Phylogeography of the endangered rosewood Dalbergia nigra (Fabaceae): insights into the evolutionary history and conservation of the Brazilian Atlantic Forest

The Brazilian rosewood (Dalbergia nigra) is an endangered tree endemic to the central Brazilian Atlantic Forest, one of the world''s most threatened biomes. The population diversity, phylogeographic structure and demographic history of this species were investigated using the variation in the chloroplast DNA (cpDNA) sequences of 185 individuals from 19 populations along the geographical range of the species. Fifteen haplotypes were detected in the analysis of 1297 bp from two non-coding sequences, trnV-trnM and trnL. We identified a strong genetic structure (F_ST=0.62, P<0.0001), with a latitudinal separation into three phylogeographic groups. The two northernmost groups showed evidence of having maintained historically larger populations than the southernmost group. Estimates of divergence times between these groups pointed to vicariance events in the Middle Pleistocene (ca. 350 000–780 000 years ago). The recurrence of past climatic changes in the central part of the Atlantic forest, with cycles of forest expansion and contraction, may have led to repeated vicariance events, resulting in the genetic differentiation of these groups. Based on comparisons among the populations of large reserves and small, disturbed fragments of the same phylogeographic group, we also found evidence of recent anthropogenic effects on genetic diversity. The results were also analysed with the aim of contributing to the conservation of D. nigra. We suggest that the three phylogeographic groups could be considered as three distinct management units. Based on the genetic diversity and uniqueness of the populations, we also indicate priority areas for conservation.     

Development of polymorphic microsatellite markers for Dalbergia nigra (Papilionoideae), an endangered tree from the Brazilian Atlantic Forest

Dalbergia nigra is an endangered tree restricted to the Brazilian Atlantic Forest. Ten polymorphic microsatellite loci were isolated and characterized in 47 trees from two populations. The total number of alleles per locus ranged from three to 12 alleles. The levels of observed and expected heterozygosities ranged from 0.304 to 0.740 and from 0.278 to 0.872, respectively. Significant deviations from Hardy-Weinberg were detected for only three loci in each population. No pair of loci exhibited significant linkage disequilibrium. These microsatellites provide an efficient tool to investigate genetic structure in forest remnants with the purpose of conservation of this species.     

Genetic variation in remnant populations of Dalbergia nigra (Papilionoideae), an endangered tree from the Brazilian Atlantic Forest

BACKGROUND AND AIMS: Dalbergia nigra, known as Brazilian rosewood, is an endangered tree species restricted to the Brazilian Atlantic Forest and has been intensively logged for five centuries due to its high-quality wood. The objective of the present study was to assess the genetic variation and structure in adults and saplings of the species from a large reserve of the Atlantic Forest, the Rio Doce State Park, and from two small surrounding fragments, one better preserved and another with a high degree of anthropogenic disturbance. METHODS: Analyses of genetic variation and structure were conducted by studying allozyme markers. Seven putative enzymatic loci were resolved, five of them being polymorphic. KEY RESULTS: The mean numbers of alleles per locus (A) were 1.93 and 1.73, while the percentages of polymorphic loci (P) were 93 and 73 % for adults and saplings, respectively. Saplings from the fragment with high anthropogenic disturbance exhibited the lowest values of A and P. The fragment that constitutes a conservation area exhibited genetic variation similar to the population from the large reserve. The observed (H(o)) and expected (H(e)) heterozygosities were not significantly different among the three populations. Only sapling populations showed F(ST) values (divergence among populations) significantly different from zero over all studied loci. The fragment with high anthropogenic disturbance exhibited considerable genetic divergence in relation to the above-cited populations. CONCLUSIONS: The evaluated populations displayed mean levels of genetic variation intermediate to those expected for narrow and widespread species. The results suggest that fragments with similar area and geographical distance from a large protected reserve can exhibit different levels of genetic variation, depending on the degree of anthropogenic disturbance. The considerable genetic variation in the protected fragment points to the importance of adequate conservation of small fragments for the preservation of genetic variation in D. nigra.     

