Genetic differences between wild and hatchery populations of Korean spotted sea bass (Lateolabrax maculatus) inferred from microsatellite markers

The spotted sea bass, Lateolabrax maculatus, is popular in recreational fishing and aquaculture in Korea. Its natural population has declined during the past two decades; thus, beginning in the early 2000s stock-enhancement programs were introduced throughout western and southern coastal areas. In this study, genetic similarities and differences between wild and hatchery populations were assessed using multiplex assays with 12 highly polymorphic microsatellite loci; 96 alleles were identified. Although many unique alleles were lost in the hatchery samples, no significant reductions were found in heterozygosity or allelic diversity in the hatchery compared to the wild population. High genetic diversity (He = 0.724–0.761 and Ho = 0.723–0.743), low inbreeding coefficient (F _IS = 0.003–0.024) and Hardy–Weinberg equilibrium were observed in both wild and hatchery populations. However, the genetic heterogeneity between the populations was significant. Therefore, genetic drift likely promoted inter-population differentiation, and rapid loss of genetic diversity remains possible. Regarding conservation, genetic variation should be monitored and inbreeding controlled in a commercial breeding program.     

The development of 10 novel polymorphic microsatellite markers through next generation sequencing and a preliminary population genetic analysis for the endangered Glenelg spiny crayfish, Euastacus bispinosus

The Glenelg spiny crayfish, Euastacus bispinosus, is an iconic freshwater invertebrate of south eastern Australia and listed as ‘endangered’ under the Environment Protection and Biodiversity Conservation Act 1999, and ‘vulnerable’ under the International Union for Conservation of Nature’s Red List. The species has suffered major population declines as a result of over-fishing, low environmental flows, the introduction of invasive fish species and habitat degradation. In order to develop an effective conservation strategy, patterns of gene flow, genetic structure and genetic diversity across the species distribution need to be clearly understood. In this study we develop a suite of polymorphic microsatellite markers by next generation sequencing. A total of 15 polymorphic loci were identified and 10 characterized using 22 individuals from the lower Glenelg River. We observed low to moderate genetic variation across most loci (mean number of alleles per locus = 2.80; mean expected heterozygosity = 0.36) with no evidence of individual loci deviating significantly from Hardy–Weinberg equilibrium. Marker independence was confirmed with tests for linkage disequilibrium, and analyses indicated no evidence of null alleles across loci. Individuals from two additional sites (Crawford River, Victoria; Ewens Ponds Conservation Park, South Australia) were genotyped at all 10 loci and a preliminary investigation of genetic diversity and population structure was undertaken. Analyses indicate high levels of genetic differentiation among sample locations (F _ST  = 0.49), while the Ewens Ponds population is genetically homogeneous, indicating a likely small founder group and ongoing inbreeding. Management actions will be needed to restore genetic diversity in this and possibly other at risk populations. These markers will provide a valuable resource for future population genetic assessments so that an effective framework can be developed for implementing conservation strategies for E. bispinosus.     

Contrasting spatial genetic structure in Annona crassiflora populations from fragmented and pristine savannas

In continuous populations, fine-scale genetic structure tends to be stronger in species with restricted pollen and seed dispersal. However, habitat fragmentation and disturbances can affect genetic diversity and spatial genetic structure due to disruption in ecological processes, such as plant reproduction and seed dispersal. In this study, we compared the genetic diversity and fine-scale spatial genetic structure (SGS) in two populations of Annona crassiflora (Annonaceae) in a pristine savanna Reserve (ESECAE) and in a fragmented disturbed savanna area (PABE), both in Cerrado biome in Central Brazil. The analyses were based on the polymorphism at 10 microsatellite loci. Our working hypothesis was that SGS is stronger and genetic diversity is lower in population at fragmented area (PABE) than at pristine area (ESECAE). Both populations presented high levels of polymorphism and genetic diversity and showed no sign of bottleneck for both Wilcoxon sign-rank test for heterozygosity excess (p > 0.05) and coalescent analyses (growth parameter g not different from zero), but population at fragmented area showed higher fixation index and stronger SGS. Besides, populations are significantly differentiated (F _ST = 0.239, R _ST = 0.483, p < 0.001 for both). Coalescent analyses showed high historical effective population sizes for both populations, high gene flow between ESECAE and PABE and recent time to most recent common ancestor (~37 k year BP). Our results suggest that despite the high genetic diversity, fragmentation and disturbance may have been affecting populations of this species increasing mating between closely related individuals leading to high fixation index and strong SGS.     

