The origin of clonal diversity and structure of Populus alba in Sardinia: evidence from nuclear and plastid microsatellite markers

Background and Aims

Populus alba is a thermophilic forest tree present in the Mediterranean basin. Its habitat is highly fragmented and its distribution range has been subject to long-term human interference, resulting in debate surrounding whether certain populations are native or exotic in origin. In particular, populations from the islands of Corsica and Sardinia are of uncertain origin. While populations of P. alba mainly reproduce sexually, clonal reproduction is also common. The aims of this study were to locate and molecularly characterize the poorly studied island populations of P. alba and compare these with samples from various spatial scales, in order to provide information on the genetic structure and phylogeography of this species. This information will provide evidence on whether the species is native to Sardinia, which is important for the development of conservation strategies.

Methods

DNA extracts were obtained from the following P. alba trees: 159 from Sardinia, 47 from Ticino regional park (northern Italy), 15 acquired from an Italian Germoplasm Bank (IRC; Italian Reference Collection) and 28 from the Mediterranean basin (MB). Genetic polymorphisms were revealed at nuclear and chloroplast DNA (cpDNA) microsatellite loci, both at the island scale (Sardinia) and at broader scales, for comparative assessment of the genetic and genotypic diversity and phylogeography.

Key Results

Based on nuclear microsatellite loci, Sardinian white poplar consists of a small number of genets (26), each of which is represented by several ramets. Despite the uniqueness of the Sardinian haplotypes and the very low value of genetic diversity at the cpDNA level (v_K = 0·15), the H_T (0·60) and the A_R (3·61) values, estimated at the nuclear level for Sardinia, were comparable with those of the other populations and collections.

Conclusions

The uniqueness of the cpDNA haplotypes, the prevalence of clonality and the restricted number of genets recorded suggest that Sardinian white poplar could be a floristic relict of the native flora of the island, which has spread through available habitats on the island mainly by means of vegetative propagation and human activities.Key words: Populus alba, Sardinia, genets, ramets, phylogeography, native forest species     

Spatial genetic structure in the Laperrine's olive (Olea europaea subsp. laperrinei), a long-living tree from the central Saharan mountains

The Laperrine's olive (Olea europaea subsp. laperrinei) is an emblematic species of the Sahelo-Saharan Mountains. Populations of this tree are locally threatened by extinction due to climatic vicissitudes and human activities, particularly in Niger and Algeria. In order to study the spatial genetic structure and the dynamics of O. e. laperrinei populations, we sampled trees in four isolated mountain ranges (Tassili n'Ajjer and Hoggar (Algeria), Tamgak and Bagzane (Niger)). A total of 237 genets were identified using nuclear microsatellites. Phylogenetic reconstruction based on plastid DNA data supported a maternal origin of O. e. laperrinei populations in South Algeria, where a higher allelic richness was observed. Based on nuclear microsatellite data, two levels of structure were revealed: first, individuals from Niger and Algeria were separated in two distinct groups; second, four less differentiated clusters corresponded to the four studied mountain ranges. These results give support to the fact that desert barriers have greatly limited long distance gene flow. Within populations, pairwise kinship coefficients were significantly correlated to geographical distance for Niger populations but not for Algerian mountains. Historical factors and habitat heterogeneity may explain the differences observed. We conclude that the Hoggar acts as an important genetic reservoir that has to be taken into account in future conservation programmes. Moreover, very isolated endangered populations (for example, Bagzane) displaying evident genetic particularities have to be urgently considered for their endemism.     

On chloroplast DNA variations in the olive ( Olea europaea L.) complex: comparison of RFLP and PCR polymorphisms

Previous papers have dealt with olive chloroplastic DNA (cpDNA) variation revealed using several methods (RFLPs, PCR-RFLPs and microsatellites) and have led to different conclusions. This paper aims to reconsider these divergences. A Southern approach was applied to reveal polymorphism. We used chloroplast DNA of Phillyrea media as a probe. Based on these data, only four chlorotypes were identified in the olive complex. The number of detected lineages was lower than reported in the literature using a direct cpDNA RFLP approach, and was insufficient to distinguish the North African subspecies europaea, maroccana, guanchica and laperrinei. Furthermore, one individual considered belonging to the subspecies laperrinei was questionable. Using other cpDNA and mitochondrial DNA (mtDNA) polymorphisms - based on PCR and RFLP methods, respectively - we showed that this individual displays the cytoplasmic lineage CE1-ME1 characteristic of most Eastern mediterranean cultivars and of Olea europaea subsp. laperrinei from Hoggar. However, based on RAPDs, this individual appeared as mislabelled and probably corresponded to a Mediterranean cultivar or a feral form. In addition, we checked O. e. subsp. laperrinei herbarium samples using two cpDNA microsatellites, which revealed polymorphisms. These also supported that both populations from Niger and Algeria displayed a chlorotype related to CE1. Consequently, based on cpDNA, the relationships of O. e. subsp. laperrinei from Hoggar with a Mediterranean lineage appeared well supported, whereas the South West Moroccan and Macaronesian olives appeared in a different clade using both mtDNA and cpDNA polymorphisms. We conclude that methods based on PCR reveal more polymorphisms in the cpDNA and lead to more-reliable results that the classical RFLP method.     

