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.     

Genetic variation of wild and cultured populations of the Kuruma prawn Marsupenaeus japonicus (Bate 1888) using microsatellites

The microsatellite DNA technique was used to detect the genetic variations between wild and cultured populations of Kuruma prawn Marsupenaeus japonicus Bate 1888. All the six microsatellite loci screened in this study showed high polymorphism for their PIC (0.6701–0.8989), which was much more than the standard value of 0.5. A total of 73 alleles were observed over six loci from 93 shrimps. The mean number of allele locus ranged from 9.83 (cultured) to 11.83 (wild). The number of effective alleles varied from 6.86 (cultured) to 8.58 (wild). The average of observed heterozygosity (H_o) of populations varied from 0.6935 (cultured) to 0.7370 (wild), and that of expected heterozygosity (He) was 0.8169 (wild) and 0.8209 (cultured). Tests of Hardy–Weinberg showed that these loci deviated significantly or highly significantly in one or both populations. Compared with the wild population, the cultured population showed little reduction in genetic variation. The total number of alleles (71, 59) was not significantly (P=0.296) different between wild and cultured populations. The paired‐samples t test of observed heterozygosity and expected heterozygosity implied that there was no significant difference (P=0.572 and 0.891 respectively) between wild and cultured populations. However, some rare allele loss might have occurred in the cultured population. A total of 14 unique alleles were found in the wild population, but only two unique alleles were observed in the cultured population. Therefore, there is a need to monitor genetic variability of cultured population, and to improve the hatchery program for the conservation of wild Kuruma prawn resources.     

Microsatellite and mitochondrial DNA analyses reveal no genetic difference between two pufferfish species torafugu <Emphasis Type="Italic">Takifugu rubripes</Emphasis> and karasu <Emphasis Type="Italic">T. chinensis</Emphasis>

It has been reported that the nucleotide sequences of the mitochondrial and nuclear genes of Takifugu pufferfish torafugu T. rubripes and karasu T. chinensis show 99–100% sequence identity, indicating a very close relationship between the two species. To further investigate this genetic relationship, we compared genetic variation at four microsatellite loci and at the mitochondrial control region (CR) (561 bp) between groups of T. rubripes caught at two locations [TrG, caught in the Genkai Sea off Tsushima Island in 2003 (n = 50); TrS, caught in the Suwo Sea off Kita-Kyushu in 2008 (n = 50)] and T. chinensis caught at one location (TcK, caught off the east coast of Korea in 2004; n = 50). Analyses using microsatellite loci showed that genetic diversity index values of the TrG, TrS and TcK groups were 0.9505, 0.9350 and 0.9335, respectively, while values of genetic distance and genetic differentiation between TrG and TcK (0.0543 and 0.0189, respectively) were smaller than those between TrG and TrS (0.0857 and 0.0194, respectively). Analyses using CR for the same specimens showed that genetic distances were consistent with those obtained using microsatellite loci. These results, together with our previous observations, suggest that T. rubripes and T. chinensis are very closely related and possibly can be regarded as the same species.     

棘头梅童鱼七个野生群体遗传多样性的微卫星分析

为研究中国沿海地区棘头梅童鱼群体的遗传多样性,利用微卫星标记技术,采用9对微卫星引物对中国连云港(LYG)、大丰(DF)、崇明(CM)、舟山(ZS)、温州(WZ)、宁德(ND)、厦门(XM)棘头梅童鱼7个野生群体的遗传多样性和遗传结构进行了分析。结果显示,实验检测到63个等位基因,各位点等位基因数为3~13个,有效等位基因数为1.7510~8.0317;各位点的观测杂合度(Ho)为0.3596~0.7854,期望杂合度(He)为0.4300~0.8780;多态信息含量(PIC)为0.3604~0.8631,其中有2个位点表现为中度多态(0.25PIC0.5),7个位点表现为高度多态(PIC0.5),具有较丰富的遗传多样性水平。Hardy-Weinber平衡分析显示,7个群体的大部分位点未偏离平衡。基于群体间Nei氏标准遗传距离构建的7个野生群体UPGMA系统进化树结果显示,ND和WZ群体遗传关系最近,ZS和WZ群体遗传关系最远,WZ和ND聚为一支,但总体上没有显著的遗传分化。     

Genetic Diversity and Population Structure in the Heavily Exploited Korean Rockfish,Sebastes schlegeli,in Korea