Genotyping of microsatellite markers to study genetic structure of the wild striped snakehead Channa striata in Malaysia

Genetic variability and differences in wild striped snakehead Channa striata from Malaysia were analysed by genotyping nine novel nuclear microsatellite loci. Analysis revealed moderate‐to‐high genetic diversity in most of the populations, indicative of large effective population sizes. The highly diversified populations are admixed populations and, therefore, can be recommended as potential candidates for selective breeding and conservation since they each contain most of the alleles found in their particular region. Three homogenous groups of the wild populations were identified, apparently separated by effective barriers, in accordance with contemporary drainage patterns. The highest population pairwise F_ST found between members of the same group reflects the ancient population connectivity; yet prolonged geographical isolation resulted in adaptation of alleles to local contemporary environmental change. A significant relationship between genetic distance and geographical isolation was observed (r = 0·644, P < 0·01). Anthropogenic perturbations indicated apparent genetic proximity between distant populations.     

Comparative Analysis of Genetic Diversity and Population Structure of Sipunculus nudus as Revealed by Mitochondrial COI Sequences

Genetic diversity and population structure of Sipunculus nudus were evaluated using a 652 base pair fragment of the mitochondrial cytochrome oxidase I gene. The populations were collected from Beihai, Sanya, and Xiamen. A total of 71 polymorphic sites defined 16 distinct haplotypes. The mean haplotype diversity and nucleotide diversity of the three populations were 0.9354 ± 0.0168 and 0.0035 ± 0.0018, respectively. Analysis at the intrapopulation level showed that the Beihai population had the greatest haplotype and nucleotide diversity, followed by the Xiamen and Sanya populations. Analysis of molecular variance showed significant genetic differentiation among the three populations (Fst = 0.0796, P < 0.05). The present results revealed that S. nudus populations had a high level of genetic diversity and distinct population structures.     

Microsatellite Polymorphisms in Cassava Landraces from the Cerrado Biome, Mato Grosso do Sul, Brazil

Using nine microsatellite loci, we investigated genetic structure and diversity in 83 Brazilian cassava accessions, including several landraces, in the Cerrado biome in Mato Grosso do Sul, Brazil. All nine loci were polymorphic, averaging 6.00 alleles per locus. Treating each of seven municipalities as a cassava group or population, they averaged 3.5 alleles per locus, with 97% polymorphic loci, high values for observed heterozygosity (0.32) and gene diversity (0.56). Total genetic variability was high (0.668), and most of this genetic variability was concentrated within municipalities (0.577). Cluster and structure analyses divided accessions into two major clusters or populations (K = 2). Also, a significant genetic versus geographic correlation was found (r = 0.4567; P < 0.0260). Migratory routes in the Cerrado are considered main contributors to the region’s high cassava diversity and spatial genetic structure, amplifying interactions between traditional farmers and the evolutionary dynamics of this crop.     

Comparative Genetic Diversity of Wild and Captive Populations of the Bare-Faced Curassow (Crax fasciolata) Based on Cross-Species Microsatellite Markers: Implications for Conservation and Management

The bare-faced curassow (Crax fasciolata) is a large Neotropical bird that suffers anthropogenic pressure across much of its range. A captive population is maintained for conservation management, although there has been no genetic screening of stocks. Based on the six microsatellite markers developed for Crax globulosa, the genetic variability of C. fasciolata and possible differences between a wild and a captive population were investigated. Only three loci were polymorphic, with a total of 27 alleles. More than half of these alleles were private to the wild (n = 8) or captive (n = 7) populations. Significant deviations from Hardy–Weinberg equilibrium were restricted to the captive population. Despite the number of private alleles, genetic drift has probably promoted differentiation between populations. Our results indicate that wild C. fasciolata populations are genetically impoverished and structured, but species-specific microsatellite markers will be necessary for a more reliable assessment of the species’ genetic diversity.     

