The origin of Cardamine flexuosa (Wavy Bittercress) has been a conundrum for more than six decades. Here we identify its parental species, analyse its genome structure in comparison to parental genomes and describe intergenomic structural variations in C. flexuosa. Genomic in situ hybridization (GISH) and comparative chromosome painting (CCP) uncovered the parental genomes and the chromosome composition of C. flexuosa and its presumed diploid progenitors. Cardamine flexuosa is an allotetraploid (2n = 4x = 32), originating from two diploid species, Cardamine amara and Cardamine hirsuta (2n = 2x = 16). The two parental species display almost perfectly conserved chromosomal collinearity for seven out of the eight chromosomes. A 13 Mb pericentric inversion distinguishes chromosome CA1 from CH1. A comparative cytomolecular map was established for C. flexuosa by CCP/GISH. Whereas conserved chromosome collinearity between the C. amara and C. hirsuta subgenomes might have promoted intergenomic rearrangements through homeologous recombination, only one reciprocal translocation between two homeologues has occurred since the origin of C. flexuosa. The genome of C. flexuosa demonstrates that allopolyploids can maintain remarkably stable subgenomes over 10(4) -10(5) yr throughout a wide distribution range. By contrast, the rRNA genes underwent genome-specific elimination towards a diploid-like number of loci.

• Klíčová slova
• Brassicaceae, Cardamine flexuosa (Wavy Bittercress), GISH (genomic in situ hybridization), chromosome painting, genome collinearity, intergenome translocation, polyploidy,
• MeSH
• Cardamine genetika   MeSH
• chromozomy rostlin genetika   MeSH
• délka genomu genetika   MeSH
• diploidie MeSH
• druhová specificita MeSH
• genetické lokusy genetika   MeSH
• genom rostlinný genetika   MeSH
• hybridizace in situ MeSH
• karyotypizace MeSH
• konzervovaná sekvence * MeSH
• malování chromozomů MeSH
• polyploidie * MeSH
• ribozomální DNA genetika   MeSH
• translokace genetická MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ribozomální DNA MeSH

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.

• Klíčová slova
• Allopolyploidy, Asian Cardamine, Brassicaceae, GISH (genomic in situ hybridization), autopolyploidy, centromere repositioning, chromosome rearrangements, comparative chromosome painting, diploidization, genome collinearity, hybridization, invasive species,
• MeSH
• Brassicaceae * MeSH
• Cardamine * MeSH
• genom rostlinný MeSH
• lidé MeSH
• polyploidie MeSH
• zavlečené druhy MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Dálný východ MeSH
• Evropa MeSH

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.

• Klíčová slova
• Cardamine, Allopolyploidy, Anatolia, Caucasus, Hyb-Seq, ecological niche, endemism, hygrophytic flora, phylogenomics,
• MeSH
• Cardamine * genetika   MeSH
• fylogeneze MeSH
• genetická variace MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Evropa MeSH
• Turecko MeSH

While the phenomenon of uniparental silencing of 35S rDNA in interspecific hybrids and allopolyploids is well documented, there is a notable absence of information regarding whether such silencing extends to the 5S RNA component of ribosomes. To address this gap in knowledge, we analyzed the 5S and 35S rDNA expression in Cardamine (Brassicaceae) allopolyploids, namely C. × insueta (2n = 3x = 24, genome composition RRA), C. flexuosa (2n = 4x = 32, AAHH), and C. scutata (2n = 4x = 32, PPAA) which share a common diploid ancestor (AA). We employed high-throughput sequencing of transcriptomes and genomes and phylogenetic analyses of 5S rRNA variants. The genomic organization of rDNA was further scrutinized through clustering and fluorescence in situ hybridization. In the C. × insueta allotriploid, we observed uniparental dominant expression of 5S and 35S rDNA loci. In the C. flexuosa and C. scutata allotetraploids, the expression pattern differed, with the 35S rDNA being expressed from the A subgenome, whereas the 5S rDNA was expressed from the partner subgenome. Both C. flexuosa and C. scutata but not C. × insueta showed copy and locus number changes. We conclude that in stabilized allopolyploids, transcription of ribosomal RNA components occurs from different subgenomes. This phenomenon appears to result in the formation of chimeric ribosomes comprising rRNA molecules derived from distinct parental origins. We speculate that the interplay of epigenetic silencing and rDNA rearrangements introduces an additional layer of variation in multimolecule ribosomal complexes, potentially contributing to the evolutionary success of allopolyploids.

