Annals of Botany

1. Stomata and leaf traits are key regulators of plant water use efficiency and are expected to have changed in response to rising atmospheric CO2 concentrations and climate warming over the past centuries. However, long-term data documenting such changes are rare. 2. We leveraged herbarium collections to track changes in stomatal characteristics and leaf traits in 656 individuals of invasive Japanese knotweed and its hybrid Bohemian knotweed collected across their European range and spanning 160 years of invasive spread. 3. We found that several functional traits including stomatal density and maximum anatomical stomatal conductance did not show significant changes over time but that plants adjusted their stomatal size and shape over time, and that these changes were associated with increased atmospheric CO2 levels. Interestingly, Reynoutria japonica showed increases in stomatal size and stomatal elongation, while the hybrid R. x bohemica showed a reduction in stomatal size. Traits also varied systematically with climates of origin. Plants from warmer origins with higher evaporative demands during the growing season had thicker leaves, lower SLA, smaller stomata and higher stomatal density, indicating more conservative water-use strategies. Stomatal density and gas exchange capacity co-varied with leaf structural traits, and there was a trade-off between stomatal size and number. Overall, fast leaf economic traits were associated with slow physiological traits. 4. Our results suggest that stomatal anatomical plasticity may enhance climate resilience by maintaining a stable maximum gas exchange capacity across environmental gradients. Herbarium collections provide a unique resource for reconstructing plant responses to historical environmental changes and understanding intraspecific trait variation.

Keel flowers are bilaterally symmetrical, pentamerous flowers with the reproductive organs enclosed by keel petals. Within Fabales, keel flowers are dominant in two species-rich lineages, tribe Polygaleae (Polygalaceae) and subfamily Fabaceae (Papilionoideae); however, independent events are also observed, such as in the genus Cercis. Before phylogenetic advancements were available (i.e., in contrast to more recent studies), most of the studies hypothesized a non-keeled origin for the Faboideae, although a detailed investigation has never been carried out. In this study, using the results of Aygoren Uluer et al. (2020, 2022a), the origin of some important morphological characters of the keel flower are examined, namely floral symmetry, perianth heteromorphism (i.e., three distinct petal/+sepal types), and the presence of enclosed reproductive organs. These characters are analysed within the Fabales using three different ancestral state analyses based on a phylogeny constructed for 678 taxa using published matK, rbcL and trnL plastid gene regions. The analyses show that symmetry probably originated in the (Fabaceae+Polygalaceae) clade, while the enclosed reproductive organs and three-types of petals appear to have evolved independently multiple times. Interestingly, not only the enclosed reproductive organs but also petal heteromorphism probably did not evolve in the MRCA of the Faboideae, but rather in a very early stage of the evolution of the subfamily. While future homology assessments and/or evolutionary developmental genetic (evo-devo) studies will be required to more clearly elucidate the evolutionary processes, the current study is the first attempt to investigate the origin of some important characteristics of keel flowers within order Fabales.

Understanding evolutionary relationships in hyperdiverse plant groups remains a major challenge in systematics. The orchid genus Bulbophyllum, the second largest genus of flowering plants, represents an exceptional example of phylogenetic and morphological complexity. Relationships, particularly within the species-rich Asian clade, have remained poorly resolved due to extensive morphological variation and limited resolution in previous phylogenetic studies. Here, we reconstructed phylogenetic relationships using 63 plastid genes from 355 specimens representing 322 species and 65 of the 97 recognised sections of Bulbophyllum. Our analyses confirmed that the genus comprises five major evolutionary lineages comprised of species predominantly from Australasia, Madagascar, Continental Africa, Neotropics, and Asia. We provide the first robust phylogenetic evidence for a dichotomous split within the Asian clade into two well-supported lineages: the Asian-Malesian clade and the Malesian-Papuasian clade, with the latter containing a strongly supported Papuasian subclade. Additionally, this study supports the monophyly of several currently recognised sections while clarifying relationships in previously problematic groups. This study provides the most comprehensive plastid-based phylogenomic framework for Bulbophyllum to date and establishes a foundation for future taxonomic revision and integrative analyses of diversification and trait evolution within this hyperdiverse genus.

