Low genetic diversity and limited gene flow in a dominant mangrove tree species (Rhizophora stylosa) at its northern biogeographical limit across the chain of three Sakishima islands of the Japanese archipelago as revealed by chloroplast and nuclear SSR analysis

We examined the genetic diversity, population structure and gene flow in a dominant mangrove tree (Rhizophora stylosa) at its northern biogeographical limit in Sakishima islands of the Japanese archipelago. Simple sequence repeat (SSR) markers from chloroplast (cpSSR) and nuclear DNA were used to analyze 16 populations recovered from various river basins across the chain of three Sakishima islands—Iriomote, Ishigaki and Miyako. The average number of alleles (1.7–2.7) and observed heterozygosities (0.031–0.216) at nuclear SSR and haploid diversity (0.000–0.489) at cpSSR across the populations suggested low genetic diversity in R. stylosa in Sakishima islands. cpSSR analysis identified two haplotypes, and Bayesian clustering analysis (nuclear SSR) revealed two genetic clusters. Analysis of molecular variance (nuclear SSR) showed significant population differentiations. Pairwise tests consistently revealed significant differentiation between most of the population pairs; however, the degrees of differentiations are generally correspondent to the relative geographical distances as suggested from pairwise F _ST and cpSSR genetic distances. Moreover, Mantel tests showed some signals of correlations between genetic distances (nuclear and chloroplast) and geographical distances. These results suggest that combined contribution of gene flow via pollen and propagule dispersal in R. stylosa mostly occurred between neighboring river basins. The appearances of two cpSSR haplotypes (maternal lineages) as well as two nuclear genetic clusters (putative ancestral lineages) at various river basins support the hypothesis that present-day R. stylosa populations across the Sakishima islands were established from few identical founders; however, significant differentiations among various river basins most likely resulted from the limited gene flow and high inbreeding.     

Genetic structure of disjunct Argentinean populations of the subtropical tree Anadenanthera colubrina var. cebil (Fabaceae)

Anadenanthera colubrina var. cebil is a native South American tree species inhabiting seasonally dry tropical forests (SDTFs). Its current disjunct distribution presumably represents fragments of a historical much larger area of this forest type, which has also been highly impacted by human activities. In this way the hypothesis of this study is that the natural populations of A. colubrina var. cebil from Northern Argentina represent vestiges of ancient fragmentation, but they are additionally influenced by a certain degree of gene flow among them. We aimed to analyze the genetic structure of both nuclear and chloroplast DNA to evaluate the relative role of ancient and recent fragmentation on intraspecific diversity patterns. Sixty-nine individuals of four natural populations were analyzed using eight nuclear microsatellites (ncSSR) and four chloroplast microsatellite loci (cpSSR). The level and distribution of genetic variation were estimated by standard population genetic parameters and Neighbor Joining as well as Bayesian analyses. The eight ncSSR loci were highly polymorphic, while genetic diversity of cpSSRs was low. Nuclear SSRs displayed lower genetic differentiation among populations than cpSSR haplotypes (F _ST 0.11 and 0.95, respectively). However, high differentiation between phytogeographic provinces was observed in both genomes. The high genetic differentiation detected emphasizes the role of ancient fragmentation. However, the Paranaense province also shows the effects of recent fragmentation on genetic structure, whereas gene flow by pollen preserves the effects of genetic drift in the Yungas province.     

Small effect of fragmentation on the genetic diversity of Dalbergia monticola, an endangered tree species of the eastern forest of Madagascar, detected by chloroplast and nuclear microsatellites

Background and Aims

The oriental forest ecosystem in Madagascar has been seriously impacted by fragmentation. The pattern of genetic diversity was analysed on a tree species, Dalbergia monticola, which plays an important economic role in Madagascar and is one of the many endangered tree species in the eastern forest.

Methods

Leaves from 546 individuals belonging to 18 small populations affected by different levels of fragmentation were genotyped using eight nuclear (nuc) and three chloroplast (cp) microsatellite markers.

