利用COI基因序列分析长江与澜沧江水系日本沼虾群体的遗传结构

测定了我国长江水系和澜沧江水系的日本沼虾9个群体,共79个个体的线粒体COI基因序列片段(约450bp),结果发现有89个变异位点,共计有46个单倍型。其中云南昆明(KM)群体具有较丰富的遗传多样性(h=1.000,π=0.028),而重庆(CQ)群体的遗传多样性最小(h=0.700,π=0.008)。AMOVA分析表明,群体间的遗传变异占总遗传变异的9.66%,而90.34%的遗传变异源于群体内。采用邻接法(NJ)构建的分子系统树显示,46个单倍型明显地聚为长江中下游和长江上游与澜沧江两个族群。其结果可以为合理开发和利用日本沼虾自然野生资源,以及建立和保护日本沼虾种质资源库及基因库提供必要的参考。     

RNA-seq Provides Novel Insights into Response to Acute Salinity Stress in Oriental River Prawn Macrobrachium nipponense

Marine Biotechnology - The oriental river prawn Macrobrachium nipponense is an important aquaculture species in China, Vietnam, and Japan. This species could survive in the salinity ranging from 7...     

日本沼虾AFLP反应体系的建立

目的:建立一个适于日本沼虾研究用的 AFLP 反应体系.方法:以日本沼虾 DNA 为材料,对基因组酶切时间、选择性扩增中Mg2 、dNTP浓度、预扩增产物稀释倍数及选扩性引物M 3/E 3配比等进行了比较分析.结果:酶切5h,选扩 25ul PCR 反应体系中Mg2 2mmol/L,dNTP 1.2rmnol/L,预扩产物稀释 40 倍,选扩引物M 3/E 3配比为8:1,所得产物在毛细管电泳中可得剑稳定的结果.结论:该体系的构建为 AFLP 技术在日本沼虾相关研究中的应用奠定了基础.     

Twelve polymorphic microsatellites in Oriental river prawn, Macrobrachium nipponense

Oriental river prawn, Macrobrachium nipponense, is a commercially important freshwater prawn species in China, Japan, Korea and Vietnam. Due to overfishing for food, the wild stocks M. nipponense are endangered. Twenty microsatellite loci were isolated from the M. nipponense. Twelve of these loci were polymorphic (seven to 16 alleles per locus), with expected heterozygosity ranging from 0.68 to 0.86 (n = 48). These polymorphic loci provide a valuable tool for assessing genetic diversity of wild and cultured populations.     

日本沼虾脑神经元中微绒毛小管的观察

甲壳动物中的神经细胞是调节其生命活动的重要结构,国内外对其超微结构已进行了一些研究[1～5].笔者在用透射电镜观察日本沼虾的脑神经分泌细胞时,在神经元中观察到一种微绒毛小管,在甲壳动物文献上尚未见此类报道.     

日本沼虾脑神经元中微绒毛小管的观察

甲壳动物中的神经细胞是调节其生命活动的重要结构,国内外对其超微结构已进行了一些研究[1～5].笔者在用透射电镜观察日本沼虾的脑神经分泌细胞时,在神经元中观察到一种微绒毛小管,在甲壳动物文献上尚未见此类报道.     

Anthropogenic water conditions amplify predatory impact of the non-native Oriental river prawn Macrobrachium nipponense

Biological Invasions - Investigating non-native species’ impacts on recipient communities in the context of local habitat conditions is vital to understanding their potential invasiveness....     

铅在日本沼虾体内的分布和积累

应用组织化学、透射电镜和原子吸收光谱分析等方法,研究了Pb在日本沼虾各主要器官和组织中的分布和积累情况。结果表明,在浓度为0．625mg·L^-1Pb溶液暴露10d后的日本沼虾触角腺内,具有大量的电子密度较高的Pb颗粒．在电镜下细胞内的溶酶体中沉积有大量的Pb颗粒,这些Pb通过积聚,在细胞顶端部位逐渐增多,从而出现外排现象．中肠细胞内含有Pb颗粒,细胞质出现空泡化,核膜和线粒体内嵴部分解体．肝胰脏细胞内除分布有少量Pb颗粒外,细胞结构基本完整．在沼虾鳃细胞内未发现Pb颗粒,但在鳃丝之间发现少量Pb颗粒吸附在鳃丝的表面原子吸收光谱分析结果表明,触角腺中Pb含量最高,达637．6mg·kg^-1;触角腺在Pb的解毒方面起着重要作用。     

