Genetic variability and the effect of habitat fragmentation in spotted suslik Spermophilus suslicus populations from two different regions

Endangered species worldwide exist in remnant populations, often within fragmented landscapes. Although assessment of genetic diversity in fragmented habitats is very important for conservation purposes, it is usually impossible to evaluate the amount of diversity that has actually been lost. Here, we compared population structure and levels of genetic diversity within populations of spotted suslik Spermophilus suslicus, inhabiting two different parts of the species range characterized by different levels of habitat connectivity. We used microsatellites to analyze 10 critically endangered populations located at the western part of the range, where suslik habitat have been severely devastated due to agriculture industrialization. Their genetic composition was compared with four populations from the eastern part of the range where the species still occupies habitat with reasonable levels of connectivity. In the western region, we detected extreme population structure (F _ST = 0.20) and levels of genetic diversity (Allelic richness ranged from 1.45 to 3.07) characteristic for highly endangered populations. Alternatively, in the eastern region we found significantly higher allelic richness (from 5.09 to 5.81) and insignificant population structure (F _ST = 0.03). As we identified a strong correlation between genetic and geographic distance and a lack of private alleles in the western region, we conclude that extreme population structure and lower genetic diversity is due to recent habitat loss. Results from this study provide guidelines for conservation and management of this highly endangered species.     

Population genetic structure of striped mullet, Mugil cephalus, along the coast of China, inferred by AFLP fingerprinting

The striped mullet, Mugil cephalus, is an economically important species for both aquaculture and commercial fisheries in China. In this study, the amplified fragment length polymorphism (AFLP) technique was employed to analyze population genetic diversity and genetic distance between different populations with the aim of elucidating the population structure and gene flow of M. cephalus along the coast of China. A total of 230 fragments with 150–500 bp were identified from 118 individuals by five AFLP primer combinations. The polymorphic loci within populations varied from 46.52% to 64.78%, with an average of 53.91%, and the average heterozygosity from 0.1829 to 0.2282, with an average of 0.2074. The UPGMA phenograms of 118 individuals were constructed based on Dice similarity coefficients and four clusters were recognized. AMOVA analysis revealed that 60.7% of genetic variations were within populations and 39.3% between populations. The estimated genetic distance (φ_ST) value over all polymorphic loci across the six populations was 0.393 (p < 0.0010), indicating a strong population structuring. The pairwise φ_ST value ranged from 0.1112 to 0.5358, with an average of 0.3693. The population pairwise gene flows (average N_m = 0.73) are low. In addition, the result of the Mantel test showed that there was a significant correlation between geographic and genetic distances (r = 0.5434, p = 0.0050). It was speculated that there exist at least four distinct geographic populational subdivisions of M. cephalus along the Chinese coast. This research has provided new molecular data which will aid our understanding of the genetic structure of this species.     

Low levels of quantitative and molecular genetic differentiation among natural populations of Medicago ciliaris Kroch. (Fabaceae) of different Tunisian eco-geographical origin

In this paper, we analyze the genetic variability in four Tunisian natural populations of Medicago ciliaris using 19 quantitative traits and six polymorphic microsatellite loci. We investigated the amplification transferability of 30 microsatellites developed in the model legume M. truncatula to M. ciliaris. Results revealed that about 56.66% of analyzed markers are valuable genetic markers for M. ciliaris. The most genetic diversity at quantitative traits and microsatellite loci was found to occur within populations (>80%). Low differentiations among populations at quantitative traits Q _ST  = 0.146 and molecular markers F _ST  = 0.18 were found. The majority of measured traits exhibited no significant difference in the level of Q _ST and F _ST . Furthermore, significant correlations established between these traits and eco-geographical factors suggested that natural selection should be invoked to explain the level of phenotypic divergence among populations rather than drift. There was no significant correlation between population differentiation at quantitative traits and molecular markers. Significant spatial genetic structure consistent with models of isolation by distance was detected within all studied populations. The site-of-origin environmental factors explain about 9.07% of total phenotypic genetic variation among populations. The eco-geographical factors that influence more the variation of measured traits among populations are the soil texture and altitude. Nevertheless, there were no consistent pattern of associations between gene diversity (He) and environmental factors.     

