Genetic variation in farmed populations of the gilthead sea bream Sparus aurata in Greece using microsatellite DNA markers

Genetic variation in seven reared stocks of gilthead sea bream Sparus aurata, originating from Greek commercial farms, was assessed using five polymorphic microsatellite markers and was compared with that of two natural populations from the Ionian and the Adriatic Seas. The total number of alleles per marker ranged from 11 to 19 alleles, and hatchery samples showed the same levels of observed heterozygosity with samples from the wild but substantially smaller allelic diversity and expected heterozygosity. The global genetic differentiation for the cultivated samples was significant as indicated by F_st analysis, which might indicate random genetic drift and inbreeding events operating in the hatcheries. On the contrary, no significant difference was found between the two wild populations. Population pairwise tests between farmed and wild stocks were also significant, with the exception of one hatchery sample, the Central Greece 1, which was not significantly different from the two wild samples perhaps due to its recent use in aquaculture from wild‐caught animals. The UPGMA tree topology grouped the wild samples together with the Central Greece 1 stock, and showed a clear division between wild and farmed sample sets for the six remaining hatchery samples. Knowledge of the genetic variation in S. aurata cultured populations compared with that in the wild ones is essential for setting up appropriate guidelines for the proper monitoring and management of the stocks either under traditional practices or for the implementation of selective breeding programmes.     

Residual genetic variability in domesticated populations of the Pacific blue shrimp (Litopenaeus stylirostris) of New Caledonia,French Polynesia and Hawaii and some management recommendations

The Latin American shrimp Litopenaeus stylirostris was introduced in three different Pacific islands (Tahiti, New Caledonia via Tahiti, and Hawaii) and hatchery-propagated for 7–25 generations to develop shrimp farming based on these domesticated stocks. Three microsatellite markers have been used in an attempt to assess the genetic bases of the populations available to start a selective breeding program. The comparison of eight hatchery stocks (five New Caledonian, two Hawaiian and one Tahitian stocks) and one wild Ecuadorian population showed a much lower variability in the domesticated stocks than in the wild population, especially in New Caledonia and Tahiti (2–3.7 vs. 14–27 alleles per locus; 20–60% vs. 90% expected heterozygosity). The Tahitian and the New Caledonian stocks share the same alleles, suggesting that the loss of alleles occurred during the common past of these populations. On the contrary, New Caledonian and Hawaiian populations share only one common allele at the three loci studied. Although the low genetic variability and the resulting inbreeding of the New Caledonian stocks do not seem to affect their present performance, the results of this study demonstrate the usefulness of the introduction of new stocks in order to increase the potential responses to new controlled or uncontrolled selective pressures. The introduction in New Caledonia of the Hawaiian domesticated stocks, which would provide the local shrimp industry with 40% of the allelic diversity of the species, is advised and preferred to the one of wild animals in order to take advantage (i) of the spontaneous selection which occurred during domestication and (ii) of their favourable sanitary “specific pathogen free” status (no presence of four viruses: WSV, YHV, IHHNV, TSV) which limits the risk of introduction of pathogens.     

Genetic Introgression Between Arctic Charr (Salvelinus Alpinus) and Brook Trout (Salvelinus Fontinalis) in Bavarian Hatchery Stocks Inferred from Nuclear and Mitochondrial DNA Markers

Nuclear insulin-like growth factor 2 gene (IGF-2), growth hormone 1 gene (GH-1) and internal transcribed spacer 1 (ITS-1) of the ribosomal DNA as well as the mitochondrial NADH-3 and NADH-4 dehydrogenase genes (ND-3/4) exhibited species-specific restriction fragment patterns and three microsatellite loci (Sfo18, Ssa85 and Ssa197) had non-overlapping allele size ranges in Arctic charr and brook trout and were used as diagnostic markers for testing genetic purity of hatchery stocks and wild populations of Arctic charr and brook trout in Bavaria, Germany. Screening of four wild populations (three in Arctic charr and one in brook trout) revealed only a single hybrid (back-cross to brook trout) individual in L. Starnberg. In contrast, in three (out of five) hatchery stocks of Arctic charr and in both hatchery stocks of brook trout hybrids were detected with the frequency from 3 to 100%. Three hatchery stocks (SS2, SA and BS1) represent a hybrid swarm because they contained a very high proportion of hybrids (from 83 to 100%) and most or all hybrid individuals had alien alleles at only one or a few of six unlinked diagnostic loci, indicating that post-F₁ hybrids represent the majority of individuals in these stocks and introgression has taken place. Release or escape of introgressed individuals from hatcheries into natural water bodies should be avoided in order to protect the biological diversity and genetic integrity of native fish populations.     

