从线粒体16S rDNA序列探讨绒螯蟹类的系统发生关系

测定了绒螯蟹类各物种的线粒体16SrDNA部分片段的序列,构建了NJ树、ML树和MP树。序列歧异数据比较和各系统发生树都支持新绒螯蟹属(Neoeriocheir)为一个独立的属。在3种系统发生树中,直额绒螯蟹(Eriocheir recta)都是绒螯蟹属(Eriocheir)所有其它成员的姐妹群,并且广东珠江1只直额绒螯蟹标本的16SrDNA部分序列与台湾产台湾绒螯蟹(Eriocheir formasa)的相应序列相同。这些结果不支持平绒螯蟹属(Platyeriocheir)是一个有效的属,并表明E．formosa是E．recta的同物异名。绒螯蟹属(Eriocheir)所有其它成员聚为一个单系的分支,支持中华绒螯蟹、合浦绒螯蟹与日本绒螯蟹属于同一个物种Eriocheir japonica。16SrDNA部分序列的比对表明,产于台湾的日本绒螯蟹的此段序列与合浦绒螯蟹的相同,产于崇明岛的和产于美国旧金山海湾的中华绒螯蟹的此段序列与中华绒螯蟹单元型B的序列相同。     

四种十足目甲壳动物遗传差异的RAPD分析

采用ＲＡＰＤ技术,对分类阶元不同的四种十足目甲壳动物：克氏原螯虾、汉水华溪蟹、日本绒螯蟹合浦亚种和中华绒螯蟹,进行了遗传多态性的研究。四个物种ＤＮＡ库的扩增结果表明,随着亲缘关系由近变远,物种间的相似率依次减小,由５６．１％下降至２２．３％．用ＵＰＧＭＡ方法作聚类分析构建出物种的系统树图、反映出四个物种对应的种、科间、科间以上遗传差异逐步增大,这与以形态标记为主的分类结果相一致。在同工酶分析难以揭     

利用RAPD技术对三种绒螯蟹亲缘关系的研究

采用ＲＡＰＤ技术结合一定的形态学研究手段,对中华绒螯蟹,日本绒螯蟹及其合浦亚种进行亲缘关系的研究,为三种绒螯蟹的种及亚种分析重要资料,三种绒螯蟹的第四步足前节长宽比值,与其分类标本吻合或基本吻合,用一组引物,对每种各１０个个体的基因组ＤＮＡ进行扩增,得到一批特异,可重复的扩增图谱,扩增区带的相似率分析表明,中华绒螯蟹与日本绒螯蟹的亲缘关系远,而日本绒螯蟹合浦亚种与日本绒螯蟹近,最后,对文中ＲＡＰＤ     

中华绒螯蟹与合浦绒螯蟹两地理亚种的线粒体DNA序列变异

根据线粒体16SrDNA的PCR／RFLP鉴定和对Cyt b基因全序列的分析对中国大陆7水系绒螯蟹地理种群的遗传分化进行探讨。结果表明：中华绒螯蟹与合浦绒螯蟹两亚种在17条Cyt b全序列上以40个固定的碱基变异位点相区别。前者分布在瓯江及其以北的水系;后者主要分布在瓯江及其以南的水系。发现了4种在鸭绿江以南的北方水系中生活的合浦亚种单元型,对这些单元型的形成提出了3种可能的解释[动物学报51（5）：862—866,2005]。     

Mitochondrial DNA-inferred population structure and demographic history of the mitten crab (Eriocheir sensu stricto) found along the coast of mainland China

