Does urbanisation lead to parallel demographic shifts across the world in a cosmopolitan plant?

Urbanisation is occurring globally, leading to dramatic environmental changes that are altering the ecology and evolution of species. In particular, the expansion of human infrastructure and the loss and fragmentation of natural habitats in cities is predicted to increase genetic drift and reduce gene flow by reducing the size and connectivity of populations. Alternatively, the 'urban facilitation model' suggests that some species will have greater gene flow into and within cities leading to higher diversity and lower differentiation in urban populations. These alternative hypotheses have not been contrasted across multiple cities. Here, we used the genomic data from the GLobal Urban Evolution project (GLUE), to study the effects of urbanisation on non-adaptive evolutionary processes of white clover (Trifolium repens) at a global scale. We found that white clover populations presented high genetic diversity and no evidence of reduced Ne linked to urbanisation. On the contrary, we found that urban populations were less likely to experience a recent decrease in effective population size than rural ones. In addition, we found little genetic structure among populations both globally and between urban and rural populations, which showed extensive gene flow between habitats. Interestingly, white clover displayed overall higher gene flow within urban areas than within rural habitats. Our study provides the largest comprehensive test of the demographic effects of urbanisation. Our results contrast with the common perception that heavily altered and fragmented urban environments will reduce the effective population size and genetic diversity of populations and contribute to their isolation.

The low-copy nuclear gene Agt1 as a novel DNA barcoding marker for Bromeliaceae.

BACKGROUND: The angiosperm family Bromeliaceae comprises over 3.500 species characterized by exceptionally high morphological and ecological diversity, but a very low genetic variation. In many genera, plants are vegetatively very similar which makes determination of non flowering bromeliads difficult. This is particularly problematic with living collections where plants are often cultivated over decades without flowering. DNA barcoding is therefore a very promising approach to provide reliable and convenient assistance in species determination. However, the observed low genetic variation of canonical barcoding markers in bromeliads causes problems. RESULT: In this study the low-copy nuclear gene Agt1 is identified as a novel DNA barcoding marker suitable for molecular identification of closely related bromeliad species. Combining a comparatively slowly evolving exon sequence with an adjacent, genetically highly variable intron, correctly matching MegaBLAST based species identification rate was found to be approximately double the highest rate yet reported for bromeliads using other barcode markers. CONCLUSION: In the present work, we characterize Agt1 as a novel plant DNA barcoding marker to be used for barcoding of bromeliads, a plant group with low genetic variation. Moreover, we provide a comprehensive marker sequence dataset for further use in the bromeliad research community.

Climatic differentiation in polyploid apomictic Ranunculus auricomus complex in Europe.

BACKGROUND: Polyploidy and apomixis are important factors influencing plant distributions often resulting in range shifts, expansions and geographical parthenogenesis. We used the Ranunculus auricomus complex as a model to asses if the past and present distribution and climatic preferences were determined by these phenomena. RESULTS: Ecological differentiation among diploids and polyploids was tested by comparing the sets of climatic variables and distribution modelling using 191 novel ploidy estimations and 561 literature data. Significant differences in relative genome size on the diploid level were recorded between the "auricomus" and "cassubicus" groups and several new diploid occurrences were found in Slovenia and Hungary. The current distribution of diploids overlapped with the modelled paleodistribution (22 kyr BP), except Austria and the Carpathians, which are proposed to be colonized later on from refugia in the Balkans. Current and historical presence of diploids from the R. auricomus complex is suggested also for the foothills of the Caucasus. Based on comparisons of the climatic preferences polyploids from the R. auricomus complex occupy slightly drier and colder habitats than the diploids. CONCLUSIONS: The change of reproductive mode and selection due to competition with the diploid ancestors may have facilitated the establishment of polyploids within the R. auricomus complex in environments slightly cooler and drier, than those tolerated by diploid ancestors. Much broader distribution of polyploid apomicts may have been achieved due to faster colonization mediated by uniparental reproductive system.

Ecological range shift in the polyploid members of the South American genus Fosterella (Bromeliaceae).