Key Results

For nuclear microsatellites, allelic richness (R) and heterozygosity (H_e,nuc) differed between types of forest: R = 7·36 and R = 9·55, H_e,nuc = 0·64 and H_e,nuc = 0·80 in fragmented and non-fragmented forest, respectively, but the differences were not significant. Only the mean number of alleles (N_a,nuc) and the fixation index F_IS differed significantly: N_a,nuc = 9·41 and N_a,nuc = 13·18, F_IS = 0·06 and F_IS = 0·15 in fragmented and non-fragmented forests, respectively. For chloroplast microsatellites, estimated genetic diversity was higher in non-fragmented forest, but the difference was not significant. No recent bottleneck effect was detected for either population. Overall differentiation was low for nuclear microsatellites (F_ST,nuc = 0·08) and moderate for chloroplast microsatellites (F_ST,cp = 0·49). A clear relationship was observed between genetic and geographic distance (r = 0·42 P < 0·01 and r = 0·42 P = 0·03 for nuclear and chloroplast microsatellites, respectively), suggesting a pattern of isolation by distance. Analysis of population structure using the neighbor-joining method or Bayesian models separated southern populations from central and northern populations with nuclear microsatellites, and grouped the population according to regions with chloroplast microsatellites, but did not separate the fragmented populations.

Conclusions

Residual diversity and genetic structure of populations of D. monticola in Madagascar suggest a limited impact of fragmentation on molecular genetic parameters.     

Genetic diversity and integrity of German wildcat (Felis silvestris) populations as revealed by microsatellites,allozymes, and mitochondrial DNA sequences

As a consequence of persecution and habitat fragmentation, wildcats (Felis silvestris silvestris) in Western Europe have experienced a severe reduction in population numbers and sizes. The remaining wildcat populations are considered to be endangered by losses of genetic variability and by hybridisation with free-ranging domestic cats. To investigate genetic diversity within and among wild and domestic cat populations in Germany and to estimate the extent of gene flow between both forms, we analysed a total of 266 individuals. PCR-amplification and sequencing of 322 base pairs of a highly variable part of the mitochondrial control region (HV1) of 244 specimens resulted in 41 haplotypes with 31 polymorphic sites. Additionally, eight microsatellite loci were examined for those 244 cats. Moreover, a total of 46 wildcats and 22 domestic cats could be genotyped for 13 polymorphic out of 31 enzyme loci. Genetic variability in both groups was generally high. Variability in domestic cat populations was higher than in wildcat populations. Almost no differentiation between domestic cat populations could be found (F_ST for microsatellites=3%). In contrast, wildcat populations differed significantly from one another (F_ST for microsatellites=9.55%) Within the smaller wildcat populations, a reduction of genetic diversity was detectable with regard to the nuclear DNA. Wildcat and domestic cat mitochondrial haplotypes were separated, suggesting a very low level of maternal gene flow between both forms. In microsatellites and to a somewhat lesser extent in allozymes, wildcats and domestic cats showed distinct differentiation, suggesting an only low extent of past hybridisation in certain populations. The microsatellite data set indicated a significantly reduced effective population size (bottleneck) in the recent past for one German wildcat population.     

Large‐scale connectivity,cryptic population structure,and relatedness in Eastern Pacific Olive ridley sea turtles (Lepidochelys olivacea)

Endangered species are grouped into genetically discrete populations to direct conservation efforts. Mitochondrial control region (mtCR) haplotypes are used to elucidate deep divergences between populations, as compared to nuclear microsatellites that can detect recent structuring. When prior populations are unknown, it is useful to subject microsatellite data to clustering and/or ordination population inference. Olive ridley sea turtles (Lepidochelys olivacea) are the most abundant sea turtle, yet few studies have characterized olive ridley population structure. Recently, clustering results of olive ridleys in the Eastern Tropical Pacific Ocean suggested weak structuring (F_ST = 0.02) between Mexico and Central America. We analyzed mtCR haplotypes, new microsatellite genotypes from Costa Rica, and preexisting microsatellite genotypes from olive ridleys across the Eastern Tropical Pacific, to further explore population structuring in this region. We subjected inferred populations to multiple analyses to explore the mechanisms behind their structuring. We found 10 mtCR haplotypes from 60 turtles nesting at three sites in Costa Rica, but did not detect divergence between Costa Rican sites, or between Central America and Mexico. In Costa Rica, clustering suggested one population with no structuring, but ordination suggested four cryptic clusters with moderate structuring (F_ST = 0.08, p < .001). Across the Eastern Tropical Pacific, ordination suggested nine cryptic clusters with moderate structuring (F_ST = 0.103, p < .001) that largely corresponded to Mexican and Central American populations. All ordination clusters displayed significant internal relatedness relative to global relatedness (p < .001) and contained numerous sibling pairs. This suggests that broadly dispersed family lineages have proliferated in Eastern Tropical Pacific olive ridleys and corroborates previous work showing basin‐wide connectivity and shallow population structure in this region. The existence of broadly dispersed kin in Eastern Tropical Pacific olive ridleys has implications for management of olive ridleys in this region, and adds to our understanding of sea turtle ecology and life history, particularly in light of the natal‐homing paradigm.     