Genetic variation within the endangered mangrove species Sonneratia paracaseolaris (Sonneratiaceae) in China detected by inter-simple sequence repeats analysis

Sonneratia paracaseolaris, is a critically endangered mangrove species in China. Using inter-simple sequence repeats (ISSR) markers, we compared the genetic variation of introduced populations with that of natural populations to check whether the genetic diversity has been conserved. At the species level, genetic diversity was relatively high (P = 81.37%, He = 0.2241, and SI = 0.3501). Genetic variation in introduced populations (P = 75.78%, He = 0.2291, and SI = 0.3500) was more than that in natural populations (P = 70.81%, He = 0.1903, and SI = 0.2980). Based on Nei's G_ST value, more genetic differentiation among natural populations was detected (G_ST = 0.3591). Our data show that the genetic diversity of S. paracaseolaris was conserved in introduced populations to some extent, however, owing to the small natural populations and the threats they encountered, more plants should be planted to enlarge and restore the populations.     

Genetic analyses of the federally endangered Echinacea laevigata using amplified fragment length polymorphisms (AFLP)—Inferences in population genetic structure and mating system

Echinacea laevigata (Boynton and Beadle) Blake is a federally endangered flowering plant species restricted to four states in the southeastern United States. To determine the population structure and outcrossing rate across the range of the species, we conducted AFLP analysis using four primer combinations for 22 populations. The genetic diversity of this species was high based on the level of polymorphic loci (200 of 210 loci; 95.24%) and Nei’s gene diversity (ranging from 0.1398 to 0.2606; overall 0.2611). There was significant population genetic differentiation (G_ST = 0.294; Ө^II = 0.218 from the Bayesian f = 0 model). Results from the AMOVA analysis suggest that a majority of the genetic variance is attributed to variation within populations (70.26%), which is also evident from the PCoA. However, 82% of individuals were assigned back to the original population based on the results of the assignment test. An isolation by distance analysis indicated that genetic differentiation among populations was a function of geographic distance, although long-distance gene dispersal between some populations was suggested from an analysis of relatedness between populations using the neighbor-joining method. An estimate of the outcrossing rate based on genotypes of progenies from six of the 22 populations using the multilocus method from the program MLTR ranged from 0.780 to 0.912, suggesting that the species is predominantly outcrossing. These results are encouraging for conservation, signifying that populations may persist due to continued genetic exchange sustained by the outcrossing mating system of the species.     

Genetic variation amongst viviparid snails in the genus Mekongia (Prosobranchia: Viviparidae) in Thailand

Allozyme variation in viviparid snails from the genus Mekongia in Thailand were examined across the different species, subspecies and geographical locations (river drainage systems). Using horizontal starch gel electrophoresis, 11 presumed allozyme loci (eight polymorphic) from eight enzyme systems were screened. Heterozygosity was moderately low (Hexp = 0.000–0.109, mean = 0.037). One population of Mekongia pongensis (Nong Khai) was monomorphic at all 11 examined loci in contrast to the other two populations, suggesting bottleneck within this population. Populations were more differentiated in the Mekongia sphaericula complex (F_ST = 0.587) than in either the Mekongia swainsoni complex (F_ST = 0.161) or M. pongensis species (F_ST = 0.073). Mekongia sphaericula sphaericula and Mekongia sphaericula extensa exhibited fixed allele differences at two loci, a high genetic distance (D = 0.265–0.300) and a potential polyphyletic relationship, suggesting two distinct lineages (species).     

Effects of landscape structure on genetic diversity of Geum urbanum L. populations in agricultural landscapes