The Korean rockfish, Sebastes schlegeli, is a valuable and intensively exploited species in Korea. We discuss the genetic diversity and genetic structure of four Korean rockfish populations using eight microsatellite loci. In total, 161 different alleles from 138 individuals were observed. Average allele number per locus ranged from 2.5 to 23 and allelic richness varied from 13.38 to 14.63 within a population. Despite a long history of stocking practices, we found very high levels of polymorphism (mean heterozygosity = 0.810), which is comparable to other congeneric species. No significant difference in genetic diversity and molecular genetic variance (F_ST and R_ST) was observed among four local samples (P > 0.1). Little indication of contemporary inbreeding (F_IS= 0.051) or population structure (K = 1) was detected. This absence of differentiation may reflect high levels of gene flow along the coast of Korea. Our study demonstrates that rockfish in Korea should be managed as a single unit. Currently, the species does not appear to be genetically threatened, but the potential for a rapid loss of genetic diversity remains. This information on the genetic characteristics of Korean rockfish populations has important implications for fisheries management and conservation efforts, and will aid in the sustainable exploitation of the fishing resources and the preservation of biodiversity.     

Genetic diversity and differentiation of sea trout (<Emphasis Type="Italic">Salmo trutta</Emphasis>) populations in Lithuanian rivers assessed by microsatellite DNA variation

The genetic diversity and differentiation of sea trout were studied in three river basins in Lithuania: Akmena-Dane, Bartuva, and Nemunas. A total of 282 individuals were genotyped at eight microsatellite loci. A similar level of genetic diversity was found in all of the populations studied: mean allelic richness ranged from 3.64 to 5.03, and average expected heterozygosity ranged from 0.588 to 0.721. Significant genetic divergence was observed among the different river basins as well as between populations within the drainages. All pairwise F _ST values were highly significant, ranging from 0.027 to 0.197. The analysis of molecular variance showed rather weak hierarchical population structuring within the Nemunas basin, which may be explained by extensive gene flow among different river basins or, alternatively, reflect the influence of artificial breeding. Information on genetic diversity and differentiation of the Lithuanian sea trout populations will be useful for future management decisions.     

Population structure and stock identification of chum salmon Oncorhynchus keta from Japan determined by microsatellite DNA variation

Variation at 14 microsatellite loci was surveyed in 26 chum salmon Oncorhynchus keta populations from Japan, one population from West Kamchatka and three populations from North America to determine population structure. Microsatellites were then applied to estimate stock composition of chum salmon in mixed-stock fisheries. The genetic differentiation index (F _st) over all populations and loci was 0.031, with individual locus values ranging from 0.010 to 0.081. Seven regional populations were observed in Japanese chum salmon, with late-run populations from the Pacific Coast of Honshu the most distinct. Japanese populations displayed greater genetic diversity than did those in North America. Transplantation history in some Japanese river populations influenced their present genetic characteristics. Analysis of simulated mixtures from fishery sampling suggested that accurate and precise regional estimates of stock composition should be produced when the microsatellites were used to estimate stock compositions. Stock compositions for a 2005 sample of maturing, migrating chum salmon off the north-west coast of Hokkaido near the border of the Sea of Japan and the Sea of Okhotsk indicated that this region may be a migration corridor for Hokkaido populations from the Sea of Japan coast. Microsatellites have the ability to provide fine-scale resolution of stock composition in Japanese coastal fisheries.     

Genetic diversity of cultured and wild populations of the giant freshwater prawn Macrobrachium rosenbergii (de Man, 1879) based on microsatellite analysis

Freshwater prawn (Macrobrachium rosenbergii) culture in the Western Hemisphere is primarily, if not entirely, derived from 36 individual prawns originally introduced to Hawaii from Malaysia in 1965 and 1966. Little information is available regarding genetic variation within and among cultured prawn stocks worldwide. The goal of the current study was to characterize genetic diversity in various prawn populations with emphasis on those cultured in North America. Five microsatellite loci were screened to estimate genetic diversity in two wild (Myanmar and India‐wild) and seven cultured (Hawaii‐1, Hawaii‐2, India‐cultured, Israel, Kentucky, Mississippi and Texas) populations. Average allelic richness ranged from 3.96 (Israel) to 20.45 (Myanmar). Average expected heterozygosity ranged from 0.580 (Israel) to 0.935 (Myanmar). Many of the cultured populations exhibited reduced genetic diversity when compared with the Myanmar and the India‐cultured populations. Significant deficiency in heterozygotes was detected in the India‐cultured, Mississippi and Kentucky populations (overall F_is estimated of 0.053, 0.067 and 0.108 respectively) reflecting moderate levels of inbreeding. Overall estimate of fixation index (F_st = 0.1569) revealed moderately high levels of differentiation among the populations. Outcome of this study provide a baseline assessment of genetic diversity in some available strains that will be useful for the development of breeding programmes.     

