Species radiation in the Alps: multiple range shifts caused diversification in Ringlet butterflies in the European high mountains

The distributions of European high mountain species are often characterised by small and geographically isolated populations and, in many cases, have highly complex biogeographic histories. The butterfly genus Erebia represents one of the best examples for small-scale diversification in the European high mountain systems and therefore to understand speciation processes and associated range dynamics of high mountain species. In this study, we analysed 17 polymorphic allozyme loci of 1731 individuals from 49 populations representing four species, one of which has three subspecies: Erebia nivalis; Erebia tyndarus; Erebia ottomana; and Erebia cassioides cassioides, Erebia cassioides arvernensis, and Erebia cassioides neleus. Samples were collected in the high mountain systems of Europe (i.e. Pyrenees, Massif Central, Alps, Apennines, Carpathians, Balkan high mountains). Genetic analyses supported all previously accepted species. However, the genetic differentiation within E. cassioides sensu lato into three geographically delimited groups is justifying species rank: E. arvernensis distributed in the Pyrenees, Massif Central and western Alps; E. cassioides sensu stricto in the eastern Alps and Apennines; and E. neleus in the Balkan mountains and the south-western Carpathians. While the differentiation between western Alps and Massif Central as well as eastern Alps and Apennines was low, the Pyrenees as well as the south-western Carpathians were significantly differentiated from the other regions within the respective taxon. In general, the differentiation among the populations of E. neleus was stronger than between populations of the other taxa. Within E. cassioides, we found a west-east gradient of genetic similarity over the eastern Alps. Based on the obtained genetic structures, we are able to delineate glacial refugia and interglacial range modifications. Based on the genetic structures and genetic diversity patterns, we conclude that, triggered by the glacial-interglacial cycles, repeated range modifications have taken place with subsequent differentiation and speciation in the region of the Alps and Balkans. Colonisations to Pyrenees (E. arvernensis pseudomurina, E. arvernensis pseudocarmenta), Massif Central (E. ottomana tardenota, E. a. arvernensis) and Apennines (E. cassioides majellana) appear to be recent and most probably not older than the last interglacial period.     

The genetic structure of the mountain forest butterfly Erebia euryale unravels the late Pleistocene and postglacial history of the mountain coniferous forest biome in Europe

The distribution of the mountain coniferous forest biome in Europe throughout time is not sufficiently understood. One character species of this habitat type is the large ringlet, Erebia euryale well reflecting the extension of this biome today, and the genetic differentiation of this species among and within mountain systems may unravel the late Pleistocene history of this habitat type. We therefore analysed the allozyme pattern of 381 E. euryale individuals from 11 populations in four different European mountain systems (Pyrenees, Alps, Carpathians, Rila). All loci analysed were polymorphic. The mean F(ST) over all samples was high (20%). Furthermore, the mean genetic distance among samples was quite high (0.049). We found four different groups well supported by cluster analyses, bootstraps and hierarchical variance analyses: Pyrenees, western Alps, eastern Alps and southeastern Europe (Carpathians and Rila). The genetic diversity of the populations was highest in the southeastern European group and stepwise decreased westwards. Interestingly, the populations from Bulgaria and Romania were almost identical; therefore, we assume that they were not separated by the Danube Valley, at least during the last ice age. On the contrary, the differentiation among the three western Alps populations was considerable. For all these reasons, we assume that (i) the most important refugial area for the coniferous mountain forest biome in Europe has been located in southeastern Europe including at least parts of the Carpathians and the Bulgarian mountains; (ii) important refugial areas for this biome existed at the southeastern edge of the Alps; (iii) fragments of this habitat types survived along the southwestern Alps, but in a more scattered distribution; and (iv) relatively small relicts have persisted somewhere at the foothills of the Pyrenees.     