钱塘江日本沼虾野生群体遗传变异的SSR分析

利用微卫星标记分析了钱塘江干流水域闻堰、富阳、场口、桐庐等7个野生日本沼虾群体的遗传多样性和遗传结构。结果表明:10个微卫星位点呈现高度多态性;闻堰、富阳、场口、新安江等中下游野生群体的遗传多样性水平有高于歙县和休宁两个上游群体遗传多样性水平的趋势。符号检验和Wilcoxon符号秩次检验的结果表明,钱塘江日本沼虾群体近期没有发生瓶颈效应,群体数量也没有下降。FST的范围介于0.0201～0.1069。分子方差分析结果显示,大部分的遗传变异(93.48%)存在于个体间,少部分遗传变异(6.52%)存在于群体之间。群体间FST及AMOVA分析表明,群体处于中等遗传分化水平;基于DA遗传距离构建的NJ聚类树显示,地理位置相邻的群体聚在一起。STRUCTURE分析413份参试的日本沼虾样本被分为2个理论种群,即上游歙县和休宁群体为一个理想种群,其余中下游的5个群体为另一个理想种群。日本沼虾的遗传多样性和遗传结构与所生存的地理位置具有相关性。     

Age Structure,Changing Demography and Effective Population Size in Atlantic Salmon (Salmo salar)