Background and Aims: The distribution of polyploidy along a relatively steep Andean elevation and climatic gradient is studied using the genus Fosterella L.B. Sm. (Bromeliaceae) as a model system. Ecological differentiation of cytotypes and the link of polyploidy with historical biogeographic processes such as dispersal events and range shift are assessed. Methods: 4',6-Diamidino-2-phenylindole (DAPI) staining of nuclei and flow cytometry were used to estimate the ploidy levels of 159 plants from 22 species sampled throughout the distribution range of the genus. Ecological differentiation among ploidy levels was tested by comparing the sets of climatic variables. Ancestral chromosome number reconstruction was carried out on the basis of a previously generated phylogeographic framework. Key Results: This study represents the first assessment of intrageneric, intraspecific and partially intrapopulational cytotype diversity in a genus of the Bromeliaceae family. In Fosterella , the occurrence of polyploidy was limited to the phylogenetically isolated penduliflora and rusbyi groups. Cytotypes were found to be ecologically differentiated, showing that polyploids preferentially occupy colder habitats with high annual temperature variability (seasonality). The combined effects of biogeographic history and adaptive processes are presumed to have shaped the current cytotype distribution in the genus. Conclusions: The results provide indirect evidence for both adaptive ecological and non-adaptive historical processes that jointly influenced the cytotype distribution in the predominantly Andean genus Fosterella (Bromeliaceae). The results also exemplify the role of polyploidy as an important driver of speciation in a topographically highly structured and thus climatically diverse landscape.

Origin and genetic differentiation of pink-flowered Sorbus hybrids in the Western Carpathians.

Background and Aims: Diversity of the genus Sorbus has been affected by interspecific hybridizations. Pink-flowered hybrid species have been insufficiently studied so far. They comprise bigenomic hybrid species derived from crosses S. aria s.l. × S. chamaemespilus and trigenomic ones, where S. aucuparia was involved as well. The main objective of the present study was to reconstruct their hybrid origins as well as to assess genetic distinction among several morphologically recognized hybrid species. Methods: Samples from putative maternal species and eight pink-flowered and two white-flowered hybrid species were collected in the Western Carpathians and the Sudetes. In total, 370 specimens were analysed. Six chloroplast microsatellites were used to infer parentage, whereas nuclear amplified fragment length polymorphism (AFLP) markers were employed for the identification of clones and patterns of genetic variation. Ploidy levels were estimated by flow cytometry on a subset of 140 individuals. Key Results: Genetic data supported their hybrid origins proposed based on flower and leaf morphology, and chloroplast DNA (cpDNA) revealed recurrent origins ( S. caeruleomontana , S. haljamovae ), even from bidirectional hybridization events ( S. zuzanae ). All bigenomic and trigenomic hybrid species (except triploid S. zuzanae ) were found to be tetraploid. In addition to polyploidy, low genetic variation and the presence of clones within and among populations were observed, suggesting predominantly apomictic reproduction of the hybrid species. Most of the described hybrid species appeared also genetically distinct. Conclusions: The data suggest that multiple hybridization events in the Western Carpathian Sorbus have led to the formation of separate, partially reproductively isolated genetic lineages, which may or may not be discriminated morphologically. Even bidirectional hybridization can produce individuals classified to the same taxon based on phenotype. For some hybrid taxa, hybridization pathways were proposed based on their genetic proximity to parental species and differences in genome sizes.

Prospects and limits of the flow cytometric seed screen--insights from Potentilla sensu lato (Potentilleae, Rosaceae).