Premise of studyUnder increasingly frequent pollinator-limited environments, plants need to rely on modes of reproductive assurance such as selfing and cloning. However, few studies investigate the interplay between selfing and cloning in plants that can do both. Here, we explore mechanisms determining the relative expression of selfing and cloning throughout the European distribution of the wild woodland strawberry (Fragaria vesca) under a pollinator-free environment. MethodsWe established an outdoor common garden with 121 woodland strawberry genotypes from across Europe and excluded them from pollinators. For each genotype, we recorded reproductive traits and performed hand-pollination treatments. Key ResultsWe found a weak trade-off between cloning and selfing, driven by increased seed and fruit provisioning rather than flower production. The capacity to autonomously self-fertilize was determined by the lateral proximity of the anthers to the pistils (lateral herkogamy), but not by early inbreeding depression. Genotypes sampled at lower latitudes and altitudes were better at self-fertilizing and had smaller petals. The propensity to clone increased towards the east, where genotypes also had smaller petals, particularly at higher latitudes. ConclusionAt the species level, we detected a trade-off between the propensity for clonal reproduction and the capacity for self-fertilization. At a continental scale, the capacity to self-fertilize varied along a north-south gradient, whereas clonal propensity varied along an east-west gradient. Our results suggest that these two modes of reproductive assurance may compensate for reduced pollinator attractiveness (smaller petals) in regions where each mode is most strongly expressed.

The orchid genus Dipodium R.Br. (Epidendroideae) comprises leafy autotrophic and leafless mycoheterotrophic species, the latter confined to sect. Dipodium. This study examined plastome degeneration in Dipodium in a phylogenomic and temporal context. Whole plastomes were reconstructed and annotated for 24 Dipodium samples representing 14 species and two putatively new species, encompassing over 80% of species diversity in sect. Dipodium. Phylogenomic analysis based on 68 plastid loci including a broad outgroup sampling across Orchidaceae found sect. Leopardanthus as sister lineage to sect. Dipodium. Dipodium ensifolium, the only leafy autotrophic species in sect. Dipodium was found sister to all leafless, mycoheterotrophic species, supporting a single evolutionary origin of mycoheterotrophy in the genus. Divergence time estimations found that Dipodium arose ca. 33.3 Ma near the lower boundary of the Oligocene and crown diversification commenced in the late Miocene, ca. 11.3 Ma. Mycoheterotrophy in the genus was estimated to have evolved in the late Miocene, ca. 7.3 Ma, in sect. Dipodium. The comparative assessment of plastome structure and gene degradation in Dipodium revealed that plastid ndh genes were pseudogenised or physically lost in all Dipodium species, including in leafy autotrophic species of both Dipodium sections. Levels of plastid ndh gene degradation were found to vary among species as well as within species, providing evidence of relaxed selection for retention of the NADH dehydrogenase complex within the genus. Dipodium exhibits an early stage of plastid genome degradation as all species were found to have retained a full set of functional photosynthesis-related genes and housekeeping genes. This study provides important insights into plastid genome degradation along the transition from autotrophy to mycoheterotrophy in a phylogenomic and temporal context.

In style length polymorphism, morph-specific sequence of encountering male and female sex organs within a flower by pollinators can cause differences in the need to avoid self-pollination and encourage inter-morph pollination. We asked if this difference can lead to disparity in stigma-anther separation (herkogamy) between morphs and spatial match between sex organs of complementary morphs (reciprocity). Further, we tested if herkogamy, and hence the level of selfing, is fairly constant among individuals of a population. Additionally, we examined the relationship between herkogamy and reciprocity among individuals of a population to understand functional interactions between these two morphological traits. Using data on sex organ heights for >200 heterostylous species from the literature, we observed that the short-styled morph had higher herkogamy as compared to the long-styled morph indicating a higher need to avoid selfing. Reciprocity did not show a consistent difference between the upper and lower sex organs implying a strong influence of local ecological factors. In most populations, allometric relationships suggested that herkogamy and hence the level of selfing remains constant. Finally, we observed that herkogamy and reciprocity can be related among individuals of a population, sometimes indicating a potential trade-off between avoidance of self-pollination and facilitation of inter-morph pollination.

