Phylogenetic challenges in a recently diversified and polyploid-rich Alyssum (Brassicaceae) lineage: low divergence, reticulation, and parallel polyploid speciation.

Elucidating the evolution of recently diverged and polyploid-rich plant lineages may be challenging even with high-throughput sequencing, both for biological reasons and bioinformatic difficulties. Here, we apply target enrichment with genome skimming (Hyb-Seq) to unravel the evolutionary history of the Alyssum montanum-A. repens species complex. Reconstruction of phylogenetic relationships in diploids supported recent and rapid diversification accompanied by reticulation events. Of the 4 main clades identified among the diploids, 3 clades included species from the Alps, Apennine, and Balkan peninsulas, indicating close biogeographic links between these regions. We further focused on the clade distributed from the Western Alps to the Iberian Peninsula, which comprises numerous polyploids as opposed to a few diploids. Using a recently developed PhyloSD (phylogenomic subgenome detection) pipeline, we successfully tracked the ancestry of all polyploids. We inferred multiple polyploidization events that involved 2 closely related diploid progenitors, resulting into several sibling polyploids: 2 autopolyploids and 6 allopolyploids. The skewed proportions of major homeolog-types and the occurrence of some minor homeolog-types, both exhibiting geographic patterns, suggest introgression with the progenitors and other related diploids. Our study highlights a unique case of parallel polyploid speciation that was enhanced by ecological and geographic separation and provides an excellent resource for future studies of polyploid evolution.

Evolution of hygrophytic plant species in the Anatolia-Caucasus region: insights from phylogenomic analyses of Cardamine perennials.

BACKGROUND AND AIMS: Southwestern Asia is a significant centre of biodiversity and a cradle of diversification for many plant groups, especially xerophytic elements. In contrast, little is known about the evolution and diversification of its hygrophytic flora. To fill this gap, we focus on Cardamine (Brassicaceae) species that grow in wetlands over a wide altitudinal range. We aimed to elucidate their evolution, assess the extent of presumed historical gene flow between species, and draw inferences about intraspecific structure. METHODS: We applied the phylogenomic Hyb-Seq approach, ecological niche analyses and multivariate morphometrics to a total of 85 Cardamine populations from the target region of Anatolia-Caucasus, usually treated as four to six species, and supplemented them with close relatives from Europe. KEY RESULTS: Five diploids are recognized in the focus area, three of which occur in regions adjacent to the Black and/or Caspian Sea (C. penzesii, C. tenera, C. lazica), one species widely distributed from the Caucasus to Lebanon and Iran (C. uliginosa), and one western Anatolian entity (provisionally C. cf. uliginosa). Phylogenomic data suggest recent speciation during the Pleistocene, likely driven by both geographic separation (allopatry) and ecological divergence. With the exception of a single hybrid (allotetraploid) speciation event proven for C. wiedemanniana, an endemic of southern Turkey, no significant traces of past or present interspecific gene flow were observed. Genetic variation within the studied species is spatially structured, suggesting reduced gene flow due to geographic and ecological barriers, but also glacial survival in different refugia. CONCLUSIONS: This study highlights the importance of the refugial regions of the Black and Caspian Seas for both harbouring and generating hygrophytic species diversity in Southwestern Asia. It also supports the significance of evolutionary links between Anatolia and the Balkan Peninsula. Reticulation and polyploidization played a minor evolutionary role here in contrast to the European relatives.

Phylogenomics and genome size evolution in Amomum s. s. (Zingiberaceae): Comparison of traditional and modern sequencing methods.

