Intraspecific divergences of Rhodiola alsia (Crassulaceae) based on plastid DNA and internal transcribed spacer fragments

In the present study, we used two maternally inherited plastid DNA intergenic spacers, rpl20‐rps12 and trnS‐trnG, and the biparentally inherited nuclear ribosomal internal transcribed spacer (ITS) region to explore genetic variation and phylogeographical history of Rhodiola alsia, a herb endemic to the Qinghai‐Tibetan Plateau (QTP). Based on range‐wide sampling (18 populations and 227 individuals), we detected 45 plastid DNA haplotypes and 19 ITS sequence types. Only three plastid DNA haplotypes were widespread; most haplotypes were restricted to single sites or to neighbouring populations. Analysis of molecular variance revealed that most of the genetic variance was found within populations (51.24%) but that populations were also distinct (F_ST = 0.48759). We found three areas with relatively high plastid DNA diversity and these could further be recognized as potentially isolated divergence centres based on the ITS sequence type distribution. These represent three potentially isolated glacial refugia for R. alsia: one of them has long been recognized as an important refugium on the south‐eastern edge of the QTP, whereas the others are new and located in the north and south of the Tanggula Mountains on the plateau platform. Divergence time estimates based on ITS suggest that the main lineages of R. alsia diverged from each other 0.35–0.87 Mya, indicating that climatic oscillations during the Pleistocene may have been an important driver of intraspecific divergence in R. alsia. Rhodiola alsia probably experienced a phylogeographical history of retreat to isolated glacial refugia during Quaternary glaciations that led to different degrees of allopatric intraspecific divergence. © 2011 The Linnean Society of London, Botanical Journal of the Linnean Society, 2012, 168 , 204–215.     

Multiple introgression events and range shifts in Schizocodon (Diapensiaceae) during the Pleistocene

Range shifts during the Pleistocene shaped the unique phylogeographical structures of numerous species. Accompanying species migration, sister taxa may have experienced multiple introgression events. Here, we report the signature of introgression events in multiple areas in Schizocodon, herbs endemic to Japan, using amplified fragment length polymorphism (AFLP) fingerprinting and plastid DNA haplotyping in 48 populations. Although the present distributions of S. soldanelloides and S. ilicifolius are mainly allopatric, the species share plastid DNA haplotypes in each region (north‐eastern, north‐central, south‐central and south‐western Japan); in contrast, the specific groups were highly supported by AFLP analyses. These results support the occurrence of multiple introgression events in Schizocodon. Notably, the disjunct plastid haplotypes found only in S. ilicifolius var. intercedens suggest complete plastid DNA replacement at local areas from S. soldanelloides into S. ilicifolius var. ilicifolius. Furthermore, we found that S. soldanelloides experienced range contraction and expansion during glacial and interglacial cycles based on mismatch distribution analysis and ecological niche modelling. Based on several pieces of evidence, our study supports the idea that historical range shifts associated with Pleistocene climatic oscillations favoured multiple and regional introgression events in Schizocodon. © 2013 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 173 , 46–63.     

Phylogeography of the genus Dasiphora (Rosaceae) in the Qinghai‐Tibetan Plateau: divergence blurred by expansion

Plateau uprisings and climatic oscillations are considered to have caused extensive allopatric divergences that account for the rich species diversity of the Qinghai‐Tibetan Plateau (QTP). However, secondary contact during range shifts in the Quaternary glacial cycles or inter‐uplift stages may have restored the gene flow between species and so counteracted these divergences, particularly in rapidly‐adapting dominant elements. We tested this hypothesis by determining the phylogeographical history of Dasiphora (Rosaceae), a genus of two species that are widely distributed on the QTP and co‐exist in numerous localities. We sequenced two chloroplast DNA fragments (rbcL, trnT‐L) for 559 individuals from 87 populations. Bayesian methods were used to identify phylogenetic relationships and to estimate divergence times. Demographic histories were inferred using neutrality tests, mismatch distribution analysis, and coalescent simulation. A total of 112 haplotypes that clustered into three major groups were identified. The formation of these groups and their subgroups was dated to between the Pliocene and the late Pleistocene. In addition, we found that some groups underwent multiple extensive expansions. Species‐specific haplotypes were identified for each species, although these haplotypes phylogenetically intermixed. These results suggest that recent plateau uplifts and climatic oscillations might have caused the deep divergences observed within this genus. However, later range expansions probably blurred these divergences and possible species boundaries. Our results shed new light on the complex evolutionary history of the QTP alpine plants. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 111 , 777–788.     