Elucidating the origin of carpel has been a challenge in botany for a long time. The Unifying Theory suggested that the carpel originate from a composite organ comprising an ovule-bearing shoot and a foliar part enclosing the shoot. A logical inference from this theory is that placenta in angiosperms should have radiosymmetrical vasculature, just like that in a young branch. Anaxagorea is the most basal genus of the primitive angiosperm family, Annonaceae. The conspicuous carpel stipe makes it an ideal material for exploring the carpel vasculature. In this study, serial sections of flower and carpel were delineated in Anaxagorea luzonensis and A. javanica, and a three-dimensional model of the carpel vasculature was reconstructed. The results show that (1) vascular bundles at both the carpel stipe and the ovule/placenta are in a radiosymmetrical pattern, (2) the amphicribral bundles would develop into ring-arranged bundle complex with the carpel maturation, (3) the ovule/placenta bundles were separated from the bundles of the carpel wall, and, (4) all the radiosymmetrical vasculature (including amphicribral bundles and ring-arranged bundle complexes) in the carpel were fed by a larger radiosymmetrical bundle system. These results suggest that the radiosymmetrical pattern of carpel vasculature are in line with the Unifying Theory.

PREMISEHedychium J.Koenig (ginger lilies: Zingiberaceae) is endemic to the Indo-Malayan Realm (IMR) and is known for its fragrant flowers. Two different pollination syndromes characterize the genus: diurnal or bird pollination and nocturnal or moth pollination systems. To date, no attempt has been undertaken to understand the evolution of floral traits in this genus. METHODSWe estimated ancestral character-states, phylogenetic signals, and character correlations for thirteen discrete and eight continuous floral traits representing 75% species diversity of Hedychium. Diversification rate estimation analyses were also employed to understand trait-dependent diversification in the genus. RESULTSInflorescence structure, cincinnus capacity, and curvature of floral tubes revealed strong phylogenetic dependence, whereas number of open flowers per inflorescence per day, color of the labellum, and exertion of the stigma characterized higher ecological effects. Diversification rate estimations suggested that the labellum width, floral tube length, and labellum color played a major role in the evolutionary diversification of Hedychium. CONCLUSIONSWe identified bract type and cincinnus capacity as synapomorphies for Hedychium, while the island-specific clade III was characterized by slender cylindrical inflorescence, coiling of floral tubes, and longer bract to calyx ratio. The circum-Himalayan clade IV is the most speciose, derived, and with most variable floral traits. Although floral color and size lacked any association with pollinator-specific traits (moth and bird pollination), pale colored flowers were most common in the early diverging clades (clade I, II-el., and II-de.), indicating their ancestral nature, when compared to brightly colored flowers.

Serjania is the only genus of Paullinieae that exhibits all types of vascular variants in stems and includes S. piscatoria with a complex vascular structure that has intrigued botanists for centuries. Here, we analyzed the stem development of S. piscatoria in an evolutionary context and determined its phylogenetic position within the genus. We studied four individuals using standardized anatomical techniques and employed DNA sequencing and phylogenetic analysis to determine the species phylogenetic position. Additionally, we employed ancestral state reconstruction to explore the pattern of evolution of vascular variants. We find that the stem development in S. piscatoria is determined by various ontogenetic processes that result in vascular variants that occur through modifications during primary and/or secondary growth, or ectopic cambia formation. These various patterns are classified into distinct categories of vascular variants, highlighting the lability of vascular meristems and the polymorphism within the species, which manifests across different individuals. Serjania piscatoria belongs to a clade composed of species with compound stems, from which the fissured stems observed in the species would have evolved. The findings provide evidence for the diverse stem vasculature in Serjania, and the importance of studying vascular variant diversity from a developmental and evolutionary perspective.