• Klíčová slova
• Cardamine, 35S rDNA, 5S rDNA, chromosome evolution, gene silencing, polyploidy, rRNA genes,
• MeSH
• Cardamine * genetika   MeSH
• fylogeneze * MeSH
• genom rostlinný genetika   MeSH
• hybridizace in situ fluorescenční MeSH
• polyploidie * MeSH
• regulace genové exprese u rostlin MeSH
• ribozomální DNA genetika   MeSH
• RNA ribozomální 5S * genetika   MeSH
• umlčování genů * MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• ribozomální DNA MeSH
• RNA ribozomální 5S * MeSH

BACKGROUND AND AIMS: Sexual reproduction is known to drive plant diversification and adaptation. Here we investigate the evolutionary history and spatiotemporal origin of a dodecaploid (2n = 12x = 96) Eurasian deciduous woodland species, Cardamine bulbifera, which reproduces and spreads via vegetative bulb-like structures only. The species has been among the most successful range-expanding understorey woodland plants in Europe, which raises the question of the genetic architecture of its gene pool, since its hexaploid (2n = 6x = 48) but putatively outcrossing closest relative, C. quinquefolia, displays a smaller distribution range in Eastern Europe towards the Caucasus region. Cardamine bulbifera belongs to a small monophyletic clade of four species comprising also C. abchasica (2n = 2x = 16) and C. bipinnata (unknown ploidy) from the Caucasus region. METHODS: We sequenced the genomes of the two polyploids and their two putative ancestors using Illumina short-read sequencing technology (×7-8 coverage). Covering the entire distribution range, genomic data were generated for 67 samples of the two polyploids (51 samples of C. bulbifera, 16 samples of C. quinquefolia) and 6 samples of the putative diploid taxa (4 samples of C. abchasica, 2 samples of C. bipinnata) to unravel the evolutionary origin of the polyploid taxa using phylogenetic reconstructions of biparentally and maternally inherited genetic sequence data. Ploidy levels of C. bulbifera and C. quinquefolia were analysed by comparative chromosome painting. We used genetic assignment analysis (STRUCTURE) and approximate Bayesian computation (ABC) modelling to test whether C. bulbifera represents genetically differentiated lineages and addressed the hypothesis of its hybrid origin. Comparative ecological modelling was applied to unravel possible niche differentiation among the two polyploid species. KEY RESULTS: Cardamine bulbifera was shown to be a non-hybridogenous, auto-dodecaploid taxon of early Pleistocene origin, but with a history of past gene flow with its hexaploid sister species C. quinquefolia, likely during the last glacial maximum in shared refuge areas in Eastern Europe towards Western Turkey and the Crimean Peninsula region. The diploid Caucasian endemic C. abchasica is considered an ancestral species, which also provides evidence for the origin of the species complex in the Caucasus region. Cardamine bulbifera successfully expanded its distribution range postglacially towards Central and Western Europe accompanied by a transition to exclusively vegetative propagation. CONCLUSIONS: A transition to vegetative propagation in C. bulbifera is hypothesized as the major innovation to rapidly expand its distribution range following postglacially progressing woodland vegetation throughout Europe. Preceding and introgressive gene flow from its sister species C. quinquefolia in the joint refuge area is documented. This transition and ecological differentiation may have been triggered by preceding introgressive gene flow from its sister species in the joint East European refuge areas.

• Klíčová slova
• Cardamine bulbifera, chromosome painting, clonal reproduction, demographic history, ecological modelling, evolutionary history, genomics, polyploidy, postglacial expansion, reproductive shift,
• MeSH
• Bayesova věta MeSH
• Cardamine * genetika   MeSH
• fylogeneze MeSH
• polyploidie MeSH
• rozmnožování MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