BACKGROUND AND AIMS: A targeted enrichment NGS approach was used to construct the phylogeny of Amomum Roxb. (Zingiberaceae). Phylogenies based on hundreds of nuclear genes, the whole plastome and the rDNA cistron were compared with an ITS-based phylogeny. Trends in genome size (GS) evolution were examined, chromosomes were counted and the geographical distribution of phylogenetic lineages was evaluated. METHODS: In total, 92 accessions of 54 species were analysed. ITS was obtained for 79 accessions, 37 accessions were processed with Hyb-Seq and sequences from 449 nuclear genes, the whole cpDNA, and the rDNA cistron were analysed using concatenation, coalescence and supertree approaches. The evolution of absolute GS was analysed in a phylogenetic and geographical context. The chromosome numbers of 12 accessions were counted. KEY RESULTS: Four groups were recognised in all datasets though their mutual relationships differ among datasets. While group A (A. subulatum and A. petaloideum) is basal to the remaining groups in the nuclear gene phylogeny, in the cpDNA topology it is sister to group B (A. repoeense and related species) and, in the ITS topology, it is sister to group D (the Elettariopsis lineage). The former Elettariopsis makes a monophyletic group. There is an increasing trend in GS during evolution. The largest GS values were found in group D in two tetraploid taxa, A. cinnamomeum and A. aff. biphyllum (both 2n = 96 chromosomes). The rest varied in GS (2C = 3.54-8.78 pg) with a constant chromosome number 2n = 48. There is a weak connection between phylogeny, GS and geography in Amomum. CONCLUSIONS: Amomum consists of four groups, and the former Elettariopsis is monophyletic. Species in this group have the largest GS. Two polyploids were found and GS greatly varied in the rest of Amomum.

Morphological and environmental differentiation as prezygotic reproductive barriers between parapatric and allopatric Campanula rotundifolia agg. cytotypes.

BACKGROUND AND AIMS: Reproductive isolation and local establishment are necessary for plant speciation. Polyploidy, the possession of more than two complete chromosome sets, creates a strong postzygotic reproductive barrier between diploid and tetraploid cytotypes. However, this barrier weakens between polyploids (e.g. tetraploids and hexaploids). Reproductive isolation may be enhanced by cytotype morphological and environmental differentiation. Moreover, morphological adaptations to local conditions contribute to plant establishment. However, the relative contributions of ploidy level and the environment to morphology have generally been neglected. Thus, the extent of morphological variation driven by ploidy level and the environment was modelled for diploid, tetraploid and hexaploid cytotypes of Campanula rotundifolia agg. Cytotype distribution was updated, and morphological and environmental differentiation was tested in the presence and absence of natural contact zones. METHODS: Cytotype distribution was assessed from 231 localities in Central Europe, including 48 localities with known chromosome counts, using flow cytometry. Differentiation in environmental niche and morphology was tested for cytotype pairs using discriminant analyses. A structural equation model was used to explore the synergies between cytotype, environment and morphology. KEY RESULTS: Tremendous discrepancies were revealed between the reported and detected cytotype distribution. Neither mixed-ploidy populations nor interploidy hybrids were detected in the contact zones. Diploids had the broadest environmental niche, while hexaploids had the smallest and specialized niche. Hexaploids and spatially isolated cytotype pairs differed morphologically, including allopatric tetraploids. While leaf and shoot morphology were influenced by environmental conditions and polyploidy, flower morphology depended exclusively on the cytotype. CONCLUSIONS: Reproductive isolation mechanisms vary between cytotypes. While diploids and polyploids are isolated postzygotically, the environmental niche shift is essential between higher polyploids. The impact of polyploidy and the environment on plant morphology implies the adaptive potential of polyploids, while the exclusive relationship between flower morphology and cytotype highlights the role of polyploidy in reproductive isolation.

MorphoTools2: an R package for multivariate morphometric analysis.

SUMMARY: The package MorphoTools2 is intended for multivariate analyses of morphological data. Commonly used tools are missing or scattered across several R packages. The new package, in order to make the workflow convenient and fast, wraps available statistical and graphical tools and provides a comprehensive framework for checking and manipulating input data, core statistical analyses and a wide palette of functions designed to visualize results. AVAILABILITY AND IMPLEMENTATION: Stable version is available from CRAN: https://cran.r-project.org/package=MorphoTools2. The development version is available from the following GitHub repository: https://github.com/MarekSlenker/MorphoTools2. The software is distributed under the GNU General Public Licence (v.3). SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Allele Sorting as a Novel Approach to Resolving the Origin of Allotetraploids Using Hyb-Seq Data: A Case Study of the Balkan Mountain Endemic Cardamine barbaraeoides.