Using genetic diversity and mating system parameters estimated from genetic markers to determine strategies for the conservation of <Emphasis Type="Italic">Araucaria angustifolia</Emphasis> (Bert.) O. Kuntze (Araucariaceae)

In order to understand the impacts of forest fragmentation on Araucaria angustifolia populations, we evaluated the genetic diversity and mating system using SSR markers and open-pollinated seeds from four populations of varying sizes and spatial isolation, in and around one of the best-conserved Araucaria Forest remnants in Southern Brazil. The four population types of A. angustifolia include: (1) a continuous forest; (2) a physically isolated cluster located 2 km from the continuous forest; (3) an open population in a field located between the cluster and continuous forest; and (4) a fragment on a private property located 5 km from the cluster. Approximately 28 seeds were collected from ten reproductive trees in each population. We found higher amounts of alleles (113) and exclusive alleles (25) in the continuous forest than in the other populations. The multilocus paternity correlation was significantly higher and effective number of pollen donors was significantly lower in the private population, decreasing the diversity and consequently the variance effective size of families sampled from that population. However, despite its isolation from the other studied fragments, the private population had the second highest number of alleles as well as unique alleles from the other populations. Therefore, strategies for A. angustifolia conservation should focus not only on larger populations, such as those found in protected areas, but also include smaller and isolated fragments on private properties as these populations are able to maintain high levels of genetic diversity and functional connectivity between isolated stands across a landscape.     

Molecular Diversity and Population Structure of a Worldwide Collection of Cultivated Tetraploid Alfalfa (Medicago sativa subsp. sativa L.) Germplasm as Revealed by Microsatellite Markers

Information on genetic diversity and population structure of a tetraploid alfalfa collection might be valuable in effective use of the genetic resources. A set of 336 worldwide genotypes of tetraploid alfalfa (Medicago sativa subsp. sativa L.) was genotyped using 85 genome-wide distributed SSR markers to reveal the genetic diversity and population structure in the alfalfa. Genetic diversity analysis identified a total of 1056 alleles across 85 marker loci. The average expected heterozygosity and polymorphism information content values were 0.677 and 0.638, respectively, showing high levels of genetic diversity in the cultivated tetraploid alfalfa germplasm. Comparison of genetic characteristics across chromosomes indicated regions of chromosomes 2 and 3 had the highest genetic diversity. A higher genetic diversity was detected in alfalfa landraces than that of wild materials and cultivars. Two populations were identified by the model-based population structure, principal coordinate and neighbor-joining analyses, corresponding to China and other parts of the world. However, lack of strictly correlation between clustering and geographic origins suggested extensive germplasm exchanges of alfalfa germplasm across diverse geographic regions. The quantitative analysis of the genetic diversity and population structure in this study could be useful for genetic and genomic analysis and utilization of the genetic variation in alfalfa breeding.     

Population genetic structure of the tongue sole (Cynoglossus semilaevis) in Korea based on multiplex PCR assays with 12 polymorphic microsatellite markers

The tongue sole, Cynoglossus semilaevis, is an important fishery resource in Korea. About 100 tongue sole sampled from three major habitats along the western coast of Korea were assessed using multiplex assays with 12 highly polymorphic microsatellite loci to explore the population genetic structure of the species; 151 alleles and similar high levels of gene diversity (mean number of alleles (N_A) = 10.42, mean expected heterozygosity (He) = 0.78) were detected. Three populations showed significant Hardy–Weinberg equilibrium deviations at four loci. Although a significant difference in the number of unique alleles was observed among populations, genetic population subdivision was low by F-statistics (overall F _ST = 0.007, p < 0.05). However, this substructure was not supported by analysis of molecular variance or analyses of isolation by distance. The results suggest a lack of genetic structure among the tongue sole populations in Korean waters and that the populations should be managed as a single unit. The lack of genetic differentiation among samples may be due to high levels of larval dispersal in ocean currents. Alternatively, the populations may have diverged too recently for significant genetic differentiation to have become evident. Given the intensity of tongue sole aquaculture activity in China, which adjoins the western coast of Korea, the possibility that aquaculture may have partially contributed to the population genetic characteristics detected cannot be excluded. This study provides the basic information on nature population structure of C. semilaevis that may help to preserve and manage tongue soles in Korea.     