Microsatellite genetic variation in wild and hatchery populations of the sea cucumber (Apostichopus japonicus Selenka) from northern China

Farming of the sea cucumber Apostichopus japonicus (Selenka) started 20 years ago and is still in rapid expansion in China. In order to assess the genetic status of both wild and cultivated stocks of this species, we used eight microsatellite markers to estimate the level of genetic diversity within five hatchery stocks and two wild populations of A. japonicus, and compared the degree of genetic differentiation between them. High levels of polymorphism were observed over all loci. The mean alleles and expected heterozygosities over the seven stocks were 10.4–12.3 and 0.735–0.783 respectively. The results of the microsatellite survey provide no evidence to show that hatchery practice of the sea cucumber in China to date has significantly affected the genetic variability of the cultured stocks. Significant differentiation was found between most pairs of the hatchery stocks and wild populations (F_st range: 0.008–0.036), and no obvious difference was detected between the wild populations (F_st=0.008). The information on the genetic variation and differentiation obtained in this study can be applied for future genetic monitoring of A. japonicus aquaculture stocks and will be useful for future genetic improvement by selective breeding, and for designing suitable management guidelines for these genetic materials.     

Genetic diversity and relatedness estimates for captive barramundi (Lates calcarifer,Bloch) broodstock informs efforts to form a base population for selective breeding

Aquaculture of barramundi or Asian seabass (Lates calcarifer) is growing in both Australia and Southeast Asia and there is substantial interest to improve production efficiency through selective breeding. The establishment of a large and genetically diverse base population is a prerequisite for a sustainable and long‐term productive breeding program. Before selective breeding programs can begin for Australian barramundi it is important to assess the overall genetic diversity of current captive broodstock populations. To address this question, 407 captive barramundi broodstock from eight separate Australian broodstock populations were genotyped using 16 polymorphic microsatellite DNA markers. A Bayesian STRUCTURE analysis indicated that captive Australian broodstock are broadly divided into two genetic stocks. Multivariate analysis between broodstock individuals and pairwise F_ST between broodstock populations also supported the existence of two stocks. Comparisons with data obtained from natural stocks suggested that hatchery individuals were either sourced from the two stocks or represented an admixture between them. Genetic diversity was low within each broodstock population (allelic richness ranged from 2.67 to 3.42 and heterozygosity ranged from 0.453 to 0.537) and relatedness estimates within hatcheries were generally low (average r was equal to 0.141). We recommend sourcing captive individuals according to high levels of neutral genetic diversity and low levels of relatedness for the establishment of a base population. We also make recommendations about including genetically diverse wild individuals.     

近交及其对水产养殖的影响

因大多数野生物种的捕捞已接近甚至超过了可开发的限度,因此必须大量增加水产品的养殖以满足人们对水产品的需要。目前,许多水产养殖种类的苗种来源于野生亲体或野生种苗,但是当野生种群被过度捕捞或数量大量减少时,越来越多的苗种将来源于人工蓄养的亲体群体。     