Mountains of the Balkan Peninsula are significant biodiversity hotspots with great species richness and a large proportion of narrow endemics. Processes that have driven the evolution of the rich Balkan mountain flora, however, are still insufficiently explored and understood. Here we focus on a group of Cardamine (Brassicaceae) perennials growing in wet, mainly mountainous habitats. It comprises several Mediterranean endemics, including those restricted to the Balkan Peninsula. We used target enrichment with genome skimming (Hyb-Seq) to infer their phylogenetic relationships, and, along with genomic in situ hybridization (GISH), to resolve the origin of tetraploid Cardamine barbaraeoides endemic to the Southern Pindos Mts. (Greece). We also explored the challenges of phylogenomic analyses of polyploid species and developed a new approach of allele sorting into homeologs that allows identifying subgenomes inherited from different progenitors. We obtained a robust phylogenetic reconstruction for diploids based on 1,168 low-copy nuclear genes, which suggested both allopatric and ecological speciation events. In addition, cases of plastid-nuclear discordance, in agreement with divergent nuclear ribosomal DNA (nrDNA) copy variants in some species, indicated traces of interspecific gene flow. Our results also support biogeographic links between the Balkan and Anatolian-Caucasus regions and illustrate the contribution of the latter region to high Balkan biodiversity. An allopolyploid origin was inferred for C. barbaraeoides, which highlights the role of mountains in the Balkan Peninsula both as refugia and melting pots favoring species contacts and polyploid evolution in response to Pleistocene climate-induced range dynamics. Overall, our study demonstrates the importance of a thorough phylogenomic approach when studying the evolution of recently diverged species complexes affected by reticulation events at both diploid and polyploid levels. We emphasize the significance of retrieving allelic and homeologous variation from nuclear genes, as well as multiple nrDNA copy variants from genome skim data.

Parallel Alpine Differentiation in Arabidopsis arenosa.

Parallel evolution provides powerful natural experiments for studying repeatability of evolution and genomic basis of adaptation. Well-documented examples from plants are, however, still rare, as are inquiries of mechanisms driving convergence in some traits while divergence in others. Arabidopsis arenosa, a predominantly foothill species with scattered morphologically distinct alpine occurrences is a promising candidate. Yet, the hypothesis of parallelism remained untested. We sampled foothill and alpine populations in all regions known to harbor the alpine ecotype and used SNP genotyping to test for repeated alpine colonization. Then, we combined field surveys and a common garden experiment to quantify phenotypic parallelism. Genetic clustering by region but not elevation and coalescent simulations demonstrated parallel origin of alpine ecotype in four mountain regions. Alpine populations exhibited parallelism in height and floral traits which persisted after two generations in cultivation. In contrast, leaf traits were distinctive only in certain region(s), reflecting a mixture of plasticity and genetically determined non-parallelism. We demonstrate varying degrees and causes of parallelism and non-parallelism across populations and traits within a plant species. Parallel divergence along a sharp elevation gradient makes A. arenosa a promising candidate for studying genomic basis of adaptation.

Niche similarity in diploid-autotetraploid contact zones of Arabidopsis arenosa across spatial scales.