基于SSR-seq技术评估中间球海胆多代选育群体和普通养殖群体的遗传多样性

为明确不同选育群体中间球海胆的遗传多样性和遗传结构,利用SSR-seq技术和15个微卫星位点,对1个家系选育群体(FP)、1个群体选育群体(IP)和1个未经选育的普通养殖群体(CP)的遗传多样性及遗传结构进行了分析。结果显示,15个微卫星位点共检测出112个等位基因,FP、IP、CP 3个群体的平均观测等位基因数(N_a)分别为5.077、5.133和6.133个,平均有效等位基因(N_e)分别为2.816、2.873和3.638个,平均观测杂合度(H_o)分别为0.522、0.441和0.501,平均期望杂合度(H_e)分别为0.595、0.599和0.667,平均多态性信息含量(PIC)分别为0.546、0.543和0.623。家系选育群体(FP) H_e与H_o的差值(0.073)低于IP (0.158)和CP (0.166),平均固定指数(F)(0.115)低于IP (0.248)和CP (0.246)。3个群体间遗传分化系数(F_st)介...     

Development of 25 Novel Microsatellite Loci and Genetic Variation Analysis in Breeding Populations of the Pearl Oyster,Pinctada fucata

In this study, we screened 103,050 gene sequences of Pinctada fucata and identified 1450 simple sequence repeats (SSRs) distributing in 1355 sequences, in which 97 have sufficient flanking sequences for primer design, and 25 novel microsatellite loci were stable and polymorphic with N_a which varied from 2 to 6. Six microsatellite loci were found in the open reading frame of their corresponding sequences. With the 25 SSR loci, five breeding populations and a wild population, a total of 240 individuals from wild population (W), three consecutive selected populations (G₁, G_2, and G₃), and two backcross populations (D_o, D_r) (40 individuals per population) were assayed. In these populations, we detected 108 alleles of 111 bp to 362 bp. Each population had mean H_o of 0.5718–0.7366 and mean H_e of 0.5830–0.6954. Except the F_st = 0.0697 (W vs. G₃), pairwise F_st values ranged from 0.0131 to 0.0473, the genetic identity coefficients varied from 0.8688 to 0.9663, and genetic distance ranged from 0.0343 to 0.1307, all of these suggested that the breeding populations had genetic differentiation but at a low level generally. Besides, all the genetic index values, backcross populations (D_o, D_r) were higher than corresponding G₃ values, indicating that backcross could make the offspring incline to the recurrent parents.     

Genetic diversity and differentiation of Nile tilapia (Oreochromis niloticus) revealed by DNA microsatellites

To assess the genetic diversity of Nile tilapia Oreochromis niloticus, a total of 250 fish from five Egyptian populations were genotyped using six microsatellite markers. Heterozygosity and Wright's F‐statistics (F_IS, F_ST, and F_IT) were calculated to determine the genetic variation within and between these populations. Observed heterozygosities were in the range of 0.4 (Burullus) to 0.96 (Qena), with F_IS values ranging from 0.082 to 0.282. The mean F_ST showed that approximately 96.5% of the genetic variation was within‐population and 3.5% was among populations. Standard genetic distances were used to classify the five populations into two major groups. The deeper lotic river Nile populations of Assuit and Cairo formed one group and the shallow less lotic Delta lakes populations of Manzalla and Burullus formed the second group, with the upstream Nile Qena population being an outgroup. The findings from the current study help understanding of the broad‐scale population structuring of the Nile tilapia (O. niloticus) allowing the population groupings identified to act as potential sources of genetic variation. These populations could be included in future Marker‐Assisted‐Selection programs for economically desired production traits.     