Phylogeographic sampling guided by species distribution modeling reveals the Quaternary history of the Mediterranean–Canarian Cistus monspeliensis (Cistaceae)

Accurate inference in phylogeography requires appropriate sampling strategies. Complex questions demand a large sample size at both the population and genetic levels to obtain precise reconstructions. This is the case of the phylogeographic history of Cistus monspeliensis, a plant that displays low plastid (cpDNA) diversity in the Mediterranean Basin but high diversity in the Canary Islands. Here, we aimed to identify Mediterranean refugial areas and to accurately quantify inter‐island colonization events in the Canaries. Using a previous study as starting point, we increased sample size in two ways: (i) additional sampling of plastid genetic markers (from 1041 to 1899 bp); and (ii) additional sampling of populations (from 47 to 69) in long‐term persistence areas suggested by species distribution modeling (SDM). The synergy between SDM and extended population sampling helped find higher genetic diversity. Our deeper phylogeographic sampling of C. monspeliensis revealed the following: (i) potential refugia in long‐term persistence areas with high cpDNA diversity in western Europe and the Canary Islands; and (ii) a significant increase (from 7 to 12) in the number of inferred inter‐island colonization events across the archipelago. Our results stress the usefulness of SDM to identify the genetic signature associated with potential refugial areas. We herein propose a field sampling approach based on SDM that, in combination with a larger cpDNA sampling, can help answer a wide array of phylogeographic questions, such as the location of Quaternary refugia and number of colonizations across archipelagos.     

Population structure and genetic diversity in tristylous Narcissus triandrus: insights from microsatellite and chloroplast DNA variation

We investigated cpDNA sequence and nuclear microsatellite variation among populations of the wild daffodil Narcissus triandrus to examine the role of historical vs. contemporary forces in shaping population structure, morphological differentiation and sexual-system evolution. This wide-ranging heterostylous species of the Iberian Peninsula is largely composed of two allopatric varieties (vars. cernuus and triandrus), and populations with either stylar trimorphism or dimorphism. Dimorphic populations only occur in var. triandrus, are mainly restricted to the northwestern portion of the species range, and uniformly lack the mid-styled morph (M-morph). Chloroplast DNA (cpDNA) sequence variation revealed strong geographical structuring and evidence for a fragmentation event associated with differentiation of the two varieties. In var. triandrus, population fragmentation, restricted gene flow and isolation-by-distance were also inferred. Significant differences in genetic diversity and population structure between the two varieties likely reflect historical and contemporary differences in demography and gene flow among populations. Discordance between cpDNA markers and both microsatellite and morphological variation indicate that hybridization has occurred between the two varieties at contact zones. There were no differences in genetic diversity or population structure between dimorphic and trimorphic populations, and chloroplast haplotypes were not associated with either sexual system, indicating transitions in morph structure within each maternal lineage. M-morph frequencies were positively correlated with differentiation at microsatellite loci, indicating that the evolutionary processes influencing these neutral markers also influence alleles controlling the style morphs.     

Pollinating fig wasps: genetic consequences of island recolonization

The levels of genetic diversity and gene flow may influence the long-term persistence of populations. Using microsatellite markers, we investigated genetic diversity and genetic differentiation in island (Krakatau archipelago, Indonesia) and mainland (Java and Sumatra, Indonesia) populations of Liporrhopalum tentacularis and Ceratosolen bisulcatus, the fig wasp pollinators of two dioecious Ficus (fig tree) species. Genetic diversity in Krakatau archipelago populations was similar to that found on the mainland. Population differentiation between mainland coastal sites and the Krakatau islands was weak in both wasp species, indicating that the intervening 40 km across open sea may not be a barrier for wasp gene flow (dispersal) and colonization of the islands. Surprisingly, mainland populations of the fig waSPS may be more genetically isolated than the islands, as gene flow between populations on the Javan mainland differed between the two wasp species. Contrasting growth forms and relative 'immunity' to the effects of deforestation in their host fig trees may account for these differences.     