Geographically congruent large‐scale patterns of plastid haplotype variation in the European herbs Silene dioica and S. latifolia (Caryophyllaceae)

The closely related dioecious herbs Silene latifolia and Silene dioica are widespread and predominantly sympatric in Europe. The species are interfertile, but morphologically and ecologically distinct. A study of large‐scale patterns of plastid DNA (polymerase chain reaction–restriction fragment length polymorphism) haplotypes in a sample of 198 populations from most of the European ranges of both species revealed extensive interspecific haplotype sharing. Four of the 28 detected haplotypes were frequent (found in > 40 populations) and widespread. Three of these frequent haplotypes occurred in both species and the geographic distribution of each haplotype was broadly congruent in both species. Each of these three, shared and widespread haplotypes is likely to have colonized central and/or northern Europe after the last glaciation from one or more of refugial areas in southern Europe. Interspecific hybridization and plastid introgression within refugial regions and/or during the early stages of postglacial expansion is the most plausible explanation for the broadly similar distribution patterns of the shared, frequent chloroplast haplotypes in the two species. The fourth frequent, widespread haplotype was absent from S. latifolia and almost entirely restricted to Nordic S. dioica. It is most likely that this haplotype spread into the Nordic countries from a central or northern European source or from a refugial area in Russia. © 2009 The Linnean Society of London, Botanical Journal of the Linnean Society, 2009, 161 , 153–170.     

Mosaic variation in allozyme and plastid DNA markers in the European ranges of Silene vulgaris and its partially sympatric relative S. uniflora (Caryophyllaceae)

The perennial herbs Silene vulgaris and S. uniflora are closely related, partially sympatric and interfertile, yet morphologically distinct. We used nuclear (allozyme) and plastid (polymerase chain reaction–restriction fragment length polymorphism) DNA markers to investigate whether these species have a shared history of postglacial colonization and hybridization in Europe, as inferred from large‐scale patterns of geographic variation. The majority of plastid haplotypes and allozyme alleles were widespread and patchily distributed within both species and there was no geographic structure in the distributions of shared allozymes or haplotypes. The mosaic variation is consistent with a scenario in which repeated episodes of interspecific hybridization pre‐dated the largely allopatric range expansion of the two species during the postglacial period. Our overall results are not consistent with a scenario of extensive hybridization and introgression during the postglacial range expansion of the species or within their current areas of sympatry, but we found some evidence for local, postglacial evolution and hybridization in the Baltic region. © 2011 The Linnean Society of London, Botanical Journal of the Linnean Society, 2011, 166 , 127–148.     

Genetic diversity,genetic structure and phylogeography of the Iberian endemic Gypsophila struthium (Caryophyllaceae) as revealed by AFLP and plastid DNA sequences: connecting habitat fragmentation and diversification

Iberian gypsum outcrops are highly fragmented and ecologically challenging environments for plant colonization. As gypsophytes occur exclusively in such habitats, they are ideal models for the study of both the effects of habitat fragmentation and selection on population genetic diversity and structure. In this study, we used amplified fragment length polymorphism (AFLP) and plastid DNA sequences to investigate the phylogeographical history of the Iberian plant Gypsophila struthium (Caryophyllaceae), a widespread endemic restricted to Iberian gypsum outcrops. Gypsophila struthium consists of two subspecies that differ in the architecture of their inflorescence and have mostly allopatric ranges. Gypsophila struthium subsp. struthium occurs in central, eastern and south‐eastern Iberia, whereas G. struthium subsp. hispanica occurs in northern and eastern areas. AFLPs revealed low but significant genetic differentiation between the subspecies, probably as a result of a recent diversification during the Pliocene–Pleistocene. In the geographical contact zone between the taxa, the Bayesian analyses revealed populations with mixed ancestries and genetic clusters predominantly of one or the other subspecies, indicating incomplete reproductive barriers between them. Plastid DNA haplotypes revealed strong geographical structure and testified to processes of isolation by distance and continuous range expansion for some haplotype clades. The Bayesian analyses of the population structure of AFLP data and nested clade phylogeographical analysis (NCPA) of plastid haplotypes revealed that the putative ancestral range corresponded to central and eastern populations of G. struthium subsp. struthium, with those lineages contributing through more recent expansion to increased genetic diversity and structure of the south‐eastern and eastern ranges of this subspecies and to the diversification of G. struthium subsp. hispanica in northern and eastern gypsum outcrops. © 2013 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 173 , 654–675.     