PREMISE: Whole genome duplication is a major evolutionary event, but its role in ecological divergence remains equivocal. When populations of different ploidy (cytotypes) overlap in space, "contact zones" are formed, allowing the study of evolutionary mechanisms contributing toward ploidy divergence. Multiple contact zones per species' range are often described but rarely leveraged as natural replicates. We explored whether the strength of niche differentiation of diploid and autotetraploid Arabidopsis arenosa varies over distinct contact zones and if the frequency of triploids decreases from seedling to adult stage. METHODS: We characterized ploidy composition and habitat preferences in 264 populations across three contact zones using climatic niche modeling. Ecological differences of cytotypes were also assessed using local vegetation surveys at 110 populations within two contact zones, and at the finer scale within five mixed-ploidy sites. This was complemented by flow cytometry of seedlings. RESULTS: We found no niche differences between diploid and tetraploid populations within contact zones for either climatic or local environmental variables. Comparisons of cytotypes within mixed-ploidy sites found weak niche differences that were inconsistent in direction. Triploid individuals were virtually absent (0.14%) in the field, and they were at a similarly low frequency (0.2%) in ex situ germinated seedlings. CONCLUSIONS: This study demonstrates the strength in investigating different spatial scales across several contact zones when addressing ecological niche differentiation between ploidies. The lack of consistent habitat differentiation of ploidies across the scales and locations supports the recently emerging picture that processes other than ecological differentiation may underlie ploidy coexistence in diploid-autopolyploid systems.

So Closely Related and Yet So Different: Strong Contrasts Between the Evolutionary Histories of Species of the Cardamine pratensis Polyploid Complex in Central Europe.

Recurrent polyploid formation and weak reproductive barriers between independent polyploid lineages generate intricate species complexes with high diversity and reticulate evolutionary history. Uncovering the evolutionary processes that formed their present-day cytotypic and genetic structure is a challenging task. We studied the species complex of Cardamine pratensis, composed of diploid endemics in the European Mediterranean and diploid-polyploid lineages more widely distributed across Europe, focusing on the poorly understood variation in Central Europe. To elucidate the evolution of Central European populations we analyzed ploidy level and genome size variation, genetic patterns inferred from microsatellite markers and target enrichment of low-copy nuclear genes (Hyb-Seq), and environmental niche differentiation. We observed almost continuous variation in chromosome numbers and genome size in C. pratensis s.str., which is caused by the co-occurrence of euploid and dysploid cytotypes, along with aneuploids, and is likely accompanied by inter-cytotype mating. We inferred that the polyploid cytotypes of C. pratensis s.str. are both of single and multiple, spatially and temporally recurrent origins. The tetraploid Cardamine majovskyi evolved at least twice in different regions by autopolyploidy from diploid Cardamine matthioli. The extensive genome size and genetic variation of Cardamine rivularis reflects differentiation induced by the geographic isolation of disjunct populations, establishment of triploids of different origins, and hybridization with sympatric C. matthioli. Geographically structured genetic lineages identified in the species under study, which are also ecologically divergent, are interpreted as descendants from different source populations in multiple glacial refugia. The postglacial range expansion was accompanied by substantial genetic admixture between the lineages of C. pratensis s.str., which is reflected by diffuse borders in their contact zones. In conclusion, we identified an interplay of diverse processes that have driven the evolution of the species studied, including allopatric and ecological divergence, hybridization, multiple polyploid origins, and genetic reshuffling caused by Pleistocene climate-induced range dynamics.

Diversification and independent polyploid origins in the disjunct species Alyssum repens from the Southeastern Alps and the Carpathians.

PREMISE: Disjunct distributions have been commonly observed in mountain plant species and have stimulated phylogeographic and phylogenetic research. Here we studied Alyssum repens, a member of the polyploid species complex A. montanum-A. repens, which exhibits SE Alpine-Carpathian disjunctions with a large elevational span and consists of diploid and tetraploid populations. We aimed to investigate the species' genetic and cytotype structure in the context of its distribution patterns, to elucidate the polyploid origins and to propose an appropriate taxonomic treatment. METHODS: We combined AFLP fingerprinting markers, sequence variation of the highly repetitive ITS region of rDNA and the low-copy DET1 nuclear gene, genome size, and morphometric data. RESULTS: We identified four geographically structured genetic lineages. One consisted of diploid populations from the foothills of the Southeastern Alps and neighboring regions, and the three others were allopatric montane to alpine groups comprising diploids and tetraploids growing in the Southeastern Carpathians and the Apuseni Mts. in Romania. CONCLUSIONS: We inferred a vicariance scenario associated with Quaternary climatic oscillations, accompanied by one auto- and two allopolyploidization events most likely involving a northern Balkan relative. Whereas genetic differentiation and allopatric distribution would favor the taxonomic splitting of this species, the genetic lineages largely lack morphological distinguishability, and their ecological, cytotype and genome size divergence is only partial. Even though we probably face here a case of incipient speciation, we propose to maintain the current taxonomic treatment of Alyssum repens as a single, albeit variable, species.