活血丹小尺度空间遗传结构

活血丹(Glechoma lonituba)是唇形科活血丹属的多年生克隆草本植物。采用简单重复序列区(ISSR)分子标记技术,比较分析了3个不同斑块活血丹的遗传多样性、克隆多样性以及小尺度空间遗传结构,并探讨其与生境异质性、繁殖体传播和人为干扰的相关性。结果表明:1)活血丹物种水平的遗传多样性很低,各斑块的遗传多样性较低,以水渠边斑块最高,平葛村斑块次之,竹林下斑块最低。2)活血丹物种水平的克隆多样性较高,各斑块活血丹的克隆多样性以水渠边斑块最大,平葛村斑块次之,竹林下斑块最低。3)遗传分化系数Gst为0.7129,表明活血丹的遗传变异大部分存在于斑块间;斑块间的基因流较小,仅为0.2004。4)空间自相关分析表明活血丹一定的空间距离下存在显著的空间遗传结构,竹林下斑块在100cm时存在显著性正相关,其X轴截矩为205.994cm;平葛村斑块在200cm时存在显著性正相关,其X轴截矩为235.388cm;水渠边斑块在150cm时存在显著性正相关,其X轴截矩为240.336cm。应用软件SPAGeDi 1.2软件对各斑块的空间遗传结构进行量化,表明平葛村斑块具有最强的空间遗传结构,水渠边和竹林下斑块的空间遗传结构较弱。活血丹的遗传结构、克隆结构及空间分布格局受到其繁殖体传播特征、人为干扰和繁殖权衡的影响,是其对生境异质性的适应结果。     

Fine-scale genetic structure among genetic individuals of the clone-forming monotypic genus Echinosophora koreensis (Fabaceae)

• Background and Aims For rare endemics or endangered plantspecies that reproduce both sexually and vegetatively it iscritical to understand the extent of clonality because assessmentof clonal extent and distribution has important ecological andevolutionary consequences with conservation implications. Asurvey was undertaken to understand clonal effects on fine-scalegenetic structure (FSGS) in two populations (one from a disturbedand the other from an undisturbed locality) of Echinosophorakoreensis, an endangered small shrub belonging to a monotypicgenus in central Korea that reproduces both sexually and vegetativelyvia rhizomes. • Methods Using inter-simple sequence repeats (ISSRs) asgenetic markers, the spatial distribution of individuals wasevaluated using Ripley's L(d)-statistics and quantified thespatial scale of clonal spread and spatial distribution of ISSRgenotypes using spatial autocorrelation analysis techniques(join-count statistics and kinship coefficient, F_ij) for totalsamples and samples excluding clones. • Key Results A high degree of differentiation betweenpopulations was observed (_ST(g) = 0·184, P < 0·001).Ripley's L(d)-statistics revealed a near random distributionof individuals in a disturbed population, whereas significantaggregation of individuals was found in an undisturbed site.The join-count statistics revealed that most clones significantlyaggregate at 6-m interplant distance. The Sp statistic reflectingpatterns of correlograms revealed a strong pattern of FSGS forall four data sets (Sp = 0·072–0·154), butthese patterns were not significantly different from each other.At small interplant distances (2 m), however, jackknifed 95% CIs revealed that the total samples exhibited significantlyhigher F_ij values than the same samples excluding clones. • Conclusion The strong FSGS from genets is consistentwith two biological and ecological traits of E. koreensis: bee-pollinationand limited seed dispersal. Furthermore, potential clone matesover repeated generations would contribute to the observed highF_ij values among genets at short distance. To ensure long-termex situ genetic variability of the endangered E. koreensis,individuals located at distances of 10–12 m should becollected across entire populations of E. koreensis.     