The flow cytometric seed screen allows for identification of reproductive modes of seed formation and inference of the ploidy of contributing gametes. However, the lack of a mathematical formalization to infer male/female genomic contributions, and the prerequisite of a binucleate female contribution to the endosperm limits its applicability. We evaluated this assumption combining a DNA-based progeny survey with a comparison of the cytology of reproductive pathways co-occurring within single individuals representing 14 Potentilleae species from six phylogenetic lineages. A numerical framework valid for sexual and pseudogamous taxa was developed, enabling quantification of female and male genomes contributing to embryo and endosperm independent of gametophyte origins, numbers of sperm involved and ploidy of parents. The inference strongly depended on accurate peak index estimation. The endosperm of Potentilleae species received a binucleate female contribution in five evolutionary lineages whereas endosperm formation remained uncertain in the Tormentillae. A modified flow cytometric seed screen protocol was developed to cope with low endosperm contents. Evolutionary conservation of a binucleate female contribution to the endosperm suggested wide applicability of flow cytometric seed screen - at least in the Potentilleae. However, alternative progeny surveys and precise embryo/endosperm ploidy estimates are required for a comprehensive understanding of the cytology of seed formation.

The Data Portal "Chromosome Numbers of the Flora of Germany": Progress After Five Years, Recent Developments, and Future Strategies.

Chromosome numbers and information on ploidy level are of great importance for better understanding of plant evolution and taxonomy but also for species identification. Technical developments in flow cytometry dramatically improved the measurement of genome size and triggered a renewed interest in chromosome numbers. Web-based portals make these kind of data accessible for a wide audience in both academia and citizen science. The specialised database of German plant chromosome counts and ploidy estimates comprises to date more than 14,000 records covering 52% of the German taxa. The database is accessed about 70 times per month and became an integral part of several more comprehensive initiatives. One example is the Rothmaler Flora of Germany, which now for the first time features chromosome data with verified origin from the region of interest. Another is the German plant information hub FloraWeb currently amended by several new data sources with the chromosome number database as a major component.

Cryptic invasion suggested by a cytogeographic analysis of the halophytic Puccinellia distans complex (Poaceae) in Central Europe.

Introduction: Despite the wealth of studies dealing with the invasions of alien plants, invasions of alien genotypes of native species (cryptic invasions) have been vastly neglected. The impact of cryptic invasions on the biodiversity of plant communities can, however, be significant. Inland saline habitats and halophytes (i.e., salt-tolerant plant species) are especially threatened by this phenomenon as they inhabit fragmented remnants of largely destroyed habitats, but at the same time some of these halophytic species are rapidly spreading along salt-treated roads. To study potential cryptic invasion of halophytes, the patterns of genome size and ploidy variation in the Puccinellia distans complex (Poaceae), the most rapidly spreading roadside halophyte in Central Europe, were investigated. Methods: DNA flow cytometry with confirmatory chromosome counts were employed to assess ploidy levels of 1414 individuals from 133 populations of the P. distans complex. In addition, climatic niche modelling was used to predict the distributions of selected cytotypes. Results: Eight groups differing in ploidy level and/or genome size were discovered, one diploid (2x; 2n = 14), two tetraploid (4xA, 4xB; 2n = 28), one pentaploid (5x; 2n = 35), three hexaploid (6xA, 6xB, 6xC; 2n = 42), and one heptaploid (7x; 2n = 49). The hexaploids (mostly the 6xC cytotype) were widespread through the study area, spreading intensively in both anthropogenic and natural habitats and probably hybridizing with the natural habitat dwelling tetraploids. In contrast, the non-hexaploid cytotypes rarely spread and were predominantly confined to natural habitats. Discussion: The extensive spread of the hexaploid cytotypes along roadsides has most likely facilitated their incursion into natural habitats. The colonization of new natural habitats by the hexaploids may pose a threat to the indigenous Puccinellia populations by compromising their genetic integrity and/or by outcompeting them.

Implications of hybridisation and cytotypic differentiation in speciation assessed by AFLP and plastid haplotypes--a case study of Potentilla alpicola La Soie.