Phylogeography of the Qinghai–Tibet Plateau endemic alpine herb Pomatosace filicula (Primulaceae)

In order to trace the response of alpine plants on the Qinghai–Tibet Plateau (QTP) to the Quaternary climate oscillations, the phylogeographic history of Pomatosace filicula Maxim. was investigated in the present study. Based on sequence variations of two maternally inherited plastid markers, matK and trnH-psbA, and the biparentally inherited nuclear ribosomal internal transcribed spacer (nrITS), we estimated the population genetic structure, lineage divergence timescale, and population dynamics of P. filicula. Seven plastid haplotypes and two nrITS genotypes were identified across the range-wide sampling of 200 individuals from 24 populations. Although AMOVA revealed a high level of differentiation among populations (F_ST = 0.560), no significant phylogeographic structure was detected (N_ST = 0.503, G_ST = 0.518, P > 0.05). Molecular dating suggested that the divergences between major plastid lineages and nrITS genotypes occurred during the early and middle Pleistocene, and the middle Pleistocene, respectively. This species most likely survived at multiple unglaciated sites on the QTP during the Last Glacial Maximum, with most of these sites located above 4000 m a.s.l. The species probably experienced range expansion at its distribution fringe, but demographic tests did not suggest significant population size changes. We proposed that pronounced effective gene flow (N_em = 0.393) and short generation time may have obscured the phylogeographic and demographic patterns of this species. Our findings will shed new light on the Quaternary evolutionary history of the alpine flora of the QTP.     

Intraspecific molecular phylogeny and phylogeography of the Meriones meridianus (Rodentia: Cricetidae) complex in northern China reflect the processes of desertification and the Tianshan Mountains uplift

The phylogeographical patterns and demographic history of mitochondrial DNA (cytochrome b, N = 327; D‐loop, N = 252) and nuclear DNA (IRBP gene, N = 235) haplotypes were studied for the Meriones meridianus complex in northern China, a desert‐dwelling gerbil species complex. The phylogenetic analyses, which were performed on the separate and combined (mitochondrial + nuclear) datasets, revealed two divergent clades (Clade A and Clade B) corresponding to distinct geographical regions. Clade A contained the haplotypes found mostly in individuals from the Tianshan Mountains area. Clade B contained haplotypes from populations located in other deserts in northern China. The divergence times indicated that the history of the M. meridianus complex was influenced by the uplift of the Tianshan Mountains and climate‐induced habitat fluctuations. In the Pleistocene, the expansion of forests and grasslands during interglacial period led to the isolation of the M. meridianus complex, which preferred to inhabit deserts. Hence, long geological isolation and the M. meridianus complex adaptation to local ecological conditions led to its genetic divergence. Clade A had long‐lasting demographic stability, most likely because the populations of this clade remained in a stable desert environment for a long time. However, the extension of other deserts and disappearance of palaeolakes during the last glacial period resulted in demographic expansion of Clade B. Furthermore, our genetic data indicated that two subspecies may exist within the M. meridianus complex. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 110 , 362–383.     

Phylogeographic study of Chinese seabuckthorn (Hippophae rhamnoides subsp. sinensis Rousi) reveals two distinct haplotype groups and multiple microrefugia on the Qinghai‐Tibet Plateau