Mitochondrial DNA Diversity of Orchid Bee Euglossa fimbriata (Hymenoptera: Apidae) Populations Assessed by PCR-RFLP

Euglossa fimbriata is a euglossine species widely distributed in Brazil and occurring primarily in Atlantic Forest remnants. In this study, the genetic mitochondrial structure of E. fimbriata from six Atlantic Forest fragments was studied by RFLP analysis of three PCR-amplified mtDNA gene segments (16S, COI-COII, and cyt b). Ten composite haplotypes were identified, six of which were exclusive and represented singleton mitotypes. Low haplotype diversity (0.085–0.289) and nucleotide diversity (0.000–0.002) were detected within samples. AMOVA partitioned 91.13% of the overall genetic variation within samples and 8.87% (?_st = 0.089; P < 0.05) among samples. Pairwise comparisons indicated high levels of differentiation among some pairs of samples (?_st = 0.161–0.218; P < 0.05). These high levels indicate that these populations of E. fimbriata, despite their highly fragmented landscape, apparently have not suffered loss of genetic variation, suggesting that this particular population is not currently endangered.     

Intrapopulational genetic diversity of Araucaria angustifolia (Bertol.) Kuntze is different when assessed on the basis of chloroplast or nuclear markers

A natural population of Araucaria angustifolia (Bertol.) Kuntze was evaluated to quantify genetic diversity, using nucleotide sequences of chloroplast maturase K genes (matK) and six specific nuclear polymorphic microsatellite loci (nuSSR). Estimates of nucleotide diversity assessed from matK sequences were low (π = 0.001), but they indicated the potential of this gene for evaluating divergence among widespread populations. The matK sequence (1,565 bp) was used for analysis of phylogenetic relationships, enabling clustering of the South American species (A. angustifolia and A. araucana). For analysis of intraspecific genetic diversity, nuSSR were more informative than matK, resulting in a polymorphic loci percentage of 69, a number average of alleles per locus of 11.6, expected genetic diversity 0.741, and significant spatial genetic structure up to 25 m. Our data can help in the selection of genetic sources for conservation of this species, which is at risk of extinction.     

Rangewide Genetic Diversity in Natural Populations of Chinese Pine (Pinus tabulaeformis)

Thirteen natural populations from throughout the range of the Chinese pine (Pinus tabulaeformis Carr.) were examined using inter-simple sequence repeat markers to characterize the genetic structure at the species level and to compare the extent and distribution of genetic variation among central, intermediate, and marginal populations. Although the total genetic variation in the Chinese pine was mainly maintained within populations, the genetic differentiation among populations was significant (P < 0.001). The genetic divergence was significantly correlated with geographic distance (P < 0.05). Genetic diversity tended to decrease from the central to intermediate and marginal populations. The marginal populations had significantly lower intrapopulation genetic diversity than central populations (P < 0.05). Cluster analysis based on Nei’s unbiased genetic distances confirmed the difference among four central populations and the rest. Both historical and contemporary factors may have played key roles in shaping the spatial genetic structure of this species.     

Genetic diversity and structure of an endemic and critically endangered stream river salamander (Caudata: Ambystoma leorae) in Mexico

Small or isolated populations are highly susceptible to stochastic events. They are prone and vulnerable to random demographic or environmental fluctuations that could lead to extinction due to the loss of alleles through genetic drift and increased inbreeding. We studied Ambystoma leorae an endemic and critically threatened species. We analyzed the genetic diversity and structure, effective population size, presence of bottlenecks and inbreeding coefficient of 96 individuals based on nine microsatellite loci. We found high levels of genetic diversity expressed as heterozygosity (H_o = 0.804, H_e = 0.613, H_e* = 0.626 and H_Nei = 0.622). The population presents few alleles (4–9 per locus) and genotypes (3–14 per locus) compared with other mole salamanders species. We identified three genetically differentiated subpopulations with a significant level of genetic structure (F_ST = 0.021, R_ST = 0.044 y D_est = 0.010, 95 % CI). We also detected a reduction signal in population size and evidence of a genetic bottleneck (M = 0.367). The effective population size is small (Ne = 45.2), but similar to another mole salamanders with restricted distributions or with recently fragmented habitat. The inbreeding coefficient levels detected are low (F_IS = ?0.619–0.102) as is gene flow. Despite, high levels of genetic diversity A. leorae is critically endangered because it is a small isolated population.     