Plant species in fragmented populations are affected by landscape structure because persistence within and migration among inhabited patches may be influenced by the identity and configuration of surrounding habitat elements. This may also be true for species of the semi-natural vegetation in agricultural landscapes. To determine the effect of landscape elements we analyzed Wood Avens (Geum urbanum L.) populations within three 4×4 km² agricultural landscapes in Germany, Switzerland and Estonia, which differ in levels of land use intensity and habitat fragmentation. Genetic variation was determined in 15 randomly selected populations in each landscape using 10 microsatellite loci. The landscape structure was assessed at two circles around each population, with radii defined by the range limits of spatial genetic autocorrelation. Multiple regression analysis was used to determine the influence of landscape structure variables for inter- and intrapopulation genetic diversity. Gene diversity was equally high in Germany (H_e=0.27) and Switzerland (H_e=0.26) but lower in Estonia (H_e=0.16). A high overall inbreeding coefficient (F_IS=0.89) was found, as expected for a selfing breeding system in G. urbanum. Genetic differentiation among populations was high (overall F_ST=0.43, 0.48, and 0.45 in Estonia, Switzerland and Germany, respectively), and did not differ among the three landscapes. Only a moderate influence of individual land use types on genetic diversity within and among populations was found with some idiosyncratic relationships. Genetic variation within populations was correlated to the amount of hedgerows positively in Estonia but negatively in Switzerland. The study demonstrates that the distribution of individual land use types affects the genetic pattern of a common plant species. However, different variables were identified to influence the genetic structure in three different landscapes. This indicates a major influence of landscape-specific land use history and stochastic processes determining gene flow and plant population structure.     

Drift happens: Molecular genetic diversity and differentiation among populations of jewelweed (Impatiens capensis Meerb.) reflect fragmentation of floodplain forests

Landscape features often shape patterns of gene flow and genetic differentiation in plant species. Populations that are small and isolated enough also become subject to genetic drift. We examined patterns of gene flow and differentiation among 12 floodplain populations of the selfing annual jewelweed (Impatiens capensis Meerb.) nested within four river systems and two major watersheds in Wisconsin, USA. Floodplain forests and marshes provide a model system for assessing the effects of habitat fragmentation within agricultural/urban landscapes and for testing whether rivers act to genetically connect dispersed populations. We generated a panel of 12,856 single nucleotide polymorphisms and assessed genetic diversity, differentiation, gene flow, and drift. Clustering methods revealed strong population genetic structure with limited admixture and highly differentiated populations (mean multilocus F_ST = 0.32, F_ST’ = 0.33). No signals of isolation by geographic distance or environment emerged, but alleles may flow along rivers given that genetic differentiation increased with river distance. Differentiation also increased in populations with fewer private alleles (R² = 0.51) and higher local inbreeding (R² = 0.22). Populations varied greatly in levels of local inbreeding (F_IS = 0.2–0.9) and F_IS increased in more isolated populations. These results suggest that genetic drift dominates other forces in structuring these Impatiens populations. In rapidly changing environments, species must migrate or genetically adapt. Habitat fragmentation limits both processes, potentially compromising the ability of species to persist in fragmented landscapes.     

Population structure and genetic diversity distribution in wild and cultivated populations of the traditional Chinese medicinal plant Magnolia officinalis subsp. biloba (Magnoliaceae)

Magnolia officinalis subsp. biloba, a traditional Chinese medicinal plant, experienced severe declines in the number of populations and the number of individuals in the late 20th century due to the widespread harvest of the subspecies. A large-scale cultivation program was initiated and cultivated populations rapidly recovered the loss in individual plant numbers, but wild populations remained small as a consequence of cutting. In this study, the levels of genetic variation and genetic structure of seven wild populations and five domestic populations of M. officinalis subsp. biloba were estimated employing an AFLP methodology. The plant exhibited a relatively high level of intra-population genetic diversity (h = 0.208 and H _j = 0.268). The cultivated populations maintained approximately 95% of the variation exhibited in wild populations, indicating a slight genetic bottleneck in the cultivated populations. The analysis of genetic differentiation revealed that most of the AFLP diversity resided within populations both for the wild group (78.22%) and the cultivated group (85.92%). Genetic differentiation among populations in the wild group was significant (F _ST = 0.1092, P < 0.005), suggesting wild population level genetic structure. Principal coordinates analysis (PCO) did not discern among wild and cultivated populations, indicating that alleles from the wild population were maintained in the cultivated gene pool. Results from the present study provide important baseline data for effectively conserving the genetic resources of this medicinal subspecies.     