Historical climate change can shape the genetic pattern of a species. Studies on this phenomenon provide great advantage in predicting the response of species to current and future global climate change. Chinese seabuckthorn (Hippophae rhamnoides subsp. sinensis) is one of the most important cultivated plants in Northwest China. However, the subspecies history and the potential genetic resources within the subspecies range remain unclear. In this study, we utilized two intergenic chloroplast regions to characterize the spatial genetic distribution of the species. We found 19 haplotypes in total, 12 of which were unique to the Chinese seabuckthorn. The populations observed on the Qinghai‐Tibet Plateau (QTP) consisted of most of the haplotypes, while in the northeast of the range of the subspecies, an area not on the QTP, only four haplotypes were detected. Our study also revealed two distinct haplotype groups of the subspecies with a sharp transition region located in the south of the Zoige Basin. 89.96% of the genetic variation located between the regions. Mismatch analysis indicated old expansions of these two haplotype groups, approximately around the early stage of Pleistocene. Additional morphological proofs from existing studies and habitat differentiation supported a long independent colonization history among the two regions. Potential adaptation probably occurred but needs more genome and morphology data in future. Chinese seabuckthorn have an older population expansion compared with subspecies in Europe. The lack of large land ice sheets and the heterogeneous landscape of the QTP could have provided extensive microrefugia for Chinese seabuckthorn during the glaciation period. Multiple localities sustaining high‐frequency private haplotypes support this hypothesis. Our study gives clear insight into the distribution of genetic resources and the evolutionary history of Chinese seabuckthorn.     

The Quaternary evolutionary history,potential distribution dynamics,and conservation implications for a Qinghai–Tibet Plateau endemic herbaceous perennial,Anisodus tanguticus (Solanaceae)

Various hypotheses have been proposed about the Quaternary evolutionary history of plant species on the Qinghai–Tibet Plateau (QTP), yet only a handful of studies have considered both population genetics and ecological niche context. In this study, we proposed and compared climate refugia hypotheses based on the phylogeographic pattern of Anisodus tanguticus (three plastid DNA fragments and nuclear internal transcribed spacer regions from 32 populations) and present and past species distribution models (SDMs). We detected six plastid haplotypes in two well‐differentiated lineages. Although all haplotypes could be found in its western (sampling) area, only haplotypes from one lineage occurred in its eastern area. Meanwhile, most genetic variations existed between populations (F_ST = 0.822). The SDMs during the last glacial maximum and last interglacial periods showed range fragmentation in the western area and significant range contraction in the eastern area, respectively, in comparison with current potential distribution. This species may have undergone intraspecific divergence during the early Quaternary, which may have been caused by survival in different refugia during the earliest known glacial in the QTP, rather than geological isolation due to orogenesis events. Subsequently, climate oscillations during the Quaternary resulted in a dynamic distribution range for this species as well as the distribution pattern of its plastid haplotypes and nuclear genotypes. The interglacial periods may have had a greater effect on A. tanguticus than the glacial periods. Most importantly, neither genetic data nor SDM alone can fully reveal the climate refugia history of this species. We also discuss the conservation implications for this important Tibetan folk medicine plant in light of these findings and SDMs under future climate models. Together, our results underline the necessity to combine phylogeographic and SDM approaches in future investigations of the Quaternary evolutionary history of species in topographically complex areas, such as the QTP.     

Multiple autopolyploidizations and range expansion of Allium przewalskianum Regel. (Alliaceae) in the Qinghai‐Tibetan Plateau

We used the Allium przewalskianum diploid–tetraploid complex on the Qinghai‐Tibetan Plateau (QTP) as a model to examine how this complex responded to the Quaternary climatic oscillations, and whether multiple autopolyploidizations have occurred. We sequenced five chloroplast DNA (cpDNA) fragments (accD‐psaI, trnH‐psbA, trnL‐trnF, trnS‐trnG and rpl16‐intron) in 306 individuals (all of known ploidy level) from 48 populations across the distribution of this species complex. We identified a total of 32 haplotypes—11 in diploids only, 13 in tetraploids only, and 8 found in both cytotypes. This, plus network analyses, indicated that tetraploids have arisen independently from diploids at least eight times. Most populations in the eastern QTP contained multiple haplotypes, but only a single haplotype was found for 17 tetraploid populations on the western QTP, suggesting a recent colonization of the western QTP. We further found that this species complex underwent an earlier range expansion around 5–150 thousand years ago (kya), after the largest glacial period (800–170 kya) in the QTP. In addition, the high frequencies of tetraploids in the QTP suggested that the tetraploid A. przewalskianum cytotype has evolutionary advantages over diploids in colonizing and/or surviving the arid habitats of the QTP.     

