Polymorphic microsatellite loci for population studies of the razor clam, <Emphasis Type="Italic">Sinonovacula constricta</Emphasis>

The razor clam (Sinonovacula constricta) is an important aquacultured bivalve in China. The natural populations of this species are decreasing quickly. To facilitate studies on genetic diversity and population structure of wild populations, microsatellites were isolated from a CA enriched genomic library. Eight microsatellite loci were polymorphic in 30 individuals from Chongming in Shanghai, China. The number of alleles per polymorphic locus varied from 6 to 13 and the values of observed heterozygosity and expected heterozygosity ranged from 0.350 to 1.000 and from 0.602 to 0.902, respectively. These microsatellites are being used in studying population differentiation and genetic diversity for effective conservation and management genetic resources of S. constricta.     

Novel polymorphic microsatellites for studying genetic diversity of red Asian arowanas

Asian arowana (Scleropages formosus) is a highly endangered fish species listed in Appendix 1 of CITES since 1980. Fourteen novel polymorphic microsatellites were isolated from a CA-enriched partial genomic DNA library, and were used for studying genetic diversity of 41 red arowanas from the wild population. Surprisingly the average allele number of the 14 polymorphic microsatellites was as high as 12.4/locus. The average observed heterozygosity was 0.78 ranging from 0.51 to 0.95, and the gene diversity was quite high (0.78). All these data suggest that high level of genetic diversity existed in the red Asian arowana population.     

Isolation and characterization of twelve novel microsatellites in yellow catfish, Pelteobagrus fulvidraco

The yellow catfish Pelteobagrus fulvidraco is a freshwater fish species. Due to overfishing and pollution of freshwater ecosystems, the wild stocks of this fish reduced substantially. We isolated and characterized 12 polymorphic microsatellites of this species. The number of alleles at the 12 microsatellite loci ranged from four to eight, with an average of 6.6/locus. The average observed heterozygosity was 0.72, whereas the expected heterozygosity ranged from 0.60 to 0.86 (average: 0.80). All 12 microsatellites conformed to Hardy–Weinberg Equilibrium and were in linkage equilibrium. These 12 novel microsatellites could facilitate studies of genetic diversity and population structure of the yellow catfish to supply necessary information of conservation of the yellow catfish.     

Comparing RADseq and microsatellites for estimating genetic diversity and relatedness — Implications for brown trout conservation

The conservation and management of endangered species requires information on their genetic diversity, relatedness and population structure. The main genetic markers applied for these questions are microsatellites and single nucleotide polymorphisms (SNPs), the latter of which remain the more resource demanding approach in most cases. Here, we compare the performance of two approaches, SNPs obtained by restriction‐site‐associated DNA sequencing (RADseq) and 16 DNA microsatellite loci, for estimating genetic diversity, relatedness and genetic differentiation of three, small, geographically close wild brown trout (Salmo trutta) populations and a regionally used hatchery strain. The genetic differentiation, quantified as F_ST, was similar when measured using 16 microsatellites and 4,876 SNPs. Based on both marker types, each brown trout population represented a distinct gene pool with a low level of interbreeding. Analysis of SNPs identified half‐ and full‐siblings with a higher probability than the analysis based on microsatellites, and SNPs outperformed microsatellites in estimating individual‐level multilocus heterozygosity. Overall, the results indicated that moderately polymorphic microsatellites and SNPs from RADseq agreed on estimates of population genetic structure in moderately diverged, small populations, but RADseq outperformed microsatellites for applications that required individual‐level genotype information, such as quantifying relatedness and individual‐level heterozygosity. The results can be applied to other small populations with low or moderate levels of genetic diversity.     