Effective population size (N_e) is a central evolutionary concept, but its genetic estimation can be significantly complicated by age structure. Here we investigate N_e in Atlantic salmon (Salmo salar) populations that have undergone changes in demography and population dynamics, applying four different genetic estimators. For this purpose we use genetic data (14 microsatellite markers) from archived scale samples collected between 1951 and 2004. Through life table simulations we assess the genetic consequences of life history variation on N_e. Although variation in reproductive contribution by mature parr affects age structure, we find that its effect on N_e estimation may be relatively minor. A comparison of estimator models suggests that even low iteroparity may upwardly bias N_e estimates when ignored (semelparity assumed) and should thus empirically be accounted for. Our results indicate that N_e may have changed over time in relatively small populations, but otherwise remained stable. Our ability to detect changes in N_e in larger populations was, however, likely hindered by sampling limitations. An evaluation of N_e estimates in a demographic context suggests that life history diversity, density-dependent factors, and metapopulation dynamics may all affect the genetic stability of these populations.THE effective size of a population (N_e) is an evolutionary parameter that can be informative on the strength of stochastic evolutionary processes, the relevance of which relative to deterministic forces has been debated for decades (e.g., Lande 1988). Stochastic forces include environmental, demographic, and genetic components, the latter two of which are thought to be more prominent at reduced population size, with potentially detrimental consequences for average individual fitness and population persistence (Newman and Pilson 1997; Saccheri et al. 1998; Frankham 2005). The quantification of N_e in conservation programs is thus frequently advocated (e.g., Luikart and Cornuet 1998; Schwartz et al. 2007), although gene flow deserves equal consideration given its countering effects on genetic stochasticity (Frankham et al. 2003; Palstra and Ruzzante 2008).Effective population size is determined mainly by the lifetime reproductive success of individuals in a population (Wright 1938; Felsenstein 1971). Variance in reproductive success, sex ratio, and population size fluctuations can reduce N_e below census population size (Frankham 1995). Given the difficulty in directly estimating N_e through quantification of these demographic factors (reviewed by Caballero 1994), efforts have been directed at inferring N_e indirectly through measurement of its genetic consequences (see Leberg 2005, Wang 2005, and Palstra and Ruzzante 2008 for reviews). Studies employing this approach have quantified historical levels of genetic diversity and genetic threats to population persistence (e.g., Nielsen et al. 1999b; Miller and Waits 2003; Johnson et al. 2004). N_e has been extensively studied in (commercially important) fish species, due to the common availability of collections of archived samples that facilitate genetic estimation using the temporal method (e.g., Hauser et al. 2002; Shrimpton and Heath 2003; Gomez-Uchida and Banks 2006; Saillant and Gold 2006).Most models relating N_e to a population''s genetic behavior make simplifying assumptions regarding population dynamics. Chiefly among these is the assumption of discrete generations, frequently violated in practice given that most natural populations are age structured with overlapping generations. Here, theoretical predictions still apply, provided that population size and age structure are constant (Felsenstein 1971; Hill 1972). Ignored age structure can introduce bias into temporal genetic methods for the estimation of N_e, especially for samples separated by time spans that are short relative to generation interval (Jorde and Ryman 1995; Waples and Yokota 2007; Palstra and Ruzzante 2008). Moreover, estimation methods that do account for age structure (e.g., Jorde and Ryman 1995) still assume this structure to be constant. Population dynamics will, however, likely be altered as population size changes, thus making precise quantifications of the genetic consequences of acute population declines difficult (Nunney 1993; Engen et al. 2005; Waples and Yokota 2007). This problem may be particularly relevant when declines are driven by anthropogenic impacts, such as selective harvesting regimes, that can affect age structure and N_e simultaneously (Ryman et al. 1981; Allendorf et al. 2008). Demographic changes thus have broad conservation implications, as they can affect a population''s sensitivity to environmental stochasticity and years of poor recruitment (Warner and Chesson 1985; Ellner and Hairston 1994; Gaggiotti and Vetter 1999). Consequently, although there is an urgent need to elucidate the genetic consequences of population declines, relatively little is understood about the behavior of N_e when population dynamics change (but see Engen et al. 2005, 2007).Here we focus on age structure and N_e in Atlantic salmon (Salmo salar) river populations in Newfoundland and Labrador. The freshwater habitat in this part of the species'' distribution range is relatively pristine (Parrish et al. 1998), yet Atlantic salmon in this area have experienced demographic declines, associated with a commercial marine fishery, characterized by high exploitation rates (40–80% of anadromous runs; Dempson et al. 2001). A fishery moratorium was declared in 1992, with rivers displaying differential recovery patterns since then (Dempson et al. 2004b), suggesting a geographically variable impact of deterministic and stochastic factors, possibly including genetics. An evaluation of those genetic consequences thus requires accounting for potential changes in population dynamics as well as in life history. Life history in Atlantic salmon can be highly versatile (Fleming 1996; Hutchings and Jones 1998; Fleming and Reynolds 2004), as exemplified by the high variation in age-at-maturity displayed among and within populations (Hutchings and Jones 1998), partly reflecting high phenotypic plasticity (Hutchings 2004). This diversity is particularly evident in the reproductive biology of males, which can mature as parr during juvenile freshwater stages (Jones and King 1952; Fleming and Reynolds 2004) and/or at various ages as anadromous individuals, when returning to spawn in freshwater from ocean migration. Variability in life history strategies is further augmented by iteroparity, which can be viewed as a bet-hedging strategy to deal with environmental uncertainty (e.g., Orzack and Tuljapurkar 1989; Fleming and Reynolds 2004). Life history diversity and plasticity may allow salmonid fish populations to alter and optimize their life history under changing demography and population dynamics, potentially acting to stabilize N_e. Reduced variance in individual reproductive success at low breeder abundance (genetic compensation) will achieve similar effects and might be a realistic aspect of salmonid breeding systems (Ardren and Kapuscinski 2003; Fraser et al. 2007b). Little is currently known about the relationships between life history plasticity, demographic change and N_e, partly due to scarcity of the multivariate data required for these analyses.Our objective in this article is twofold. First, we use demographic data for rivers in Newfoundland to quantify how life history variation influences age structure in Atlantic salmon and hence N_e and its empirical estimation from genetic data. We find that variation in reproductive contribution by mature parr has a much smaller effect on the estimation of N_e than is often assumed. Second, we use temporal genetic data to estimate N_e and quantify the genetic consequences of demographic changes. We attempt to account for potential sources of bias, associated with (changes in) age structure and life history, by using four different analytical models to estimate N_e: a single-sample estimator using the linkage disequilibrium method (Hill 1981), the temporal model assuming discrete generations (Nei and Tajima 1981; Waples 1989), and two temporal models for species with overlapping generations (Waples 1990a,b; Jorde and Ryman 1995) that differ principally in assumptions regarding iteroparity. A comparison of results from these different estimators suggests that iteroparity may often warrant analytical consideration, even when it is presumably low. Although sometimes limited by statistical power, a quantification and comparison of temporal changes in N_e among river populations suggests a more prominent impact of demographic changes on N_e in relatively small river populations.     