Contrasting responses to Pleistocene climate changes: a case study of two sister species Allium cyathophorum and A. spicata (Amaryllidaceae) distributed in the eastern and western Qinghai–Tibet Plateau

It has been hypothesized that species occurring in the eastern and the western Qinghai–Tibet Plateau (QTP) responded differently to climate changes during the Pleistocene. Here, we test this hypothesis by phylogeographic analysis of two sister species, Allium cyathophorum and A. spicata. We sequenced two chloroplast DNA (cpDNA) fragments (accD‐psaI and the rpl16 intron) of 150 individuals, and the nuclear (ITS) region of 114 individuals, from 19 populations throughout the distributional ranges of these species. The divergence between the two species was dated at 779 ‐ 714 thousand years before the present and was likely initiated by the most major glaciation in the QTP. Analysis of chlorotype diversity showed that A. spicata, the species occurring in the western QTP, contains much lower genetic diversity (0.25) than A. cyathophorum (0.93), which is distributed in the eastern QTP. Moreover, multiple independent tests suggested that the A. spicata population had expanded recently, while no such expansion was detected in A. cyathophorum, indicating a contrasting pattern of responses to Pleistocene climate changes. These findings highlight the importance of geographical topography in determining how species responded to the climate changes that took place in the QTP during the Pleistocene.     

Extensive population expansion of Pedicularis longiflora (Orobanchaceae) on the Qinghai‐Tibetan Plateau and its correlation with the Quaternary climate change

The Qinghai‐Tibetan Plateau (QTP) is thought to be more strongly affected by the Quaternary glaciations than most other regions of the same latitude. It would be of great interest to investigate the population genetic structure of organisms distributed on the platform and its correlation with the Quaternary climatic oscillations. Here we used the chloroplast (cp)DNA trnT‐trnF sequence to study genetic variation and phylogeography of Pedicularis longiflora, an alpine herb with extensive distribution on the QTP. Based on a range‐wide sampling comprising 41 populations and 910 individuals, we detected 30 cpDNA haplotypes that were divided into five clades by phylogenetic and network analyses and a strong phylogeographical structure. All haplotypes but one in the three basal clades occur exclusively in the southeast QTP, whereas haplotypes in the young clade V occupy almost the whole species range. In particular, the young haplotype H18 occurs in 420 individuals, even at a frequency of 100% in some QTP platform populations and the Altai population. The haplotype distribution pattern, together with molecular clock estimation and mismatch distribution analysis, suggests that the southeast QTP was either a refuge for P. longiflora during the Quaternary climatic change or is the place of origin of the species. The present wide distribution of the species on the QTP platform has resulted from recent population expansions which could be dated back to 120 000–17 000 years ago, a period mostly before the last glacial maximum. The possible relationships among geographic genetic structure, climatic change and species diversification in Pedicularis are also discussed.     

Pliocene intraspecific divergence and Plio‐Pleistocene range expansions within Picea likiangensis (Lijiang spruce), a dominant forest tree of the Qinghai‐Tibet Plateau

A knowledge of intraspecific divergence and range dynamics of dominant forest trees in response to past geological and climate change is of major importance to an understanding of their recent evolution and demography. Such knowledge is informative of how forests were affected by environmental factors in the past and may provide pointers to their response to future environmental change. However, genetic signatures of such historical events are often weak at individual loci due to large effective population sizes and long generation times of forest trees. This problem can be overcome by analysing genetic variation across multiple loci. We used this approach to examine intraspecific divergence and past range dynamics in the conifer Picea likiangensis, a dominant tree of forests occurring in eastern and southern areas of the Qinghai‐Tibet Plateau (QTP). We sequenced 13 nuclear loci, two mitochondrial DNA regions and three plastid (chloroplast) DNA regions in 177 individuals sampled from 22 natural populations of this species, and tested the hypothesis that its evolutionary history was markedly affected by Pliocene QTP uplifts and Quaternary climatic oscillations. Consistent with the taxonomic delimitation of the three morphologically divergent varieties examined, all individuals clustered into three genetic groups with intervariety admixture detected in regions of geographical overlap. Divergence between varieties was estimated to have occurred within the Pliocene and ecological niche modelling based on 20 ecological variables suggested that niche differentiation was high. Furthermore, modelling of population‐genetic data indicated that two of the varieties (var. rubescens and var. linzhiensis) expanded their population sizes after the largest Quaternary glaciation in the QTP, while expansion of the third variety (var. likiangensis) began prior to this, probably following the Pliocene QTP uplift. These findings point to the importance of geological and climatic changes during the Pliocene and Pleistocene as causes of intraspecific diversification and range shifts of dominant tree species in the QTP biodiversity hot spot region.     