基于微卫星评估草鱼放流亲本对野生群体遗传多样性的影响

利用筛选的13对草鱼多态性微卫星标记,开展了2011至2015年长江中游草鱼亲本增殖放流对野生群体遗传多样性的影响评估。通过对各位点的遗传多样性分析,13个微卫星位点的多态信息含量为0.8622(0.657~0.950),基因多样度为0.8555(0.675~0.936)。15个群体的有效等位基因数为7.4503~10.1536,等位基因丰度为11.483~15.204,说明15个草鱼群体的遗传多样性水平总体较高。遗传分化指数分析表明,群体间不存在显著遗传分化(FST5%)。通过贝叶斯聚类分析和主成分分析可将草鱼群体分为4个组群,根据分组结果以及来源划分分别对草鱼群体进行AMOVA分析,发现遗传变异大部分来自于群体内个体间,组间及组内群体间的分化水平较低(FCT5%,FSC5%),与FST分析结果一致。研究表明,当前草鱼亲本增殖放流模式对野生群体遗传结构影响不明显。     

Genetic diversity in the Indian population of Penaeus monodon (Fabricius, 1798) as revealed by mtDNA sequence analysis

Genetic diversity in the Indian population of the tiger shrimp Penaeus monodon was determined by using partial sequence data of the mitochondrial DNA (mtDNA) control region (D‐loop) and the 16S rRNA gene. Eight populations from different geographical locations (Mumbai, Kochi, Mangalore, Kakinada, Gopalpur, Chilika, Paradeep and Andaman) were collected and analysed. The amplified polymerase chain reaction products of size 577 bp for the control region and 472 bp for 16S rRNA were sequenced in both directions and data were analysed through clustal , arlequin , mega and phylip . A significant genetic structure was found among the Indian populations. The mtDNA control region proved to be a powerful marker in comparison with 16S rRNA for population studies of this species. The east coast population was more genetically diverse than the west coast. The Andaman population was found to be the most diverse among all the populations. The populations on the west coast were found to be genetically more structured and differentiated than the populations on the east coast. The results revealed a high level of genetic diversity and also distinct population structuring of P. monodon, suggesting great possibilities of genetic improvement for growth and other economic traits.     

Wild stock structure of <Emphasis Type="Italic">Girella punctata</Emphasis> in Japan revealed shallow genetic differentiation but subtle substructure in subsidiary distributions

Twelve populations of Girella punctata, from widespread locations of the species’ range in Japan and Korea, were screened for sequence variability within the mitochondrial DNA (mtDNA) control region (n = 128) and at five polymorphic microsatellite loci (n = 547) to determine the genetic structure maintaining population integrity. mtDNA variability of 132 variable sites within a 334-bp region reveals shallow genetic differentiation across populations. The weak differentiation of G. punctata was partly supported by the screening of five polymorphic microsatellite loci. However, hierarchical analysis of molecular variance and principal component analysis on the basis of allele frequencies in microsatellite loci extracted a subtle substructure in a subsidiary population and in near-subsidiary populations in the semi-enclosed Seto Inland Sea.     

用微卫星DNA技术对中国对虾人工选育群体遗传多样性的研究

利用微卫星技术对中国对虾人工选育群体第1代和第6代群体的遗传多样性进行了分析。对10个微卫星位点进行了扩增,共产生74个等位基因,每个位点产生的等位基因数从3到13不等。在两个群体中,所观察到的等位基因数都比有效等位基因数多。多态信息含量PIC值0．5567～0．8877,说明这10个微卫星位点在中国对虾中具有较高的信息含量。两个群体的平均杂合度分别为0．6400(CP1)、0．6300(CP6),并通过计算基因型的P值,确定了对Hardy-Weinberg平衡的偏离情况。对Fis值的计算表明两个群体内共有5个微卫星位点存在杂合度观察值过剩的现象。两个群体的Shannon多样性指数分别为1．6830、1．7382,整个选育群体(两个群体作为一个群体)的遗传多样性指数为1．7742。从遗传多样性所占的比例来看,96．415％的遗传变异是来自群体内,只有3．585％的遗传变异是来自群体间。两个群体间的相似性系数高达0．9187,彼此间的遗传距离仅为0．0848,体现出人工选育群体的遗传分化程度较低。结果均说明第6代群体还有较大的选育潜力,可以继续保持遗传效应,最终保证选种育种工作的成功。     

Genetic diversity and structure of domesticated strains of red sea bream,Pagrus major,inferred from microsatellite DNA markers