Spatial and temporal population genetic variation and structure of Nothotsuga longibracteata (Pinaceae), a relic conifer species endemic to subtropical China

Nothotsuga longibracteata, a relic and endangered conifer species endemic to subtropical China, was studied for examining the spatial-temporal population genetic variation and structure to understand the historical biogeographical processes underlying the present geographical distribution. Ten populations were sampled over the entire natural range of the species for spatial analysis, while three key populations with large population sizes and varied age structure were selected for temporal analyses using both nuclear microsatellites (nSSR) and chloroplast microsatellites (cpSSR). A recent bottleneck was detected in the natural populations of N. longibracteata. The spatial genetic analysis showed significant population genetic differentiation across its total geographical range. Notwithstanding, the temporal genetic analysis revealed that the level of genetic diversity between different age class subpopulations remained constant over time. Eleven refugia of the Last Glacial Maximum were identified, which deserve particular attention for conservation management.     

High levels of genetic differentiation and selfing in the Brazilian cerrado fruit tree Dipteryx alata Vog. (Fabaceae)

Dipteryx alata is a native fruit tree species of the cerrado (Brazilian savanna) that has great economic potential because of its multiple uses. Knowledge of how the genetic variability of this species is organized within and among populations would be useful for genetic conservation and breeding programs. We used nine simple sequence repeat (SSR) primers developed for Dipteryx odorata to evaluate the genetic structure of three populations of D. alata located in central Brazil based on a leaf sample analysis from 101 adults. The outcrossing rate was evaluated using 300 open-pollinated offspring from 25 seed-trees. Pollen dispersal was measured by parentage analysis. We used spatial genetic structure (SGS) to test the minimal distance for harvesting seeds in conservation and breeding programs. Our data indicate that the populations studied had a high degree of genetic diversity and population structure, as suggested by the high level of divergence among populations . The estimated outcrossing rate suggested a mixed mating system, and the intrapopulation fixation index was influenced by SGS. We conclude that seed harvesting for genetic conservation and breeding programs requires a minimum distance between trees of 196 m to avoid collecting seeds from related seed-trees.     

Fine-scale spatial genetic structure and gene flow in a small, fragmented population of Sinojackia rehderiana (Styracaceae), an endangered tree species endemic to China

Populations of Sinojackia rehderiana are highly threatened and have small and scattered distribution due to habitat fragmentation and human activities. Understanding changes in genetic diversity, the fine-scale spatial genetic structure (SGS) at different life stages and gene flow of S. rehderiana is critical for developing successful conservation strategies for fragmented populations of this endangered species. In this study, 208 adults, 114 juveniles and 136 seedlings in a 50 × 100-m transect within an old-growth forest were mapped and genotyped using eight microsatellite makers to investigate the genetic diversity and SGS of this species. No significant differences in genetic diversity among different life-history stages were found. However, a significant heterozygote deficiency in adults and seedlings may result from substantial biparental inbreeding. Significant fine-scale spatial structure was found in different life-history stages within 19 m, suggesting that seed dispersal mainly occurred near a mother tree. Both historical and contemporary estimates of gene flow (13.06 and 16.77 m) indicated short-distance gene dispersal in isolated populations of S. rehderiana. The consistent spatial structure revealed in different life stages is most likely the result of limited gene flow. Our results have important implications for conservation of extant populations of S. rehderiana. Measures for promoting pollen flow should be taken for in situ conservation. The presence of a SGS in fragmented populations implies that seeds for ex situ conservation should be collected from trees at least 19-m apart to reduce genetic similarity between neighbouring individuals.     

Fine-scale spatial genetic structure in mixed oak stands with different levels of hybridization

Oaks are model species for the study of natural introgressive hybridization. High interfertility among oak taxa might result in collective evolution, through transpecific spread of advantageous alleles, challenging the standard concept of species. Nine highly polymorphic microsatellite (nuSSR) loci were analysed in three mixed oak populations of Quercus pyrenaica and Quercus petraea (Montejo, Somosierra and Robregordo) with different density and hybridization rates. Both leaf morphology and molecular markers were used to assess individual admixture rates. Insights about the relative effect of density and hybridization rates on fine-scale spatial genetic structure (SGS) were obtained from autocorrelograms and Sp statistics. Differences in SGS among populations were higher than between species. These differences cannot be attributed solely to census densities but also relate to other factors, such as the spatial configuration of the population. Hybridization was an important factor shaping within-population spatial genetic structure, and an interspecific component of SGS was found in Somosierra. Indirect estimates of historical gene flow in Montejo were compared with actual values of gene dispersal assessed by parentage analysis in a former study. Similar values were found for current and historical gene flow in both species, which might reflect demographical stability.     