Role of ploidy in colonization of alpine habitats in natural populations of Arabidopsis arenosa.

BACKGROUND AND AIMS: Polyploidy is an important driver of plant diversification and adaptation to novel environments. As a consequence of genome doubling, polyploids often exhibit greater colonizing ability or occupy a wider ecological niche than diploids. Although elevation has been traditionally considered as a key driver structuring ploidy variation, we do not know if environmental and phenotypic differentiation among ploidy cytotypes varies along an elevational gradient. Here, we tested for the consequences of genome duplication on genetic diversity, phenotypic variation and habitat preferences on closely related diploid and tetraploid populations that coexist along approx. 2300 m of varying elevation. METHODS: We sampled and phenotyped 45 natural diploid and tetraploid populations of Arabidopsis arenosa in one mountain range in Central Europe (Western Carpathians) and recorded abiotic and biotic variables at each collection site. We inferred genetic variation, population structure and demographic history in a sub-set of 29 populations genotyped for approx. 36 000 single nucleotide polymorphisms. KEY RESULTS: We found minor effects of polyploidy on colonization of alpine stands and low genetic differentiation between the two cytotypes, mirroring recent divergence of the polyploids from the local diploid lineage and repeated reticulation events among the cytotypes. This pattern was corroborated by the absence of ecological niche differentiation between the two cytotypes and overall phenotypic similarity at a given elevation. CONCLUSIONS: The case of A. arenosa contrasts with previous studies that frequently showed clear niche differentiation between cytotypes. Our work stresses the importance of considering genetic structure and past demographic processes when interpreting the patterns of ploidy distributions, especially in species that underwent recent polyploidization events.

The story of promiscuous crucifers: origin and genome evolution of an invasive species, Cardamine occulta (Brassicaceae), and its relatives.

BACKGROUND AND AIMS: Cardamine occulta (Brassicaceae) is an octoploid weedy species (2n = 8x = 64) originated in Eastern Asia. It has been introduced to other continents including Europe and considered to be an invasive species. Despite its wide distribution, the polyploid origin of C. occulta remained unexplored. The feasibility of comparative chromosome painting (CCP) in crucifers allowed us to elucidate the origin and genome evolution in Cardamine species. We aimed to investigate the genome structure of C. occulta in comparison with its tetraploid (2n = 4x = 32, C. kokaiensis and C. scutata) and octoploid (2n = 8x = 64, C. dentipetala) relatives. METHODS: Genomic in situ hybridization (GISH) and large-scale CCP were applied to uncover the parental genomes and chromosome composition of the investigated Cardamine species. KEY RESULTS: All investigated species descended from a common ancestral Cardamine genome (n = 8), structurally resembling the Ancestral Crucifer Karyotype (n = 8), but differentiated by a translocation between chromosomes AK6 and AK8. Allotetraploid C. scutata originated by hybridization between two diploid species, C. parviflora and C. amara (2n = 2x = 16). By contrast, C. kokaiensis has an autotetraploid origin from a parental genome related to C. parviflora. Interestingly, octoploid C. occulta probably originated through hybridization between the tetraploids C. scutata and C. kokaiensis. The octoploid genome of C. dentipetala probably originated from C. scutata via autopolyploidization. Except for five species-specific centromere repositionings and one pericentric inversion post-dating the polyploidization events, the parental subgenomes remained stable in the tetra- and octoploids. CONCLUSIONS: Comparative genome structure, origin and evolutionary history was reconstructed in C. occulta and related species. For the first time, whole-genome cytogenomic maps were established for octoploid plants. Post-polyploid evolution in Asian Cardamine polyploids has not been associated with descending dysploidy and intergenomic rearrangements. The combination of different parental (sub)genomes adapted to distinct habitats provides an evolutionary advantage to newly formed polyploids by occupying new ecological niches.