Intraspecific allozymatic differentiation reveals the glacial refugia and the postglacial expansions of European Erebia medusa (Lepidoptera: Nymphalidae)

Allozyme analysis of Erebia medusa over large regions of Europe revealed a significant population differentiation (F_ST: 0.149 ± 0.016). A UPGMA-analysis showed a division into four major lineages with mean inter-group genetic distances ranging from 0.051 (±0.010) to 0.117 (±0.024). An AMOVA revealed that rather more than two-thirds of the variance between samples was being between these lineages and less than one-third within lineages. An eastern group included the samples from the Czech Republic, Slovakia and north-eastern Hungary. This genetic lineage expressed significantly higher genetic diversity than the other three. A second lineage was formed by the samples from France and Germany. The two samples from western Hungary represent a third delimited lineage and the sample from northern Italy a fourth. We suppose that this genetic differentiation took place during the last ice-age in four disjunct refugia. The genetically more diverse eastern genetic lineage might have evolved in a relatively large refugium in south-eastern Europe. We assume that the other three lineages developed in relatively small relict areas around the Alps. It is likely for the western lineage that its ice-age distribution showed at least one disjunction in late Würm with the consequence of further genetic differentiation. Most probably, the eastern lineage colonized postglacial Central Europe using two alternative routes: one north and one south of the Carpathians. Up to now, neither similar glacial refugia, nor comparable secondary disjunctions in late Würm, are reported for any other animal or plant species.     

Multiple differentiation centres of a non-Mediterranean butterfly species in south-eastern Europe

Aim  The analysis of the phylogeographical structures of many European species reveals the importance of Mediterranean glacial refugia for many thermophilic species, but also underlines the relevance of extra-Mediterranean glacial differentiation centres for a number of temperate species. In this context, phylogeographical analyses of species from south-eastern Europe are highly important for a comprehensive understanding of Europe as a whole.
Location  Romania and Bulgaria.
Methods  We analysed 19 allozyme loci for 615 individuals of the temperate butterfly species Erebia medusa from 28 populations.
Results  These populations had an intermediate genetic diversity, but the Bulgarian populations were significantly more diverse than the ones north of the Danube in Romania. The differentiation among populations was strong, and 52.1% of the genetic variance among populations was distributed between these two countries. The genetic differentiation was considerably stronger in Romania than in Bulgaria, but several sublineages were distinguished within each of these countries.
Main conclusions  The observed genetic structure is so strong that it is most probably the result of glacial differentiation processes in south-eastern Europe and not a post-glacial structure. The strong differentiation into the two groups north and south of the Danube suggests a separating effect by this river valley. The strong differentiation accompanied with genetic impoverishment in Romania suggests the existence of several differentiation centres: at least two small ones on the southern slopes of the southern Carpathians and one in the eastern Carpathian Basin. The considerably weaker differentiation among the Bulgarian samples and their significantly higher genetic diversity imply that gene flow occurred among different regions of Bulgaria during the last ice age.     

Effects of recent and past climatic shifts on the genetic structure of the high mountain Yellow‐spotted ringlet butterfly Erebia manto (Lepidoptera,Satyrinae): a conservation problem

Mountain species have evolved important genetic differentiation due to past climatic fluctuations. The genetic uniqueness of many of these lineages is now at risk due to global warming. Here, we analyse allozyme polymorphisms of 1306 individuals (36 populations) of the mountain butterfly Erebia manto and perform Species Distribution Models (SDMs). As a consensus of analyses, we obtained six most likely genetic clusters: (i) Pyrenees with Massif Central; (ii) Vosges; (iii–v) Alps including the Slovakian Carpathians; (vi) southern Carpathians. The Vosges population showed the strongest genetic split from all other populations, being almost as strong as the split between E. manto and its sister species Erebia eriphyle. The distinctiveness of the Pyrenees‐Massif Central group and of the southern Carpathians group from all other groups is also quite high. All three groups are assumed to have survived more than one full glacial–interglacial cycle close to their current distributions with up‐hill and down‐slope shifts conforming climatic conditions. In contrast with these well‐differentiated groups, the three groups present in the Alps and the Slovakian Carpathians show a much shallower genetic structure and thus also should be of a more recent origin. As predicted by our SDM projections, rising temperatures will strongly impact the distribution of E. manto. While the populations in the Alps are predicted to shrink, the survival of the three lineages present here should not be at risk. The situation of the three other lineages is quite different. All models predict the extinction of the Vosges lineage in the wake of global warming, and also the southern Carpathians and Pyrenees‐Massif Central lineages might be at high risk to disappear. Thus, albeit global warming will therefore be unlikely to threaten E. manto as a species, an important proportion of the species’ intraspecific differentiation and thus uniqueness might be lost.     