Background and AimsThe evolution of reproductive isolation between previously interbreeding populations is a fundamental driver of plant speciation. Within Amaryllidaceae, Nothoscordum represents an evolutionarily complex genus, characterized by an unusually high incidence of chromosomal rearrangements. During fieldwork, Nothoscordum montevidense and Nothoscordum bonariense were found growing in sympatry, along with individuals exhibiting intermediate morphological traits, suggesting a putative hybrid origin. To test this hypothesis, we employed an integrative approach to characterize the morphologically intermediate specimens and the two sympatric populations. Materials and methodsTo characterize the putative hybrids we have combined morphological, cytogenetic analyses (chromosome counts, CMA/DAPI banding, and FISH) and flow cytometry-based genome size estimation. Phylogenetic relationships and genomic structure were also investigated through Genotyping-by-Sequencing (GBS), complete chloroplast genome assembly, and comparative repetitive DNA analysis. We also performed species distribution modeling and phenological analyses of the putative parental species. Key ResultsMultiple lines of evidence confirm the hybrid origin of the studied plants. Cytogenetic analyses revealed specimens with 2n = 21 (1C {approx} 33 pg = 32.274 Mbp) and 2n = 25 (1C {approx} 37 pg = 36.186 Mbp), accompanied by meiotic irregularities consistent with interspecific hybridization. Chloroplast genome phylogeny identified N. montevidense (2n = 16, 1C {approx} 25 pg) as the maternal lineage, while GBS data confirmed N. bonariense (2n = 26, 1C {approx} 41 pg) as the paternal contributor and revealed evidence of subsequent backcrossing. Comparative analysis of repetitive DNA showed reduced 35S rDNA diversity in the hybrid, indicative of post-hybridization genomic restructuring. Despite the observed genomic complexity, no clear morphological differentiation was detected among hybrid individuals. Phenological analyses and species distribution models demonstrated broad overlap between parental species. ConclusionsOur findings highlight the role of hybridization in shaping genome architecture in cytogenetically labile plant lineages. Furthermore, our results underscore that morphological similarity can mask profound genomic complexity, reinforcing the value of integrative approaches to understand genera characterized by reticulate evolution and genomic instability.

Male reproductive investment, in particular pollen production, is a crucial and ecologically relevant component of a plants phenotype and reproductive success. Its evolutionary trajectory, however, remains understudied, partly due to a lack of convenient methods to assess it. We developed a protocol for pollen quantification by flow cytometry and applied it to 107 flowers from 38 Papaveraceae species differing widely in floral traits (e.g., floral symmetry, stamen number), pollination syndromes (e.g., wind and insect pollination) and reproductive systems (e.g., degree of autogamy). We phylogenetically tested whether pollen number evolved in association with ovule, carpel, stamen and flower numbers per inflorescence, and if there were interacting effects between floral symmetry and/or self-compatibility with pollen and ovule production. Compared to manual counts, results using flow cytometry were similar, but much faster to obtain and more precise. Pollen and ovule numbers per flower varied > 39,000x and > 550x, respectively, among species. Pollen production correlated positively with ovule, carpel and stamen numbers. Lineage-specific trajectories to pollen-to-ovule ratio reduction (to values < 300) are observed. One involved increased female investment in ruderal species belonging to the subfamily Papaveroideae, while the other occurs through decreased male investment and is associated with the evolution of floral traits towards greater specialisation. The impact of reproductive systems on male and female investment is limited to ovule production in non-actinomorphic flowers. Taken together, these results revealed that the evolutionary associations between reproductive systems, floral traits, and pollen and ovule production are lineage-specific. Given the profound contrasts at the subfamily level of Papaveraceae, broader surveys across the diversity of flowering plants are clearly needed to better understand factors driving the evolution of reproductive investment. Such studies will certainly be facilitated by our new high-throughput pollen counting method outlined here.

BackgroundPlants are exposed to various environmental challenges. With ongoing climate change, droughts and insect outbreaks are expected to become more frequent. Thus, a better understanding is needed of how different plant species respond to such single and combined challenges. This study investigated common versus species-specific responses to environmental challenges in three perennial plant species of different growth forms and whether responses differ intraspecifically among accessions. Clones of different accessions of the herbaceous species Tanacetum vulgare, the woody vine Solanum dulcamara, and the tree Populus nigra were subjected to similar control, herbivory, drought, and combined (drought and herbivory) treatments for the same periods. After the exposure, concentrations of foliar phytohormones and various morphological traits were measured. ResultsAcross all species, several foliar phytohormones and one of ten morphological traits responded consistently to the environmental challenges. Jasmonoyl-isoleucine was induced by herbivory and the combined treatment, abscisic acid (ABA) by drought and the combined treatment, and indole acetic acid by the combined treatment in all species. Root mass remained unchanged in all species. However, structural equation models (SEMs) revealed a shared regulatory pathway across species in which ABA connected treatment and root mass, indicating a common hormonal response potentially linking challenges to growth responses. Despite these common patterns, species-specific responses were pronounced. In P. nigra, a unique induction of salicylic acid was found under the combined treatment, while aboveground mass and root-shoot ratio remained unaffected by any treatment, in contrast to the other two species. Species-specific SEMs further indicated distinct phytohormone-mediated pathways underlying morphological variation. Phenotypic plasticity reflected these species-specific patterns, with none of the phytohormones or morphological traits exhibiting uniform plasticity across species. Intraspecific variation further shaped responses, as phytohormone and morphological trait plasticity depended on accession, indicating substantial accession-specific plant responses. ConclusionsOur results indicate that some responses to comparable challenges may be conserved across species, while others are species-specific. The combined treatment elicited the most pronounced responses, and such complex responses may become more frequent under current global change. Our study highlights that comprehensive understanding of plant responses requires systematic comparisons at both interspecific and intraspecific scales.