Genetic structure and conservation of Mountain Lions in the South-Brazilian Atlantic Rain Forest

The Brazilian Atlantic Rain Forest, one of the most endangered ecosystems worldwide, is also among the most important hotspots as regards biodiversity. Through intensive logging, the initial area has been reduced to around 12% of its original size. In this study we investigated the genetic variability and structure of the mountain lion, Puma concolor. Using 18 microsatellite loci we analyzed evidence of allele dropout, null alleles and stuttering, calculated the number of allele/locus, PIC, observed and expected heterozygosity, linkage disequilibrium, Hardy-Weinberg equilibrium, F(IS), effective population size and genetic structure (MICROCHECKER, CERVUS, GENEPOP, FSTAT, ARLEQUIN, ONESAMP, LDNe, PCAGEN, GENECLASS software), we also determine whether there was evidence of a bottleneck (HYBRIDLAB, BOTTLENECK software) that might influence the future viability of the population in south Brazil. 106 alleles were identified, with the number of alleles/locus ranging from 2 to 11. Mean observed heterozygosity, mean number of alleles and polymorphism information content were 0.609, 5.89, and 0.6255, respectively. This population presented evidence of a recent bottleneck and loss of genetic variation. Persistent regional poaching constitutes an increasing in the extinction risk.     

Fine-scale genetic structure of the threatened rosewood Dalbergia nigra from the Atlantic Forest: comparing saplings versus adults and small fragment versus continuous forest

Assessing the population genetic structure of threatened species is important for developing successful conservation strategies. In this study, we evaluated the fine-scale spatial genetic structure (SGS) of Dalbergia nigra from a regenerating secondary forest fragment and compared it with previous data from a primary forest of a large reserve. A total of 107 adult and 111 saplings were mapped and genotyped for seven microsatellite loci. The genetic diversity was high and similar in adults (H _e?=?0.682) and saplings (H _e?=?0.680). The spatial extent of SGS was higher in adults than in saplings. Overlapping generations in the potentially reproductive individuals is the likely explanation for the higher SGS in adults (Sp?=?0.016) in relation to the saplings (Sp?=?0.010). The SGS in the adults from the secondary forest fragment was similar to that found in the primary forest. Considering the SGS found in adults, from both the secondary and primary forests, seeds for ex situ conservation should be collected from trees at least 80 m apart to reduce the genetic similarity between samples. These results highlight the importance of preserving small forest fragments to allow successful regeneration and maintenance of the genetic diversity in D. nigra.     

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, population structure, and conservation of Sophora moorcroftiana (Fabaceae), a shrub endemic to the Tibetan Plateau

Sophora moorcroftiana is a perennial leguminous low shrub endemic to the middle reaches of Yarlung Zangbo River in Tibet. It is an important species to fix sand dunes and to avoid the formation of shifting sands; therefore, its progressive over-exploitation may enhance land desertification. The levels and distribution of genetic variability of this species were evaluated from 10 natural populations at 24 loci encoding 13 enzymes, using allozyme analysis by starch gel electrophoresis. Data obtained revealed moderate levels of genetic variation within populations (Pp=27.5%, Ap=1.5, Hep=0.122) and a considerable divergence among populations (FST=0.199). Significant positive correlations (r2=0.49, p<0.05; r2=0.46, p<0.05) were found between elevation and both mean number of alleles per locus (A) and gene diversity (He) in the studied populations of S. moorcroftiana. Lower genetic diversity in lower elevation populations might be due to the negative effects of human pressures and habitat fragmentation, to adaptation to high altitudes as a consequence of a peripatric speciation process, or to directional gene flow along the river basin from the source populations located in the west at higher altitudes. The evaluation of the degree of threat has led to the inclusion of this species in the category of EN ("endangered"), and conservation strategies for this endemic species are discussed on the basis of these findings.     