Limited genetic diversity and high differentiation among the remnant adder (Vipera berus) populations in the Swiss and French Jura Mountains

Although the adder (Vipera berus) has a large distribution area, this species is particularly threatened in Western Europe due to high habitat fragmentation and human persecution. We developed 13 new microsatellite markers in order to evaluate population structure and genetic diversity in the Swiss and French Jura Mountains, where the species is limited to only a few scattered populations. We found that V. berus exhibits a considerable genetic differentiation among populations (global F_ST = 0.269), even if these are not geographically isolated. Moreover, the genetic diversity within populations in the Jura Mountains and in the less perturbed Swiss Alps is significantly lower than in other French populations, possibly due to post-glacial recolonisation processes. Finally, in order to minimize losses of genetic diversities within isolated populations, suggestions for the conservation of this species in fragmented habitats are proposed.     

Genetic diversity of fragmented populations of<Emphasis Type="Italic"> Polygala reinii</Emphasis> (Polygalaceae), a perennial herb endemic to Japan

Genetic variation at 10 allozyme loci was analyzed in 14 populations of Polygala reinii (Polygalaceae), a perennial herb endemic to central Honshu, Japan, with a fragmented geographical distribution. The levels of genetic variation within species (P=80.0, A=3.10, H_E=0.303) and within populations (P=42.1, A=1.61, H_E=0.163) were considerably higher than the mean for other endemic plants or short-lived perennial herbs. Genetic differentiation among populations was also high (G_ST=0.404). The genetic distance phenogram tended to show a clustering of the populations reflecting the fragmentation of the species range. A principal component analysis revealed the same tendency, as well as three groupings of populations in the Tokai district, on the Kii Peninsula and in the northern Kinki district. A negative correlation was obtained between the levels of gene flow and geographical distance among the populations (r=–0.745, P<0.0001). These results indicated limited gene flow among populations in P. reinii, presumably due to the geographical isolation accompanying the fragmented distribution. On the other hand, the geographical differentiation between the Japan Sea and Pacific Ocean sides was found in P. reinii, suggesting the influence of postglacial migration on the establishment of the genetic structure of this species.     

Genetic Diversity and Geographic Differentiation in the Threatened Species Dysosma pleiantha in China as Revealed by ISSR Analysis

Dysosma pleiantha, an important threatened medicinal plant species, is restricted in distribution to southeastern China. The species is capable of reproducing both sexually and asexually. In this study, inter-simple sequence repeat marker data were obtained and analyzed with respect to genetic variation and genetic structure. The extent of clonality, together with the clonal and sexual reproductive strategies, varied among sites, and the populations under harsh ecological conditions tended to have large clones with relatively low clonal diversity caused by vegetative reproduction. The ramets sharing the same genotype show a clumped distribution. Across all populations surveyed, average within-population diversity was remarkably low (e.g., 0.111 for Nei’s gene diversity), with populations from the nature reserves maintaining relatively high amounts of genetic diversity. Among all populations, high genetic differentiation (AMOVA: Φ_ST = 0.500; Nei’s genetic diversity: G _ST = 0.465, Bayesian analysis: Φ_B = 0.436) was detected, together with an isolation-by-distance pattern. Low seedling recruitment due to inbreeding, restricted gene flow, and genetic drift are proposed as determinant factors responsible for the low genetic diversity and high genetic differentiation observed.     

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.     

Genetic diversity of the black mangrove (Avicennia germinans L.) in Colombia

This study analyzed the genetic diversity and patterns of genetic structure in Colombian populations of Avicennia germinans L. using microsatellite loci. A lower genetic diversity was found on both the Caribbean (Ho = 0.439) and the Pacific coasts (Ho = 0.277) than reported for the same species in other locations of Central American Pacific, suggesting the deterioration of genetic diversity. All the populations showed high inbreeding coefficients (0.131–0.462) indicating heterozygotes deficience. The genetic structure between the Colombian coasts separated by Central American Isthmus was high (F_RT = 0.39) and the analyses of the genetic patterns of A. germinans revealed a clear differentiation of populations and no-recent gene flow evidence between coasts. Genetic structure was found within each coast (F_ST = 0.10 for the Caribbean coast and F_ST = 0.22 for the Pacific coast). The genetic patterns along the two coasts appear to reflect a forcing by local geomorphology and marine currents. Both coasts constitute a different Evolutionary Significant Unit, so we suggest for future transplantations plans that propagules or saplings of the populations of the Caribbean coast should not be mixed with those of the Pacific Colombian coast. Besides, we suggest that reforestation efforts should carefully distinguish propagules sources within each coast.     