Genetic diversity and population genetic structure of wild banana Musa ornata (Musaceae) in Mexico

The wild banana Musa ornata is an inhabitant of the tropical regions of Mexico characterized by patches of tropical rainforest. The overexploitation of its habitat has caused the extinction of several populations affecting diversity and population genetic structure of remaining ones. We used microsatellite markers to determine the genetic diversity and the population’s genetic structure of all extant populations. The thirty-two microsatellite loci previously characterized for M. acuminata and M. balbisiana were tested in M. ornata. Only twelve amplified. From these seven were polymorphic and were used for genetic analyses. The Nei’s diversity estimator shows low levels of genetic diversity (H _e = 0.263) with a mean of 4.40 alleles per locus. Excess homozygosity was evident in all populations indicating high levels of inbreeding. F _ST pairwise analyses and AMOVA indicated low genetic differentiation. However, 28 % of private alleles were registered, suggesting limited gene flow. Genetic distances, Jaccard’s coefficient and principal component analysis showed a good correspondence to geographical locations. The Mantel test performed was not significant. The results support the hypothesis of recent fragmentation events; therefore, not enough time has passed to detect differences between populations. However, it is also likely that results are caused by factors such as bottleneck, decline in pollinator populations, self-pollination and/or a tendency towards clonal reproduction. It is proposed that the preservation strategy focuses on maintaining all the remaining populations and ensuring their connectivity, so as to maintain gene flow and increase the genetic diversity of this species.     

Population structure and genetic diversity of the only extant Baroninae swallowtail butterfly,Baronia brevicornis,revealed by ISSR markers

Due to its relict nature, the unique Baroninae swallowtail, Baronia brevicornis, is considered a “living fossil”. It is also one of the most enigmatic butterfly species with contentious origins and peculiar ecological characteristics. The aim of this study is to evaluate the genetic diversity and population structure of this endemic species of butterfly in Mexico. We sampled populations in two areas within its restricted geographical range in central Mexico and the isolated subspecies population in the state of Chiapas. Three ISSR primers produced 66 loci, indicating a high genetic diversity (P = 100 %, H _e  = 0.22) and variation range in these populations (62 % < P < 85 %, 0.18 < H _e  < 0.25). The Chiapas population presented the lowest values. The observed high values can be explained by the population dynamic of this species characterized by a very high density of individuals over very limited areas. Variation between populations appears to reflect both the age of colonization and locality perturbation level. Two methods of genetic structure analysis (Self-Organizing Map and Structure analysis) match to define three clusters. Natural and anthropogenic barriers may explain the separation between two clusters (cluster 1 and 2) of central Mexico but an unexpected result revealed that the Chiapas population is not genetically distinguishable from the central Mexico populations (cluster 3) leading us to hypothesize a possible “recent” separation or anthropogenic introduction. Habitat and host plant specificity probably limits the exchange of individuals between populations thus increasing fragmentation and leading to a complex genetic structure. We should put in place population monitoring schemes at different spatial scales, combining field occurrences and genetic tools, in order to reduce extinction susceptibility and keep track of recolonization events for this enigmatic species.     

Evaluation of genetic diversity of Panicum turgidum Forssk from Saudi Arabia

The genetic diversity of 177 accessions of Panicum turgidum Forssk, representing ten populations collected from four geographical regions in Saudi Arabia, was analyzed using amplified fragment length polymorphism (AFLP) markers. A set of four primer-pairs with two/three selective nucleotides scored 836 AFLP amplified fragments (putative loci/genome landmarks), all of which were polymorphic. Populations collected from the southern region of the country showed the highest genetic diversity parameters, whereas those collected from the central regions showed the lowest values. Analysis of molecular variance (AMOVA) revealed that 78% of the genetic variability was attributable to differences within populations. Pairwise values for population differentiation and genetic structure were statistically significant for all variances. The UPGMA dendrogram, validated by principal coordinate analysis-grouped accessions, corresponded to the geographical origin of the accessions. Mantel’s test showed that there was a significant correlation between the genetic and geographical distances (r = 0.35, P < 0.04). In summary, the AFLP assay demonstrated the existence of substantial genetic variation in P. turgidum. The relationship between the genetic diversity and geographical source of P. turgidum populations of Saudi Arabia, as revealed through this comprehensive study, will enable effective resource management and restoration of new areas without compromising adaptation and genetic diversity.     