Phylogeography of Podocarpus matudae (Podocarpaceae): pre‐Quaternary relicts in northern Mesoamerican cloud forests

Aim Cloud forests of northern Mesoamerica represent the northern and southern limit of the contact zone between species otherwise characteristic of North or South America. Several phylogeographic studies featuring temperate conifer species have improved our understanding of species responses to environmental changes. In contrast, conifer species that presumably colonized northern Mesoamerica from South America are far less studied. A phylogeographic study of Podocarpus matudae (Podocarpaceae) was conducted to identify any major evolutionary divergences or disjunctions across its range and to determine if its current distribution is associated with pre‐Quaternary climatic and/or long‐distance dispersal events. Location Northern Mesoamerica (Mexico and Guatemala). Methods Sixteen populations (157 individuals) of P. matudae were screened for variation at two plastid DNA markers. The intra‐specific phylogenetic relationships among haplotypes were reconstructed using Bayesian inference. Population genetic analyses were undertaken to gain insight into the evolutionary history of these populations. To test whether genetic divergence among populations occurred at different time‐scales plastid DNA sequence data and fossil‐ and coalescent‐based calibrations were integrated. Results The combination of plastid markers yielded 11 haplotypes. Differentiation among populations based on DNA variation (G_ST) (0.707, SE 0.0807) indicated a clear population structure in P. matudae. Differentiation for ordered alleles (N_ST) (0.811, SE 0.0732) was higher than that for G_ST, indicating phylogeographical structure in P. matudae. Most of the total variation (81.3%, P < 0.0001) was explained by differences among populations. The estimated divergence time between the unique haplotypes from a Guatemalan population and the two most common haplotypes from the Sierra Madre Oriental in Mexico was between 10 and 20 Ma, and further haplotype divergence in the poorly resolved clade of the Sierra Madre Oriental occurred between 3 and 0.5 Ma. Main conclusions Divergence estimations support the hypothesis that extant Podocarpus matudae populations are pre‐Quaternary relicts. This finding is consistent with fossil and pollen data that support a Miocene age for temperate floristic elements in Mesoamerican cloud forests, whereas further haplotype divergence within the Sierra Madre Oriental, Chiapas and Guatemala occurred more recently, coinciding with Pleistocene cloud forest refugia.     

Genetic diversity and evolutionary history of the Schizothorax species complex in the Lancang River (upper Mekong)

The genus Schizothorax (Cyprinidae), one of the most diverse genera of ichthyofauna of the Qinghai‐Tibetan Plateau (QTP), is a good candidate for investigating patterns of genetic variation and evolutionary mechanisms. In this study, sequences from the mitochondrial control region, the cytochrome b gene, and two nuclear genes were used to re‐examine the genetic diversity and investigate the evolutionary history of the Schizothorax species complex inhabiting the Lancang River. Three maternal clades were detected in the Schizothorax species complex, but frequent nuclear allele sharing also occurred among the three maternal clades. A discrepancy between topologies of mitochondrial and nuclear loci might result from introgression or/and incomplete lineage sorting. The divergence of the clades of the Schizothorax species complex was closely related to the Late Pliocene and Early Pleistocene orogenesis of the QTP and Southwest Mountains of China. Demographic analyses indicated that the species complex subsequently persisted in situ with stable populations during Pleistocene glacial cycling, which suggested that Pleistocene climate changes did not exert a remarkable influence on the species complex. Our study provides a comprehensive analysis of the genetic diversity and evolutionary history of the Schizothorax species complex in the Lancang River.     