Attracting specific pollinators can be favoured by natural selection to avoid reproductive interference between sympatric plant species. However, the ways in which fine differences in floral traits lead to the attraction of specific pollinators are diverse and unknown in many pollination interactions. We surveyed pollinators on three sympatric Aristolochia species (A. clematitis, A. pistolochia and A. rotunda) pollinated by Diptera to investigate if specific pollination occurs. To decipher if specific pollination may be mediated by different floral odours, we characterized the volatile organic compounds (VOCs) emitted by flowers and highlighted those VOCs electrophysiologically detected by pollinators in A. rotunda and A. pistolochia. Among the most abundant pollinators, Forcipomyia monilicornis was a specific pollinator of A. pistolochia while two Dasyhelea species were specific pollinators of A. clematitis. Forcipomyia aristolochiae and T. ruficeps were non-specific pollinators of A. rotunda, although they were more frequently found in A. rotunda flowers. The floral odours of A. rotunda and A. pistolochia differed significantly from each other and elicited specific electrophysiological responses in their respective pollinators. Although several pollinator species visit more than one Aristolochia species, those pollinators are preferentially found in one Aristolochia species. Selective attraction is likely mediated by specific VOCs.

Aristolochia subgenus Siphisia constitutes a monophyletic lineage of predominantly lianescent species, with occasional shrubs or herbs, and is characterized by remarkable diversity in perianth morphology. Members of Siphisia serve as larval hosts for endangered Lepidoptera and are widely used in traditional medicine. Despite its ecological and ethnobotanical significance, Siphisia systematics remains unresolved due to limited genomic resources and insufficient phylogenetic signal across previously sampled loci. Here, we present a phylogenomically informed framework for Siphisia, integrating 46 newly collected accessions and seven public datasets across 44 taxa. Using genome skimming ([~]30x coverage) and HybPiper, we recovered Angiosperms353 nuclear loci, including supercontigs, for phylogenetic reconstruction via concatenation and coalescent approaches. The resulting species trees resolve seven strongly supported monophyletic clades, each defined by distinct biogeographic patterns and morphological synapomorphies. Comparative plastome analyses from de novo assemblies explored quadripartite structure, plastid phylogeny, GC content, and gene synteny. Cytonuclear discordance was concentrated in species-rich Asian clades, while hybridization signals were rare and limited to deep backbone nodes in North American lineages. These results indicate that incomplete lineage sorting, rather than introgression, accounts for most gene-species tree conflicts and likely reflects strong reproductive isolation following speciation. We also revise a previously taxonomically ambiguous complex--the now well-supported A. versicolor species group--based on integrated phylogenomic and morphological evidence. Within this group, we describe five previously unrecognized cryptic species and identify a novel pollination syndrome involving floral adaptations for pollinator filtering. This syndrome may contribute to prezygotic isolation and recent diversification, and it challenges the prevailing assumption that Aristolochia pollination is universally governed by a trapping-release mechanism--suggesting this model may not apply to subg. Siphisia.