Disjunct distributions during glacial and interglacial periods in mountain butterflies: Erebia epiphron as an example

Over several decades, the distribution patterns and evolution of alpine disjunct species has become an increasingly discussed subject. Large scale genetic analysis has allowed the resolution of the past range changes and intraspecific evolution of many species, in Europe especially of Mediterranean origin. However, the phylogeographic structures of species with arctic-alpine disjunct distribution patterns are relatively poorly studied. The existing phylogeographic analysis (mostly of alpine plant species) supports disjunct distributions during glacial as well as post-glacial periods for a number of species. However, several questions still remain unresolved and we therefore analysed the Mountain Ringlet Erebia epiphron as a model for such alpine disjunct species. We found strong differentiation into five different lineages supporting five differentiation centres: (i) the eastern Pyrenees, (ii) the mountain ranges between the central Pyrenees and south-western Alps, (iii, iv) two areas along the southern Alps margin and (v) the northern Alps margin. We propose that these patterns evolved due to the humidity requirements of this species, which did not allow survival in the dry glacial steppes, but along the margins of the wetter glaciated high mountain ranges.     

Cytotype distribution and phylogeography of Hieracium intybaceum (Asteraceae)

Using flow cytometry and amplified fragment length polymorphism (AFLP), we explored the cytogeography and phylogeography of Hieracium intybaceum, a silicicolous species distributed in the Alps and spatially isolated in the Vosges Mountains and the Schwarzwald Mountains. We detected two ploidies, diploid and tetraploid, but no triploid or mixed‐ploidy populations. Whereas diploids are sexual and distributed all across the Alps, tetraploids are apomictic and seem to be confined to the western Alps and the Vosges. We detected a low level of genetic variation. Bayesian clustering identified four clusters/genetic groups, which are partly congruent with the ploidal pattern. The first two groups consisting exclusively of diploids dominate the whole distribution range in the Alps and show east–west geographical separation with a diffuse borderline running from eastern Switzerland to the eastern part of North Tyrol. The third genetic group lacks a defined geographical range and includes diploid and tetraploid plants. The last genetic group comprises tetraploid plants in the French Alps and the Vosges. We suppose that diploids colonized the deglaciated areas from source populations most likely located mainly in the southern part of the recent distribution range and occasionally also in the western Alps. Gene flow and further differentiation likely took place. Apomictic tetraploids most likely originated in the western Alps or in the refugium at the south‐western foot of the Alps. Their rather limited geographical range (partly contrasting with the theory of geographical parthenogenesis) can be explained by their rather recent origin. © 2015 The Linnean Society of London, Botanical Journal of the Linnean Society, 2015, 179 , 487–498.     

Genetic differentiation among populations of Cicerbita alpina (L.) Wallroth (asteraceae) in the western Alps

In this study we analyzed the genetic population structure of the hygrophilous tall-herb Cicerbita alpina in the western Alps because this group of mountain plants is underrepresented in the biogeographical literature. AFLP (amplified fragment length polymorphism) fingerprints of 40 samples were analyzed from four populations situated in a transect from the southwestern Alps to the eastern part of the western Alps and one population from the Black Forest outside the Alps. Two genetic groups can be distinguished. The first group (A) comprises the populations from the northern and eastern parts of the western Alps, and the second group (B) comprises the populations from the southwestern Alps and the Black Forest. Group A originates most likely from at least one refugium in the southern piedmont regions of the Alps. This result provides molecular evidence for a humid climate at the southern margin of the Alps during the Würm glaciation. Group B originates presumably from western or northern direction and we discuss two possible scenarios for the colonization of the Alps, i.e. (1) long-distance dispersal from southwestern refugia and (2) colonization from nearby refugia in the western and/or northern Alpine forelands. The study demonstrates that the target species harbours considerable genetic diversity, even on a regional scale, and therefore is a suitable model for phylogeographic research.     