基于SSR标记的长江下游原良种场鳙亲本和后备亲本种质资源现状分析

分析长江下游原良种场鳙(Aristichthys nobilis)亲本和后备亲本的遗传多样性和遗传结构,评估鳙育种群体的种质资源现状,可为鳙的种质资源管理和活体资源库建设提供科学的参考依据。本研究利用已发表的10对SSR荧光引物结合毛细管电泳技术,对7个亲本群体和1个后备亲本群体共638份DNA进行基因分型。主要应用GenAlEx 6.501、Cervus 3.0和Structure 2.3.4等软件进行遗传多样性参数计算和群体遗传结构分析。结果表明,8个群体总体遗传多样性水平较高,但存在一定程度的近亲繁殖风险。其平均等位基因数(N_a)为14.83±1.45,观测杂合度(H_o)和期望杂合度(H_e)分别为0.76±0.05、0.82±0.02,Shannon's信息指数(I)为2.08±0.09,近交系数(F)为0.08±0.05。各亲本群体之间遗传多样性水平存在差异,后备亲本遗传多样性水平最高。鳙的贝叶斯群体遗传结构分析图(Structure)与主坐标分析(PCoA)结果具有一致性,将所有样本划分为两个类群,大部分亲本群体之间并无明显的遗传分化,而后备亲本和亲本之间则表现明显的遗传分化。分子方差分析(AMOVA)显示,11%的遗传变异来自群体间,群体间的遗传分化处于中等水平,而89%的遗传变异来自群体内。本研究探究了鳙亲本和后备亲本的遗传差异性,对野生原种引进和保护提供了建议,以期为构建健康负责的增殖放流种苗繁育体系提供理论支持和数据参考。     

Use of Microsatellite DNA Profiling to Identify Japanese Flounder,Paralichthys olivaceus of Hatchery Origin

Japanese flounder, Paralichthys olivaceus, is an economically important marine fish species in Asia. A suite of 18 microsatellite markers chosen from published genetic linkage maps was used to carry out parentage assignments of 188 hatchery‐reared juveniles from a small number of breeders. The probabilities of exclusion for the 18 microsatellite markers were 0.604–0.913, and the effectiveness of combined probability of exclusion reached 100% when using the eight microsatellite markers with higher Excl 1 probabilities. The cultured and wild stocks (WSs) were differentiated in a release‐recapture population based on these markers. Of the 321 recaptured offspring, 28.34% were assigned to their parental pairs in our broodstock, whereas the remaining offspring could not be traced back to a possible sire or dam. Significant reduction in genetic diversity of the cultured stock (CS) had not been found compared with that of the WS. The results suggest that CSs released into the wild will not adversely affect the genetic structure of natural populations. Our results demonstrate that these markers provide an efficient tool for parentage assignments and genetic analysis of Japanese flounder.     

AFLP and microsatellites as genetic tags to identify cultured gilthead seabream escapees: data from a simulated floating cage breaking event

Genetic discrimination using DNA fingerprinting is rapidly developing for cultured stock and wild fish populations. Microsatellites and AFLPs are being widely used in aquaculture to assign fish or processed fish products, to their claimed origin, paternity or strain. In the present study, 147 AFLP and 4 microsatellite markers were used as genetic tags in gilthead seabream, Sparus auratus. Specimens from two different hatchery broodstocks (one of Atlantic and one of Mediterranean origin) and wild fishes from a natural population were fingerprinted. Putative offspring from these broodstocks were computer-generated, and the confidence in the parentage assignment of their genetic profiles to the hatchery broodstock assessed. The virtual offspring were then mixed with specimens from a natural population to simulate an accidental escape from a floating cage. The risk of false paternity inclusion was evaluated to test the ability to identify either Atlantic or Mediterranean hatchery offspring among wild fish. The method proved to be reliable, and could therefore be used to forecast the impact of fish farm escapees.     