PremiseThe relationships between reproductive investment, phenotype and fitness have been broadly studied in cross-pollinated plants in contrast to selfing species, which are considered less interesting in this area because they are supposed to be a dead-end in any evolutionary pathway. Still, selfing plants are unique systems to study these questions since the position of reproductive structures and traits related to flower size play an important role in female and male pollination success. MethodErysimum incanum s.l. is a selfing species complex exhibiting three levels of ploidy: diploids, tetraploids and hexaploids. This species complex shows traits typically associated with the selfing syndrome. Here, we used 1609 plants belonging to these three ploidies to characterize floral phenotype and spatial configuration of reproductive structures, reproductive investment (pollen and ovules production) and plant fitness. Then, we explored the relationship between all these variables using structural equation modelling across ploidy levels. Key ResultsAn increase in ploidy level leads to bigger flowers with more exerted stamens and a greater amount of pollen and ovules. In addition, hexaploid plants exhibit higher absolute values for herkogamy which is positively correlated with fitness. Phenotypic traits and pollen production are indirectly selected by the relationship among ovules and fitness, maintained across ploidies. ConclusionsChanges in floral phenotypes, reproductive investment and fitness with the ploidy level suggests that genome duplication can be a driver for the reproductive strategy transitions by modifying the investment in pollen and ovules and linking them with plant phenotype and fitness.

PREMISERhododendron L. (Ericaceae Juss.), is the most species-rich genus of woody flowering plants with > 1000 species. Despite the interest in the genus and numerous previous phylogenetic analysis, the infrageneric classification for the genus is still debated, partly due to its huge diversity, partly due to homoplasy in key characters and partly due to incongruence between phylogenetic markers. Here, we provide a broad coverage of representative species of all Rhododendron subgenera, sections, and most subsections to resolve its infrageneric phylogeny or highlight areas of incongruence, support previous analyses of diversification patterns and establish a relationship between genome size evolution and its diversification. METHODSWe generated sequences of two plastid (trnK and trnL-F) and two nuclear (ITS and rpb2-i) markers for a total of 259 Rhododendron species, and used likelihood and Bayesian statistics to analyze the data. We analyzed the markers separately to discuss and understand incongruence among the data sets and among previous studies. RESULTSWe found that the larger a subgenus, the more strongly it is supported as monophyletic. However, the smaller subgenera pose several problems, e.g., R. subgen. Azaleastrum consists of two sections inferred to be polyphyletic. The main shift to higher diversification in the genus occurred in the Himalayan/SE Asian clade of R. subgen. Hymenanthes. We found that polyploidy occurs in almost all subgenera but most polyploid species are within R. subgen. Rhododendron sections Rhododendron and Schistanthe. CONCLUSIONWhereas previous reports stated that genome sizes of tropical plants are lower than those of colder and temperate regions in angiosperms in general, our study provides evidence for such a shift to small genome-tropical species within a genus. Taken together, we see the merit in the recognition of the five major clades at the sub generic level but given the amount of incongruence a large amount of species cannot be confidently assigned to one of these five clades. Further, genome-wide data will be necessary to assess whether these currently unassignable taxa are independent taxa, assignable to one of the five major clades or whether they are inter-subgeneric hybrids.

Global coffee production faces increasing threats from climate change, including rising temperatures, prolonged droughts, and spreads of diseases. Wild coffee species, notably those growthing in highly constrained environments in Madagascar, offer a critical genetic resource to address these challenges. Understanding adaptive traits allowing these species to establish into arid environments is essential to implement guid breeding strategies into create resilient coffee varieties. Here, we hypothesize that these wild Coffee species display unique traits compared to other Coffea species, reflecting adaptations to arid and seasonally dry environments. We used the architectural analysis to describe three Baracoffea species and compare them to known cultivated Coffee. Field studies were conducted in two contrasting sites in Madagascar, focusing on three wild coffee species, natively growing into arid environments: Coffea ambongensis, C. bissetiae, and C. boinensis. Structural traits at the whole plant scale were measured across developmental stages using morphological and architectural analyses. Our results suggest that unique traits characterize Barracoffea species, such as rhythmic growth, terminal flowering on short shoots, and species-specific developmental strategies. These findings highlight the architectural diversity of Baracoffea, identify potential key drivers of ecological adaptation and therefore highlights the potential of this group of species for breeding climate- resilient coffee varieties.

