Annals of Botany

Background and AimsWhile genome size limits the minimum sizes and maximum numbers of cells that can be packed into a given leaf volume, mature cell sizes can be substantially larger than their meristematic precursors and vary in response to abiotic conditions. Mangroves are iconic examples of how abiotic conditions can influence the evolution of plant phenotypes. MethodsHere, we examined the coordination between genome size, leaf cell sizes, and cell packing densities, and leaf size in 13 mangrove species across four sites. Four of these species occurred at more than one site, allowing us to test the effect of climate on leaf anatomy. ResultsWe found that genome sizes of mangroves were very small compared to other angiosperms, and, like other angiosperms, mangrove cells were always larger than the minimum size defined by genome size. Increasing mean annual temperature of a growth site led to higher packing densities of veins (Dv) and stomata (Ds) and smaller epidermal cells but had no effect on stomatal size. Contrary to other angiosperms, mangroves exhibited (1) a negative relationship between guard cell size and genome size; (2) epidermal cells that were smaller than stomata, and (3) coordination between Dv and Ds that was not mediated by epidermal cell size. Furthermore, mangrove epidermal cell sizes and packing densities covaried with leaf size. ConclusionsWhile mangroves exhibited coordination between veins and stomata and attained a maximum theoretical stomatal conductance similar to other angiosperms, the tissue-level tradeoffs underlying these similar relationships across species and environments was markedly different, perhaps indicative of the unique structural and physiological adaptations of mangroves to their stressful environments.

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 study of morphological diversity (i.e., disparity) offers unique opportunities to understand evolutionary patterns and processes. Plant disparity studies reveal that morphological disparification can be related to factors such as secondary woodiness or to pollination niche, for example. Similarly, some pollen traits are known to be shaped by environmental pressures, but this influence has only been evaluated in monads, never in multi-grained dispersal units. In this study, we investigated the disparity of aggregated dispersal units in two lineages of Neotropical mimosoid legumes. The Mimosa and Stryphnodendron clades are independent lineages that share similarities in pollen morphology and biome shifts. In this context, we asked: What are the patterns of pollen disparity in these lineages, and are these patterns similar between lineages occurring in the same biomes? MethodsTo answer these questions, we compiled data from the literature on pollen morphology and biomes of occurrence for a phylogenetically representative set of taxa in the Mimosa and Stryphnodendron clades. With these data, we calculated morphospaces and disparity metrics, and tested whether the pollen morphology of distinct lineages occurring in the same biome differs significantly. Key ResultsOur results show that Mimosa and Stryphnodendron clades exhibit distinct patterns of pollen disparity, as do independent lineages occurring in the same biomes. Additionally, we observed that certain biomes support greater or lesser levels of morphological disparity. ConclusionsWe conclude that (1) the Mimosa clade has greater disparity, possibly due to evolution of novel pollen morphologies in the genus Mimosa, (2) there is a maintenance of similarities in the pollen of the Stryphnodendron clade, Adenopodia and Piptadenia, and (3) the evolution of pollen grains in these groups appears to be primarily shaped by phylogeny and developmental constraints, with environmental pressures playing a comparatively smaller role.

Summer drought represents one of the main stress sources stress for plant communities in the Mediterranean region. Plants can adopt several response strategies to cope with stress, reflected in the adoption of specific Plant Functional Traits (PFTs). Trait-based approaches commonly meet three critical issues: they overlook Intraspecific Variability (ITV), they focus on a large spatial scale, or they focus on single trait responses to stress. In this study, we present evidence for a significant amount of ITV in morphological and anatomical trait syndromes observed between three local populations of Phyllirea latifolia, Pistacia lentiscus and Quercus ilex, distributed along an aridity gradient. Thicker, more physiologically expensive leaves and lower heights found in the drier sites mainly conform to drought-resistance strategies. Interestingly, PFTs from Cistus salviifolius were found not to vary between sites. This implies that not all species vary at the same geographical scale, possibly depending on their different successional role. The main implication behind our findings is that climate can easily drive significant ITV in multiple traits among plant populations, even at a local scale, although trait responsiveness might be species-specific. Different plant populations hailing from the same geographical regions might thus respond differently to climate change. HIGHLIGHTVariations in Plant Functional Traits from several Mediterranean species found along an aridity gradient on a local (<60Km radius) scale; responses consistent to reported drought adaptations.

