直聚丙烯酰胺凝胶电泳方法(PAGE)和RAPD技术对象山港网箱养殖大黄鱼群体遗传多样性进行检测。结果表明，所分析的12种同工酶共记录了27个基因座位，其中3个基因座位Est-2、Est-3和m-Adh-2为多态，其多态座位比例为11．111％，平均杂合度为0．0279。16个RAPD随机引物共检测出119个位点，其中多态位点20个(16.81％)，个体间的遗传相似系数为0．844～0．972，遗传变异度为0．0927，Shannon多样性指数为6．326，多样性值为0．0532。不论是同工酶电泳分析结果还是RAPD分析结果，均表明象山港网箱养殖大黄鱼遗传多样性水平较低。     

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).     

Evaluation of a Fast Method Based on the Presence of Two Restriction Sites in the Mitochondrial ND5 (mt ND5) Gene for the Identification of Scomber Species

The purpose of this work was to evaluate the suitability of a method based on the presence of two restriction sites (for Hae III and Hindf I) in the mitochondrial NADH dehydrogenase subunit 5 (mt ND5) gene to identify Scomber species. The evaluation was performed on 144 reference and market samples by sequencing of the entire 505-bp fragment of the mt ND5 gene and of a 464-bp fragment of the Kocher fragment of the cytochrome b gene (mt Cytb). Sequence analysis of any of the two fragments allows the identification of each of the four Scomber species, but S. japonicus and S. colias had the same restriction sites at the ND5 amplicon and would not have been differentiated by this analysis. Similarly, loss of the Hae III site in some S. scombrus individuals would have misidentified them as not being Scomber. All the market products were correctly labeled except one acquired in Spain labeled as originating in the Atlantic and containing S. japonicus.     

Imported ornamental fish are colonized with antibiotic‐resistant bacteria

There has been growing concern about the overuse of antibiotics in the ornamental fish industry and its possible effect on the increasing drug resistance in both commensal and pathogenic organisms in these fish. The aim of this study was to carry out an assessment of the diversity of bacteria, including pathogens, in ornamental fish species imported into North America and to assess their antibiotic resistance. Kidney samples were collected from 32 freshwater ornamental fish of various species, which arrived to an importing facility in Portland, Oregon from Colombia, Singapore and Florida. Sixty‐four unique bacterial colonies were isolated and identified by PCR using bacterial 16S primers and DNA sequencing. Multiple isolates were identified as bacteria with potential to cause disease in both fish and humans. The antibiotic resistance profile of each isolate was performed for nine different antibiotics. Among them, cefotaxime (16% resistance among isolates) was the antibiotic associated with more activity, while the least active was tetracycline (77% resistant). Knowing information about the diversity of bacteria in imported ornamental fish, as well as the resistance profiles for the bacteria will be useful in more effectively treating clinical infected fish, and also potential zoonoses in the future.     

Effect of iodophor disinfection of non‐hardened Salmo trutta eggs on their bacterial and fungus load

This study investigated the efficiency of iodophor disinfection (135 ppm active iodine for 15–30 min) of non‐hardened Salmo trutta eggs against different groups of bacteria and against fungus. Egg samples were taken from non‐disinfected and from disinfected eggs, microorganisms were cultured on specific nutrient media and their mass was measured by turbidimetric methods. Bacteria and fungus mass of non‐hardened eggs could be reduced but not eliminated by iodophor disinfection with 135 ppm active iodine for 15 min. The extent of reduction was 47–65% (Experiment 1). The efficiency of disinfection increased with disinfection time as the reduction in bacteria and fungus mass was 40–55% after 15 min and 58–74% after 30 min (Experiment 2). Disinfection efficiency of iodophor solution diluted in water (reduction 49–57%) and of iodophor solution diluted in sodium chloride solution iso‐osmolar to the oocytes (reduction 52–61%) was similar (Experiment 3). The reduction in bacteria and fungus mass was persistent as it was 39–72% lower in embryos deriving from disinfected eggs than in embryos deriving from non‐disinfected ones (Experiment 4). In conclusion, the tested disinfection method is inadequate to eliminate pathogens completely but it could positively influence immune defence of eggs and embryos.     

Nitrogenous waste excretion and accumulation of urea and ammonia inChalcalburnus tarichi (Cyprinidae), endemic to the extremely alkaline Lake Van (Eastern Turkey)

The endemic, anadromous cyprinidChalcalburnus tarichi is the only fish species known to occur in alkaline Lake Van (Eastern Anatolia, Turkey). EightC. tarichi were maintained individually in Lake Van water (17 – 19°C; pH 9.8; 153 mEq·I^–1 total alkalinity; 22 total salinity) and tank water samples analyzed for 24 h in 2 to 4 h intervals. At zero time, < 1µM ammonia was present and urea was undetectable in the tank water; at 24 h, total ammonia and urea made up 114±32 and 35±25µM, respectively. Over the experimental period, ammonia-N and urea-N excretion averaged 1041±494 and 607±169moles·kg^–1 fish·h^–1, respectively. The extent of urea excretion was highly variable between specimens. Uric acid excretion was not detectable.Urea was present at high concentrations in all tissues and plasma (25 – 35moles·g^–1·ml^–1) of freshly caughtC. tarichi; total ammonia content of the tissues was by a factor of 1.9 (liver) to 3.0 (brain) lower. High arginase activity (2.4±0.2 U·min^–1·g^–1) was detected in the liver ofC. tarichi but ornithine carbamoylphosphate transferase, a key enzyme of the ornithine-urea-cycle, was absent. Ureagenesis is likely through degradation of arginine and/or uricolysis. High glutamine synthetase activity (11±0.6 U·min^–1·g^–1) and low ammonia content in brain suggest that, like other teleosts,C. tarichi has an efficient ammonia detoxification in the brain, but in no other tissue.Nitrogenous waste excretion at alkaline pH is discussed. The ability ofC. tarichi to excrete high levels of ammonia at extremely alkaline pH is unique among teleosts studied so far. The mechanism of ammonia excretion under Lake Van conditions remains to be elucidated.