Pervasive population genomic consequences of genome duplication in Arabidopsis arenosa.

Ploidy-variable species allow direct inference of the effects of chromosome copy number on fundamental evolutionary processes. While an abundance of theoretical work suggests polyploidy should leave distinct population genomic signatures, empirical data remains sparse. We sequenced ~300 individuals from 39 populations of Arabidopsis arenosa, a naturally diploid-autotetraploid species. We find that the impacts of polyploidy on population genomic processes are subtle yet pervasive, such as reduced efficiency of purifying selection, differences in linked selection and rampant gene flow from diploids. Initial masking of deleterious mutations, faster rates of nucleotide substitution and interploidy introgression likely conspire to shape the evolutionary potential of polyploids.

Growth form evolution and hybridization in Senecio (Asteraceae) from the high equatorial Andes.

Changes in growth forms frequently accompany plant adaptive radiations, including páramo-a high-elevation treeless habitat type of the northern Andes. We tested whether diverse group of Senecio inhabiting montane forests and páramo represented such growth form changes. We also investigated the role of Andean geography and environment in structuring genetic variation of this group. We sampled 108 populations and 28 species of Senecio (focusing on species from former genera Lasiocephalus and Culcitium) and analyzed their genetic relationships and patterns of intraspecific variation using DNA fingerprinting (AFLPs) and nuclear DNA sequences (ITS). We partitioned genetic variation into environmental and geographical components. ITS-based phylogeny supported monophyly of a Lasiocephalus-Culcitium clade. A grade of herbaceous alpine Senecio species subtended the Lasiocephalus-Culcitium clade suggesting a change from the herbaceous to the woody growth form. Both ITS sequences and the AFLPs separated a group composed of the majority of páramo subshrubs from other group(s) comprising both forest and páramo species of various growth forms. These morphologically variable group(s) further split into clades encompassing both the páramo subshrubs and forest lianas, indicating independent switches among the growth forms and habitats. The finest AFLP genetic structure corresponded to morphologically delimited species except in two independent cases in which patterns of genetic variation instead reflected geography. Several morphologically variable species were genetically admixed, which suggests possible hybrid origins. Latitude and longitude accounted for 5%-8% of genetic variation in each of three AFLP groups, while the proportion of variation attributed to environment varied between 8% and 31% among them. A change from the herbaceous to the woody growth form is suggested for species of high-elevation Andean Senecio. Independent switches between habitats and growth forms likely occurred within the group. Hybridization likely played an important role in species diversification.

Actionable, long-term stable and semantic web compatible identifiers for access to biological collection objects.

With biodiversity research activities being increasingly shifted to the web, the need for a system of persistent and stable identifiers for physical collection objects becomes increasingly pressing. The Consortium of European Taxonomic Facilities agreed on a common system of HTTP-URI-based stable identifiers which is now rolled out to its member organizations. The system follows Linked Open Data principles and implements redirection mechanisms to human-readable and machine-readable representations of specimens facilitating seamless integration into the growing semantic web. The implementation of stable identifiers across collection organizations is supported with open source provider software scripts, best practices documentations and recommendations for RDF metadata elements facilitating harmonized access to collection information in web portals. Database URL: : http://cetaf.org/cetaf-stable-identifiers.

On the origins of Balkan endemics: the complex evolutionary history of the Cyanus napulifer group (Asteraceae).