Random amplified polymorphic DNA assessment of diversity in western Mediterranean populations of the seagrass Posidonia oceanica

Posidonia oceanica is an endemic Mediterranean seagrass species that has often been assumed to contain low levels of genetic diversity. Random amplified polymorfic DNA (RAPD) markers were used to assess genetic diversity among five populations from three geographical regions (north, central, and south) of the western Mediterranean Sea. Stranded germinating seeds from one of the central populations were also included in the analysis. Forty-one putative genets were identified among 76 ramets based on 28 RAPD markers. Genotypic diversity strongly depended on the spatial structure, age, and maturity of the meadows. The lowest clonal diversity was found in the less structured and youngest prairies. Conversely, a high genotypic diversity was found in the highly structured meadows. The genotypic diversity in these meadows was at the same level as in P. australis and higher than previously reported data for P. oceanica populations in the Tyrrhenian Sea near the coast of Italy.     

Habitat loss other than fragmentation per se decreased nuclear and chloroplast genetic diversity in a monoecious tree

Generally, effect of fragmentation per se on biodiversity has not been separated from the effect of habitat loss. In this paper, using nDNA and cpDNA SSRs, we studied genetic diversity of Castanopsis sclerophylla (Lindl. & Paxton) Schotty populations and decoupled the effects of habitat loss and fragmentation per se. We selected seven nuclear and six cpDNA microsatellite loci and genotyped 460 individuals from mainland and island populations, which were located in the impoundment created in 1959. Number of alleles per locus of populations in larger habitats was significantly higher than that in smaller habitats. There was a significant relationship between the number of alleles per locus and habitat size. Based on this relationship, the predicted genetic diversity of an imaginary population of size equaling the total area of the islands was lower than that of the global population on the islands. Re-sampling demonstrated that low genetic diversity of populations in small habitats was caused by unevenness in sample size. Fisher's α index was similar among habitat types. These results indicate that the decreased nuclear and chloroplast genetic diversity of populations in smaller habitats was mainly caused by habitat loss. For nuclear and chloroplast microsatellite loci, values of F(ST) were 0.066 and 0.893, respectively, and the calculated pollen/seed dispersal ratio was 162.2. When separated into pre-and post-fragmentation cohorts, pollen/seed ratios were 121.2 and 189.5, respectively. Our results suggest that habitat loss explains the early decrease in genetic diversity, while fragmentation per se may play a major role in inbreeding and differentiation among fragmented populations and later loss of genetic diversity.     

Protecting evolutionary significant units for the remnant populations of <Emphasis Type="Italic">Berchemiella wilsonii</Emphasis> var. <Emphasis Type="Italic">pubipetiolata</Emphasis> (Rhamnaceae)

Berchemiella wilsonii var. pubipetiolata (Rhamnaceae) is an endangered tree in eastern China. Habitat destruction has resulted in fragmentation of remnant populations and extinction of local populations. AFLP and cpDNA markers were used to determine the population structure of remnant populations of B. wilsonii var. pubipetiolata. Moderate nuclear genomic diversity was found within each of the four remnant populations (H _S = 0.141–0.172), while the cpDNA haplotype diversity in each population ranged from 0.356 to 0.681. Six haplotypes were identified by a combined cpRFLP and cpSSR analysis in a total of 89 individuals. AMOVA revealed significantly AFLP genetic differentiation within and between regions (Φ_SC = 0.196, Φ_CT = 0.396, respectively), and a high cpDNA haplotype differentiation between regions (Φ_CT = 0.849). The results suggest low gene flow between populations of B. wilsonii var. pubipetiolata. Strong genetic divergence between two regional populations as revealed by both AFLP and cpDNA markers provided convincing evidence that two distinct evolutionary lineages existed, and should be recognized as ‘evolutionary significant units’ (ESUs) for conservation concerns.     