Pedigree analysis of recaptured fish in the stock enhancement program of spotted halibut Verasper variegatus

ABSTRACT:   Spotted halibut Verasper variegatus hatchery juveniles produced in 2002 were genotyped using three microsatellite DNA markers (msDNA) and then released into natural waters. Subsequently, recaptured individuals were examined using msDNA. In order to evaluate the effectiveness of the stock enhancement program, from the genetic point of view, a pairwise F _ST test was implemented to estimate the genetic divergence between the wild captive broodstock, the hatchery offspring and the recaptured samples. The analysis showed significant differentiation between the broodstock and recaptured samples. Pedigree determination using msDNA was used to calculate the effective population size of the recaptured stock, which was found to be very low ( N _e ≈ 8). Equal family survivability was observed between the two recaptured stocks, but not between the released and recaptured stock. The number of identified families was higher and more equalized in the hatchery offspring compared to the recaptured samples, where the number of families declined. This fact was caused by an unequal family survivability just before or just after release. Separately, the number of contributing parents to the hatchery offspring was lower than the broodstock census number. Consequently, these two facts caused the genetic divergence of the recaptured stock from the broodstock.     

Survey of genetic variation at three microsatellite loci in captive populations of endangered Japanese minnow <Emphasis Type="Italic">Aphyocypris chinensis</Emphasis> with implications for reduction of inbreeding

The Japanese population of the cyprinid minnow Aphyocypris chinensis is nearing extinction in the wild. The genetic diversity of three microsatellite loci in five captive populations was investigated, and an effective breeding strategy to reduce inbreeding from pairwise relatedness (R _xy ) between each captive line is discussed. The average number of alleles ranged 2.33–4.67 and the average heterozygosity ranged 0.283–0.602. The pairwise relatedness observed in most combinations showed a significant decrease between the populations. It is suggested that exchange of individuals between different breeding lines should effectively stop inbreeding. Studies show that the effective population size (N _e ) estimated from the number of parental individuals was 8.54 in one captive population, which is insufficient to maintain genetic diversity. It is recommended that more parental individuals should be used, and to exchange fish in a rotating mating mode between institutions participating in captive breeding of A. chinensis.     

驯食配合饲料的大口黑鲈3个选育世代的遗传多样性分析

为了检测驯食配合饲料的大口黑鲈(Micropterus salmoides)3个选育世代群体遗传多样性水平变化,利用微卫星标记技术对驯食配合饲料大口黑鲈选育基础群体(Sp0)和第二、三和四代选育群体(Sp2、Sp3和Sp4)共240尾样品进行检测。结果显示,18个微卫星位点共获得44个等位基因。Sp0、Sp2、Sp3和Sp4的平均观测杂合度(H_o)分别为0.4895、0.4802、0.4579和0.4206,平均期望杂合度(H_e)分别为0.4615、0.4454、0.4621和0.3916,平均多态信息含量(PIC)分别为0.3791、0.3659、0.3764和0.3257。4个群体间的配对比较群体间遗传分化指数(F_(st))值在0.01612～0.16162之间、遗传距离(D_a)在0.0249～0.1434之间。遗传变异来源(AMOVA)分析显示,只有8.38%的变异来自于群体间,其余遗传变异均来自于个体间。研究表明,经连续多代选育之后,易驯食配合饲料的快长大口黑鲈选育群体具有中度遗传多样性,具备选育潜力,可继续进行选育。     

Microsatellite DNA markers of Ezo abalone (Haliotis discus hannai): a preliminary assessment of natural populations sampled from heavily stocked areas

Reproductive success of released animals becomes a growing interest in the context of stock enhancement program and conservation biology. In the case of the Ezo abalone (Pacific abalone; Haliotis discus hannai), limited information is available about the extent of interbreeding between natural and released hatchery populations. This study aimed to develop microsatellite DNA markers from Ezo abalone to address the issue. We developed 38 microsatellite markers that worked well in three full-sib families established from wild caught abalone. Of these, nine markers, with which alleles could be scored easily and accurately, were applied to two population samples derived from coastal areas where intensive releases of hatchery-produced abalone seedlings have been carried out. Multilocus estimates of genetic population differentiation calculated based on the nine markers (F_ST analogue θ and R_ST analogue ρ) rejected a null hypothesis of genetic homogeneity between the populations (θ=0.048, P<0.00001; ρ=0.029, P=0.0051). Although both populations conformed to Hardy Weinberg's equilibrium at almost all of the nine loci, the results of simulation analysis for variance of relatedness coefficient provided evidence of nonrandom mating in each population, possibly caused by the cumulative effect of stocking on the genetic make-up. The population analysis presented here is preliminary; but we consider that the microsatellite markers are potentially an efficient means to examine the reproductive contribution of released abalone to natural resources.     