The genetic diversity and structure of nine domesticated strains of red sea bream used in a private hatchery were studied and compared to a wild population. A total of 313 individuals were genotyped at eight microsatellite loci. Average number of alleles per locus ranged from 5.5 to 9.4 in domesticated strains, but that of the wild population was 28.4. Heterozygosity of domesticated strains (ranged 0.697–0.804) was also lower compared to the wild population (0.952). Estimated Ne also decreased in all domesticated strains (ranged 10.3–126.0) compared to the wild population (1422.5). The UPGMA tree and 3‐D FCA showed that there were two main clusters containing domesticated strains, and the wild population was at the middle of both of the domesticated clusters. The STRUCTURE analysis also supported the phylogenetic analysis, and revealed three sub‐clusters in the domesticated strains. Pairwise F_ST revealed that all domesticated strains were statistically different from the wild population, and also the differentiation between domesticated strains was all statistically significant. Information on genetic diversity and structure of domesticated strains of red sea bream obtained in this study will be useful for future broodstock management and selective breeding programmes.     

Genetic Diversity and Differentiation in Broodstocks of the Endangered Chinese Sucker,Myxocyprinus asiaticus,Using Microsatellite Markers

Chinese Sucker, Myxocyprinus asiaticus, is a valuable and endangered freshwater species in China. We investigated genetic diversity and differentiation of four M. asiaticus broodstock populations using 10 microsatellite loci. Three populations showed weak genetic differentiation among them. The other and the most distant population exhibited the lowest genetic diversity. It was inferred that wild M. asiaticus ancestry in the Yangtze River had sufficient gene flow among localities, while under the situation of population decline and fragmentation, restocking programs without genetic management might aggravate loss of genetic diversity and increase genetic differentiation. Maintenance of population connectivity among localities requires real attention in future restocking efforts.     

Genetic and morphological divergences between wild and captive‐bred populations of Salmo trutta abanticus

Salmo trutta abanticus is a non‐anadromous trout species native to Lake Abant and Seven Lakes in Turkey. A restocking programme by captive breeding was initiated in 1999 to support S. trutta abanticus population. Reared 2‐year‐old juveniles from randomly caught wild parental individuals in Maçka breeding farm were introduced into Lake Abant. We aimed to compare genetic and morphological divergences between wild‐ and captive‐bred populations using seven microsatellite loci and geometric morphometric measurements. A significant genetic and morphological divergences were detected between all population in F_st and canonical variate analysis based on geometric morphometric with 10 homolog landmark. Eighty‐six microsatellites alleles were recorded across loci. Number of private alleles, observed alleles and observed heterozygosity are statistically significant higher in Maçka captive‐bred population than Lake Abant and Seven Lakes populations. Of 42 tests, three departures from Hardy–Weinberg equilibrium were detected in all populations after Bonferroni correction. Two pairs of loci (Ssa85 – Str73 and Str73‐Str543) in Maçka, one pairs of loci (Ssa85‐Str73) in Abant and two pairs of loci (Ssa85‐Str60 and Str73‐Str543) in Seven Lakes populations show linkage disequilibrium. Population structure analysed with Structure software showed three genetic groups (?K = 3) in our studied populations. Relatedness estimates show higher mean relatedness values (r = 0.220 ± 0.230) for Maçka captive‐breed population than wild populations of Abant Lake and Seven Lakes (r = 0.140 ± 0.210 and r = 0.170 ± 0.200 respectively).     

Genetic divergence in Indian populations of M. rosenbergii using microsatellite markers

Macrobrachium rosenbergii, known as the giant freshwater prawn or Malaysian prawn, is the sixth largest aquaculture species in Asia. Knowledge of genetic diversity of M. rosenbergii is important to support management and conservation programmes, which will subsequently help in sustainable production of this economically important species. This study aimed to analyse the genetic diversity and population structure of five M. rosenbergii populations using 11 microsatellite loci. In analysing 240 samples, the number of alleles, observed heterozygosity (H_O) and expected heterozygosity (H_E) ranged from 3 to 20, from 0.250 to 0.978 and from 0.556 to 0.944 respectively. The five stocks of M. rosenbergii displayed high level of genetic diversity. Both the F_ST and amova analyses showed that there was significant genetic differentiation among all populations. The UPGMA dendrogram based on Nei's genetic distance matrix revealed that the Narmada and Mahi populations were in one cluster and Mahanadi and Subarnarekha populations in another single major branch, whereas the Kerala population clearly showed a separate cluster. This information on genetic variation will be useful for genetic improvement and conservation of Indian populations of giant freshwater prawn M. rosenbergii.