Contemporary habitat loss reduces genetic diversity in an ecologically specialized butterfly

Aim This study investigated the influence of contemporary habitat loss on the genetic diversity and structure of animal species using a common, but ecologically specialized, butterfly, Theclinesthes albocincta (Lepidoptera: Lycaenidae), as a model. Location South Australia. Methods We used amplified fragment length polymorphism (AFLP) and allozyme datasets to investigate the genetic structure and genetic diversity among populations of T. albocincta in a fragmented landscape and compared this diversity and structure with that of populations in two nearby landscapes that have more continuous distributions of butterflies and their habitat. Butterflies were sampled from 15 sites and genotyped, first using 363 informative AFLP bands and then using 17 polymorphic allozyme loci (n = 248 and 254, respectively). We complemented these analyses with phylogeographic information based on mitochondrial DNA (mtDNA) haplotype information derived from a previous study in the same landscapes. Results Both datasets indicated a relatively high level of genetic structuring across the sampling range (AFLP, F_ST = 0.34; allozyme, F_ST = 0.13): structure was greatest among populations in the fragmented landscape (AFLP, F_ST = 0.15; allozyme, F_ST = 0.13). Populations in the fragmented landscape also had significantly lower genetic diversity than populations in the other two landscapes: there were no detectable differences in genetic diversity between the two continuous landscapes. There was also evidence (r² = 0.33) of an isolation by distance effect across the sampled range of the species. Main conclusions The multiple lines of evidence, presented within a phylogeographic context, support the hypothesis that contemporary habitat fragmentation has been a major driver of genetic erosion and differentiation in this species. Theclinesthes albocincta populations in the fragmented landscape are thus likely to be at greater risk of extinction because of reduced genetic diversity, their isolation from conspecific subpopulations in other landscapes, and other extrinsic forces acting on their small population sizes. Our study provides compelling evidence that habitat loss and fragmentation have significant rapid impacts on the genetic diversity and structure of butterfly populations, especially specialist species with particular habitat preferences and poor dispersal abilities.     

Genetic variation and population structure in Korean populations of sand dune speciesSalsola komarovi (Chenopodiaceae)

Salsola komarovi lljin is a herbaceous annual native to the sand dunes and beaches of Japan, northern China, Sakhalln and Korea. Starch-gel electrophoresis was conducted on leaves and stems collected from 300 plants in eight Korean populations. The mean number of alleles per locus (A _p=1.51), mean expected heterozygosity (He _p=0.116), and total genetic diversity (H _T=0.279) were comparable with those for species with similar life history and ecological traits. A general conformance of genotype frequencies to Hardy-Weinberg expectations (meanF _IS=−0.030) indicates thatS. komarovi is an outcrossing species. Slightly more than 20% of the genetic variation was found among populations (F _ST=0.204). In addition, significant differences in allele frequency were detected between populations at all 11 polymorphic loci (P<0.001). Nei's genetic identities range from 0.885 to 0.985 with a mean of 0.942. However, indirect estimates of the number of migrant per generation (0.97, calculated fromF _ST and 0.31, calculated from seven private alleles) indicate that the levels of gene flow is low among Korean populations. Although the species maintains a moderate level of genetic variation within populations, the small, isolated natural populations of the species have been severely destructed by human activities, particularly in summer season. If this is true, conservation efforts should be focused on those populations that currently maintain the most genetic diversity (e.g., populations of Cheju Island and coast of the southwestern Korean Peninsula).     

Concordance between life history traits, invasion history, and allozyme diversity of the Everglades invader Melaleuca quinquenervia

During the century following its initial introduction in 1886, the Australian tree Melaleuca quinquenervia (Myrtaceae) dispersed from a few introduction points to occupy over 200,000 ha, primarily in historic Everglades wetlands of southern Florida. Cellulose acetate gel electrophoresis (CAGE) was used to investigate the allozyme diversity and population genetic structure of 208 individuals in a dozen populations resulting from this invasion. The analyses showed that these populations have a high (82%) rate of polymorphic loci and an average of 2 alleles/locus. There was substantial heterozygosity (mean H_e = 0.356), which concords well with recent studies reporting a greater number of introduction events and sources than generally recognized. The introduction history and distributional patterns within Florida have led to geographic structuring (G_ST = 0.419) in which the Gulf Coast metapopulation has a greater effective number of alleles and greater heterozygosity than the Atlantic Coast metapopulation. The gene diversity in M. quinquenervia was comparable to other tropical woody species. Its strong population divergence was reminiscent of pioneer species and consistent with its status as a plant invader in Florida.     