Isolation and characterization of polymorphic microsatellites from red coral grouper (Plectropomus maculatus)

The red coral grouper, Plectropomus maculatus, is a high value marine food fish species. We isolated the first set of 10 polymorphic microsatellites and examined their polymorphism in two related species: the Malabar grouper Epinephelus malabaricus and the brown‐marbled grouper Epinephelus fuscoguttatus. The average allele number of these 10 microsatellite DNA loci is 9.1/locus with a range of four to 17, whereas the expected heterozygosity ranged from 0.18 to 0.94, averaging 0.76. Five of the 10 markers amplified polymorphic and specific products in E. malabaricus, whereas only four were amplified in E. fuscoguttatus. These microsatellites could be applicable to captive breeding and to the studies on genetic diversity and population structure of grouper wild stocks.     

Population genetic inferences using immune gene SNPs mirror patterns inferred by microsatellites

Single nucleotide polymorphisms (SNPs) are replacing microsatellites for population genetic analyses, but it is not apparent how many SNPs are needed or how well SNPs correlate with microsatellites. We used data from the gopher tortoise, Gopherus polyphemus—a species with small populations, to compare SNPs and microsatellites to estimate population genetic parameters. Specifically, we compared one SNP data set (16 tortoises from four populations sequenced at 17 901 SNPs) to two microsatellite data sets, a full data set of 101 tortoises and a partial data set of 16 tortoises previously genotyped at 10 microsatellites. For the full microsatellite data set, observed heterozygosity, expected heterozygosity and F_ST were correlated between SNPs and microsatellites; however, allelic richness was not. The same was true for the partial microsatellite data set, except that allelic richness, but not observed heterozygosity, was correlated. The number of clusters estimated by structure differed for each data set (SNPs = 2; partial microsatellite = 3; full microsatellite = 4). Principle component analyses (PCA) showed four clusters for all data sets. More than 800 SNPs were needed to correlate with allelic richness, observed heterozygosity and expected heterozygosity, but only 100 were needed for F_ST. The number of SNPs typically obtained from next‐generation sequencing (NGS) far exceeds the number needed to correlate with microsatellite parameter estimates. Our study illustrates that diversity, F_ST and PCA results from microsatellites can mirror those obtained with SNPs. These results may be generally applicable to small populations, a defining feature of endangered and threatened species, because theory predicts that genetic drift will tend to outweigh selection in small populations.     

PCR primers for trinucleotide and tetranucleotide microsatellites in greater amberjack,Seriola dumerili

Eighteen nuclear‐encoded microsatellites from a genomic DNA library of greater amberjack, Seriola dumerili, were isolated and characterized. The microsatellites include 13 perfect (five tetranucleotide and eight trinucleotide) and five imperfect (three tetranucleotide, one trinucleotide and one combination dinucleotide/trinucleotide) repeat motifs. The number of alleles at the 18 microsatellites among a sample of 29 fish ranged from two to 20; gene diversity (expected heterozygosity) ranged from 0.068 to 0.950, whereas observed heterozygosity ranged from 0.069 to 0.966. Following Bonferroni correction, genotypes at all 18 microsatellites fit expectations of Hardy–Weinberg equilibrium, and all pairwise comparisons of microsatellites did not deviate significantly from genotypic equilibrium. Greater amberjack support commercial and recreational fisheries along both the Atlantic and the Gulf coasts of the USA and represent a species with potential for worldwide aquaculture. The microsatellites developed will be useful for population genetic studies of ‘wild’ populations and breeding studies of domesticated populations.     

Effect of microsatellite selection on individual and population genetic inferences: an empirical study using cross‐specific and species‐specific amplifications