O_LIHydraulic, stomatal and anatomical traits strongly influence drought survival and productivity, but the extent to which these trait-types are coordinated and relate to climate-of-origin in herbaceous species remains poorly understood. C_LIO_LIWe quantified hydraulic, stomatal and anatomical traits in eight populations of Themeda triandra selected from diverse temperature and precipitation regimes across Australia, grown under common conditions and exposed to drought. Trait - climate-of-origin relationships were quantified using multiple linear mixed models. C_LIO_LILeaf xylem embolism resistance (P50) was unrelated to climate-of-origin precipitation. Instead, plants from drier climates exhibited lower maximum stomatal conductance, earlier stomatal closure during drought and higher stomatal safety margins (the difference between water potential at stomatal closure and P50). Plants from warmer climates exhibited greater embolism resistance and higher hydraulic conductance, along with wider vessels and lower vein density, suggesting adaptation to high evaporative demand. C_LIO_LIOur findings highlight two primary hydraulic strategies related to climate-of-origin: 1) conservative water use and drought avoidance in drier climates, and 2) embolism resistance and hydraulic efficiency in warmer climates. These findings provide new insights into the intraspecific differences in traits and adaptive strategies used by C4 grasses to persist in contrasting climates. C_LI

Vulnerability to cavitation in leaves is the result of highly adaptive anatomical and physiological traits that can be linked to water availability in a species climate of origin. Despite similar gross leaf morphology, eucalypt species are often confined to specific climate envelopes across the variable rainfall environments of Australia. In this study, we investigate how the progression of cavitation differs among eucalypts and whether this is related to other hydraulic and physical leaf traits. We used the Optical Visualisation technique to capture cavitation progression across the leaves of eight eucalypt species (Angophora crassifolia, Corymbia tessellaris, Eucalyptus atrata, Eucalyptus grandis, Eucalyptus laevopinea, Eucalyptus longifolia, Eucalyptus macrandra, Eucalyptus tereticornis) from a wide range of climates and grown in a common garden setting. Vulnerability to cavitation, represented by the leaf water potential required for 50% cavitation of leaf vessels, varied significantly among species (-3.48 MPa to -8.25 MPa) and correlated linearly with home climate precipitation and leaf SLA (R2 of 0.64 and 0.75, respectively). P12-P88, the range of water potentials between which 12% to 88% of cavitation occurs, was decoupled from P50 but also correlated with leaf SLA (R2 of 0.72). We suggest the magnitude of P12-P88 may be representative of a species drought strategy - a large P12-P88 signifying leaves that exhibit drought tolerance (retention of leaves under drought conditions) and a small P12-P88 signifying drought avoidance (leaf shedding after a threshold of drought is reached). Our results agree with other studies that highlight these cavitation metrics as genetically fixed traits. Turgor loss point, on the other hand, may be more plastic, as evidenced by the low variability of this trait across these eucalypt species grown in a common garden environment. Further study will help to establish the SLA-related anatomical traits that impart cavitation resistance and to extend these conclusions to a greater number of species and home climates.