Genetic structure of the Eurasian lynx population in north-eastern Poland and the Baltic states

We analyzed the genotypes of Eurasian lynx (Lynx lynx) from three populations in the westernmost part of the species main range. One population was situated at the distribution edge (NE Poland) and the two other (Latvia and Estonia) were located within the main, contiguous range of the species. The aim was to determine if the genetic composition varied among these populations and if there was evidence of isolation among them. Based on microsatellite allele frequencies, we found the allelic richness in Polish lynx to be lower than that in lynx from Latvia and Estonia. We also found significant differentiation among the lynx populations, with the NE Poland population forming a distinct genetic group relative to the two other populations (R _ST = 0.15 and 0.22, P < 0.0001). We suggest that genetic differentiation among lynx populations is the result of habitat insularisation that limits gene flow. This finding emphasizes the necessity to consider the lynx genetic differentiation in conservation planning of this species in Poland.     

Genetic structure of wild populations of the endangered Desert Pupfish complex (Cyprinodontidae: Cyprinodon)

Data from 10 microsatellite DNA loci were used to describe the genetic structure of the two extant species (Cyprinodon macularius and C. eremus) of the endangered Desert Pupfish complex of southwestern United States and northwestern Mexico. Variation at microsatellite loci was significantly correlated (Mantel test) with that of previous mtDNA results, both for the complex and for the relatively wide-ranging C. macularius alone. Both species showed unusually high levels of microsatellite diversity for non-marine fish (H _e = 0.84–0.93; A_R = 11.9–17.0). There was evidence (R _ST > F _ST) that the two extant populations of C. eremus have been isolated sufficiently long for mutation to contribute significantly to genetic divergence, whereas divergence among the nine assayed populations of C. macularius could be attributed to genetic drift alone. Correspondingly, 10% of the diversity in C. eremus was attributable to differences between the two populations, whereas, for C. macularius, only 2.7% was attributable to among-population variation. Within C. macularius, a small (0.8%), but statistically significant, portion was attributable to differences between populations in the Salton Sea area and those on the lower Colorado River delta. The two populations of C. eremus and five groups of populations of C. macularius are recommended as management units for conservation genetics management of the two species.     

Genetic differentiation between two varieties of Oreocharis benthamii (Gesneriaceae) in sympatric and allopatric regions

The pattern of genetic differentiation between diverging species receives much attention as one of the key observable features of speciation. It has often been suggested that introgression between closely related species occurs commonly where their distributions overlap, leading to their becoming more morphologically and genetically similar, but there are a few opposite results. However, most of these studies have been carried out with animals and separate species; few have looked at intraspecific cases, especially in plants. Here, we conduct a comparative study on patterns of genetic differentiation among populations of two varieties of Oreocharis benthamii in allopatry and sympatry based on ISSR data for 754 individuals from 26 populations, in order to understand the processes leading to speciation. Contrary to expectations, the facultative xenogamy (mixed mating) species O. benthamii has a relatively low genetic diversity within populations (H = 0.1014, I = 0.1528) and high genetic differentiation among populations (G_ST = 0.5867, Ф_ST = 0.659), as is typically found for selfing species. Genetic variance between the two varieties in sympatric populations (44%, Ф_ST = 0.444) is significantly more than that in allopatric populations (14%, Ф_ST = 0.138). Consistent with the taxonomical delimitation of the two varieties, all sampled individuals of O. benthamii clustered into two genetic groups. Moreover, the genetic structures of populations of both varieties are correlated with their different geographical origins. Our studies show that significant divergence between sympatric populations of the two varieties could be attributed primarily to reinforcement by genetic divergent selection in sympatry where secondary contact had occurred. The major proportion of the genetic variation in outcrossing and mixed mating plants may exist among populations when the populations are distributed in fragmented habitats, due to the paucity of suitable habitat combined with inefficient seed dispersal mechanism and limited pollinator foraging area that may limit the gene flow.