Background and aimsThe way plants cope with biotic and abiotic selective pressures determines their success in the colonization of remote oceanic islands, which ultimately depends on the phylogenetic constrains and ecological response of the lineage. In this study we aim to evaluate the relative role of geographical and ecological forces in the origin and evolution of the Madeiran ivy (H. maderensis). MethodsTo determine the phylogenetic placement of H. maderensis within the western polyploid clade of Hedera (three species), we analysed 40 populations (92 individuals) using genotyping-by-sequencing and including H. helix as outgroup. Climatic niche differences among the four study species were evaluated using a database with 706 records representing the entire species ranges. To test species responses to climate, a set of 19 vegetative and regenerative functional traits were examined for 70 populations (335 individuals). Key resultsPhylogenomic results revealed a nested pattern with H. maderensis embedded within H. iberica. Gradual niche differentiation from the coldest and most continental populations of H. iberica to the warm and stable coastal population sister to H. maderensis parallels the geographical pattern observed in the phylogeny. Similarity in adaptive traits is observed for H. maderensis and H. iberica. The two species show leaves with higher SLA, lower LDMC and thickness and smaller fruits than those of H. hibernica. ConclusionsAcquisition of the Macaronesian climatic niche and the associated functional syndrome in mainland European ivies (small fruits, leaves high SLA, and low LDMD and thickness) was a key step in the colonization of Madeira by the H. iberica/H. maderensis lineage, which points to climatic pre-adaptation as a driver of island colonization (dispersal and establishment). Once in Madeira, speciation was driven by geographical isolation, while ecological processes are regarded as secondary forces with a putative impact in the lack of further in situ diversification.

Self-fertilization has recurrently evolved from outcrossing. Self-fertilization provides an advantage in the short-term as individuals do not require a mate to reproduce, but self-fertilization is also associated with both decreased genetic diversity and accumulation of weakly deleterious mutations, which could, however, be alleviated in polyploid selfers. If pollinators are not limited, individual fitness is thus expected to be higher in outcrossers than in selfers. We measured several life history traits in four Capsella species under two different treatments (disturbed and undisturbed) to assess the effects of mating system and ploidy level on reproductive, vegetative and phenological traits. The experiment was carried out outdoor in Northwest Greece, within the range of the obligate outcrossing species, C. grandiflora, so it could be naturally pollinated and its fitness directly compared to that of its self-fertilizing relatives. Disturbance of the environment did not affect the phenotype in any of the four species. However, for most traits the obligate outcrossing species performed better than all selfing ones. In contrast, polyploidy did not seem to confer an advantage in terms of survival or reproduction compared to diploidy. Finally, plants from Asia and northern Europe had lower performances than accessions from southern Europe and the Middle-East.

Leaf shape varies widely across plant taxa and has repeatedly been shown to affect ecophysiology, interspecific interactions, and fitness. We used population genomics, genome wide association studies (GWAS), and comparative genomics to determine the genetic basis and evolutionary history of an uncharacterized Mendelian leaf shape polymorphism in Ipomoea hederacea. To do so, we assembled a reference genome and generated whole genome sequencing for 123 individuals from 55 populations. We identified a 117 kb indel that perfectly co-segregates with leaf shape by conducting a GWAS and assessing differences in coverage. Syntenic orthologs for genes on the indel were present in five other Ipomoea species with various leaf shapes, indicating the indel is newly arisen deletion in I. hederacea despite similar leaf shape phenotypes in the other species. More broadly, these results illustrate how a range of leaf shape phenotypes can be produced by distinct genetic mechanisms even in closely related species. Although none of the genes on the indel itself are known leaf shape candidates, there are multiple leaf shape candidate genes in close proximity that are involved in the auxin biosynthesis pathway. Additionally, the genes within the indel have gene functions that could affect other potentially ecologically relevant traits that have previously been shown to be associated with leaf shape in I. hederacea. Therefore, the pleiotropic effect of the indel polymorphism can have important implications for understanding the ecological mechanisms influencing a well-documented leaf shape latitudinal cline in I. hederacea. Significance StatementIpomoea hederacea has been used to investigate the ecological and evolutionary effects of leaf shape because of a well-documented latitudinal leaf shape cline governed by an uncharacterized Mendelian polymorphism in this species. We identified a 117 kb indel that perfectly co-segregates with leaf shape in I. hederacea. The indel appears to be a distinct genetic mechanism than those governing similar leaf shapes in other closely related Ipomoea species. Additionally, possible pleiotropic effects of the indel on other traits has important implications for understanding the ecology and evolution of leaf shape variation in I. hederacea. Therefore, both the results and the new genomic resources that we developed will help facilitate future work understanding the genetic, developmental, and ecological mechanisms governing leaf shape variation.