Population genetic structure of the endangered Eastern Bristlebird, Dasyornis brachypterus; implications for conservation

For species that are habitat specialists or sedentary, population fragmentation may lead to genetic divergence between populations and reduced genetic diversity within populations, with frequent inbreeding. Hundreds of kilometres separate three geographical regions in which small populations of the endangered Eastern Bristlebird, Dasyornis brachypterus, a small, ground-dwelling passerine that occurs in fire-prone bushland in eastern Australia, are currently found. Here, we use mitochondrial and microsatellite DNA markers to: (i) assess the sub-specific taxonomy designated to northern range-edge, and central and southern range-edge D. brachypterus, respectively, and (ii) assess levels of standing genetic variation and the degree of genetic subdivision of remnant populations. The phylogenetic relationship among mtDNA haplotypes and their spatial distribution did not support the recognised subspecies boundaries. Populations in different regions were highly genetically differentiated, but in addition, the two largest, neighboring populations (located within the central region and separated by ~50 km) were moderately differentiated, and thus are likely closed to migration (microsatellites, F _ST = 0.06; mtDNA, F _ST = 0.12, ?? _ST = 0.08). Birds within these two populations were genotypically diverse and apparently randomly mating. A long-term plan for the conservation of D. brachypterus??s genetic diversity should consider individual populations as separate management units. Moreover, managers should avoid actively mixing birds from different populations or regions, to conserve the genetic integrity of local populations and avoid outbreeding depression, should further translocations be used as a recovery tool for this species.     

Genetic diversity and differentiation of Siberian spruce populations at nuclear microsatellite loci

The results of the study of 21 populations of Siberian spruce (Picea obovata Ledeb.) from different parts of the species natural range by microsatellite (SSR) analysis of nuclear DNA are presented. Using nine loci developed for Picea abies (L.) Karst. and Picea glauca (Moench) Voss and detecting variation in Picea obovata, the parameters of intra- and interpopulation genetic diversity, as well as the degree of population differentiation, were determined. It was demonstrated that the population of Siberian spruce in the study was characterized by a relatively high average level of intrapopulation variability (H_o = 0.408; H_e = 0.423) and low interpopulation differentiation (F_st = 0.048, P = 0.001) at this class of DNA markers. The genetic distance between populations ranged from 0.009 to 0.167, averaging 0.039. The isolated Magadan population, located in the extreme Northeast of Russia at a considerable distance from the main species range and characterized by the lowest genetic diversity among the studied populations, was maximally differentiated from the rest of the spruce populations. In addition, the steppe Ubukun population from Buryatia and the population from the Bogd Khan Uul Biosphere Reserve, Mongolia, were considerably different in the genetic structure from most populations of Siberian spruce, although to a lesser extent than the Magadan population. These findings are consistent with the results of previous studies of this species carried out using allozyme and microsatellite loci of chloroplast DNA and point to the prospects of using nuclear microsatellites as DNA markers to analyze the population genetic structure of Siberian spruce.     

Population structure and genetic variation in the genus Dipteronia Oliv. (Aceraceae) endemic to China as revealed by cpSSR analysis