Some butterflies do not care much about topography: a single genetic lineage of Erebia euryale (Nymphalidae) along the northern Iberian mountains

The phylogeography of montane species often reveals strong genetic differentiation among mountain ranges. Both classic morphological and genetic studies have indicated distinctiveness of Pyrenean populations of the butterfly Erebia euryale. This hypothesis remained inconclusive until data from the westernmost populations of the distribution area (Cantabrian Mountains) were analysed. In the present study, we set out to describe the population structure of Erebia euryale in western Cantabria, where the species occurs in scattered localities. For this goal, we estimate the genetic diversity and differentiation found in 218 individuals from six Cantabrian (North Spain) localities genotyped by 17 allozyme loci. We also sequence 816 bp of the cytochrome oxidase subunit I mitochondrial gene in 49 individuals from Cantabrian localities and 41 specimens from five other European sites. Mitochondrial data support the recognition of four major genetic groups previously suggested for the European populations based on allozyme polymorphisms. Both mitochondrial and nuclear markers reveal genetic distinctiveness of a single Pyrenean–Cantabrian lineage of E. euryale. The lack of geographical structure and the star‐like topology displayed by the mitochondrial haplotypes indicate a pattern of demographic expansion in northern Iberia, probably related to Upper Pleistocene climatic oscillations. By contrast, within the Pyrenean–Cantabrian lineage, Cantabrian samples are genetically structured in nuclear datasets. In particular, San Isidro is significantly differentiated from the other five populations, which cluster into two groups. We recognize an evolutionary significant unit for Pyrenean–Cantabrian populations of Erebia euryale. Our results also illustrate that the evolutionary history of a species may be shaped by processes undetectable by using mtDNA alone.     

Phylogeography of a subalpine tall‐herb Ranunculus platanifolius (Ranunculaceae) reveals two main genetic lineages in the European mountains

A phylogeographical analysis of Ranunculus platanifolius, a typical European subalpine tall‐herb species, indicates the existence of two main genetic lineages based on amplified fragment length polymorphism (AFLP) markers. One group comprises populations from the Balkan Peninsula and the south‐eastern Carpathians and the other includes the remaining part of the range of the species, encompassing the western Carpathians, Sudetes, Alps, Pyrenees and Scandinavia. The main phylogeographical break observed in this species runs across the Carpathians and separates the main parts of this range (western and south‐eastern Carpathians), supporting a distinct glacial history of populations in these areas. The high genetic similarity of the Balkan Peninsula and south‐eastern Carpathian populations could indicate a common glacial refugium for these contemporarily isolated areas of species distribution. The western and northern part of the species range displays an additional weak differentiation into regional phylogeographical groups, which could have been shaped by isolation in glacial refugia or even by a postglacial isolation. The observed weak phylogeographical structure could also be linked with ecological requirements, allowing survival along streams in relatively low, forested mountain ranges. © 2013 The Linnean Society of London     

Survival of cold-adapted species in isolated mountains: the population genetics of the Sudeten ringlet,Erebia sudetica sudetica,in the Jeseník Mts., Czech Republic

Relic populations of cold-adapted species, trapped in isolated mountain pockets within the temperate zone, are predicted to suffer considerably due to ongoing climate warming. The butterfly Erebia sudetica sudetica is an example restricted to the Eastern Sudety Mts. Here, the butterfly forms permanent populations on subalpine tall-herb grasslands, but also occupies woodland clearings and hay meadows at lower altitudes. We assume differences among the genetic diversities of the populations due to differences in the temporal continuity of these habitats. Therefore, 17 allozyme loci were analysed for 276 individuals from 13 different localities (six tall-herb stands, two meadows, five forest clearings) in the Jeseník Mts. with a maximum distance of 20 km among them. We obtained a significantly higher genetic diversity for the subalpine populations than for the forest clearing populations. The genetic differentiation among the forest clearing populations was higher than among the subalpine ones. They also showed a significant isolation-by-distance system. These findings support the idea that the lower-elevation populations might have been founded by more than one dispersal event from the subalpine sites, but also secondary colonisations and gene flow in the forest belt. Due to founder effects and possibly further subsequent bottlenecks, these forest clearing populations did not harbour the entire genetic diversity of the taxon. Therefore, conservation actions should focus on the subalpine tall-herb formation.     

