水产养殖动物基因组研究的现状及其应用前景

The genomic resources from human and several model organisms have been increased very fast since 1990. The techniques for developing genomic resources have already been very advanced and smart. These could make scientists see and improve organism in genomic level. For Chinese aquaculture scientists and aquatic industry, developing genomic resources and genetic tools for the native species are most important in the genomic era. The genomic resources and genetic tools for several aquatic species have been developed and some of them have been used in the marker based selection and other researches. The genome research work on aquaculture species was reviewed in this paper, especially a USDA genome project was focused. Some functional genomic research for aquatic animal was also discussed here. The importance and necessity of China aquaculture species genome project were discussed. Common carp and other cultured fishes in Cyprinidae such as grass carp, silver carp, bighead carp etc were recommended as the candidate species for genome research, because the output of all carps is almost up to 1/3 of total fisheries output in China. Common carp with another virtue for genome research is that there are much more families and strains in common carp than those in other cultured species in China, and those families and strains are the basis for genome research and mapping quantitative trait loci associated with important economic trait. Although the first linkage map of common carp made by Sun needs to be added with more markers for mapping QTL and Type I markers ,it has laid the groundwork for QTL mapping and markerassisted selection in common carp. Because the model organism zebrafish and common carp, grass carp and other carps cultured in China all belong to Cyprinidae, the China carp genome research will obtain a lot of useful information from zebrafish genome research. How the China carp genome program will be conducted and what kinds of strategy involved in this program were all suggested. How the results of the genome research of aquaculture species will be used in the aquaculture industry was reviewed and analyzed here.     

鱼类几种新型免疫因子的研究进展

Cytokines are low molecular weight proteins that serve as chemical messengers within the innate and adaptive immune systems. To date, great progresses have been made in fish cytokine researches. A number of cytokine genes have been cloned and sequenced in fish. This review will focus on a number of novel immune-related cytokines including interleukin, interferon, interferon regulatory factors, Myxovirus resistance proteins, transforming growth factor, tumor necrosis factor, chemokines (CC and CXC chemokines), NK dell enhancement factor, MHCⅠ, MHCⅡ and some of their receptors, which have been identified in many fish species recently. Their genes and molecular structures are clarified. These cytokines are evolutionary well conserved. They share high identities at both the nucleotide and amino acid levels with the high vertebrate cytokines, and maintain characteristic structural motifs of those higher vertebrates. The function of some cytokine genes are analyzed in conventional manner by production of recombinant molecules. Several fish cytokines have been identified based on functional similarity to, or cross-reactivity with, mammalian cytokines. Moreover, molecular techniques, such as suppression subtractive hybridization, PCR and cDNA library screening, have recently enabled the identification of fish cytokine genes. Because of fish phylogenetic position and the fact that their immune systems have not been elaborated to the extent seen in mammals, progresses in this field will deepen our understanding of the molecular origins of cytokine genes and extend our knowledge on their mechanisms conferring disease resistance and the recombinant cytokines to control fish diseases.     

水产食品特定腐败菌与货架期的预测和延长

Fresh fish and lightly preserved fish products are welcome by the global market, however, they are also among the most perishable food products. The research on specific spoilage organisms (SSO) reveals the spoilage process of aquatic product. This paper reviews the current knowledge (past ten years) on SSO of fresh fish and lightly preserved fish products with particular emphasis on characteristics of SSO and how to apply this concept to determine, predict and extend the shelf life of aquatic product. During storage, the microflora changes owing to different abilities of the microorganisms to tolerate the preservation conditions. SSO is defined as special microorganisms which can increase rapidly during preservation and has the ability to produce off- odours and off- flavours associated with spoilage, and spoilage metabolites. Identification of an SSO relies on comparison of the sensory and chemical characteristics of spoiled product with those of isolates from the spoilage microflora. Generally, the SSO of fresh fish may be a single species or genus, but the ones of lightly preserved fish products will be more complex. One exciting area for use of SSO aims to obtain quantitative knowledge about probable behavior of SSO and their function during the progression of spoilage. Thus mathematical models on the growth of SSO are established to evaluate the quality lost degree of product, which provide a sound information for the rational development of devices to monitor loss of products shelf life. Models for the growth of Pseudomonas spp, S. putrefaciens, P. phosphoreum have been established, and validated for shelf life prediction of seafood successfully. Another application field of SSO intends to develop the techniques to prolong the shelf life of food products by inhibiting SSO targetedly. Targeted inhibition of spoilage bacteria during preservation reduces their growth and results in a significant extension of shelf life in despite of the activity of non - spoilage organisms has not been influenced. Such techniques have been applied in perversion field of fresh fish and lightly preserved fish.     