Although whole‐genome sequencing is becoming more accessible and feasible for nonmodel organisms, microsatellites have remained the markers of choice for various population and conservation genetic studies. However, the criteria for choosing microsatellites are still controversial due to ascertainment bias that may be introduced into the genetic inference. An empirical study of red deer (Cervus elaphus) populations, in which cross‐specific and species‐specific microsatellites developed through pyrosequencing of enriched libraries, was performed for this study. Two different strategies were used to select the species‐specific panels: randomly vs. highly polymorphic markers. The results suggest that reliable and accurate estimations of genetic diversity can be obtained using random microsatellites distributed throughout the genome. In addition, the results reinforce previous evidence that selecting the most polymorphic markers leads to an ascertainment bias in estimates of genetic diversity, when compared with randomly selected microsatellites. Analyses of population differentiation and clustering seem less influenced by the approach of microsatellite selection, whereas assigning individuals to populations might be affected by a random selection of a small number of microsatellites. Individual multilocus heterozygosity measures produced various discordant results, which in turn had impacts on the heterozygosity‐fitness correlation test. Finally, we argue that picking the appropriate microsatellite set should primarily take into account the ecological and evolutionary questions studied. Selecting the most polymorphic markers will generally overestimate genetic diversity parameters, leading to misinterpretations of the real genetic diversity, which is particularly important in managed and threatened populations.     

Development and characterization of microsatellites in Vanda varieties

The orchid genus Vanda includes more than 70 monopodial species and numerous hybrids. The aim of this study was to develop microsatellite markers for this horticultural valuable genus. Thirteen polymorphic microsatellite markers were isolated from the variety Vanda Miss Joaquim and were characterized in four Vanda species, 11 Vanda hybrids, and one Aranda intergenus hybrid. Populations from three species were also analysed. Number of alleles ranged from two to 19. For the V. sumatrana population, the expected heterozygosity ranged from 0 to 0.76 (mean 0.31) and the observed heterozygosity ranged from 0 to 1 (mean 0.36). All the varieties tested were distinct from one another (similarity index < 0.8). These microsatellites could be used for studying genetic diversity and population structure of wild populations within the orchid genus Vanda, as well as for distinguishing cultured Vanda varieties.     

Isolation and characterization of highly polymorphic microsatellite loci for the garden tiger moth Arctia caja (Lepidoptera: Arctiidae)

A set of microsatellite markers was isolated from Arctia caja, a species that has been declining in UK for several decades. As has been found with other Lepidoptera species, this proved to be a difficult task. Eight enriched libraries identified 103 positive clones, and these yielded only seven polymorphic microsatellites. Allelic diversity ranged from 10 to 23 alleles per locus in a population of 30 individuals. Significant heterozygosity deficits were shown by three of these loci, presumably due to null alleles. The remaining four loci were in Hardy–Weinberg equilibrium and will be used to investigate the population genetic structure of A. caja.     

Isolation and characterization of microsatellite loci in the xishishe clam <Emphasis Type="Italic">Coelomactra antiquata</Emphasis> (Bivalvia: Veneroida)

The xishishe clam Coelomactra antiquata is a commercially important bivalve species, but has been suffering from severe population decline due to over-exploitation and the deterioration of environmental conditions. To investigate the genetic variation and structures of this clam, nine polymorphic microsatellites were developed. The number of alleles ranged from two to 28 per locus with the observed heterozygosity ranging from 0.083 to 0.921. This set of microsatellites will be useful for the study of the genetic structures of C. antiquata.     

Evidence of parasite-mediated disruptive selection on genetic diversity in a wild fish population

Identifying the processes maintaining genetic variability in wild populations is a major concern in conservation and evolutionary biology. Parasite-mediated selection may strongly affect genetic variability in wild populations. The inbreeding depression theory predicts that directional selection imposed by parasites should act against the most inbred hosts, thus favouring genetic diversity in wild populations. We have tested this prediction by evaluating the strength and shape of the relationship between the load of a harmful fin-feeder ectoparasite ( Tracheliastes polycolpus ) and the genome-wide genetic diversity (i.e. heterozygosity measured at a set of 15 microsatellites) of its fish host, the rostrum dace ( Leuciscus leuciscus ). Contrary to expectation, we found a nonlinear relationship between host genetic diversity and ectoparasite load, with hosts that were either homozygous or heterozygous harbouring significantly fewer parasites than hosts with an intermediate level of heterozygosity. This relationship suggests that parasites could increase the variance of global heterozygosity in this host population through disruptive selection on genetic diversity. Moreover, when genetic diversity was measured at each locus separately, we found two very strong positive associations between host genetic diversity and the ectoparasite load. This latter result has three main implications: (i) genome-wide effect cannot alone explain the nonlinear relationship between global heterozygosity and ectoparasite load, (ii) negative non-additive allelic interactions (i.e. underdominance) may be a mechanism for resisting ectoparasite infection, and (iii) ectoparasites may favour homozygosity at some loci in this host population.     