BACKGROUND: Hybridisation is presumed to be an important mechanism in plant speciation and a creative evolutionary force often accompanied by polyploidisation and in some cases by apomixis. The Potentilla collina group constitutes a particularly suitable model system to study these phenomena as it is morphologically extensively variable, exclusively polyploid and expresses apomixis. In the present study, the alpine taxon Potentilla alpicola has been chosen in order to study its presumed hybrid origin, identify underlying evolutionary processes and infer the discreteness or taxonomic value of hybrid forms. RESULTS: Combined analysis of AFLP, cpDNA sequences and ploidy level variation revealed a hybrid origin of the P. alpicola populations from South Tyrol (Italy) resulting from crosses between P. pusilla and two cytotypes of P. argentea. Hybrids were locally sympatric with at least one of the parental forms. Three lineages of different evolutionary origin comprising two ploidy levels were identified within P. alpicola. The lineages differed in parentage and the complexity of the evolutionary process. A geographically wide-spread lineage thus contrasted with locally distributed lineages of different origins. Populations of P. collina studied in addition, have been regarded rather as recent derivatives of the hexaploid P. argentea. The observation of clones within both P. alpicola and P. collina suggested a possible apomictic mode of reproduction. CONCLUSIONS: Different hybridisation scenarios taking place on geographically small scales resulted in viable progeny presumably stabilised by apomixis. The case study of P. alpicola supports that these processes played a significant role in the creation of polymorphism in the genus Potentilla. However, multiple origin of hybrids and backcrossing are considered to produce a variety of evolutionary spontaneous forms existing aside of reproductively stabilised, established lineages.

Genome size evolution in the diverse insect order Trichoptera.

BACKGROUND: Genome size is implicated in the form, function, and ecological success of a species. Two principally different mechanisms are proposed as major drivers of eukaryotic genome evolution and diversity: polyploidy (i.e., whole-genome duplication) or smaller duplication events and bursts in the activity of repetitive elements. Here, we generated de novo genome assemblies of 17 caddisflies covering all major lineages of Trichoptera. Using these and previously sequenced genomes, we use caddisflies as a model for understanding genome size evolution in diverse insect lineages. RESULTS: We detect a ∼14-fold variation in genome size across the order Trichoptera. We find strong evidence that repetitive element expansions, particularly those of transposable elements (TEs), are important drivers of large caddisfly genome sizes. Using an innovative method to examine TEs associated with universal single-copy orthologs (i.e., BUSCO genes), we find that TE expansions have a major impact on protein-coding gene regions, with TE-gene associations showing a linear relationship with increasing genome size. Intriguingly, we find that expanded genomes preferentially evolved in caddisfly clades with a higher ecological diversity (i.e., various feeding modes, diversification in variable, less stable environments). CONCLUSION: Our findings provide a platform to test hypotheses about the potential evolutionary roles of TE activity and TE-gene associations, particularly in groups with high species, ecological, and functional diversities.

Genome Size Dynamics in Marine Ribbon Worms (Nemertea, Spiralia).

Nemertea is a phylum consisting of 1300 mostly marine species. Nemertea is distinguished by an eversible muscular proboscis, and most of the species are venomous. Genomic resources for this phylum are scarce despite their value in understanding biodiversity. Here, we present genome size estimates of Nemertea based on flow cytometry and their relationship to different morphological and developmental traits. Ancestral genome size estimations were done across the nemertean phylogeny. The results increase the available genome size estimates for Nemertea three-fold. Our analyses show that Nemertea has a narrow genome size range (0.43-3.89 pg) compared to other phyla in Lophotrochozoa. A relationship between genome size and evolutionary rate, developmental modes, and habitat was found. Trait analyses show that the highest evolutionary rate of genome size is found in upper intertidal, viviparous species with direct development. Despite previous findings, body size in nemerteans was not correlated with genome size. A relatively small genome (1.18 pg) is assumed for the most recent common ancestor of all extant nemerteans. The results provide an important basis for future studies in nemertean genomics, which will be instrumental to understanding the evolution of this enigmatic and often neglected phylum.

Resolving Recalcitrant Clades in the Pantropical Ochnaceae: Insights From Comparative Phylogenomics of Plastome and Nuclear Genomic Data Derived From Targeted Sequencing.