O_LIAngiosperms with large genomes experience nuclear-, cellular- and organism-level constraints that may limit their phenotypic plasticity and ecological niche. These constraints have been documented to vary across lineages, life-history strategies, ecogeographic patterns and environmental conditions. Therefore, we test the hypotheses that extinction risk is higher in large-genomed compared to small-genomed species, and that the effect of genome size varies across three selected covariates: life form, endemism, and climatic zones. C_LIO_LIWe collated genome size and extinction risk information for a representative sample of angiosperms comprising 3,250 species, which we analyzed alongside life form, endemism and climate variables using a phylogenetic framework. C_LIO_LIAngiosperm genome size is positively correlated with extinction risk, a pattern driven by a signal in herbaceous but not woody species, regardless of climate and endemism. The influence of genome size is stronger in endemic herbaceous species, but is relatively homogenous across different climates. Beyond its indirect link via endemism and climate, genome size also influences extinction risk directly and significantly. C_LIO_LIGenome size may serve as a proxy for difficult-to-measure parameters associated with resilience and vulnerability in herbaceous angiosperms. Therefore, it merits further exploration as a useful biological attribute for understanding intrinsic extinction risk and augmenting plant conservation efforts. C_LI

Intraspecific variation in leaf shape affects many physiological processes but its impact on plant distribution is underexplored. Using a common garden, we studied daytime thermoregulation of lobed and entire leaf genotypes of Ipomoea hederacea, which displays a latitudinal leaf shape cline. Both leaf shapes maintained similar temperatures but entire leaf genotypes had significantly increased stomatal conductance in warmer/sunnier weather. With less potential water loss, lobed genotypes may have advantages in drier conditions. Lobed genotypes are more common in the north of the species range, which receives less summer precipitation, suggesting water availability as a potential clinally varying selective agent.

Exploring patterns and causes of intraspecific trait variation is crucial for a better understanding of the effects of climate change on plant populations and ecosystems. However, our current understanding of the intraspecific trait variation is mainly based on structural (morphological) traits, and we have limited knowledge on patterns and causes of variation in biochemical traits (e.g., leaf pigments), which are also crucial for plant adaptation. As a result, we also do not know how similar the climatic effects on structural versus biochemical traits are. Using plant traits from 110 genotypes representing 11 Festuca rubra populations grown in 4 different climates, we studied trait covariation among structural traits (linked to fitness, resource use, gas exchange, and reproduction) and biochemical traits (linked to photosynthesis, photoprotection, and oxidative stress). We also disentangled the relative role of the climate of origin and the climate of cultivation in the structural versus biochemical traits and tested for adaptive plasticity in the traits. We found that 1) biochemical traits did not covary with structural traits and represent independent photoharvesting - photoprotection strategy dimension of functional variation; 2) interactive effects of climate of origin and cultivation were more pronounced for biochemical than structural traits. 3) Trait plasticity was affected by the climate of origin (precipitation and temperature as well as their interaction); 4) F. rubra showed both adaptive and mal-adaptive plasticity, and adaptiveness depended upon trait type, cultivation climate, and climate of origin. Overall, our results suggest that structural and biochemical plant traits respond differentially to climate and thus the response of one group of traits cannot be predicted from the other. Responses are also strongly determined by interactions between the climate of origin and cultivation. Thus, more studies on variation in biochemical traits, their correspondence to other traits, and their variation with climate are needed.

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.

Plants with large geographic distributions experience varying biotic and abiotic conditions across their geographic range that can influence plant defense traits among plant populations. It is often the case that more productive regions support higher herbivore numbers. Yet whether this results in greater herbivore pressure and stronger plant defenses in these populations compared to populations inhabiting less productive regions is uncertain. Here we quantified defense traits and herbivory of Solanum carolinense L. across a large productivity gradient that spans its entire north-south range (29-44{degrees} N), using both field observations (33 populations; [~]14 plants per population) and a common garden experiment (15 populations; [~]4 plants per population). We examined the effects of productivity on 1) plant defense traits (specific leaf area, trichome density and glycoalkaloid concentration), 2) herbivory, and 3) the correlation among traits and herbivory in field and common garden plants. Trichome density and herbivory were higher for S. carolinense at the center of its range, while glycoalkaloid concentrations were negatively associated with productivity both in the field and in the common garden. In the field, plants with higher glycoalkaloid production experienced reduced herbivory, but there was no association with plant defense traits and herbivory in the common garden. Overall, these findings suggest that different types of defensive traits within a single species may follow different ecological and evolutionary trends and highlight the need for trait-specific considerations when applying plant defense hypotheses at the intraspecific level.