日本沼虾雄性生殖细胞原代培养方法

细胞培养方法对于研究虾类疾病、内分泌生理是一种有力的工具。本文介绍了日本沼虾(Macrobrachium nipponense)雄性生殖细胞原代培养的方法。实验发现精原细胞和精母细胞在体外具有较强的分裂能力。通过测定细胞的贴壁率及观察细胞生长情况,初步确定了该类细胞体外培养的最适pH范围为7.2~7.4,渗透压范围为400~500mmol/L。     

Demography and Genetic Structure of a Recovering Grizzly Bear Population

ABSTRACT Grizzly bears (brown bears; Ursus arctos) are imperiled in the southern extent of their range worldwide. The threatened population in northwestern Montana, USA, has been managed for recovery since 1975; yet, no rigorous data were available to monitor program success. We used data from a large noninvasive genetic sampling effort conducted in 2004 and 33 years of physical captures to assess abundance, distribution, and genetic health of this population. We combined data from our 3 sampling methods (hair trap, bear rub, and physical capture) to construct individual bear encounter histories for use in Huggins—Pledger closed mark—recapture models. Our population estimate, Ň = 765 (95% CI = 715–831) was more than double the existing estimate derived from sightings of females with young. Based on our results, the estimated known, human-caused mortality rate in 2004 was 4.6% (95% CI = 4.2–4.9%), slightly above the 4% considered sustainable; however, the high proportion of female mortalities raises concern. We used location data from telemetry, confirmed sightings, and genetic sampling to estimate occupied habitat. We found that grizzly bears occupied 33,480 km² in the Northern Continental Divide Ecosystem (NCDE) during 1994–2007, including 10,340 km² beyond the Recovery Zone. We used factorial correspondence analysis to identify potential barriers to gene flow within this population. Our results suggested that genetic interchange recently increased in areas with low gene flow in the past; however, we also detected evidence of incipient fragmentation across the major transportation corridor in this ecosystem. Our results suggest that the NCDE population is faring better than previously thought, and they highlight the need for a more rigorous monitoring program.     

金属铜在日本沼虾体内的分布和积累

应用组织化学、透射电镜和原子吸收光谱分析等方法,研究了铜在日本沼虾各主要器官和组织中的分布和积累情况。结果表明,在铜离子浓度为0.08 mg.L-1的溶液中暴露10 d后的日本沼虾肝胰脏内,具有大量的电子密度较高的铜颗粒,主要分布在肝胰脏的肝小管和R细胞中,在血腔中有从R细胞排出的铜颗粒;电镜观察发现铜主要分布在细胞内的溶酶体中,少量颗粒吸附在滑面内质网上;这些铜颗粒通过积聚,在细胞顶端逐渐增多,从而出现外排现象。中肠细胞内也分布有铜颗粒,细胞质出现空泡化,线粒体内嵴部分解体,并有铜颗粒附着在线粒体上。在触角腺中分布有少量的铜颗粒,触角腺细胞内的铜颗粒主要分布于溶酶体中,内质网出现弯曲。原子吸收光谱分析结果表明,肝胰脏中铜的含量最高,达到546.39 mg.kg-1,肝胰脏有储藏铜元素的功能。     

日本沼虾幼虾对碱度和pH的适应性

采用急性毒性实验法,研究日本沼虾（Macrobrachium nipponense）对碱度和DH的适应能力,探讨内陆盐碱水域养殖的可能性。结果表明,pH对幼虾的24、48、72、96h半数致死值分别为10．13、9．72、9．67和9．51,安全与适应范围分别为5．26～8．67、5．10～8．84。碱度对幼虾的24h半数有效浓度为17．96mmol／L,95％置信限14．60—22．53mmol／L;24、48、96h半数致死浓度分别为48．95、45．15和44．96mmol／L,95％置信限分别为45．72～50．60、39．46～51．67及34．34～55．38mmol／L,安全碱度11．52mmol／L。结果表明,在pH≤9．0、碱度≤20．0mmol／L的内陆盐碱水域,可以养殖日本沼虾。     

亚硝酸钠和多聚磷酸钠对日本沼虾的毒性研究

针对目前危害虾类养殖的水污染中比较严重的亚硝酸盐和多聚磷酸盐污染,主要研究亚硝酸钠和多聚磷酸钠对日本沼虾的毒性影响。通过实验得出：亚硝酸钠对日本沼虾24、48、96h的半致死浓度分别为46、26、13．33mg／L,多聚磷酸钠对日本沼虾24、48、96h的半致死浓度分别为l233．33、1180、1080mg／L。实验表明,活性氧是随着亚硝酸钠和多聚磷酸钠的浓度增加而增加,而超氧化物歧化酶的酶活力则是随着亚硝酸钠和多聚磷酸钠的浓度增加先稍微增加,后又下降,这表明虾的免疫系统对低浓度的毒物有一定的抵抗能力,但随着毒物浓度增大,虾的免疫系统会受到抑制,虾的免疫功能下降,抗病力降低,导致染病死亡。     