转Hu-IFN-α基因草鱼雌核发育F1的遗传配型分析

The Hu-IFN-α gene, which was transducted into downstream promoter of β-actin gene of common carp (Cyprinus carpio), was recombined by DNA recombination technology. These recombined genes were injected into 1-2 cell fertilized eggs of grass carp (Ctenophatyngodon idellus) by microinjection technology, we gained transgenetic fish by molecular detection methods. In order to analyse the genetic expression of tranHu-IFN-α gene gynogensis F₁, which male individualization were gained by raising methyltestosterone, molecular genetic marker technology was used. In our research, 30 random primers were picked out from 48 and were used into RAPD-PCR, the result indicated that 1 169 clear, steady and repeated DNA finger printing bands were achieved. On the basis of gentic distance matrix among tranHu-IFN-α gene gynogenesis F₁ group, the genetic relationship of gynogenesis F₁ were analysed by UPGMA, the results showed the genetic patterns are close between the 3# male gynogenesis F₁ and the the 23# female of gynogenesis F₁, 5# and 27#, 2# and 28#, 2# and 30#. The data indicated that these group could be served as parent of tranHu-IFN-α grass carp (Ctenophatyngodon idellus) pure line.     

丁鱥雌核发育鱼的异源精子诱导及其与亲本的RAPD比较分析

Gynogenesis is thought to be a useful method to generate fully inbred line and to initiate monosex culture in teleost fish. This article presents results of a study of gynogenesis of tench, Tinca tinca L. from Eltrix River of Xinjiang, China, induced by the common carp ( Cyprinus carpio ) sperm inactivated with UV irradiation. When cold shocking method was used to prevent exmasion of the second polar body in order to produce gynogenetic diploids, the optimum treated parameters were screened as cold shocking for 20min at 4℃, 5min after insemination. On this condition, 10.23% of the treated eggs survived at feeding stage. A comparative study between the gynogenesis and their parents was made by RAPD technique. 82 random primers of 10 nucleotide long sequence were used in RAPD analysis. The results showed that each primer gives 3- 12 fragments for each sample with the fragment length 330 - 2 460 bp, and the genetic similarity between gynogenesis and their mother was 97.4 %, while the genetic similarity between gynogenesis and their father was 27.4%, and there were no bands only shared by gynogenesis and their father, the common carp. It revealed that the chromosome of their father, the common carp, did not penetrate into the construction of gynogenetic tench.     

ABSTRACTS AND KEY WORDS

Productive Artificial Breeding Technique of Clam Mactra veneriformis
Hao Yong-fang et al. （1）
Abstract In order to explore the artificial breeding technique of clam Mactra veneriformis, the related research had been carried on in the 80 m^3 concrete pond in 2013. The clam Mactra veneriformis spawned until sexual maturity, alga was given priority to with Isochrysis zhanjiangensis. Chaetoceros muelleri.     

海洋贝类雌核发育研究进展和展望

The paper summarized the progresses and achievements of the studies on the gynogenesis of the marine mollusks at home and abroad in recent years,and pointed out the study direction on gynogenetic shellfishes and 6-DMAP used as an ideal medicine for raising the induced rate of marine shellfish gynogenesis.     

鱼类粘膜免疫研究进展

Fish immunology has achieved great progress in recent years. While before 1990s, most researches focused on the fish systematic immunity, and the mucosal immunity of fish had not been given enough attention. Indeed, it has been shown that fish mucosal immunity plays an important role in disease defense. Fish mucosal immunity research has made some exciting progress in this decade. This review will focus on such progress: Constitution of mucosal-associated tissues and distribution of different immune cells, including T/B lymphocytes, granules, monocytes, macrophages, goblet cells, etc, in these sites have been well described with the development of some monoclonal antibody to these cells and associated techniques. Non-specific immune response mechanism of mucosal tissues reported these years, such as secretion of non-specific anti-bacteria and anti-fungi substances in mucus, the respiratory burst, enzyme activity of immune cells and so on, is believed important for fish disease defense. The specific immunity of mucosal tissues also attracts much interest and makes great achievement in antigen presenting, MHC genes, antibody producing and antibody secreting cells, comparison of serum and mucus immunoglobulin, relationships of immune response between different mucosal immune tissues. Whether mucosal immune system is independent of systematic immune system is another interesting question and causes great concern. In recent years, some evidences from phyletic evolution and ontogenesis show that mucosal immunity is prior to systematic immunity in evolution. Dynamics of antibody producing of mucosal tissues and serum in immersion or oral vaccines immunized fish also shows immune response can be elicited in mucosal tissues independent of systematic immune system. Some researchers also begin to pay attention to factors involved in mucosal immune regulations, for instance, neuromodulators and cytokines. The level of these factors changes in fish immune response process but the mechanisms of regulation still remain unknown. Prospect of the promising future of fish mucosal immunity has also been discussed in this review.     