The genus Dipteronia Oliv. endemic to central and southern China consists of two species, D. sinensis Oliv. and D. dyerana Henry, both of them are rare and endangered. In the present study, genetic variation in 17 populations of D. sinensis and four populations of D. dyerana was estimated using ten polymorphic chloroplast microsatellite loci (cpSSR). Forty-nine chloroplast haplotypes were identified from 204 individuals analyzed. Thirty-nine haplotypes were found in D. sinensis, while ten in D. dyerana. No haplotype was shared between the species. AMOVA analysis revealed that the majority of the genetic variation was partitioned among populations within D. sinensis (F _ST = 0.7980) and D. dyerana (F _ST = 0.654). Cluster analysis grouped the 21 populations into two groups according to their species delimitation. The populations of D. sinensis were further divided into two subgroups corresponding to their geographical distributions. Correlation analysis revealed significant correlation between geographical distance and genetic distance of these populations (r = 0.326, p < 0.01 for D. sinensis vs. r = 0.777, p < 0.05 for D. dyerana). Genetic structure of these populations and a calculated pollen-to-seed flow ratio of (3.2:1 vs. 0.6:1) within the species suggested that little gene flow has occurred among the populations over an extended period. Thus, it implies that the genus Dipteronia might have experienced a genetic bottleneck and limited expansion during its evolutionary history.     

High genetic diversity and significant population structure in Cedrus brevifolia Henry, a narrow endemic Mediterranean tree from Cyprus

Endemic island plant species with a narrow distribution are often, but not always, linked to low genetic variation within populations and a lack of differentiation among populations. Cedrus brevifolia is a narrow endemic island tree species of Cyprus. Its range is restricted to a single forest, divided into five neighbouring sites. This study, using biparentally inherited nuclear microsatellites and paternally inherited plastid (chloroplast) microsatellites, assessed the genetic variation of C. brevifolia within its sole population and the level of genetic differentiation among formed sites. The results from both markers showed high diversity (nuclear H _T?=?0.70; plastid H _T?=?0.93), strongly suggesting that the species did not experience severe bottleneck events or extensive genetic drift. Besides, the maintenance of a high genetic diversity in C. brevifolia may suggest that it originates from a widespread congener species. Significant genetic differentiation at nuclear (G _ST?=?0.052) and plastid (G _ST?=?0.119) markers was found among the formed sites. Remarkably, the relatively high genetic differentiation found at plastid markers was comparable to values observed in two widespread congener cedar species. The genetic differentiation probably occurred due to fragmentation of a previously uniform population. This would lead to the shaping of different genetic groups (Bayesian analysis) and to significant population substructure. Furthermore, significant values observed for both isolation by distance and large-scale spatial genetic structure could indicate ineffective gene flow among sites and the early geographical isolation of the more isolated sites from the core population.     

Genetic diversity and population structure of the roughskin sculpin (<Emphasis Type="Italic">Trachidermus fasciatus</Emphasis> Heckel) inferred from microsatellite analyses: implications for its conservation and management

Identification of population units is crucial for management and monitoring programs, especially for endangered wild species. The roughskin sculpin (Trachidermus fasciatus Heckel) is a small catadromous fish and has been listed as a second class state protected aquatic animal since 1988 in China. To achieve sustainable conservation of this species, it is necessary to clarify the existing genetic structure both between and within populations. Here, population genetic structure among eight populations of T. fasciatus were analyzed by using 16 highly polymorphic microsatellites. High levels of genetic variation were observed in all populations. All pairwise F _ST estimates were significant after false discovery rate correction (overall average F _ST = 0.054). Furthermore, both STRUCTURE and discriminant analysis of principal components (DAPC) analysis showed that the eight populations were grouped into six clusters. BAYESASS analysis showed generally low recent and asymmetric migration among populations. All these results suggested significant genetic structure across populations. However, there was no isolation by distance relationship among populations, likely resulting from barriers to gene flow created by habitat fragmentation. Our results highlight the need for in situ conservation efforts for T. fasciatus across its entire distribution range, through maximizing habitat size and quality to preserve overall genetic diversity and evolutionary potential.     

Genetic diversity and gene flow in a Caribbean tree <Emphasis Type="Italic">Pterocarpus officinalis</Emphasis> Jacq.: a study based on chloroplast and nuclear microsatellites

We analysed the molecular diversity of Pterocarpus officinalis, a tree species distributed in Caribbean islands, South and Central America to quantify the genetic variation within island, to assess the pattern of differentiation and infer levels of gene flow; with the overall goal of defining a strategy of conservation. Two hundred two individuals of 9 populations were analysed using three chloroplast and six nuclear microsatellite markers. The observed heterozygosity varied markedly among the populations for nuclear (H _Onuc = 0.20–0.50) and chloroplast microsatellites (H _cp = 0.22–0.68). The continental population from French Guyana showed a higher value of H _Onuc than island populations, and the differences were significant in some cases. The fixation index F _IS ranged from −0.043 to 0.368; a significant heterozygote deficit was detected in 7 populations. The heterozygosity excess method suggested that two populations in Guadeloupe have undergone a recent bottleneck. Global and pairwise F _ST were high for both nuclear (F _STnuc = 0.29) and chloroplast microsatellites (F _STcp = 0.58). The neighbour-joining tree based on both markers, presented a differentiation pattern that can be explained by the seed dispersal by flotation and marine stream. The comparison of Bayesian approach and the method based on allelic frequency demonstrate a very limited number of migrants between populations.     