The mitten crab, Eriocheir sensu stricto , is economically important in East Asia, although it is an invasive species in Europe and North America. Little is known about its population structure and historical demography in its native range, especially along the Pacific coast of China. We collected mitochondrial cytochrome oxidase subunit II and cytochrome  b sequences from 154 individuals distributed in the rivers along the Chinese coast and 15 individuals from Japan. Phylogenetic analysis resulted in three major monophyletic groups: northern China, southern China and Japan. Negligible migration was detected among those groups by coalescent analysis. Hence, we support the recognition of three species: Eriocheir hepuensis in southern China, Eriocheir sinensis in northern China and Eriocheir japonica in Japan. The populations in the middle (the Oujiang and Minjiang Rivers) possess a mixture of haplotypes similar to either the northern or the southern haplotypes. We believe that secondary intergradation as the most likely cause of the clinal variation based on examining the genetic variation in the latitudinal space. The estimated divergence time between E. sinensis and E. hepuensis is 2.24 million years ago (Ma), while the divergence time between E. japonica and E. sinensis is 1.83 Ma. Both are in the late Pliocene, suggesting that land bridges associated with low sea level during that time might have severed as vicariant barriers for speciation. The divergence of the northern population and the 'northern haplotypes' in the middle population was estimated at 0.12 Ma, while the time separating the southern population and the 'southern haplotypes' in the middle populations was estimated as 0.16 Ma, implicating possible secondary contact in the late Pleistocene.     

Molecular biomarkers of the physiological state of mitten crab Eriocheir japonica (de Haan, 1835) in the estuarine zones of Peter the Great Bay (Sea of Japan)

The mitten crab Eriocheir japonica (de Haan, 1835) is used as an indicator species for the biochemical monitoring of different estuarine zones in Peter the Great Bay (Sea of Japan). The following molecular biomarkers are determined in the gills and hepatopancreas of the crabs: the activity of biotransformation of enzyme glutathione-S-transferase; antioxidative enzymes, superoxide dismutase, and catalase; and reduced glutathione concentration and lipid peroxidation level measured as malondialdehyde. The state of biochemical indicators testify to the good physiological condition of crabs from estuaries of Shkotovka and Sukhodol rivers and to a depressed physiological condition for crans from the Tesnya River. The latter crabs have lower activity of catalase but higher glutathione-S-transferase activity and lipid peroxidation level. Using the data of molecular-biomarker determination, the integral biochemical index was calculated, which made it possible to rank the estuaries by the extent of adverse effects on the biota. Researching the functional condition of the indicator species allows one to estimate and predict the biota state under anthropogenic impact.     

四种绒螯蟹分子分类与系统发育

利用PCR技术,扩增了中华绒螯解、狭额绒螯蟹线粒体16SrDNA片段,经测序,与GenBank数据库中的日本绒螯解16SrDNA同源序列进行比较。结果显示,在长为376bp的16SrDNA同源序列中有33个多态性核革酸位点（8．78％）,其中种间多态性核苷酸位点28个（7．45％）,种间差异远大于种内差异。引入方蟹科其它近缘种类厚纹蟹、相手蟹和张口蟹的16SrDNA同源序列与上述4种绒螯蟹比较分析,MP法和NJ法构建的分子系统树表明：中华绒螯蟹与日本绒螯蟹亲缘关系最近,首先聚在一起,然后与台湾绒螯蟹聚为一支,狭额绒螯蟹则为相对独立的一支,且进化速度大于前3种绒螯蟹,但最后与前者仍聚在同一组,狭额绒螯蟹只是绒螯蟹属系统进化中的一个侧支,故本研究结果不支持Sakai（1983）和Guo等（1997）把狭额绒螯蟹和台湾绒螯蟹各自立为新属的观点。     

直额绒螯蟹新模记述(甲壳纲,十足目)

形态学和分子生物学的研究表明:直额绒螯蟹Eri- 物异名.本文为直额绒螯蟹指定了新模,并对此进行了描ocheir recta(Stimpson,1858)足一有效种名,应予恢复.台述.湾绒螯蟹Eriocheir formosa Chan,Hung et Yu,1995是它的同物异名.本文为直额绒螯蟹指定了新模,并对此进行了描述.     