Plastid DNA sequence data have been traditionally widely used in plant phylogenetics because of the high copy number of plastids, their uniparental inheritance, and the blend of coding and non-coding regions with divergent substitution rates that allow the reconstruction of phylogenetic relationships at different taxonomic ranks. In the present study, we evaluate the utility of the plastome for the reconstruction of phylogenetic relationships in the pantropical plant family Ochnaceae (Malpighiales). We used the off-target sequence read fraction of a targeted sequencing study (targeting nuclear loci only) to recover more than 100 kb of the plastid genome from the majority of the more than 200 species of Ochnaceae and all but two genera using de novo and reference-based assembly strategies. Most of the recalcitrant nodes in the family's backbone were resolved by our plastome-based phylogenetic inference, corroborating the most recent classification system of Ochnaceae and findings from a phylogenomic study based on nuclear loci. Nonetheless, the phylogenetic relationships within the major clades of tribe Ochnineae, which comprise about two thirds of the family's species diversity, received mostly low support. Generally, the phylogenetic resolution was lowest at the infrageneric level. Overall there was little phylogenetic conflict compared to a recent analysis of nuclear loci. Effects of taxon sampling were invoked as the most likely reason for some of the few well-supported discords. Our study demonstrates the utility of the off-target fraction of a target enrichment study for assembling near-complete plastid genomes for a large proportion of samples.

A comprehensive chloroplast DNA-based phylogeny of the genus Potentilla (Rosaceae): implications for its geographic origin, phylogeography and generic circumscription.

A reconstruction of phylogenetic relationships based on three chloroplast DNA markers comprising 98 species of the genus Potentilla and 15 additional genera from the tribe Potentilleae (Rosaceae) is presented. The phylogeny supported the current generic concept of the subtribe Fragariinae and resolved major lineages within the subtribe Potentillinae. Resolved lineages corresponded to the combined genera Argentina, Piletophyllum, and Tylosperma, an European and Asian group of species approximately equivalent to the Trichocarpae Herbaceae sensu Th. Wolf, the series Tormentillae Th. Wolf, a North American clade comprising the genera Horkelia, Horkeliella, and Ivesia, the species Potentilla fragarioides and P. freyniana, and to a taxonomically highly diverse but molecularly little diverged core group of Potentilla, respectively. Age estimates of phylogenetic splits resolved in the Potentilleae using Bayesian inference, suggested a diversification of the tribe in the Eocene and radiation of two major evolutionary lineages corresponding to the Fragariinae and Potentillinae at approximately comparable times. Ancestral area reconstructions based on the recent distribution ranges of species and collection sites of cpDNA haplotypes suggested an Asian origin for Potentilla s.str., and explained the arrival of this still informal taxon in Europe and particularly North America by multiple dispersal events. In combining the phylogenetic and geographic data with molecularly inferred time estimates and taxonomy, strongly contrasting evolutionary patterns were identified. These evolutionary patterns included rapid speciation on a continental and worldwide scale accompanied by multiple intercontinental dispersals opposed to largely diverged lineages of limited taxonomic diversity and vicariant geographic distribution. The molecular-based phylogeographic hypothesis finally is discussed on the background of the fossil record of Potentilla.

Early Diverging and Core Bromelioideae (Bromeliaceae) Reveal Contrasting Patterns of Genome Size Evolution and Polyploidy.

The subfamily Bromelioideae is one of the most diverse groups among the neotropical Bromeliaceae. Previously, key innovations have been identified which account for the extraordinary radiation and species richness of this subfamily, especially in the so-called core Bromelioideae. However, in order to extend our understanding of the evolutionary mechanisms, the genomic mechanisms (e.g. polyploidy, dysploidy) that potentially underlie this accelerated speciation also need to be tested. Here, using PI and DAPI staining and flow cytometry we estimated genome size and GC content of 231 plants covering 30 genera and 165 species and combined it with published data. The evolutionary and ecological significance of all three genomic characters was tested within a previously generated dated phylogenetic framework using ancestral state reconstructions, comparative phylogenetic methods, and multiple regressions with climatic variables. The absolute genome size (2C) of Bromelioideae varied between 0.59 and 4.11 pg, and the GC content ranged between 36.73 and 41.43%. The monoploid genome sizes (Cx) differed significantly between core and early diverging lineages. The occurrence of dysploidy and polyploidy was, with few exceptions, limited to the phylogenetically isolated early diverging tank-less lineages. For Cx and GC content Ornstein-Uhlenbeck models outperformed the Brownian motion models suggesting adaptive potential linked to the temperature conditions. 2C-values revealed different rates of evolution in core and early diverging lineages also related to climatic conditions. Our results suggest that polyploidy is not associated with higher net diversification and fast radiation in core bromelioids. On the other hand, although coupled with higher extinction rates, dysploidy, polyploidy, and resulting genomic reorganizations might have played a role in the survival of the early diverging bromelioids in hot and arid environments.