Eight polymorphic microsatellite loci for the critically endangered crested ibis,Nipponia nippon (Ciconiiformes: Threskiornithidae)

The Chinese crested ibis, Nipponia nippon, is a critically endangered bird. The current population of this species has developed from four wild individuals rediscovered in 1981. Given its conservation status, there is considerable interest in assessing the genetic diversity and individual relatedness in this species. For this purpose, a set of eight polymorphic di‐ or trinucleotide microsatellite loci was developed for the crested ibis. The expected heterozygosity at these loci ranges from 0.01 to 0.50, with less than four alleles being observed at individual loci, a reflection of the serious population bottleneck experienced by this species.     

Genetic structure analysis of a highly inbred captive population of the African antelope Addax nasomaculatus. Conservation and management implications

The African antelope Addax nasomaculatus is a rare mammal at high risk of extinction, with no more than 300 individuals in the wild and 1,700 captive animals distributed in zoos around the world. In this work, we combine genetic data and genealogical information to assess the structure and genetic diversity of a captive population located at Parque Lecocq Zoo (N=27), originated from only two founders. We amplified 39 microsatellites previously described in other Artiodactyls but new to this species. Seventeen markers were polymorphic, with 2–4 alleles per locus (mean=2.71). Mean expected heterozygosity (He) per locus was between 0.050 (marker ETH3) and 0.650 (marker D5S2), with a global He of 0.43. The mean inbreeding coefficient of the population computed from pedigree records of all registered individuals (N=53) was 0.222. The mean coancestry of the population was 0.298 and F_IS index was ?0.108. These results reflect the importance of an adequate breeding management on a severely bottlenecked captive population, which would benefit by the incorporation of unrelated individuals. Thanks to the successful amplification of a large number of microsatellites commonly used in domestic bovids, this study will provide useful information for the management of this population and serve as future reference for similar studies in other captive populations of this species. Zoo Biol 30:399–411, 2011. © 2010 Wiley‐Liss, Inc.     

Characterization and cross-species amplification of microsatellites from the endangered Hawksbill turtle (Eretmochelys imbricate)

Twelve novel polymorphic microsatellites were isolated from the endangered Hawksbill turtle (Eretmochelys imbricate). Eight of 12 markers were used to study genetic diversity of two sea turtle species: E. imbricate and green sea turtle (C. mydas). In E. imbricate, the average allele number of the eight microsatellites was 6.25/locus with a range of 3–13. The average expected and observed heterozygosity was 0.66 and 0.63 respectively. In C. mydas, the average allele number of the eight markers was 11.63/locus. The observed heterozyosity (0.68) was lower than the expected heterozyosity (0.79). Most of 12 microsatellites amplified specific and polymorphic PCR products in other six turtle species. Hence, the developed microsatellites would facilitate studies on genetic diversity and population structure of E. imbricate and other marine turtle species.     