Chloroplast DNA haplotype diversity and postglacial recolonization of Hagenia abyssinica (Bruce) J.F. Gmel. in Ethiopia

We investigated genetic variation of 273 individuals from 25 populations of the monotypic species Hagenia abyssinica (Rosaceae) from the highlands of Ethiopia at three chloroplast microsatellite loci. The objectives were to infer the factors that shaped the genetic structure and to reconstruct the recolonization history of the species. Six haplotypes that were phylogenetically grouped into two lineages were identified. Homology of the three loci to the respective regions of the chloroplast genome was confirmed by sequencing. The chloroplast haplotypes found in Hagenia showed a clear pattern of congruence between their geographical distribution and genealogical relationships. A very low haplotype diversity within populations (h _S = 0.079, v _S = 0.058) and a very high population differentiation (G _ST = 0.899, N _ST = 0.926) was observed, reflecting very low mixing between recolonizing lineages. Restricted gene flow through seeds, rare long-distance dispersal, contiguous range expansion and mutation shaped the genetic structure of Hagenia. Fossil pollen records suggested that the trend of postglacial recolonization of Hagenia was first in the south and latter went to the north in Ethiopia.     

Population stock structure of leatherback turtles (Dermochelys coriacea) in the Atlantic revealed using mtDNA and microsatellite markers

This study presents a comprehensive genetic analysis of stock structure for leatherback turtles (Dermochelys coriacea), combining 17 microsatellite loci and 763 bp of the mtDNA control region. Recently discovered eastern Atlantic nesting populations of this critically endangered species were absent in a previous survey that found little ocean-wide mtDNA variation. We added rookeries in West Africa and Brazil and generated longer sequences for previously analyzed samples. A total of 1,417 individuals were sampled from nine nesting sites in the Atlantic and SW Indian Ocean. We detected additional mtDNA variation with the longer sequences, identifying ten polymorphic sites that resolved a total of ten haplotypes, including three new variants of haplotypes previously described by shorter sequences. Population differentiation was substantial between all but two adjacent rookery pairs, and F _ST values ranged from 0.034 to 0.676 and 0.004 to 0.205 for mtDNA and microsatellite data respectively, suggesting that male-mediated gene flow is not as widespread as previously assumed. We detected weak (F _ST = 0.008 and 0.006) but significant differentiation with microsatellites between the two population pairs that were indistinguishable with mtDNA data. POWSIM analysis showed that our mtDNA marker had very low statistical power to detect weak structure (F _ST < 0.005), while our microsatellite marker array had high power. We conclude that the weak differentiation detected with microsatellites reflects a fine scale level of demographic independence that warrants recognition, and that all nine of the nesting colonies should be considered as demographically independent populations for conservation. Our findings illustrate the importance of evaluating the power of specific genetic markers to detect structure in order to correctly identify the appropriate population units to conserve.     

Genetic diversity and differentiation of the endangered Japanese endemic tree Magnolia stellata using nuclear and chloroplast microsatellite markers