黄河干流中上游水生生物资源调查研究

In order to protect scientifically the fishery resources and eco-environment of Yellow River , nine investigatioins on hydrobios in the middle and upper reaches of main Yellow River had been carried out since 2002 according to The Standard for the investigation of inland fishery resources. The investigations showed the present status of the fishery resources in these reaches are as follows ：87species （genera）and 8 phylum of phytoplankton,42 species and 6 phylum of zooplankton, 14 species（genera） of banthic organism , 5 species of hydroplant, 38 species of fish. A comparative analysis,With the results of ＂the Yellow River Fishery Resources Survey in 1982＂,shows that the general trend of the plankton is that the average biomass is getting apparently decreased, the aboriginal fishes are endangered intensifically, and there must be some problem of aquatic biological invasion in these reaches. A granual miniaturization of fishes in main Yellow River has been noticed. The authors proposed to carry out investigation and researches on hydrobios resources in main Yellow River , and develop plans to protect the fishery resources in yellow River, which will be an important basis for rehabilitation of the Yellow River waters and it＇s ecological environment, for the maintenance of biological diversity of Yellow River＇s waters. And this work will be of great significance for people＇s life and sustainable development of economics in Yellow River basin.     

鱼类诺达病毒及其所导致的疾病

In recent years, piscine nodaviruses have emerged as major pathogens of a wide range of larval and juvenile marine finfish resulting in high mortality in aquaculture worldwide. Affected fish exhibit a range of neurological signs, such as erratic swimming behaviour with the associated microscopic lesions of necrosis and vacuolation of the central nervous tissues and retina. Numerous roundshaped, unenveloped and 25-30 nm in diameter virus particles were found in the cytoplasm of affected retinal and nerve cells. Nodaviruses have a bipartite genome of positivesense RNA,with RNA1 encoding the RNAdependent RNA polymerase and RNA2 encoding the capsid protein. Both RNA are capped, but not polyadenylated. The family Nodaviridae comprises two genera: Alphanodavirus and Betanodavirus, members of which primarily infect insects and fish, respectively. Therefore, betanodavirus is also named piscine nodavirus. At present, piscine nodaviruses are divided into four genotypes based on partial sequences of the coat protein gene. ELISA and RT-PCR amplification have been developed as specific diagnostic methods for the d etection of the virus. Antibodies to striped jack (Pseudocaranx dentex) nervous necrosis (SJNNV) were found in 65% of plasma samples collected from wild and domestic brood stocks of striped jack, suggesting that the virus is very prevalent. Viral antigens were detected in eggs, larvae, and ovaries of hatcheryreared and wild spawner fish, suggesting both horizontal and vertical modes of transmission of the virus. Selection of nodavirusfree spawners using ELISA for detection of antigens and RT-PCR techniques have successfully reduced incidences of the virus infections in juvenile sea bass (Dicentrarchus labrax),striped jack and barfin flounder (Verasper moseri). The SSN1 and GF cell lines have been successfully used in isolating piscinenodaviruses.Although there are many papers describing the molecular characteristics of betanodavirus, our knowledge of the genomic attributes of these viruses is still limited. Vaccination studies are being undertaken by a number of researchers and need to be fostered. In particular, the use of passive immunization of broodfish with homologous and heterologous, high titre antisera are worthy of investigation.     

中国沿岸几种鲍线粒体16S rRNA基因片段序列比较及鲍属系统发育

用PCR技术克隆耳鲍（Haliolis asinina）、羊鲍（H．ovina）和多变鲍（H．varia）的线粒体16S rRNA基因的片段，并将PCR产物直接进行测序，得到长度530bp左右的片段。将这些序列与杂色鲍（H．diversicolor diversicolor）、九孔鲍（H．diversicolor supertexte）、大鲍（H．gigantea）、盘鲍（H．discus discus）、皱纹盘鲍（Haliotis discus hannai）等5种鲍的相应片段进行序列比较。结果表明，不同种鲍间16S rRNA基因的序列同源性较高，同源性范围为87．36％～90．77％，这说明鲍的这段基因片段在遗传过程中比较保守。序列的A＋T含量约为56．68％，G＋C含量约为43．32％。8种鲍序列的主要核苷酸变异位点集中在1-25bp、240-290bp和320～370bp3个区域。在序列的变异碱基巾，碱基的转换大大多于碱基的颠换，转换与颠换之比达到2．33：1，遗传距离的范围为0．002-0．128。用UPGMA法和NJ法绘制出8种鲍的系统发育树。结论认为，大鲍、皱纹盘鲍和盘鲍之间的遗传差异水平为亚种间的差异水平，台湾产的九孔鲍与大陆沿岸产的杂色鲍之间的差异仅仅是种群间的差异。     