Isolation and characterization of microsatellite DNA primers for <Emphasis Type="Italic">Sinadoxa corydalifolia</Emphasis> (Adoxaceae)

Sinadoxa corydalifolia is the only species of Sinadoxa (Adoxaceae) with the aberrant morphology. This species has become extremely endangered in the Qinghai-Tibetan Plateau. To provide a population-level genetic profile for investigation and conservation of genetic diversity of this species, we developed 10 new microsatellite loci for this species by the combining biotin capture method. About 31 microsatellites were screened from the library, 10 of the screened microsatellites are polymorphic. The number of alleles per locus in 18 individuals ranged from 3 to 11, expected heterozygosity and observed heterozygosity ranged from 0.3071 to 0.6243 and from 0.1675 to 0.4357, respectively. We further performed cross-priming tests of these primers in another species of the Adoxaceae: Adoxa moschatellina and found 9 of 10 successfully amplified the targeted sequences. These newly developed loci provide a useful tool to investigate the genetic diversity and design the conversation measures of S. corydalifolia and study the genetic divergence and the initial speciation pattern between it and the related species in the Adoxaceae.     

Isolation and characterization of 17 polymorphic microsatellites in grass carp

Here we report the isolation and characterization of 17 polymorphic loci isolated from a partial genomic DNA library of grass carp (Ctenopharyngodon idellus) enriched for CA repeats. We tested variability of these microsatellites on 24 unrelated individuals collected in China. All microsatellites were polymorphic. The average allele number was 7.9 per locus, ranging from four to 13. The observed heterozygosity was from 0.46 to 0.88 with an average of 0.71, whereas the average expected heterozygosity was 0.78. Sixteen of the 17 microsatellites conformed to Hardy–Weinberg equilibrium, and inherited independently. These microsatellites can be used to study genetic diversity and population structure of wild populations, and facilitate selective breeding of cultured broodstocks.     

Isolation and characterization of microsatellites in a marine food fish species,golden trevally Gnathanodon specious

The first set of six polymorphic microsatellites was isolated from a partial genomic DNA library of an important marine food fish Gnathanodon specious, and was characterized in 28 unrelated individuals. The allele number ranged from three to 12 with an average of seven/locus; average expected heterozygosity was 0.57 ranging from 0.10 to 0.87, whereas the observed heterozygosity ranged from 0.11 to 0.96 (average: 0.57). All six markers conform to Hardy–Weinberg and linkage equilibria. These markers will be useful for accessing genetic population structure of wild populations and for selective breeding program in fish farms.     

Isolation and characterization of dinucleotide microsatellites in greater amberjack, <Emphasis Type="Italic">Seriola dumerili</Emphasis>

Thirteen nuclear-encoded dinucleotide microsatellites were characterized from a genomic DNA library of greater amberjack, Seriola dumerili. The microsatellites include 12 perfect-repeat motifs and one imperfect-repeat motif. The number of alleles at the 13 microsatellites among a sample of 29 fish ranged from 3 to 25; gene diversity (expected heterozygosity) ranged from 0.296 to 0.948, while observed heterozygosity ranged from 0.276 to 0.897. Following Bonferroni correction, genotypes at all 13 microsatellites fit expectations of Hardy-Weinberg equilibrium. One pairwise comparison of microsatellites deviated significantly from expectations of genotypic equilibrium, suggesting that these two microsatellites may be linked. Greater amberjack support commercial and recreational fisheries along both the Atlantic and Gulf coasts of the U.S. and represent a species with potential for worldwide aquaculture. The microsatellites developed will be useful for conservation and population genetic studies of ‘wild’ and domesticated populations of greater amberjack.     

Isolation and characterization of 14 polymorphic microsatellite loci in the ark shell Scapharca subcrenata (Bivalvia: Arcidae)

The ark shell Scapharca subcrenata is an important fishery resource, but has been suffering from severe population decline due to the deterioration of the coastal environment and over-exploitation. To investigate the genetic variation and structure among populations of S. subcrenata, 14 polymorphic microsatellite loci were developed. The number of alleles per polymorphic locus ranged from 11 to 29 and the observed and expected heterozygosity varied from 0.400 to 0.923 and from 0.705 to 0.965, respectively. These microsatellites will help advance the investigation of the genetic population structure and genetic diversity in this species.