Genetic diversity and differentiation were analyzed in 11 populations of Magnolia stellata (Sieb. and Zucc.) Maxim. (Magnoliaceae) in the Tokai district, Japan. Variation at four nuclear microsatellite (nSSR) loci was examined, three chloroplast microsatellite (cpSSR) markers were developed and 13 haplotypes identified. The 11 populations were divided into three groups (A, B and C). Each population within the group was separated less than 40 km. Group B harbored the highest gene diversity (H) and allelic richness (Ar) for nSSR (H=0.74 and Ar=8.02). Group C had the highest diversity of chloroplast haplotypes (H=0.79 and Ar=6.8): 2.5 times more haplotypes than the other groups. Each population contributed differently to the total diversity, with respect to nSSR and cpSSR. AMOVA revealed that 58% of haplotypic and 15% of nSSR variation was partitioned among populations within groups. A Mantel test revealed significant correlations between population pairwise geographic ln(distance) and F_ST/(1−F_ST) for both nSSR (r=0.479; P=0.001) and cpSSR (r=0.230; P=0.040). Dendrograms of populations for nSSR, based on Nei’s genetic distance, were constructed using UPGMA and the neighbor-joining method. These results suggest that populations in group C have diverged from the other populations, while those in group B are similar to each other. For group B, fragmentation between populations should be avoided in order to maintain gene flow. For group C, the uniqueness of each population should be given the highest priority when planning genetic conservation measures for the species.     

Molecular,quantitative and abiotic variables for the delineation of evolutionary significant units: case of sandalwood (<Emphasis Type="Italic">Santalum austrocaledonicum</Emphasis> Vieillard) in New Caledonia

Various approaches have been developed to define conservation units for plant and animal species. In this study we combined nuclear microsatellites (from a previous published study) and chloroplast microsatellites (assessed in the present study), leaf and seed morphology traits and abiotic variables (climate and soil) to define evolutionary significant units (ESU) of Santalum austrocaledonicum, a tree species growing in New Caledonia. Results for chloroplast microsatellites showed that the total population heterozygosity was␣high, (H _cp = 0.84) but varied between islands. Differentiation was strong in the total population (F _stcp = 0.66) but also within the main island Grande Terre (F _stcp = 0.73) and within Iles Loyauté (F _stcp = 0.52), highlighting a limited gene flow between populations. These results confirmed those obtained with nuclear microsatellites. The cluster analysis on molecular markers discriminated two main groups constituted by the populations of Grande Terre and the populations of Iles Loyauté. A principal component analysis of leaf and seed morphology traits singled out the populations of Iles Loyauté and the western populations of Grande Terre. Quantitative genetic analyses showed that the variation between populations was under genetic control (broad sense heritability close to 80%). A high correlation between rainfall and morphological traits suggested an impact of climate on this variation. The integration of these results allows to define two ESUs, one corresponding to Grande Terre and Ile des Pins and the other the Iles Loyauté archipelago. This study stresses the need to restore some populations of Grande Terre that are currently threatened by their small size.     

Patterns of genetic variation in the Chinese endemic Psilopeganum sinense (Rutaceae) as revealed by nuclear microsatellites and chloroplast microsatellites

Psilopeganum sinense is a perennial herb endemic to Three-Gorges Reservoir Area (TGRA) in China. Genetic diversity of this endangered species was assessed by using 11 nuclear microsatellites and three chloroplast microsatellite (cpSSR) markers. A total of 8 haplotypes were identified in a survey of 212 individuals sampled from nine populations encompassing most of the natural range of the species. A low level of genetic diversity was detected: H_E = 0.301 for SSR and H_E = 0.28 for cpSSR. Populations were highly differentiated from one another: an AMOVA analysis that showed that 56.3% and 68.2% genetic variation resided between populations based on SSR and cpSSR analysis, respectively, and F_ST and F_STc (0.467 for SSR and 0.644 for cpSSR, respectively) were high. Significant differences were found between estimates of haplotypic differentiation calculated by using unordered alleles (G_STc = 0.857) and ordered alleles (N_STc = 0.728), which indicated the existence of phylogeographical structure in P. sinense. The indirect ratio of pollen flow/seed flow derived from estimates of haplotypic and nuclear DNA differentiation indicated that gene flow via pollen is less efficient than via seed. Two distinct evolutionary lineages (evolutionary significant units, ESUs) were recognized for P. sinense on the basis of both the PCoA and NCA analysis. Sampling strategies for conserving this endangered plant were discussed.     

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.     