Post-glacial history of the dominant alpine sedge Carex curvula in the European Alpine System inferred from nuclear and chloroplast markers

The alpine sedge Carex curvula ssp. curvula is a clonal, dominant graminoid found in the European Alps, the Carpathians, the Pyrenees and in some of the Balkan Mountains. It is a late-successional species of acidophilous alpine meadows that occurs on sites that were covered by ice during the last glacial maximum (LGM). By applying the amplified fragment length polymorphism (AFLP) fingerprinting and chloroplast DNA (cpDNA) sequencing, we attempted to identify the recolonization routes followed by the species after the last ice retreat. We relied on the genetic diversity of 37 populations covering the entire distributional range of the species. As a wind-pollinated species, C. curvula is characterized by a low level of population genetic differentiation. Nuclear and chloroplast data both support the hypothesis of a long-term separation of Eastern (Balkans and Carpathians) and Western (Alps and Pyrenees) lineages. In the Alps, a continuum of genetic depauperation from the east to the west may be related to a recolonization wave originating in the eastern-most parts of the chain, where the main glacial refugium was likely located. The Pyrenean populations are nested within the western Alps group and show a low level of genetic diversity, probably due to recent long-distance colonization. In contrast to the Alps, we found no phylogeographical structure in the Carpathians. The combination of reduced ice extension during the Würm period and the presence of large areas of siliceous substrate at suitable elevation suggest that in contrast to populations in the Alps, the species in the Carpathians underwent a local vertical migration rather than extinction and recolonization over long distance.     

Genetic diversity in an endangered alpine plant, Eryngium alpinum L. (Apiaceae), inferred from amplified fragment length polymorphism markers

Eryngium alpinum L. is an endangered species found across the European Alps. In order to obtain base-line data for the conservation of this species, we investigated levels of genetic diversity within and among 14 populations from the French Alps. We used the amplified fragment length polymorphism (AFLP) technique with three primer pairs and scored a total of 62 unambiguous, polymorphic markers in 327 individuals. Because AFLP markers are dominant, within-population genetic structure (e.g. FIS) could not be assessed. Analyses based either on the assumption of random-mating or on complete selfing lead to very similar results. Diversity levels within populations were relatively high (mean Nei's expected heterozygosity = 0.198; mean Shannon index = 0.283), and a positive correlation was detected between both genetic diversity measurements and population size (Spearman rank correlation: P = 0. 005 and P = 0.002, respectively). Moreover, FST values and exact tests of differentiation revealed high differentiation among populations (mean pairwise FST = 0.40), which appeared to be independent of geographical distance (nonsignificant Mantel test). Founder events during postglacial colonizations and/or bottlenecks are proposed to explain this high but random genetic differentiation. By contrast, we detected a pattern of isolation by distance within populations and valleys. Predominant local gene flow by pollen or seed is probably responsible for this pattern. Concerning the management of E. alpinum, the high genetic differentiation leads us to recommend the conservation of a maximum number of populations. This study demonstrates that AFLP markers enable a quick and reliable assessment of intraspecific genetic variability in conservation genetics.     

Genetic differentiation of the marbled white butterfly, Melanargia galathea, accounts for glacial distribution patterns and postglacial range expansion in southeastern Europe

Isolation of Mediterranean species in the southern European peninsulas during the cold glacial phases often resulted in differentiation of several genetic lineages confined to the respective peninsulas. However, whilst there is good genetic evidence for multiple refugia in Iberia, there are only limited data available for the Balkans. Therefore, we wish to examine the hypothesis of a strong genetic structuring within southeastern Europe for the existence of multiple Balkan differentiation centres and/or several leading edges. As a model we use the marbled white butterfly, Melanargia galathea. We studied 18 allozyme loci of 564 individuals from 16 populations distributed over a large part of southeastern Europe. The single populations showed moderately high genetic diversity and no northward decline of genetic diversity was detected. The overall genetic differentiation between populations was considerable (F(ST) 7.0%). Cluster analysis discriminated three genetic groups: (i) a western flank in the former Yugoslavia, parts of eastern Austria and Hungary; (ii) an eastern flank with populations from Bulgaria and Romania (south of the southern Carpathians and eastern Carpathians); and (iii) the eastern Carpathian Basin. Hierarchical variance analysis distributed 53% of the variance among populations between these three groups. One sample from the Greek-Bulgarian border clustered within the eastern flank, but showed some tendency towards the eastern Carpathian Basin populations. Two populations from Carinthia clustered together with the eastern Carpathian Basin ones and a population from Styria showed an intermediate genetic composition between the three groups. Most probably, the eastern and the western flank groups are due to postglacial range expansion from the northeastern and the northwestern edges of the glacial differentiation centre (so-called leading edges). The eastern Carpathian Basin group may have resulted from postglacial expansion from northern Greece through valley systems of the central Balkan peninsula, maybe even expanding westwards north of the Balkan mountains reaching some parts of eastern Austria (e.g. Carinthia). Therefore, the Balkanic refugium of M. galathea may or may not have been continuous along the coastal areas of the Mediterranean, but must have been strongly genetically structured.     