Pollen-mediated gene flow promotes low nuclear genetic differentiation among populations of <Emphasis Type="Italic">Cycas debaoensis</Emphasis> (Cycadaceae)

Cycas debaoensis is a critically endangered cycad species endemic to China. This species is found on two kinds of habitats according to the edaphic differences, sand and karst. A previous chloroplast DNA (cpDNA) study indicated that C. debaoensis had low genetic variation within populations and high genetic differentiation among populations. Because maternally inherited cpDNA does not fully characterize genetic structure of the species, we screened seven low-copy nuclear genes and 17 nuclear microsatellite loci to detect the nuclear genetic diversity, differentiation, and the population structure of C. debaoensis. The nuclear genes revealed higher level of genetic diversity. There were both the same and region-specific haplotypes or alleles between the karst and sand regions. Nuclear gene flow among all the populations was much greater than that of cpDNA, which indicated that pollen-mediated gene flow was much greater than seed-mediated gene flow. This promoted low nuclear genetic differentiation among populations of C. debaoensis. The study suggests that both genetic and anthropogenic disturbances have resulted in the critically endangered status of C. debaoensis.     

Is Cupressus sempervirens native in Italy? An answer from genetic and palaeobotanical data

This study represents the first large-scale analysis using nuclear molecular markers to assess genetic diversity and structure of Cupressus sempervirens L.. Genetic and fossil data were combined to infer the possible role of human activity and evolutionary history in shaping the diversity of cypress populations. We analysed 30 populations with six polymorphic nuclear microsatellite markers. Dramatic reductions in heterozygosity and allelic richness were observed from east to west across the species range. Structure analysis assigned individuals to two main groups separating central Mediterranean and eastern populations. The two main groups could be further divided into five subgroups which showed the following geographical distributions: Turkey with the Greek islands Rhodes and Samos, Greece (Crete), Southern Italy, Northern Italy, Tunisia with Central Italy. This pattern of genetic structure is also supported by samova and Barrier analyses. Palaeobotanical data indicated that Cupressus was present in Italy in the Pliocene, Pleistocene and Holocene. Furthermore, our molecular survey showed that Italian cypress populations experienced bottlenecks that resulted in reduced genetic diversity and allelic richness and greater genetic differentiation. Recent colonization or introduction may also have influenced levels of diversity detected in the Italian populations, as most individuals found in this range today have multilocus genotypes that are also present in the eastern range of the species. The data reveal a new interpretation of the history of cypress distribution characterized by ancient eastern populations (Turkey and Greek islands) and a mosaic of recently introduced trees and remnants of ancient, depauperate populations in the central Mediterranean range.     

As old as the hills: Pliocene palaeogeographical processes influence patterns of genetic structure in the widespread,common shrub Banksia sessilis

The impact of Quaternary glaciation on the development of phylogeographic structure in plant species is well documented. In unglaciated landscapes, phylogeographic patterns tend to reflect processes relating to persistence and stochasticity, yet other factors, associated with the palaeogeographical history of the landscape, including geomorphological events, can also have a significant influence. The unglaciated landscape of south‐western Western Australia is an ideal location to observe these ancient drivers of lineage diversification, with tectonic activity associated with the Darling Fault in the late Pliocene attributed to patterns of deep phylogeographic divergence in a widespread tree from this region. Interestingly, other species within this region have not shown this pattern and this palaeogeographical boundary therefore presents an opportunity to examine age and historical distribution of plant species endemic to this region. In this study, we assess patterns of genetic diversity and structure across 28 populations of the widespread shrub Banksia sessilis using three cpDNA markers and nine nuclear microsatellite markers. Sixteen cpDNA haplotypes were identified, comprising two major chloroplast DNA lineages that are estimated to have diverged in the Pliocene, approximately 3.3 million years ago. This timing coincides with major geomorphological processes in the landscape, including the separation of the Darling Plateau from the adjacent Swan Coastal Plain, as well as eustatic changes on the Swan Coastal Plain that are likely to have resulted in the physical isolation of historical plant lineages. Chloroplast lineages were broadly aligned with populations associated with older lateritic soils of the Darling Plateau and Geraldton sandplains or the younger sandy soils associated with the Swan Coastal Plain and Southern Coastline. This structural pattern of lateritic versus non‐lateritic division was not observed in the nuclear microsatellite data that identified three genetic clades that roughly corresponded to populations in the North, South, and Central portions of the distributions.     