皱纹盘鲍的遗传育种研究进展

皱纹盘鲍为我国北方的重要养殖品种,但随着养殖规模与密度的不断扩大,因种质退化等原因导致暴发性病害频发。因此,皱纹盘鲍的遗传改良研究对于皱纹盘鲍养殖产业的可持续健康发展十分重要。本文对我国皱纹盘鲍（Haliotis discus hannai）的生物学特征、微卫星标记、选择育种、杂交育种、多倍体育种、雌核发育的研究现状进行了综述,并对今后皱纹盘鲍遗传育种的方法和方向作了展望。     

温度胁迫对皱纹盘鲍生理和生化活动的影响

为探讨温度胁迫下皱纹盘鲍(Haliotis discus hannai Ino)的响应机制,采用室内控温实验,通过设置4个温度梯度(5℃、10℃、20℃和25℃),设计温度骤变处理组(皱纹盘鲍从15℃暂养温度直接转移至各实验温度)和温度缓变处理组(0.5℃/12 h),分析温度剧烈变化和温度缓慢变化对皱纹盘鲍耗氧率和排氨率的影响及其差异性;并对高温和低温处理下皱纹盘鲍消化腺中超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、酸性磷酸酶(ACP)和溶菌酶(LSZ)活性的变化情况以及不同组织(血细胞和肌肉组织)中Cu/Zn–SOD基因的表达状况进行了研究。结果表明,皱纹盘鲍耗氧率和排氨率随海水温度的升高而增加,20℃达到最高值;25℃骤变处理组与缓变处理组耗氧率和排氨率间存在极显著差异(P0.01)。5℃和10℃骤变处理组皱纹盘鲍氧氮比(O/N)同缓变处理组间存在极显著差异(P0.01)。在高温胁迫后的3 h,消化腺中SOD、CAT和LSZ活性达到最高,而ACP活性在胁迫6 h后达到最高(P0.01);低温胁迫显著降低皱纹盘鲍LSZ的活性,于胁迫9 h后达到最低(P0.01)。不同温度胁迫下,皱纹盘鲍血细胞和肌肉组织中Cu/Zn–SOD基因的相对表达量均表现上调,与对照组存在显著差异性(P0.05)。本研究表明,温度胁迫能显著影响皱纹盘鲍的生理和生化活动,这将有助于探讨皱纹盘鲍夏季高死亡率的原因,为皱纹盘鲍健康养殖提供一定的理论依据。     

用流式细胞仪检测活体鲍倍性

分别取皱纹盘鲍（Ｈａｌｉｏｔｉｓ ｄｉｓｃｕｓ ｈａｎｎａｉ）足肌和血液制样，经ＤＡＰＩ染色后，用ＰａｒｔｅｃＰＡＳ－Ⅲ型流式细胞仪测定其倍性，试验证明三倍体鲍的ＤＮＡ相对含量是二保体的１．５倍，切足法和抽取血液法两种制样方法均得到满意结果，用流式细胞仪法检测鲍倍性适应鲍多倍体产业化的需要。     

Identification of juvenile abalone <Emphasis Type="Italic">Haliotis diversicolor</Emphasis> based on number of open and sealed respiratory pores

ABSTRACT:   Changes in the number of respiratory pores (open pores) and their imprints (sealed pores) in post-larvae and juveniles were observed and compared for four abalone species Haliotis diversicolor , H. discus discus , H. madaka , and H. gigantea . The first open pore was evident at a shell length (SL) of 1.5 mm in H. diversicolor , 1.9–2.0 mm SL in H. discus discus , and 2.3–2.4 mm SL in H. madaka and H. gigantea . The number of open pores in H. diversicolor gradually increased with growth, with four to five pores at 2.5–18.0 mm SL and five to six pores at 18.0–27.0 mm SL. The other three species maintained four to five open pores after they reached 3.4–4.5 mm SL. The total number of open and sealed pores (TNP) was greater in H. diversicolor than in the other species at the same SL. Juvenile H. diversicolor were identified among field-caught abalone by the difference in the relationship between SL and TNP (SL–TNP relationship) and also by the monoclonal antibody reaction method. The results of the two methods were in perfect agreement, indicating that our method using the SL–TNP relationship is reliable for the identification of H. diversicolor .     