Combining mitochondrial and nuclear evidences: a refined evolutionary history of Erebia medusa (Lepidoptera: Nymphalidae: Satyrinae) in Central Europe based on the COI gene

We aim to infer a combined scenario for the evolution of the Woodland Ringlet, Erebia medusa, in Central Europe based on analyses of part of the mitochondrial cytochrome oxydase subunit I (COI) and a published allozyme data set. We sequenced 529 bp of COI for 158 butterflies from 32 populations from almost the entire western distribution range of the species. We applied population genetic [spatial analysis of molecular variance (samova )] and phylogeographical analyses as well as coalescence simulation to test if the published allozyme scenario supports or contradicts the observed haplotype distribution. We recorded 16 haplotypes of which four represent a total of 82%. samova grouped the 32 populations into four geographically coherent groups: (1) western Central Europe, (2) Central Europe, (3) eastern Central Europe and (iv) western Pannonia. Mismatch distribution analyses and haplotype networks are in coincidence with constant population growth and reveal a relatively shallow phylogeographical structure. We evaluated the level of discordance between population histories and gene trees using Slatkins s and the deep coalescence statistics based on our mtDNA data. These estimators decline the previously published allozyme scenario of survival in different extra‐Mediterranean refugia in Europe with an onset of differentiation at the beginning of the last ice age some 70 000 years ago. However, it supports a refined scenario if we assume an onset of vicariance driven differentiation in these refugia after the end of the middle Würm interstadials some 30 000 years ago. Therefore, the general evolutionary history of this species in Europe apparently is very recent. Most probably, areas adjoining the high mountain regions as Alps and northern Carpathians were of great importance for the late Würm glacial survival of species like Erebia medusa.     

Genetic structure of Hypochaeris uniflora (Asteraceae) suggests vicariance in the Carpathians and rapid post-glacial colonization of the Alps from an eastern Alpine refugium

Aim The range of the subalpine species Hypochaeris uniflora covers the Alps, Carpathians and Sudetes Mountains. Whilst the genetic structure and post‐glacial history of many high‐mountain plant taxa of the Alps is relatively well documented, the Carpathian populations have often been neglected in phylogeographical studies. The aim of the present study is to compare the genetic variation of the species in two major European mountain systems – the Alps and the Carpathians. Location Alps and Carpathians. Methods The genetic variation of 77 populations, each consisting of three plants, was studied using amplified fragment length polymorphism (AFLP). Results Neighbour joining and principal coordinate analyses revealed three well‐supported phylogeographical groups of populations corresponding to three disjunct geographical regions – the Alps and the western and south‐eastern Carpathians. Moreover, two further clusters could be distinguished within the latter mountain range, one consisting of populations from the eastern Carpathians and the second consisting of populations from the southern Carpathians. Populations from the Apuseni Mountains had an intermediate position between the eastern and southern Carpathians. The genetic clustering of populations into four groups was also supported by an analysis of molecular variance, which showed that most genetic variation (almost 46%) was found among these four groups. By far the highest within‐population variation was found in the eastern Carpathians, followed by populations from the southern and western Carpathians. Generally, the populations from the Alps were considerably less variable and displayed substantially fewer region‐diagnostic markers than those from the south‐eastern Carpathians. Although no clear geographical structure was found within the Alps, based on neighbour joining or principal coordinate analyses, some trends were obvious: populations from the easternmost part were genetically more variable and, together with those from the south‐western part, exhibited a higher proportion of rare AFLP fragments than populations in other areas. Moreover, the total number of AFLP fragments per population, the percentage of polymorphic loci and the proportion of rare AFLP fragments significantly decreased from east to west. Main conclusions Deep infraspecific phylogeographical gaps between the populations from the Alps and the western and south‐eastern Carpathians suggest the survival of H. uniflora in three separate refugia during the last glaciation. Our AFLP data provide molecular evidence for a long‐term geographical disjunction between the eastern and western Carpathians, previously suggested from the floristic composition at the end of 19th century. It is likely that Alpine populations survived the Last Glacial in the eastern part of the Alps, from where they rapidly colonized the rest of the Alps after the ice sheet retreated. Multiple founder effects may explain a gradual loss of genetic variation during westward colonization of the Alps.     