This article describes the use of cytogenomic and molecular approaches to explore the origin and evolution of Cardamine schulzii, a textbook example of a recent allopolyploid, in its ~110-year history of human-induced hybridization and allopolyploidy in the Swiss Alps. Triploids are typically viewed as bridges between diploids and tetraploids but rarely as parental genomes of high-level hybrids and polyploids. The genome of the triploid semifertile hybrid Cardamine × insueta (2n = 24, RRA) was shown to combine the parental genomes of two diploid (2n = 2x = 16) species, Cardamine amara (AA) and Cardamine rivularis (RR). These parental genomes have remained structurally stable within the triploid genome over the >100 years since its origin. Furthermore, we provide compelling evidence that the alleged recent polyploid C. schulzii is not an autohexaploid derivative of C. × insueta. Instead, at least two hybridization events involving C. × insueta and the hypotetraploid Cardamine pratensis (PPPP, 2n = 4x-2 = 30) have resulted in the origin of the trigenomic hypopentaploid (2n = 5x-2 = 38, PPRRA) and hypohexaploid (2n = 6x-2 = 46, PPPPRA). These data show that the semifertile triploid hybrid can promote a merger of three different genomes and demonstrate how important it is to reexamine the routinely repeated textbook examples using modern techniques.

• MeSH
• biologická evoluce * MeSH
• Cardamine genetika   MeSH
• druhová specificita MeSH
• fylogeneze MeSH
• genom rostlinný genetika   MeSH
• genová dávka MeSH
• hybridizace genetická MeSH
• hybridizace in situ MeSH
• molekulární sekvence - údaje MeSH
• nestabilita genomu * MeSH
• polyploidie MeSH
• sekvence nukleotidů MeSH
• sekvenční analýza DNA MeSH
• triploidie MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

UNLABELLED: • PREMISE OF THE STUDY: Contact zones between diploids and their autopolyploid descendants represent a unique evolutionary venue for studying polyploid establishment, cytotype coexistence, and interactions. Here, we examine cytotype coexistence in a diploid-tetraploid contact zone of a perennial herb, Cardamine amara, located north of the Alps by assessing cytotype spatial patterns, ecological divergence, and genetic variation and structure.• METHODS: Flow cytometry was applied to screen DNA ploidy levels in 302 populations (3296 individuals) and the genetic variation of a selection of 25 populations was examined using microsatellite and AFLP markers. Environmental (landscape and climatic) data were analyzed to assess ecological differentiation between the cytotypes.• KEY RESULTS: A parapatric distribution of the cytotypes with a relatively wide (over 100 km in some regions) secondary contact zone was identified. Mixed-ploidy populations, documented for the first time in this species, as well as triploid individuals were found along the diploid-tetraploid borderline. Different climatic requirements of the two main cytotypes were revealed, mirrored in their altitudinal separation. The tetraploids were genetically differentiated from both the diploids and the modeled, in silico autotetraploid genotypes, in accordance with the assumed polyploid origin and spread linked to past glaciations, and largely independent evolution in allopatry.• CONCLUSIONS: The observed spatial and genetic patterns likely reflect the evolutionary and colonization history of the two cytotypes and have been maintained by multiple factors such as ecological divergence, limited gene flow between the cytotypes, and the restricted dispersal capacity.

• Klíčová slova
• AFLPs, Alps, Brassicaceae, autopolyploidy, contact zone, cytotype coexistence, environmental predictors, microsatellites, polyploidy,
• MeSH
• analýza polymorfismu délky amplifikovaných restrikčních fragmentů MeSH
• Cardamine genetika   fyziologie   MeSH
• chromozomy rostlin genetika   MeSH
• diploidie MeSH
• distribuce rostlin * MeSH
• ekosystém * MeSH
• genetická variace * MeSH
• hybridizace genetická MeSH
• mikrosatelitní repetice MeSH
• tetraploidie MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Evropa MeSH

Plant species diversity may be considerably underestimated, especially in evolutionarily complex genera and in diversity hotspots that have enabled long-term species persistence and diversification, such as the Balkan Peninsula. Here, we address the topic of underexplored plant diversity and underlying evolutionary and biogeographic processes by investigating the hygrophytic mountain species complex of Cardamine acris s.l. distributed in the Balkans (three subspecies within C. acris) and northwestern Anatolia (C. anatolica). We performed a series of phylogenetic and phylogeographic analyses based on restriction-site associated DNA sequencing (RADseq) and target enrichment (Hyb-Seq) data in combination with habitat suitability modelling. We found C. anatolica as a clade nested within the Balkan C. acris, probably resulting from a founder event, and uncovered three allopatric cryptic lineages within C. acris subsp. acris, allowing us to recognise a total of six entities in this complex. We observed the deepest genetic split within C. acris subsp. acris in the western Balkans, which was at odds with taxonomy and showed no distribution gap. We inferred vicariance as the most likely process for population divergence in the Balkans, accompanied by gene flow between the recognised entities, which was consistent with the modelled habitat suitability dynamics. Furthermore, we discovered several polyploid populations in C. acris, representing both pure intra- and inter-lineage hybrid polyploids, but detected only minor traces of hybridization with related congeners. Overall, our results illustrate that diverse evolutionary processes may influence the history of mountain plant species in the Balkan Peninsula, including vicariance, reticulation, polyploidization and cryptic diversification.