Crayfish co-introduced symbiotic ostracod found on native crab in Japan: The first record of epibiont ostracod found a new host

Ankylocythere sinuosa (Rioja, 1942), a symbiotic ostracod native to North America, was found from the Japanese mitten crab Eriocheir japonica (De Haan, 1835), a species native to Japan, collected from a pond in Shizuoka City, Shizuoka Prefecture, central Japan. Introduced North American crayfish Procambarus clarkii (Girard, 1852), which is a host of A. sinuosa in their native range, inhabits ponds sympatrically with Japanese mitten crabs, and it is thought that the ostracods transferred from the exotic crayfish to the native crabs. In recent years, along with the artificial transportation of crayfish around the world, their symbiotic ostracods also have been found on the body surfaces of exotic crayfish in Europe and Japan. However, no studies have confirmed the infestation of exotic ostracods on native crustaceans in the field. A wide range of developmental stages of A. sinuosa from juveniles to adults were found in Japanese mitten crabs, and mating individuals were also found. This strongly suggests that they can reproduce on the body surface of Japanese mitten crabs. In the future, it will be necessary to strengthen measures against alien species to prevent these exotic symbionts from infestating native ecosystems, and we also need to investigate the exact impact of this symbiont on Japanese mitten crabs.     

Molecular connectedness between self and none self-sustainable populations of Chinese mitten crab (Eriocheir sinensis, H. Milne Edwards, 1853) with focus to the Swedish Lake Vänern and the Oder and Vistula River in Poland

The highly invasive Chinese mitten crabs (Eriocheir sinensis) was studied to reveal the genetic relationship between three none self-sustainable population within the Polish Oder and Vistula River and Lake V?nern in Sweden. Crabs from self-sustainable populations were collected from the Elbe River (Germany), San Francisco Bay (USA) and five Asian river estuaries. Both parsimony and maximum likelihood analysis of the nuclear ITS 1 and ITS 2 regions reveal a similar topological pattern. The mitochondrial cytochrome c oxidase subunit I (COI) sequence had too low resolution to be informative. The Oder River haplotypes showed high sequence similarities to both the Elbe population and/or the native spawning areas in China. The Vistula River showed haplotypic similarity to specimens from the Oder River in Poland or the Elbe River. Also high sequence similarities were observed to the specimens from the Swedish Lake V?nern indicating to an origin from the River Elbe. Minor changes in evolutionary interpretation exist depending on how gaps are treated and the analytic method. There is a complex movement involving back and forth transoceanic colonization of the Chinese mitten crab haplotypes. Invasion routes are discussed in relation to ship transfer routes and canal waterways.     

绒螯蟹触角形态特征及其在分类中的应用

基于光镜和扫描电镜的观察,研究了绒螯蟹触角的形态结构,分析了绒螯蟹第1触角内、外肢节的组成,以及外肢节内侧感觉毛的特点;第2触角内肢节的形状以及极长感觉毛和表面鳞片的特征.探讨了在绒螯蟹系统分类中,触角形态特征作为新的分类性状的可能性.     

Sexual cycle properties and changes in carotenoid levels in ovary of Japanese mitten crab <Emphasis Type="Italic">Eriocheir japonica</Emphasis> from rivers of Primorye

The sexual cycle of Japanese mitten crab Eriocheir japonica (de Haan, 1835) in its northern range in water bodies of the Primorye proved to have specific properties including changed seasonal dynamics of gonad maturity stages, delayed spawning period, and reduced breeding season. The ovary color regularly changed during the gonad growth in females, which was accompanied by an increase in total carotenoid content from 1.4 to 22.2 mg/100 g tissue and by changes in the proportion of major pigments. The relative content of the carotene fraction notably decreased and that of astaxanthin slightly decreased during gonad maturation. The revealed changes in carotenoid levels in the ovary point to the activity and significance of pigment metabolism in the reproductive cycle of E. japonica.     