BACKGROUND AND AIMS: The Balkan Peninsula is one of the most important centres of plant diversity in Europe. Here we aim to fill the gap in the current knowledge of the evolutionary processes and factors modelling this astonishing biological richness by applying multiple approaches to the Cyanus napulifer group. METHODS: To reconstruct the mode of diversification within the C. napulifer group and to uncover its relationships with potential relatives with x = 10 from Europe and Northern Africa, we examined variation in genetic markers (amplified fragment length polymorphisms [AFLPs]; 460 individuals), relative DNA content (4',6-diamidino-2-phenylindole [DAPI] flow cytometry, 330 individuals) and morphology (multivariate morphometrics, 40 morphological characters, 710 individuals). To elucidate its evolutionary history, we analysed chloroplast DNA (cpDNA) sequences of the genus Cyanus deposited in the GenBank database. KEY RESULTS: The AFLPs revealed a suite of closely related entities with variable levels of differentiation. The C. napulifer group formed a genetically well-defined unit. Samples outside the group formed strongly diversified and mostly species-specific genetic lineages with no further geographical patterns, often characterized also by a different DNA content. AFLP analysis of the C. napulifer group revealed extensive radiation and split it into nine allopatric (sub)lineages with varying degrees of congruence among genetic, DNA-content and morphological patterns. Genetic admixture was usually detected in contact zones between genetic lineages. Plastid data indicated extensive maintenance of ancestral variation across Cyanus perennials. CONCLUSION: The C. napulifer group is an example of a rapidly and recently diversified plant group whose genetic lineages have evolved in spatio-temporal isolation on the topographically complex Balkan Peninsula. Adaptive radiation, accompanied in some cases by long-term isolation and hybridization, has contributed to the formation of this species complex and its mosaic pattern.

Sequencing of the genus Arabidopsis identifies a complex history of nonbifurcating speciation and abundant trans-specific polymorphism.

The notion of species as reproductively isolated units related through a bifurcating tree implies that gene trees should generally agree with the species tree and that sister taxa should not share polymorphisms unless they diverged recently and should be equally closely related to outgroups. It is now possible to evaluate this model systematically. We sequenced multiple individuals from 27 described taxa representing the entire Arabidopsis genus. Cluster analysis identified seven groups, corresponding to described species that capture the structure of the genus. However, at the level of gene trees, only the separation of Arabidopsis thaliana from the remaining species was universally supported, and, overall, the amount of shared polymorphism demonstrated that reproductive isolation was considerably more recent than the estimated divergence times. We uncovered multiple cases of past gene flow that contradict a bifurcating species tree. Finally, we showed that the pattern of divergence differs between gene ontologies, suggesting a role for selection.

Northern glacial refugia and altitudinal niche divergence shape genome-wide differentiation in the emerging plant model Arabidopsis arenosa.

Quaternary climatic oscillations profoundly impacted temperate biodiversity. For many diverse yet undersampled areas, however, the consequences of this impact are still poorly known. In Europe, particular uncertainty surrounds the role of Balkans, a major hotspot of European diversity, in postglacial recolonization of more northerly areas, and the Carpathians, a debatable candidate for a northern 'cryptic' glacial refugium. Using genome-wide SNPs and microsatellites, we examined how the interplay of historical processes and niche shifts structured genetic diversity of diploid Arabidopsis arenosa, a little-known member of the plant model genus that occupies a wide niche range from sea level to alpine peaks across eastern temperate Europe. While the northern Balkans hosted one isolated endemic lineage, most of the genetic diversity was concentrated further north in the Pannonian Basin and the Carpathians, where it likely survived the last glaciation in northern refugia. Finally, a distinct postglacial environment in northern Europe was colonized by populations of admixed origin from the two Carpathian lineages. Niche differentiation along altitude-related bioclimatic gradients was the main trend in the phylogeny of A. arenosa. The most prominent niche shifts, however, characterized genetically only slightly divergent populations that expanded into narrowly defined alpine and northern coastal postglacial environments. Our study highlights the role of eastern central European mountains not only as refugia for unique temperate diversity but also sources for postglacial expansion into novel high-altitude and high-latitude niches. Knowledge of distinct genetic substructure of diploid A. arenosa also opens new opportunities for follow-up studies of this emerging model of evolutionary biology.