The morphometrics of autopolyploidy: insignificant differentiation among sexual-apomictic cytotypes.

Polyploidization of the plant genome affects the phenotype of individuals including their morphology, i.e. size and form. In autopolyploids, we expect mainly nucleotypic effects, from a number of monoploid genomes (i.e. chromosome sets) or genome size, seen from an increase in size or dimension of the polyploids compared with the diploids (or lower ploids). To identify nucleotypic effects, confounding effects of hybridity (observed in allopolyploids), postpolyploidization processes or environmental effects need to be considered. We morphometrically analysed five ploidy cytotypes of the sexual-apomictic species Potentilla puberula cultivated ex situ under the same experimental conditions. Sexuals are mainly tetraploid, while higher ploidy (penta- to octoploidy) is typically associated with the expression of apomixis. The cytotypes likely arose via autopolyploidization although historic involvement of another species in the origin of apomicts cannot be fully ruled out, suggested by a slight molecular differentiation among reproductive modes. We (i) revisited molecular differentiation using amplified fragment length polymorphisms and performed a morphometric analysis to test (ii) if cytotypes are morphologically differentiated from each other and (iii) if the size of individuals is related to their ploidy. Weak molecular differentiation of sexual versus apomictic individuals was confirmed. Cytotypes and reproductive modes were also morphologically poorly differentiated from each other, i.e. apomicts largely resampled the variation of the sexuals and did not exhibit a unique morphology. Overall size of individuals increased moderately but significantly with ploidy (ca. 14 % in the comparison of octo- with tetraploids). The results support an autopolyploid origin of the P. puberula apomicts and suggest a nucleotypic effect on overall plant size. We discuss taxonomic consequences of the results in the context of data on reproductive relationships among cytotypes and their ecological preferences and evolutionary origin, and conclude that cytotypes are best treated as intraspecific variants within a single species.

Annotated Draft Genomes of Two Caddisfly Species Plectrocnemia conspersa CURTIS and Hydropsyche tenuis NAVAS (Insecta: Trichoptera).

Members of the speciose insect order Trichoptera (caddisflies) provide important ecosystem services, for example, nutrient cycling through breaking down of organic matter. They are also of industrial interest due to their larval silk secretions. These form the basis for their diverse case-making behavior that allows them to exploit a wide range of ecological niches. Only five genomes of this order have been published thus far, with variable qualities regarding contiguity and completeness. A low-cost sequencing strategy, that is, using a single Oxford Nanopore flow cell per individual along with Illumina sequence reads was successfully used to generate high-quality genomes of two Trichoptera species, Plectrocnemia conspersa and Hydropsyche tenuis. Of the de novo assembly methods compared, assembly of low coverage Nanopore reads (∼18×) and subsequent polishing with long reads followed by Illumina short reads (∼80-170× coverage) yielded the highest genome quality both in terms of contiguity and BUSCO completeness. The presented genomes are the shortest to date and extend our knowledge of genome size across caddisfly families. The genomic region that encodes for light (L)-chain fibroin, a protein component of larval caddisfly silk was identified and compared with existing L-fibroin gene clusters. The new genomic resources presented in this paper are among the highest quality Trichoptera genomes and will increase the knowledge of this important insect order by serving as the basis for phylogenomic and comparative genomic studies.

Hybrid Genome Assembly of a Neotropical Mutualistic Ant.