Genetic comparison of Iranian and Azeri populations of the oriental bream Abramis brama orientalis (Berg) using microsatellites

The aim of this study was to compare, levels of genetic polymorphism between Iranian and Azeri bream (Abramis brama orientalis) populations using five microsatellite loci. The reduced number of alleles in the Iranian populations compared with the Azeri population led us to infer that the genetic variability of the Iranian stocks has been reduced due to inbreeding and genetic drift. Significant population differentiation (F_st) was observed between the Azeri and the Iranian populations, which could be explained by the higher number of alleles in the Azeri population. Significant deviations from Hardy–Weinberg equilibrium were found at more loci in the Iranian populations than the Azeri population. Beyond the hypothesis of null alleles, the heterozygote deficiency may have arisen due to breeding between related individuals in the Iranian populations. This investigation represents the first approach to the knowledge of the genetic variability of Iranian and Azeri bream populations using microsatellite markers, and the reported results could be of interest for management and conservation programmes of this species in Iran.     

Parentage assignment of stocked black sea bream <Emphasis Type="Italic">Acanthopagrus schlegelii</Emphasis> in Hiroshima Bay using microsatellite DNA markers

The genetic contribution of 51 broodstock, comprising 29 females and 22 males, reared at Hiroshima City Marine Products Promotion Center for the production of stocked black sea bream was monitored during two consecutive years using seven microsatellite DNA loci. The high discrimination ability of these markers was reflected in the polymorphic identification content (PIC=0.831), the exclusion probability (Q≈1), and the low probability of identity index (I=3.635⁻¹⁰). The total number of breeders contributing to the mating process was estimated at 32 (62.7%) in 2000 and 30 (58.8%) in 2001. On pedigree reconstruction, 69.3% of the offspring were successfully assigned to a single broodstock pair. Loss of alleles accounted for 16.9% during seed production; nevertheless, 90.9% of males and 69.0% of females participated in the mating process. Based on microsatellite genetic tagging, 58.9% of the fish sampled during the two months after release were identified as hatchery stock, presenting no significant differences from wild conspecifics in either fork length or body weight.     

人工雌核发育鲢近交F2微卫星DNA变异分析

采用17对鲢微卫星引物,以野生鲢、养殖鲢、雄鲤作对照对人工雌核发育鲢近交F2及其亲本进行了微卫星分析。结果表明:人工雌核发育鲢近交F2、养殖鲢和野生鲢群体的平均等位基因数范围为2.1～4.0;平均观测杂合度范围为0.2762～0.9588;期望杂合度范围为0.2774～0.7360;遗传多样性指数范围为0.4500～1.2258,人工雌核鲢近交F2为0.4500,显著低于养殖鲢(1.0273)和野生鲢(1.2258),揭示人工雌核发育鲢近交F2遗传多样性水平较低,纯合度较高。从遗传距离来看,人工雌核发育鲢近交F2与对照群体之间的遗传距离都要大于野生鲢与养殖鲢群体之间遗传距离,表明人工雌核发育鲢近交F2发生了一定程度的遗传分化。     

Identification of differential broodstock contribution affecting genetic variability in hatchery stocks of Atlantic salmon (Salmo salar)

Supportive breeding of Atlantic salmon (Salmo salar) is commonly employed to maintain numbers of fish where the species has become locally endangered. Increasingly, one of the main aims of population management is the preservation of natural genetic diversity. If the stocks employed in supportive breeding exhibit reduced variation they can alter the natural pattern of genetic variation observed in wild populations. In northern Spain, wild adult salmon are caught every year from local rivers and artificially crossed in order to create supportive stocks. The offspring are hatchery reared until the juvenile stage, then released into the same river where their parents were caught. In the current study, our findings demonstrate that although adult broodstock exhibit a pattern of variation similar to the wild populations, variability at microsatellite loci was drastically reduced in the juveniles released into one of three rivers analyzed. The contribution of broodstock to this juvenile stock was examined by pedigree analysis. A restricted number of females contributing to the hatchery stock was identified as the main cause of loss in genetic variation, possibly due to overmaturity of some multi-sea-winter females. We suggest that better monitoring and control of parental contribution will help in solving the problem of loss of genetic diversity in hatchery populations.     