The First Internal Transcribed Spacer (ITS-1) of Ribosomal DNA as a Molecular Marker for Phylogenetic and Population Analyses in Crustacea

The objective of the present study is to explore the feasibility of using the first internal transcribed spacer (ITS-1) of ribosomal DNA as a molecular marker for studying the interspecific and intraspecific genetic variations among crustaceans. We designed primers that could amplify ITS-1 from a majority of taxonomic groups of crustaceans. The gene was found to exhibit a high degree of length polymorphism among different groups, ranging from 182 bp in the barnacle Balanus amphitrite to approximately 820 bp in the spiny lobster Panulirus japonicus. With respect to differences between congeneric species, it was found that the ITS-1 sequences of 3 mitten crabs, Eriocheir sinensis, Eriocheir leptognathus, and Eriocheir formosa, exhibit 5.4% to 16.3% nucleotide divergence, suggesting that ITS-1 is informative for phylogenetic analysis at the species level. Yet there are extensive (0.9%–2.3%) variations within individual E. formosa, so that phylogenetic analyses could be obscured. ITS-1 was found to vary between 2 geographical populations of the shrimp Penaeus japonicus. The variations involved substitutions as well as insertions/deletions between shrimp from Australia and South China Sea. These results show that ITS-1 is highly divergent among different crustaceans and could be an appropriate marker for molecular systematic studies at the species and population levels, although the presence of intragenomic variation needs to be taken into consideration. Received August 15, 2000; accepted February 9, 2001     

Effect of natural disturbance on migrations and population dynamics of a catadromous species, Eriocheir japonicus, along the Lo-Mei stream in northern Taiwan

Japanese mitten crab, Eriocheir japonicus (de Haan), migrates between stream and sea and breeds in shallow coastal areas, exhibiting a catadromous life cycle. This study investigated how the crabs adjust reproductive strategy and migratory behavior to climate fluctuation and what kind of life history strategy individual crabs adopt in a disturbed environment. We conducted a 14-year monitoring study from 1998 to 2011 in Lo-Mei Stream in northern Taiwan. Streamflow and flood timing are controlled by precipitation and typhoons. Abundance of crabs was lower after disturbances by several typhoons and by lower temperatures associated with cold winters in 2006 and 2007. Samples from 2003 showed a reversal in reproductive trends, with increasing size in the same pattern of oocytes, compared to reproductive biology from 1999 to 2005. The life history of E. japonicus can be divided into three stages: spawning migration, growth period, and upstream migration of juveniles to compensate for downstream migration of mature crabs. Reproductively, in E. japonicus the combination of semelparity and synchronous breeding can increase reproductive potential and increase survival in a less disturbed environment at the optimum time. Environmental cues play an important role as major triggers in the persistence of populations.     

患“抖抖病"中华绒螯蟹可溶性蛋白质和同工酶分析

采用垂直板聚丙烯酰胺凝胶电泳技术,对正常的和患“抖抖病”的中华绒螯蟹几种组织中的可溶性蛋白质及乳酸脱氢酶、酯酶和过氧化化物酶等同工酶进行了比较分析。结果表明,病蟹和健康蟹的蛋白质和三种同工酶分离图谱存在明显差异。     

闽江水系绒螯蟹的表型性状差异与分子遗传差异的比较与分析

表型性状差异(differentiation in phenotypictraits,PST)和分子遗传差异(differentiation at neutral molecular markers,FST)是近期进化生物学的研究热点之一。闽江水系是我国中华绒螯蟹与合浦绒螯蟹的主要混杂地域,是研究绒螯蟹遗传与进化的理想地之一。为探讨闽江水系绒螯蟹的PST和FST,以2009和2010年度闽江水系的133个绒螯蟹样本为材料,进行了14个表型数量性状差异分析和6个微卫星标记的遗传差异分析。结果发现:除3个表型性状不存在显著差异外,其他表型性状在不同年份间均存在极显著差异(P<0.01);2009年绒螯蟹的平均期望杂合度极显著高于2010年绒螯蟹(P=0.008),而平均等位基因丰富度、观测杂合度和近交系数均不存在显著差异(P=0.136～0.675);年份间的平均FST为0.1429;通过对PST与FST的比较发现,除第二步足掌节长度(F2)性状外,其他表型数量性状的PST值均高于FST值,表明这些性状均受到了较明显的选择压力。该文研究结果为绒螯蟹的分子进化研究积累了资料,也为其他水产生物的PST和FST比较研究提供了参考。     