九孔鲍人工育苗试验

报告了我国北方沿海地区利用电厂冷却海水进行九孔鲍人工育苗的试验。试验结果表明;九孔鲍在繁殖匀性及其育苗技术工艺条件等方面与我国以往的皱纹盘鲍常规育苗方法有较明显的差别,必须采取与之相应的技术手段。利用余热水在大连地区进行九孔鲍人工繁殖是可行的。     

二倍体和三倍体皱纹盘鲍精子发生过程的超微结构

比较了二倍体和三倍体皱纹盘鲍精子发生过程中细胞和细胞器的超微结构变化。结果表明，二倍体皱纹盘鲍的精子发生经历了精原细胞、初级精母细胞、次级精母细胞、精子细胞和精子5个阶段。其形态结构发生了一系列变化，主要包括：核染色质浓缩、线粒体的发达与融合、顶体形成和胞质的减少。三倍体皱纹盘鲍各种生精细胞的直径和核径均大于二倍体，精原细胞结构与二倍体相似；初级、次级精母细胞的胞质中，除溶酶体外，线粒体、内质网等细胞器少于二倍体，线粒体大小与二倍体没有差别，但形态不典型，层状嵴不发达；三倍体皱纹盘鲍的精子发生停滞在精子细胞阶段，表现出各种畸形状态，很多趋于解体，没有发现成熟精子。     

金属蛋白酶组织抑制剂在皱纹盘鲍抗弧菌免疫中的作用

由弧菌感染引起的疾病是影响鲍存活率的主要因素,实验以皱纹盘鲍为研究对象,探讨金属蛋白酶组织抑制剂(tissue inhibitor of metalloproteinase,TIMP)在皱纹盘鲍抗弧菌免疫中的功能及其与基质金属蛋白酶1 (matrix metalloproteinase 1,MMP-1)的相互作用关系....     

皱纹盘鲍4个养殖群体杂交子代生长与存活状况比较

为利用杂种优势培育皱纹盘鲍(Haliotis discus hannai)优良新品种,本研究以4个不同养殖群体[黄岛(HD)、荣成(RC)、日本(JP)、大连(DL)]的皱纹盘鲍为亲本,设计4×4完全双列杂交,建立了4个自交家系和12个正反杂交家系,在利用微卫星标记进行亲子鉴定的基础上,对各家系的F1在1、5、13、17月龄时的生长性状、杂种优势率、生长速度和存活情况进行比较,分析杂交效应。结果表明,在各个生长阶段均有部分杂交家系与自交家系相比表现出显著的生长优势;HDRC、HDDL和JPDL家系的生长速度较高;HDDL、HDJP、RCDL、JPRC及RCHD家系有着较高的存活率;在杂种优势方面,HDRC、HDDL与DLHD家系在各生长参数与生长速度上有明显的杂种优势,HDDL、RCDL、DLHD家系在存活率上表现出明显的杂种优势。本研究筛选出的具有优势的交配组合,可作为皱纹盘鲍生产上种苗来源的参考,也为利用杂种优势培育皱纹盘鲍新品种提供了基础资料。     

Genetic variability revealed with microsatellite markers in an introduced population of the abalone Haliotis discus hannai Ino

One of the challenges for the culture of any species is to control the loss of genetic variability, which may result in a decrease in the quality of commercially important traits. The goal of this study is to assess the genetic diversity of a hatchery population of the Pacific abalone ( Haliotis discus hannai ) from the Center for Abalone Production of the Universidad Católica del Norte (CAP-UCN) that is maintained under a breeding programme. We used six polymorphic microsatellite markers within the cultivated population. The loci Awb033 and Awb079 had the highest number of alleles (11 and 10 respectively) and the loci Awb022 and Awb026 the lowest (two and four respectively). The mean number of alleles per locus was 6.83. The average observed and expected heterozygosities were 0.71 and 0.70, respectively, and the average F _IS ( f ) index was −0.023. We compared the population genetic parameters of the CAP-UCN population with previously published data of wild and hatchery populations of the same species. Results indicate lower genetic diversity estimated as allelic richness in the introduced population with a loss of 11–58% alleles per locus. Despite the high allelic loss, the estimated inbreeding coefficient suggests that the breeding programme carried out in the CAP-UCN has controlled and maintained heterozygosity levels successfully. A temporal study is necessary to determine whether the genetic diversity loss detected was caused during the initial introduction of breeders or to the breeding programme actually implemented.