Monitoring of the genetic variability of the hatchery and recaptured fish in the stock enhancement program of the rare species barfin flounder <Emphasis Type="Italic">Verasper moseri</Emphasis>

ABSTRACT:   In relation to the stock enhancement program for barfin flounder, hatchery juveniles produced in 2001 were genotyped using microsatellite DNA markers (msDNA) and then released to natural waters. Subsequently, recaptured individuals, designated as 'tentative recaptured', were examined using msDNA. In order to evaluate the effectiveness of the stock enhancement program, pairwise F _ST and genic differentiation tests were used to estimate the genetic divergence between the wild samples, the hatchery broodstock, and the tentative recaptured samples. Analysis showed significant differentiation among these three groups. Pedigree determination by msDNA was used to establish the origin of the tentative recaptured individuals, in order to elucidate whether they were hatchery produced, 'real recaptured', or wild specimens. Wild individuals were not found. The effective population size of the real recaptured stock was very low ( N _e = 16.6). Equal family survivability was observed between the released and real recaptured stocks, indicating that the genetic variability of the released stock was maintained in the natural environment. Future broodstock management, breeding designs, and family contribution equalization of the offspring to be released will be required in this rare species to avoid unintended genetic differentiation between the wild population and the hatchery broodstock.     

Impacts of hatchery release on genetic structure of rock carp <Emphasis Type="Italic">Procypris rabaudi</Emphasis> in the upper Yangtze River,China

Rock carp Procypris rabaudi is a vulnerable endemic fish in the upper Yangtze River. Hatchery release has been carried out as a major stock enhancement strategy for this species. Ten microsatellite loci were chosen to compare genetic variation between one wild population and two hatchery groups to evaluate the potential impacts of hatchery release on the genetic structure of the wild population. Two different models indicated strong evidence of recent bottlenecks in all groups. The hatchery groups were lower in the mean number of alleles per locus, allelic richness, and allelic diversity compared with the wild population. The 80% membership coefficient indicated that 14% of the wild fish could be assigned as hybrids of wild and hatchery fish. Our results suggested that hatchery release will further reduce the natural genetic diversity in the wild population, change the genetic structure of the rock carp population, and may not benefit restoration of this vulnerable fish species.     

Microsatellite analysis in domesticated and wild Atlantic salmon (Salmo salar L.): allelic diversity and identification of individuals

Samples of wild and domesticated salmon in Norway were genotyped at 12 microsatellite loci to compare allelic variability and investigate the potential of microsatellite markers for identification of individuals. The following loci were amplified: Ssa20, Ssa62NVH, Ssa71NVH, Ssa90NVH, Ssa103NVH, Ssa105NVH, SsaF43; Ssa20.19; Ssa13.37; SsOSL85; Ssa197; Ssa28. All domesticated strain samples displayed reduced variability compared to wild salmon. On average 58% of the allelic richness observed within the four wild stocks were present in the samples taken from domesticated strains. No systematic differences in heterozygosity were observed between samples representing the two groups.

Pairwise genetic distances, as estimated by Fst values and Nei [1978] was 2–8 times higher among domesticated strains than among wild strains. Among the wild stocks, the highest genetic distances were observed between the river Neiden, located in northern Norway, and the other wild stocks located in the southwest of Norway.

Assignment tests indicated that the wild and domesticated salmon could be distinguished with high precision. Less than 4% of domesticated salmon were misassigned as wild salmon, and less than 3% of wild fish were misassigned as domesticated salmon. Fish from individual domesticated strains were identified with similarly high precision. Assignment to wild salmon stocks was less accurate, with the exception of the sample taken from the river Neiden, for which 93% of the individuals were correctly assigned.