The molecular population structure of the tall forb Cicerbita alpina (Asteraceae) supports the idea of cryptic glacial refugia in central Europe

There is an ongoing debate about the glacial history of non‐arctic species in central and northern Europe. The two main hypotheses are: (1) postglacial colonization from refugia outside this region; (2) glacial survival in microclimatically favourable sites within the periglacial areas. In order to clarify the glacial history of a boreo‐montane tall forb, we analysed AFLPs from populations of Cicerbita alpina through most of its range (Scandinavia, the mountains of central Europe, the Alps, the Pyrenees and the Balkan Peninsula). We found a major differentiation between the Pyrenean population and all others, supported by principal coordinate, neighbour joining and STRUCTURE analyses. Furthermore, three populations from the central and north‐eastern Alps were genetically distinct from the bulk of populations from Scandinavia, central Europe, the Alps and the Balkan Peninsula. Most populations, including those from central and northern Europe, had moderate to high levels of genetic diversity (mean Shannon index H_Sh = 0.292, mean percentage of polymorphic loci P = 54.1%, mean Nei's gene diversity H = 0.195). The results indicate separate glacial refugia in the Pyrenean region and the Italian Alps. Furthermore, they provide evidence of glacial persistence in cryptic refugia north of the Alps, from where Scandinavia and most of the Alps are likely to have been colonized following deglaciation. © 2010 The Linnean Society of London, Botanical Journal of the Linnean Society, 2010, 164 , 142–154.     

Ecology,environment and evolutionary history influence genetic structure in five mammal species from the Italian Alps

The identification and evaluation of the ecological and environmental factors shaping patterns of natural genetic variation are fundamental goals of population and conservation genetics. Many studies focus on factors affecting single species, but it is also important to test whether some influential biotic and abiotic factors are common drivers of genetic diversity across species, or if species or species groups are each affected by different forces; a multi‐species analysis is necessary for this. Here we analysed the molecular variation from five mammal species (roe deer, red deer, chamois, mountain hare and European brown hare) at mtDNA and microsatellite loci from the eastern Italian Alps. We use phylogeographical and landscape‐level analyses to test the relative influence of large‐scale geographical history and contemporary environmental characteristics of the landscape on genetic diversity and differentiation. We found: (1) all study species except brown hare are strongly differentiated into two main groups, located west and east of a major river valley; (2) significant correlations between levels of within‐population diversity at both mtDNA and microsatellite loci, and several landscape features such as alpine grassland, water courses and anthropized areas. We conclude that heterogeneous landscape has some influence on within‐population diversity, but biogeographical history has probably had the stronger influence on current genetic patterns, despite an apparently large dispersal potential of certain species. However, our results for brown hare show that management actions such as stocking may alter these large‐scale patterns.     

Disjunct distribution of Hypericum nummularium L. (Hypericaceae): molecular data suggest bidirectional colonization from a single refugium rather than survival in distinct refugia

Hypericum nummularium has a strongly disjunct, bi‐areal distribution in Europe: it is abundant in the Pyrenees and grows in a very restricted part of the Alps, more than 1000 km away. My aim was to estimate the genetic divergence between these areas and to identify the factors responsible for the disjunction: glacial relicts, bidirectional colonization from a common refugium, long‐distance dispersal and/or human introduction? Internal transcribed spacers (ITS) sequencing (680 bp) and amplified fragment length polymorphism (AFLP) fingerprinting (104 polymorphic markers) showed very low differentiation between populations in the Alps and the Pyrenees, indicating that H. nummularium probably survived in a single refugium. Moreover, levels of genetic diversity were similar in the two areas, making human introduction and long‐distance dispersal unlikely. Thus, the species probably survived in one refugium, subsequently colonizing both areas more or less simultaneously. The comparison of genetic and geographical distances suggested a step by step migration in the Alps (isolation by distance), whereas random dispersal events were more likely in the Pyrenees. Finally, I discuss possible causes for the restricted distribution area of H. nummularium in the Alps (e.g. unsuitable habitat, low dispersal capacities) and conclude that strong human disturbance is probably the major limit to the expansion of the species in this region. © 2006 The Linnean Society of London, Biological Journal of the Linnean Society, 2006, 87 , 437–447.