We investigated generic relationships in the ingoid clade (Fabaceae) (sensu Koenen & al. 2020a), with main focus on genera with a taxonomic history in Calliandra s.l. of the tribe Ingeae (i.e. Afrocalliandra, Calliandra s.s., Sanjappa, Thailentadopsis, Viguieranthus, Zapoteca), and three genera of the tribe Acacieae (i.e., Acacia, Acaciella, Senegalia). The nuclear ribosomal ETS and ITS, and the plastid matK, trnL-trnF and ycf1 DNA-regions were analysed for 246 representatives from 36 genera using maximum likelihood as implemented in IQ-tree. The results show an Ingeae-Acacia clade within the ingoid clade, resolved in three major clades. Clade 1 (Calliandra s.s. and Afrocalliandra) is sister to clades 2 and 3. Clade 2 comprises Faidherbia, Sanjappa, Thailentadopsis, Viguieranthus and Zapoteca. Clade 3 comprises the remaining genera of the Ingeae, plus Acacia. The ingoid genus Senegalia is excluded from the Ingeae-Acacia clade. Acaciella is sister to the remaining ingoid clade when nuclear ribosomal data is included in the analyses, but included in the Ingeae-Acacia clade based on plastid data. Acacia and perhaps also Acaciella are thus nested within Ingeae. Species traditionally referred to Calliandra (Calliandra s.l.) are resolved in two clades, and the "Calliandra-pod" has apparently evolved independently several times.

The North American Thermopsideae, a monophyletic group comprising the North American endemic Baptisia, and the paraphyletic Eurasian-North American disjunct Thermopsis, is nested within the Sophoreae (Fabaceae: Papilionoideae). Previous phylogenetic studies have identified two East Asian taxa within the North American Thermopsideae, suggesting two independent dispersal events between North America-East Asia. More recent studies have also placed a third taxon, Vuralia turcica, an endemic species from Turkey, among the North American Thermopsideae. The presence of three geographically distant Eurasian taxa within a relatively young clade of North American origin is perplexing, and the biogeographic implications of this observation are not clear. To investigate this matter, 1540 low-copy nuclear genes and complete plastomes were obtained from 36 taxa across the core genistoids, including 26 newly sequenced taxa. Nuclear and plastome based maximum likelihood (ML) and ASTRAL analyses were conducted based on varying degrees of taxon coverage and read mapping consensus threshold values. Additional analyses were performed to estimate divergence times and to reconstruct biogeographic history. The results strongly support a relictual Old World clade, presently composed of V. turcica and T. chinensis, which diverged from the ancestor of the North American lineage during the mid to late Miocene. A single and recent North America-East Asia dispersal involving T. lupinoides is reported. Furthermore, the traditional inclusion of the genus Ammopiptanthus among Thermopsideae, and the recent recircumscription of T. turcica to the monotypic Vuralia, are not supported. A relatively high degree of cytonuclear discordance is reported within each sub-clade of the North American Thermopsideae. This finding is likely attributable to the high degree of interspecific hybridization reported within these groups and raises the need for more rigorous genome-scale testing to disentangle their complex phylogenies. Subjects: Biodiversity, Biogeography, Evolutionary Studies, Genetics, Plant Science

AimLeaves display a remarkable variety of shapes, each with potential ecological advantages in specific climates. While relations between leaf shape and either climate or height has been relatively well studied in eudicots, the macroecological drivers of shape remain poorly known in monocots. Here, we investigated associations between climate and plant height with the evolution of leaf shape in a clade with high species and morphological diversity. LocationGlobal. Time periodCretaceous to contemporary. Major taxa studiedPalms (Arecaceae). MethodsWe apply a Bayesian phylogenetic mixed model to test for associations between climate and leaf shape (all entire-leaved, pinnate-dissected, palmate-dissected, and costapalmate). We further reconstruct the ancestral leaf shape using multistate speciation and extinction models and compare the frequency of shapes with global temperatures through time. ResultsWe find that plant height associates with dissected leaves and that annual precipitation associates with pinnate shapes. The ancestral leaf shape is unclear but early diversification was dominated by pinnate-dissected palms, which has remained the most species-rich form of leaves throughout palm history. Main conclusionsPalms that are tall and live in humid regions are more likely to have pinnate leaves. Through geological time scales, temperature did not play an obvious role in determining leaf shapes. This study contributes to our understanding of how the diversity of leaf shapes is linked to biological and climatic factors.