• Klíčová slova
• Hyb‐Seq, RAD sequencing, habitat suitability modelling, phylogeography, polyploidy, vicariance,
• MeSH
• Cardamine * genetika   klasifikace   MeSH
• DNA rostlinná genetika   MeSH
• ekosystém MeSH
• fylogeneze * MeSH
• fylogeografie * MeSH
• genetická variace MeSH
• genom rostlinný MeSH
• populační genetika MeSH
• sekvenční analýza DNA MeSH
• tok genů MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Balkánský poloostrov MeSH
• Turecko MeSH
• Názvy látek
• DNA rostlinná MeSH

Recently formed allopolyploids represent an excellent system to study the impacts of hybridization and genomic duplication on genome structure and evolution. Here we explored the 35SrRNA genes (rDNA) in the Cardamine × schulzii allohexaploid that was formed by two subsequent hybridization events within the past c. 150 yr. The rDNA loci were analyzed by cloning, next generation sequencing (NGS), RT-PCR and FISH methods. The primary C. × insueta triploid hybrid derived from C. rivularis (♀) and C. amara (♂) had gene ratios highly skewed towards maternal sequences. Similarly, C. × schulzii, originating from the secondary hybridization event involving C. × insueta (♀) and C. pratensis (♂), showed a reduction in paternal rDNA homeologs despite an excess of chromosomes inherited from C. pratensis. We also identified novel rDNA loci in C. × schulzii, suggesting that lost loci might be slowly reinstalled by translocation (but not recombination) of genes from partner genomes. Prevalent clonal propagation of allopolyploids, C. × insueta and C. × schulzii, indicates that concerted evolution of rDNA may occur in the absence of extensive meiotic cycles. Adoption of NGS in rDNA variant analysis is highly informative for deciphering the evolutionary histories of allopolyploid species with ongoing homogenization processes.

• Klíčová slova
• Brassicaceae, concerted evolution, hybridization, hybrids, polyploidy, rDNA,
• MeSH
• Cardamine genetika   MeSH
• chromozomy rostlin genetika   MeSH
• druhová specificita MeSH
• fylogeneze MeSH
• genetická variace MeSH
• genetické lokusy * MeSH
• genom rostlinný MeSH
• klonování DNA MeSH
• křížení genetické * MeSH
• mezerníky ribozomální DNA genetika   MeSH
• polyploidie * MeSH
• regulace genové exprese u rostlin MeSH
• ribozomální DNA genetika   MeSH
• sekvence nukleotidů MeSH
• vysoce účinné nukleotidové sekvenování MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• mezerníky ribozomální DNA MeSH
• ribozomální DNA MeSH

The nomenclature of Eastern Asian populations traditionally assigned to Cardamine flexuosa has remained unresolved since 2006, when they were found to be distinct from the European species Cardamine flexuosa. Apart from the informal designation "Asian Cardamine flexuosa", this taxon has also been reported under the names Cardamine flexuosa subsp. debilis or Cardamine hamiltonii. Here we determine its correct species name to be Cardamine occulta and present a nomenclatural survey of all relevant species names. A lectotype and epitype for Cardamine occulta and a neotype for the illegitimate name Cardamine debilis (replaced by Cardamine flexuosa subsp. debilis and Cardamine hamiltonii) are designated here. Cardamine occulta is a polyploid weed that most likely originated in Eastern Asia, but it has also been introduced to other continents, including Europe. Here data is presented on the first records of this invasive species in European countries. The first known record for Europe was made in Spain in 1993, and since then its occurrence has been reported from a number of European countries and regions as growing in irrigated anthropogenic habitats, such as paddy fields or flower beds, and exceptionally also in natural communities such as lake shores.

• Klíčová slova
• Asian Cardamine flexuosa, Brassicaceae, Cardamine flexuosa subsp. debilis, Cardamine hamiltonii, Cardamine occulta, China, Cruciferae, Europe, invasive species, typification,
• Publikační typ
• časopisecké články MeSH