The success of social insects is largely intertwined with their highly advanced chemical communication system that facilitates recognition and discrimination of species and nest-mates, recruitment, and division of labor. Hydrocarbons, which cover the cuticle of insects, not only serve as waterproofing agents but also constitute a major component of this communication system. Two cryptic Crematogaster species, which share their nest with Camponotus ants, show striking diversity in their cuticular hydrocarbon (CHC) profile. This mutualistic system therefore offers a great opportunity to study the genetic basis of CHC divergence between sister species. As a basis for further genome-wide studies high-quality genomes are needed. Here, we present the annotated draft genome for Crematogaster levior A. By combining the three most commonly used sequencing techniques-Illumina, PacBio, and Oxford Nanopore-we constructed a high-quality de novo ant genome. We show that even low coverage of long reads can add significantly to overall genome contiguity. Annotation of desaturase and elongase genes, which play a role in CHC biosynthesis revealed one of the largest repertoires in ants and a higher number of desaturases in general than in other Hymenoptera. This may provide a mechanistic explanation for the high diversity observed in C. levior CHC profiles.

A reference genome of the European beech (Fagus sylvatica L.).

Background: The European beech is arguably the most important climax broad-leaved tree species in Central Europe, widely planted for its valuable wood. Here, we report the 542 Mb draft genome sequence of an up to 300-year-old individual (Bhaga) from an undisturbed stand in the Kellerwald-Edersee National Park in central Germany. Findings: Using a hybrid assembly approach, Illumina reads with short- and long-insert libraries, coupled with long Pacific Biosciences reads, we obtained an assembled genome size of 542 Mb, in line with flow cytometric genome size estimation. The largest scaffold was of 1.15 Mb, the N50 length was 145 kb, and the L50 count was 983. The assembly contained 0.12% of Ns. A Benchmarking with Universal Single-Copy Orthologs (BUSCO) analysis retrieved 94% complete BUSCO genes, well in the range of other high-quality draft genomes of trees. A total of 62,012 protein-coding genes were predicted, assisted by transcriptome sequencing. In addition, we are reporting an efficient method for extracting high-molecular-weight DNA from dormant buds, by which contamination by environmental bacteria and fungi was kept at a minimum. Conclusions: The assembled genome will be a valuable resource and reference for future population genomics studies on the evolution and past climate change adaptation of beech and will be helpful for identifying genes, e.g., involved in drought tolerance, in order to select and breed individuals to adapt forestry to climate change in Europe. A continuously updated genome browser and download page can be accessed from beechgenome.net, which will include future genome versions of the reference individual Bhaga, as new sequencing approaches develop.

Heteroploid reticulate evolution and taxonomic status of an endemic species with bicentric geographical distribution.

Reticulate evolution is considered to be among the main mechanisms of plant evolution, often leading to the establishment of new species. However, complex evolutionary scenarios result in a challenging definition of evolutionary and taxonomic units. In the present study, we aimed to examine the evolutionary origin and revise the species status of Campanula baumgartenii, a rare endemic species from the polyploid complex Campanula section Heterophylla. Morphometry, flow cytometric ploidy estimation, AFLPs, as well as chloroplast and nuclear DNA sequence markers were used to assess the morphological and genetic differentiation among C. baumgartenii, C. rotundifolia and other closely related taxa. Tetra- and hexaploid C. baumgartenii is morphologically and molecularly (AFLP) differentiated from sympatric C. rotundifolia. Contrasting signals from nuclear (ITS) and chloroplast (trnL-rpl32) markers suggest a hybrid origin of C. baumgartenii with C. rotundifolia and a taxon related to the alpine C. scheuchzeri as ancestors. Additionally, hexaploid C. baumgartenii currently hybridizes with co-occurring tetraploid C. rotundifolia resulting in pentaploid hybrids, for which C. baumgartenii serves as both seed and pollen donor. Based on the molecular and morphological differentiation, we propose to keep C. baumgartenii as a separate species. This study exemplifies that detailed population genetic studies can provide a solid basis for taxonomic delimitation within Campanula section Heterophylla as well as for sound identification of conservation targets.