Population structure and colonization history of the olive fly, Bactrocera oleae (Diptera, Tephritidae)

The olive fly, Bactrocera oleae, is the major pest of olives in most commercial olive-growing regions worldwide. The species is abundant in the Mediterranean basin and has been introduced recently into California and Mexico, creating problems for quarantine protection and international trade. Here, we use nuclear microsatellite markers and mitochondrial sequences to examine the history of olive fly range expansion and colonization. Sampled populations span the current distribution of the olive fly worldwide, including South and Central Africa, Pakistan, Mediterranean Europe and Middle East, California, and Mexico. The Pakistani populations appear to be genetically well differentiated from the remaining populations, though rooting the origins of the species is problematic. Genetic similarity and assignment tests cluster the remaining populations into two genetic groups--Africa and a group including the Mediterranean basin and the American region. That Africa, and not the Mediterranean, is the origin of flies infesting cultivated olive is supported by the significantly greater genetic diversity at microsatellite loci in Africa relative to the Mediterranean area. The results also indicate that the recent invasion of olive flies in the American region most likely originated from the Mediterranean area.     

Development of microsatellite markers from Cercis chinensis (Fabaceae)

? Premise of the study: Microsatellite markers were developed to characterize the level of genetic diversity and population genetic structure of Cercis chinensis, a widely cultivated garden plant in China with congeneric species disjunctly distributed in East Asia, North America, and the Mediterranean. ? Methods and Results: Using the Fast Isolation by AFLP of Sequences COntaining repeats (FIASCO) protocol, eight microsatellite markers were developed in C. chinensis. Seven of the markers displayed polymorphism, with the number of alleles ranging from one to four in four populations of C. chinensis. Four to six microsatellite loci exhibited interspecific transferability in C. glabra, C. chuniana, and C. chingii. ? Conclusions: These are the first microsatellite markers developed for C. chinensis, which will be further used in investigation of population genetic structure and phylogeographic pattern of C. chinensis and its congeneric species.     

The coarse‐root system of mature Populus tremuloides in declining stands in Alberta,Canada

Abstract. The coarse‐root dynamics of ramets of Populus tremuloides (aspen) were investigated with respect to persistence of the original root connections (roots of parent trees from which the ramets originated), the time of establishment of new roots at the base of the stem and the fate of the communal root system after death of individual trees. Parts of the root systems of three declining stands of aspen ramets were hydraulically excavated. From each stand, sections of all structural roots were collected at the base of live and dead trees and were analysed using dendrochronology techniques. Parent roots were identified in the root system of every tree. The trees initiated new structural roots shortly after suckering. Live roots were often connected to the stump of dead and decayed trees. Grafting was common, especially at or near the stumps. Death of trees along the parent roots over time did not seem to favour the entry of significant decay, nor promote breakage of the original root connections. Instead of becoming independent of the parent root system the ramets incorporated the parent roots into their own root systems, remaining interconnected.     

Chloroplast DNA variation and phylogeography of Ligularia tongolensis (Asteraceae), a species endemic to the Hengduan Mountains region of China

In this research, we aimed to study the genetic variation and phylogeographic pattern of Ligularia tongolensis, a perennial herb endemic to the Hengduan Mountains region of China. We sequenced two chloroplast DNA (cpDNA) intergenic spacers (trnQ-5 rps16, trnL-rpl32) in 140 individuals from 14 populations of three groups (Jinshajiang vs. Yalongjiang vs. Wumeng) within this species range. High levels of haplotype diversity (Hd = 0.814) and total genetic diversity (Ht = 0.862) were detected at the species leve...     

Environmental Heterogeneity Explains the Genetic Structure of Continental and Mediterranean Populations of Fraxinus angustifolia Vahl

Tree species with wide distributions often exhibit different levels of genetic structuring correlated to their environment. However, understanding how environmental heterogeneity influences genetic variation is difficult because the effects of gene flow, drift and selection are confounded. We investigated the genetic variation and its ecological correlates in a wind-pollinated Mediterranean tree species, Fraxinus angustifolia Vahl, within a recognised glacial refugium in Croatia. We sampled 11 populations from environmentally divergent habitats within the Continental and Mediterranean biogeographical regions. We combined genetic data analyses based on nuclear microsatellite loci, multivariate statistics on environmental data and ecological niche modelling (ENM). We identified a geographic structure with a high genetic diversity and low differentiation in the Continental region, which contrasted with the significantly lower genetic diversity and higher population divergence in the Mediterranean region. The positive and significant correlation between environmental and genetic distances after controlling for geographic distance suggests an important influence of ecological divergence of the sites in shaping genetic variation. The ENM provided support for niche differentiation between the populations from the Continental and Mediterranean regions, suggesting that contemporary populations may represent two divergent ecotypes. Ecotype differentiation was also supported by multivariate environmental and genetic distance analyses. Our results suggest that despite extensive gene flow in continental areas, long-term stability of heterogeneous environments have likely promoted genetic divergence of ashes in this region and can explain the present-day genetic variation patterns of these ancient populations.     