Species‐level phylogeographical history of the endemic species Calligonum roborovskii and its close relatives in Calligonum section Medusa (Polygonaceae) in arid north‐western China

Quaternary climatic oscillations appear to have influenced the genetic diversity and evolutionary history of arid‐adapted plants. To understand the processes involved and reveal evolutionary relationships, haplotypes were examined from Calligonum roborovskii, an endemic species occurring in the arid zones across the desert regions of north‐western China, and seven other species also from Calligonum section Medusa, including C. gobicum, C. mongolicum and the narrow endemic species C. ebi‐nuricum, C. pumilum, C. taklimakanense, C. trifarium and C. yengisaricum. Forty‐three haplotypes were identified in 422 individuals from 51 natural populations, from variation of two plastid DNA intergenic spacers (rpl32‐trnL and ycf6‐psbM). A high level of total genetic diversity was found across species for which more than two populations were examined, including C. gobicum, C. mongolicum, C. pumilum and C. roborovskii. A distinct isolation‐by‐distance pattern in each of these species was suggested by the Mantel test, indicating that restricted gene flow caused high genetic differentiation among populations. Three haplotypes were shared by two or three species each, but the other 40 haplotypes were species‐specific. The 43 haplotypes split into three major clades, but not species‐specific lineages; most of the Calligonum species were not reciprocally monophyletic, probably due to incomplete lineage sorting or introgression. The identified haplotypes were dated to 1.97 Mya (95% highest posterior density: 2.95–0.99 Mya) and diverged until the late Pleistocene, possibly linked to aridification and enlargement of deserts caused by climate changes. Variation of desert habitats during the Pleistocene might play a key role in causing the divergence.     

Cladogenesis of the European brown hare (Lepus europaeus Pallas, 1778)

A substantial portion of today’s biodiversity is attributed to the climatic oscillations of the Pleistocene Ice Ages. Gradual but dramatic climate changes were accompanied by expansion, contraction, and isolation of populations, promoting the accumulation of genome differences and adaptations in refugial populations and resulting in allopatric differentiation in a variety of taxa. In the present study, partial mitochondrial DNA sequences of the widely distributed European brown hare (Lepus europaeus) were analyzed to test whether the species’ present genetic structure is the result of postglacial re-colonization of Europe from Asia Minor (clade A) and the Balkans (clade B) only, as suggested previously, or if additional refugia are likely. Analyses indicated the presence of an additional refugium (Italy, clade I). The genealogic network of Italian hares displayed the tree-like structure expected from refugial populations, whereas central European brown hare haplotypes revealed a clear star-phylogeny indicative of past-bottleneck population growth. This population size expansion, which was confirmed by mismatch analysis, was estimated to have occurred ∼50–55 thousand years ago (kya). The divergence of clade A* from the remaining matrilines is estimated at 239 kya, whereas the divergence of the ancestors of clades B* and I from A* occurred about 128 kya. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The Pleistocene demography of an alpine juniper of the Qinghai‐Tibetan Plateau: tabula rasa,cryptic refugia or something else?

Aim Numerous palaeoecological and genetic studies have shown that different tree species responded in very different ways to Pleistocene climatic oscillations. Some were forced into small refugia far from their current range, while others were able to survive in small refugia close to, or even within, their current natural range. In this study we examine the Pleistocene demography of a juniper species (Juniperus przewalskii, Cupressaceae) from the Qinghai‐Tibetan Plateau. Location The Qinghai‐Tibetan Plateau (QTP). Methods Eight nuclear loci were sequenced in 141 individuals from 20 natural populations distributed across the entire natural range of J. przewalskii, and coalescent analysis was used to test demographic hypotheses. Results The overall nucleotide diversity in the sample was low (π_sil = 0.0029), with few rare alleles and pronounced population genetic structure (F_ST = 0.181). We detected a division previously found using chloroplast DNA markers: all segregating sites in populations from the central part of the QTP appear to be a subset of those found around the edge of the plateau, confirming the relatively young age of the former. In contrast to the middle Pleistocene bottlenecks detected in boreal tree species, the coalescent‐based analyses failed to reject the standard neutral model for the juniper species considered here. Main conclusions Juniperus przewalskii did not undergo marked changes in population sizes during the Pleistocene, although this species seems to have experienced recent, post‐glacial expansion. This finding is largely consistent with the limited number of previous studies on conifer species of the QTP, but contradicts findings of studies on boreal species. These findings have wide implications for understanding plant species’ responses to past climatic oscillations on the high‐elevation QTP.