Diversification and Demography of the Oriental Garden Lizard (Calotes versicolor) on Hainan Island and the Adjacent Mainland

The Oriental garden lizard (Calotes versicolor) is one of the few non-gekkonid lizards that are geographically widespread in the tropics. We investigated its population dynamics on Hainan Island and the adjacent mainland of China and Vietnam, focusing on the impact of cyclic upheaval and submergence of land bridges during the Pleistocene. Our Bayesian phylogenetic analysis reveals two mitochondrial lineages, A and B, which are estimated to have coalesced about 0.26 million years ago (95% credibility interval: 0.05–0.61 million years ago). Lineage A contains individuals mainly from central and southern Wuzhi Mountain on Hainan Island, whereas lineage B mainly comprises individuals from other sites on the island plus the adjacent mainland. The estimated coalescence times within lineages A (0.05 million years ago) and B (0.13 million years ago) fall within a period of cyclical land-bridge formation and disappearance in the Pleistocene. A spatial analysis of molecular variance identified two distinct population groupings: I, primarily containing lineage A, and II, mainly consisting of lineage B. However, haplotypes from lineages A and B occur sympatrically, suggesting that gene flow is ongoing. Neither Wuzhi Mountain nor Qiongzhou Strait and Gulf of Tonkin act as barriers to gene flow among C. versicolor populations. Analyses of the data using mismatch distributions and extended Bayesian skyline plots provide evidence of a relatively stable population size through time for Group I, and moderate population expansions and contractions during the end of the Pleistocene for Group II. We conclude that the phylogeographical patterns of C. versicolor are the combined product of Pleistocene sea-level oscillations and nonphysical barriers to gene flow.     

青虾“软壳综合症”病原及其特性

从患软壳综合症的濒死青虾体内分离到一株细菌QXL0711B, 经人工感染试验, 其对青虾的半数致死浓度(LC50)为1.47×106 CFU/mL, 具有较强毒力。API 32E系统鉴定及16S rRNA序列分析, 该病原菌为维罗纳气单胞菌温和生物变种(Aeromonas veronii biovar sobria, 登录号: FJ808727)。其系统发育分析表明, 菌株QXL0711B与维罗纳气单胞菌(登录号: X71120)和维罗纳气单胞菌温和生物变种(登录号: AY987729)的亲缘关系最近,     

Genetic Diversity and Population Structure of the Oriental Garden Lizard,Calotes versicolor Daudin, 1802(Squamata: Agamidae) along the Mekong River in Thailand and Lao PDR

Calotes versicolor Daudin,1802,is geographically widespread along the Mekong River basin.The Mekong River is play important role as a significant natural barrier to several terrestrial animals living on different sides.This study aims to analyze the genetic diversity and population structure of C.versicolor populations collected from different sides of Mekong River using mitochondrial cytochrome c oxidase subunit 1(CO1)sequences.We obtained sequences of 200 individuals from 18 sampling localities from left and right sides of the Mekong River in Lao PDR and Thailand respectively.Overall,91 haplotypes were detected,which reflect high levels of genetic diversity in this species at the study areas.Haplotype network and phylogenetic analyses revealed that there were six major lineages(lineage C–lineage H)of C.versicolor populations within the Mekong River,whereas lineages A and B have previously been found from China and Vietnam.The genetic distance among C.versicolor was significantly related to spatial distance,however,the Mekong River had no significant effect on genetic distance.Our findings,together with previous studies,suggests that C.versicolor in Asia is a species complex with other cryptic lineages being likely but there is a need for further exploration.Thus,comprehensive genetic,biological and ecological studies of C.versicolor should be conducted throughout its entire distribution range.     

日本沼虾卵黄蛋白原合成部位的初步研究

十足目卵巢中卵黄的来源,在过去的几十年研究中一直存在争议,内源性合成和外源性合成均有报道。以雌性日本沼虾为实验材料,根据外形观察和组织学研究确定其发育阶段,可以分为:卵原细胞增殖期;卵黄发生前期;初级卵黄发生期;次级卵黄发生期;成熟期和抱卵期(消退期)。从处于不同发育期的卵巢和肝胰腺中提取总RNA,用RT-PCR方法探讨不同发育期的日本沼虾卵巢和肝胰腺卵黄蛋白原mRNA表达,确定是否有卵黄蛋白原的合成功能。检测结果可以初步判定日本沼虾卵巢和肝胰腺都具有卵黄蛋白原mRNA表达功能,都是卵黄蛋白原的合成部位,其合成的量与沼虾卵巢的发育阶段相关。