Population genetic structure of wild daffodils (Narcissus pseudonarcissus L.) at different spatial scales

We studied the population genetic and clonal structure of the endangered long-lived perennial plant Narcissus pseudonarcissus using random amplified polymorphic markers. Estimates for mean gene diversity within 15 populations of N. pseudonarcissus of three neighbouring geographical regions were high in comparison to other long-lived perennials (H _eN = 0.33). The genetic diversity of the two smallest populations (<200 plants) was significantly reduced, indicating loss of genetic variability due to drift. The analysis of the population genetic structure revealed a significant genetic differentiation both between regions (Φ_ST = 0.06) and between populations within regions (Φ_ST = 0.20). However, there was incomplete correspondence between geographical regions and the population genetic structure. In order to preserve the overall genetic variation in wild populations of N. pseudonarcissus, management measures should thus aim to protect many populations in each region. The spatial genetic structure within populations of N. pseudonarcissus was in agreement with an isolation by distance model indicating limited gene flow due to pollinator behaviour and restricted seed dispersal. The very restricted spatial extent of clonal growth (<5 cm) and the high level of clonal diversity indicate that clonal growth in N. pseudonarcissus is not an important mode of propagation and that management measures should favour sexual reproduction in order to avoid further reductions in the size and number of populations.     

Unconstrained gene flow between populations of a widespread epiphytic lichen Usnea subfloridana (Parmeliaceae,Ascomycota) in Estonia

Few studies have investigated the genetic diversity of populations of common and widespread lichenized fungi using microsatellite markers, especially the relationships between different measures of genetic diversity and environmental heterogeneity. The main aim of our study was to investigate the population genetics of a widespread and mainly clonally reproducing Usnea subfloridana at the landscape scale, focusing on the comparison of lichen populations within hemiboreal forest stands. Particular attention has been paid to the genetic differentiation of lichen populations in two geographically distinct regions in Estonia and the relationships between forest characteristics and measures of genetic diversity. We genotyped 578 Usnea thalli from eleven lichen populations using seven specific fungal microsatellite markers. Measures of genetic diversity (allelic richness, Shannon's information index, Nei's unbiased genetic diversity, clonal diversity, the number of multilocus genotypes, the number of private alleles, and the minimum number of colonization events) were calculated and compared between Usnea populations. Shared haplotypes, gene flow and AMOVA analyses suggest that unconstrained gene flow and exchange of multilocus genotypes exist between the two geographically remote regions in Estonia. Stand age, mean circumference of the host tree, size of forest site and tree species composition did not show any significant influence on allelic richness, Shannon's information index, Nei's unbiased genetic diversity, clonal diversity, the number of private alleles, and the minimum number of colonization events of U. subfloridana populations. Therefore it was concluded that other factors of habitat heterogeneity could probably have a more significant effect on population genetics of U. subfloridana populations.     

Genetic structure and diversity in relation to the recently reduced population size of the rare conifer, <Emphasis Type="Italic">Pseudotsuga japonica</Emphasis>, endemic to Japan

Rare species consisting of small populations are subject to random genetic drift, which reduces genetic diversity. Thus, determining the relationship between population size and genetic diversity would provide key information for planning a conservation strategy for rare species. We used six microsatellite markers to investigate seven extant populations of the rare conifer Pseudotsuga japonica, which is endemic to the Kii Peninsula and Shikoku Island regions that are geographically separated by the Kii Channel in southwest Japan. The population differentiation of P. japonica was relatively high (F_ST = 0.101) for a coniferous species, suggesting limited gene flow among populations. As expected, significant regional differentiation (AMOVA; p?<?0.05) indicated genetic divergence across the Kii Channel. A strong positive correlation between census population size and the number of rare alleles (r?=?0.862, p?<?0.05) was found, but correlations with major indices of genetic diversity were not significant (allelic richness: r?=?0.649, p?=?0.104, expected heterozygosity: r?=?0.361, p?=?0.426). The observed order of magnitude of correlation with three genetic diversity indices corresponded with the theoretically expected order of each index’ sensitivity (i.e., the rate of decline per generation) to the bottleneck event. Thus, features that exhibit a faster response, i.e., the number of rare alleles, would have been subject to deleterious effects of the recent decline in population size, which is presumably caused by the development of extensive artificial plantations of other tree species over the last several decades. Finally, we propose a conservation plan for P. japonica based on our findings.