Inferring recruitment history from spatial genetic structure within populations of the colonizing tree Albizia julibrissin (Fabaceae)

Comparative analyses of spatial genetic structure (SGS) among species, populations, or cohorts give insight into the genetic consequences of seed dispersal in plants. We analysed SGS of a weedy tree in populations with known and unknown recruitment histories to first establish patterns in populations with single vs. multiple founders, and then to infer possible recruitment scenarios in populations with unknown histories. We analysed SGS in six populations of the colonizing tree Albizia julibrissin Durazz. (Fabaceae) in Athens, Georgia. Study sites included two large populations with multiple, known founders, two small populations with a single, known founder, and two large populations with unknown recruitment histories. Eleven allozyme loci were used to genotype 1385 individuals. Insights about the effects of colonization history from the SGS analyses were obtained from correlograms and Sp statistics. Distinct differences in patterns of SGS were identified between populations with multiple founders vs. a single founder. We observed significant, positive SGS, which decayed with increasing distance in the populations with multiple colonists, but little to no SGS in populations founded by one colonist. Because relatedness among individuals is estimated relative to a local reference population, which usually consists of those individuals sampled in the study population, SGS in populations with high background relatedness, such as those with a single founder, may be obscured. We performed additional analyses using a regional reference population and, in populations with a single founder, detected significant, positive SGS at all distances, indicating that these populations consist of highly related descendants and receive little seed immigration. Subsequent analyses of SGS in size cohorts in the four large study populations showed significant SGS in both juveniles and adults, probably because of a relative lack of intraspecific demographic thinning. SGS in populations of this colonizing tree is pronounced and persistent and is determined by the number and relatedness of founding individuals and adjacent seed sources. Patterns of SGS in populations with known histories may be used to indirectly infer possible colonization scenarios for populations where it is unknown.     

Spatial genetic structure of Aquilegia taxa endemic to the island of Sardinia

Background and Aims

The Mediterranean Basin is one of the most important regions for the Earth''s plant biodiversity; however, the scarcity of studies on fine scale patterns of genetic variation in this region is striking. Here, an assessment is made of the spatial genetic structure of all known locations of the three Sardinian endemic species of Aquilegia in order to determine the relative roles of gene flow and genetic drift as underlying evolutionary forces canalizing the divergence of Sardinian Aquilegia taxa, and to see if the spatial genetic structure found fits the current taxonomic differentiation of these taxa.

Methods

DNA from 89 individuals from all known locations of Aquilegia across Sardinia was analysed by means of amplified fragment length polymorphism (AFLP) markers. Both principal co-ordinates analysis (PCoA) and Bayesian clustering analyses were used to determine the spatial genetic structure irrespective of any taxonomic affiliation. Historical effects of gene flow and genetic drift were assessed by checking for the existence of isolation-by-distance patterns.

Key Results

STRUCTURE and PCoA analyses revealed a pattern of genetic variation geographically structured into four spatial genetic groups. No migration–drift equilibrium was detected for Aquilegia in Sardinia, when analysed either as a whole or in individual groups. The scenario approached a Case III pattern sensu Hutchinson and Templeton, which is associated with extreme isolation conditions where genetic drift has historically played a dominant role over gene flow.

Conclusions

The pattern of genetic variation of Sardinian taxa of Aquilegia indicates that genetic drift has been historically more influential than gene flow on population structure of Sardinian species of Aquilegia. Limited seed dispersal and divergent selection imposed by habitat conditions have been probably the main causes reinforcing post-Pleistocene geographical isolation of Aquilegia populations. The spatial genetic structure found here is not fully compatible with current taxonomic affiliations of Sardinian Aquilegia taxa. This is probably a consequence of the uncoupling between morphological and genetic patterns of differentiation frequently found in recently radiated taxa.     