Identification and characterization of microsatellite markers for the population genetic structure in endemic red-tailed barb,Gonoproktopterus curmuca

Gonoproktopterus curmuca is an endangered red tailed barb found in Southern part of Western Ghat, India. As a part of stock-specific, propagation assisted rehabilitation and management program, polymorphic microsatellites markers were used to study the genetic diversity and population structure of this species from the three River systems of Southern Western Ghats, such as Periyar River, the Chalakkudy River, and the Chaliyar River. From selected eight polymorphic microsatellite markers, the number of alleles per locus ranged from 2 to 8, and the average number of alleles among 3 populations ranged from 5.0 to 5.75. The mean observed (H_ob) and expected (H_ex) heterozygosity ranged from 0.5148 to 0.5360 and from 0.5996 to 0.6067, respectively. Significant deviations from Hardy–Weinberg Equilibrium expectation were found at majority of the loci (except Gcur MFW72 and Gcur MFW19) and in all three populations in which heterozygote deficits were apparent. The analysis of molecular variance indicates that the percent of variance among populations and within populations were 6.73 and 93.27, respectively. The pairwise F_ST values between populations indicate that there were significant deviations in genetic differentiations for the red-tailed barb populations from these three Rivers of the Western Ghats, India. The microsatellites methods reported a low degree of gene diversity and lack of genetic heterogeneity in the population of G. curmuca, which strongly emphasize the need of fishery management, conservation and rehabilitation of G. curmuca.     

Chloroplast DNA phylogeography of Betula maximowicziana, a long-lived pioneer tree species and noble hardwood in Japan

Betula maximowicziana is an ecologically and economically important tree species in Japan. In order to examine the phylogeographical pattern of the species in detail, maternally inherited chloroplast (cp) DNA variations of 25 natural populations of Betula maximowicziana and a total of 12 populations of three related species were evaluated by PCR-RFLP analysis. Two main haplotypic groups of B. maximowicziana populations (northern and southern) were detected, with the main boundary passing through the Tohoku region in northeastern Japan; in addition there was high genetic differentiation among the 25 populations studied (G _ST = 0.950, G_\textST^￠ = 0. 9 7 7 G_{\text{ST}}^{\prime } = 0. 9 7 7 ). The phylogeographical pattern exhibited by B. maximowicziana was much more similar to that of alpine plants than to that of beech and oak. Comparison of the patterns of genetic structure obtained from the cpDNA with previously and newly acquired data on bi-parentally inherited nuclear DNA indicates that the nuclear genome was transferred via pollen from the northern haplotypic group to the southern group more frequently than it moved in the opposite direction. Although common haplotypes were detected among B. maximowicziana and the two related species examined, these haplotypes were not shared sympatrically, suggesting very rare hybridization among the species currently occurring in their natural populations.     

Tertiary Origin and Pleistocene Diversification of Dragon Blood Tree (Dracaena cambodiana-Asparagaceae) Populations in the Asian Tropical Forests

Background

The origin of extraordinarily rich biodiversity in tropical forests is often attributed to evolution under stable climatic conditions over a long period or to climatic fluctuations during the recent Quaternary period. Here, we test these two hypotheses using Dracaena cambodiana, a plant species distributed in paleotropical forests.

Methods

We analyzed nucleotide sequence data of two chloroplast DNA (cpDNA: atpB-rbcL and trnD-trnT) regions and genotype data of six nuclear microsatellites from 15 populations (140 and 363 individuals, respectively) distributed in Indochina Peninsular and Hainan Island to infer the patterns of genetic diversity and phylogeographic structure. The population bottleneck and genetic drift were estimated based upon nuclear microsatellites data using the software programs BOTTLENECK and 2MOD. The lineage divergence times and past population dynamics based on cpDNA data were estimated using coalescent-based isolation-with-migration (IMa) and BEAST software programs.

Results

A significant phylogeographic structure (N _ST = 0.876, G _ST = 0.796, F _ST-SSR =0.329, R _ST =0.449; N _ST>G _ST, R _ST>F _ST-SSR, P<0.05) and genetic differentiation among populations were detected. Bottleneck analyses and Bayesian skyline plot suggested recent population reduction. The cpDNA haplotype network revealed the ancestral populations from the southern Indochina region expanded to northward. The most recent ancestor divergence time of D. cambodiana dated back to the Tertiary era and rapid diversification of terminal lineages corresponded to the Quaternary period.

Conclusions

The results indicated that the present distribution of genetic diversity in D. cambodiana was an outcome of Tertiary dispersal and rapid divergence during the Quaternary period under limited gene flow influenced by the uplift of Himalayan-Tibetan Plateau and Quaternary climatic fluctuations respectively. Evolutionary processes, such as extinction-recolonization during the Pleistocene may have contributed to the fast diversification in D. cambodiana.