稻田网箱养殖辽河水系中华绒螯蟹幼蟹的个体生长

本实验在室外网箱内监测辽河水系中华绒螯蟹(Eriocheir sinensis)幼蟹阶段个体生长发育的每一次蜕壳生长情况。2013年实验选取雄蟹和雌蟹各200只,记录了每个蜕壳阶段雌蟹和雄蟹的生长,包括壳长、壳宽、总重和蜕壳间隔时长(d),并且观察雌蟹和雄蟹形态特征的变化。实验共进行111 d,幼蟹共蜕壳11次。实验结束时,雄蟹剩余34只,雌蟹剩余42只;雄蟹的特定生长率为(7.176 5±0.168 4)%/d,雌蟹的特定生长率为(7.283 3±0.174 3)%/d;雌蟹生长蜕壳过程中腹部的形态变化大,腹部由三角形变成卵圆形;雄蟹在生长蜕壳过程中螯足的增长明显较雌蟹快,并且在本实验最后一次即第11次蜕壳后螯足腹面内侧出现1小撮绒毛,外侧也出现少量绒毛,但不易被发现。     

阳澄湖原产及“洗澡”中华绒螯蟹背甲的几何形态分析

为有效确认非阳澄湖产“洗澡蟹”与阳澄湖原产中华绒螯蟹(Eriocheir sinensis)背甲形态的差异, 研究运用几何形态分析法, 比较研究了前者在阳澄湖中人为“洗澡”式养殖1个月始末时, 其与阳澄湖原产蟹的背甲形态特征。背甲的形态分析基于所建立的35个地标点系统来进行, 即背甲经图像处理后提取地标点坐标, 开展相对扭曲主成分以及判别分析, 再利用薄板样条分析和网格变形将形态变异矢量可视化。相对扭曲得分的逐步判别分析结果显示: “洗澡”式养殖前后阳澄湖原产蟹与非阳澄湖产蟹背甲形态差异均极其显著, 判别准确率均为100%。网格变形结果进一步发现, 与非阳澄湖蟹情况不同, “洗澡”式养殖前后的阳澄湖产蟹背甲形态变化不显著。研究首次定量证实, 非阳澄湖产中华绒螯蟹即使在阳澄湖“洗澡”式养殖1个月, 背甲形态仍无法与原产蟹趋同。这种差异性具有区别阳澄湖原产和“洗澡”蟹的应用潜力。     

植酸酶对中华绒螯蟹幼蟹生长、消化性能及物质利用率的影响

为探讨植酸酶对中华绒螯蟹(Eriocheir sinensis)幼蟹生长、消化性能及物质利用率的影响, 设计了6种配合饲料, 以不含植酸和植酸酶的组别为对照组(C), 在含有10 g/kg植酸的饲料中, 分别加入0、500、1000、1500 U/kg的植酸酶, 分别记为P0、P500、P1000、P1500和P2000。投喂初始体重为(4.34±0.05) g的幼蟹, 56d后称重并取样分析。结果发现: P0幼蟹增重率、特定生长率、蛋白质效率低于对照组, 饲料系数则高于对照组(P<0.05); 幼蟹增重率、特定生长率、蛋白质效率随着饲料中植酸酶含量的增加而升高, 在P2000达到最高, 且该组的饲料系数最低(P<0.05); P1500和P2000全蟹体磷含量显著高于P0 (P<0.05); 在P2000中, 幼蟹肝胰腺中胰蛋白酶、淀粉酶以及肠道胰蛋白酶活力达到最高(P<0.05); 中华绒螯蟹蛋白质消化率和磷透析率随着饲料中植酸酶含量的增加而逐渐升高, 其中P2000显著高于P0(P<0.05), 与对照组无显著差异(P>0.05); P2000幼蟹的氮、磷保留率最高(P<0.05)。以上结果表明, 在含有植酸的饲料中添加2000 U/kg的植酸酶, 能够显著提高幼蟹的生长和胰蛋白酶活力, 进而提高幼蟹对蛋白质的利用率, 降低饲料系数。此外, 植酸酶的添加也能有效提高幼蟹体磷含量和氮/磷保留率。