Low levels of realized seed and pollen gene flow and strong spatial genetic structure in a small,isolated and fragmented population of the tropical tree Copaifera langsdorffii Desf

Over the past century, the Brazilian Atlantic forest has been reduced to small, isolated fragments of forest. Reproductive isolation theories predict a loss of genetic diversity and increases in inbreeding and spatial genetic structure (SGS) in such populations. We analysed eight microsatellite loci to investigate the pollen and seed dispersal patterns, genetic diversity, inbreeding and SGS of the tropical tree Copaifera langsdorffii in a small (4.8 ha), isolated population. All 112 adult trees and 128 seedlings found in the stand were sampled, mapped and genotyped. Seedlings had significantly lower levels of genetic diversity (A=16.5±0.45, mean±95% s.e.; H_e=0.838±0.006) than did adult trees (A=23.2±0.81; H_e=0.893±0.030). Parentage analysis did not indicate any seed immigration (m_seeds=0) and the pollen immigration rate was very low (m_pollen=0.047). The average distance of realized pollen dispersal within the stand was 94 m, with 81% of the pollen travelling <150 m. A significant negative correlation was found between the frequency and distance of pollen dispersal (r=−0.79, P<0.01), indicating that short-distance pollinations were more frequent. A significant SGS for both adults (∼50 m) and seedlings (∼20 m) was also found, indicating that most of the seeds were dispersed over short distances. The results suggested that the spatial isolation of populations by habitat fragmentation can restrict seed and pollen gene flow, increase SGS and affect the genetic diversity of future generations.     

喀斯特耐阴灌木淡黄金花茶的精细空间遗传结构与基因流

淡黄金花茶(Camelia flavida)是一种具有淡黄色花瓣的金花茶植物,为喀斯特典型的耐阴灌木。基于广西弄岗喀斯特季节性雨林15 hm2(300 m×500 m)监测样地,利用13对微卫星(SSR)标记,探讨喀斯特地貌对淡黄金花茶的精细空间遗传结构(SGS)、基因流和小尺度范围遗传分化的影响。结果表明:(1)淡黄金花茶在50 m距离内产生显著的SGS,在灌木类型和依赖种子传播的物种中具有中等程度的SGS强度(Sp=0.0248);(2)淡黄金花茶种子和花粉传播平均距离较短,分别为12.47 m和29.03 m,72.2%花粉和81.0%种子的传播距离均主要是在20 m内;(3)淡黄金花茶在小尺度范围(种群间距离1 km)的4个种群,甚至相距100 m的两个斑块产生了显著的遗传分化。喀斯特生境地貌对淡黄金花茶的基因流产生阻碍作用,从而导致淡黄金花茶在小尺度范围产生遗传分化。     

Characterization of microsatellite markers for Thoropa taophora (Anura, Cycloramphidae), a frog endemic to the Brazilian Atlantic Rain Forest

Here we characterize nine microsatellite loci for Thoropa taophora collected from a coastal and an island population in southeastern Brazil. The number of alleles per locus ranged from three to 11 for the coastal population and from two to eight for the island population. Observed heterozygosities ranged from 0.308 to 0.778 for the coastal population and from zero to 0.737 for the island population. One locus did not conform to Hardy-Weinberg expectations on the coast, while two were in disequilibrium on the island. These markers are currently being used to examine genetic differentiation and patterns of dispersal in T. taophora.     

Development of microsatellite markers for Dimorphandra mollis (Leguminosae), a widespread tree from the Brazilian cerrado

? Premise of the study: Microsatellite markers were developed for Dimorphandra mollis (Leguminosae), a widespread tree in the Brazilian cerrado (a savanna-like vegetation). ? Methods and Results: Microsatellite markers were developed from an enriched library. The analyses of polymorphism were based on 56 individuals from three populations. Nine microsatellite loci were polymorphic, with the number of alleles per locus ranging from three to 10 across populations. The observed and expected heterozygosities per locus and population ranged from 0.062 to 0.850 and from 0.062 to 0.832, respectively. ? Conclusions: These microsatellites provide an efficient tool for population genetics studies and will be used to assess the genetic diversity and spatial genetic structure of D. mollis.