Population structure,seasonal genotypic differentiation,and clonal diversity of weedy dandelions in three Boston area populations (Taraxacum sp.)

Weedy dandelions have a worldwide distribution and thrive in urban environments despite a lack of sexual reproduction throughout most of its range. North American dandelions, introduced from Eurasia, are believed to be primarily, if not exclusively, apomictic triploids. In some European populations, apomicts co‐occur with diploid sexual individuals and hybridizations can create genetically unique apomicts, which may subsequently disperse and establish new populations globally. Using six nuclear microsatellite markers and a cpDNA intergenic spacer, we investigate the impact of this unusual natural history on population structure and diversity in three urban Boston area dandelion populations. Our results show high levels of genetic diversity within populations, spatial population structure, and seasonal genotypic differentiation in flowering times. We find evidence that sexual reproduction and recombination, presumably in Europe, and extensive gene flow drive these patterns of diversity and create the appearance of panmixia despite the lack of evidence for local sexual reproduction.     

Structure of Pinus nigra Arn. populations in Bulgaria revealed by chloroplast microsatellites and terpenes analysis: Provenance tests

Pinus nigra is a forest and low elevation mountain species found around the Mediterranean Sea that has had its distribution reduced and fragmented by anthropogenic disturbance. Due to commercial interest it is currently being replanted, however, the genetic structure of populations is little known and current planting strategies could threaten its genetic diversity. In the present study we investigated the genetic structure and genetic diversity of P. nigra populations in Bulgaria using chloroplast microsatellite markers and terpene analysis. Nine provenances were chosen throughout the species' range in Bulgaria. Following DNA extraction, chloroplast microsatellite (cpSSR) loci were surveyed using three primer pairs. Between 5 and 9 size variants were identified at each locus. A total of 22 size variants at the 3 loci were identified, that were combined in 68 different haplotypes, of which 7 represent 39.81% of the genetic structure. AMOVA analysis revealed that 6.06% of the variation was found among populations, while 93.94% was expressed within populations. The cpSSR analysis divided European Black pine populations into four groups, the first represented by populations located the eastern Rhodopes, Sr. Gora and St. Planina mountains, while the second group is primarily located in the Phodopes and Slavianca mountains. The populations from Pirin and Osogovo mountains show different genetic patterns. Terpene analysis revealed that most of the monoterpene pool in P. nigra was accounted for by α-pinene followed by β-pinene. The presence of four distinct terpene groups is not consistent with physical distances between populations, and a similar non-significant correlation between genetic distance determined by chloroplast microsatellites analysis and chemotype distance (determined by terpenes) was observed. Our results suggest that the structural pattern of genetic diversity of cpDNA in European Black pine populations is the consequence of historical biogeographic processes.     

Genetic structure of island populations of <Emphasis Type="Italic">Prunus lannesiana</Emphasis> var. <Emphasis Type="Italic">speciosa</Emphasis> revealed by chloroplast DNA,AFLP and nuclear SSR loci analyses

The wild flowering cherry Prunus lannesiana var. speciosa is highly geographically restricted, being confined to the Izu Islands and neighboring peninsulas in Japan. In an attempt to elucidate how populations of this species have established we investigated the genetic diversity and differentiation in seven populations (sampling 408 individuals in total), using three kinds of genetic markers: chloroplast DNA (cpDNA), amplified fragment length polymorphisms (AFLPs), and 11 nuclear SSR polymorphic loci. Eight haplotypes were identified based on the cpDNA sequence variations, 64 polymorphic fragments were scored for the AFLP markers, and a total of 154 alleles were detected at the 11 nuclear SSR loci. Analysis of molecular variance showed that among-population variation accounted for 16.55, 15.04 and 7.45% of the total detected variation at the cpDNA, AFLPs, and SSR loci, respectively. Thus, variation within populations accounted for most of the genetic variance for all types of markers, although the genetic differentiation among populations was also highly significant. For cpDNA variation, no clear structure was found among the populations, except that of the most distant island, although an “isolation by distance” pattern was found for each marker. Both neighbor-joining trees and structure analysis indicate that the genetic relationships between populations reflect geological variations between the peninsula and the islands and among the islands. Furthermore, hybridization with related species may have affected the genetic structure, and some genetic introgression is likely to have occurred.