Pollinators usually are attracted by flower displays, but whether non-green leaves display can function as a flower guide is less well studied. Furthermore, it is still largely unknown whether photosynthetic cost is paid-off by benefits afforded by attraction pollinators and how non-green leaves maintain photosynthetic capacity. Thus, white leaf and inflorescence/flower traits, pollinator visitation, fruit set and seed production, spectral properties, leaf structure, net photosynthetic rate and chlorophyll fluorescence of adaxial and abaxial surface were studied in Actinidia kolomikta (Rupr. & Maxim.) Maxim. The reproductive branches of A. kolomikta have a large number of white leaves at the exterior canopy, but white inflorescences grow at the canopys interior, and we found that showy display of white leaves attracted pollinators reach the flowers. Although incident light was reflected largely, we found that photosynthetic rate of white leaves was maintained at relatively high levels. Furtherly, spongy tissue play the vital role in maintenance photosynthetic capacity of white leaves, which effectively supports inflorescence development during flowering. Thus, white leaves of A. kolomikta enhanced reproductive fitness through attracting pollinators and decreasing reproductive cost: (1) spatially via white leaves locating at reproductive branches and temporally via synchronism between white leaves and flowers; (2) increasing white leaves display and decreasing investment in reproduction; (3) compensatory mechanisms via maintaining photosynthetic capacity of white leaves. Therefore, we propose that these dual functions of white leaves during blossom lower the plants cost of reproduction, and that this is an adaptation to the climatic challenges of the high altitudes and latitudes at which it grows.

Postmating prezygotic (PMPZ) traits play an important role in mating success, especially in species where gametes from multiple males compete. Despite this, the effect of mating system transitions, and attendant shifts in the intensity of sexual selection, on specific PMPZ traits and their underlying loci is still poorly understood. Here we assessed differences in pollen PMPZ traits and tissue-specific gene expression (in leaf, pollen, and style) between two closely related plant species with different mating systems--Solanum lycopersicum (selfing) & Solanum pennellii (outcrossing). We focused on species differences in loci with known roles in pollen tube growth rate, including pectin methylesterases (PMEs) and their inhibitors (PMEIs), and in- vitro & in-vivo pollen tube growth rates. Among the gene expression differences observed between species, we found that the expression domain of pollen-biased genes was much narrower in the selfing species S. lycopersicum compared to the outcrossing species S. pennellii, including for most reproductive PMEs and PMEIs. In addition, S. pennellii had faster pollen tube growth rates in-vivo, while S. lycopersicum had faster in-vitro pollen tube growth rates. We propose that the lower expression of pollen tube development genes in S. lycopersicum style tissue, and reduced in-vivo pollen performance, is a result of reduced allocation to stylar mechanisms that modulate pollen tube growth, potentially consistent with relaxed selection on cryptic female choice in the selfing species. Article SummaryThis study investigates the evidence for differential postmating sexual selection in plants, by analyzing reproductive gene expression and pollen performance in two Solanum species with different mating systems. It finds that species differ systematically in pollen tube growth rates, and that pollen ( male)-biased genes have higher secondary expression in stylar ( female) tissues in the outcrossing species. These observations are consistent with the outcrossing species experiencing stronger selection on female reproductive tract traits that influence male postmating performance. This study evaluates key expectations of sexual selection on postmating traits but does so in the uncommon context of flowering plants.

Patterns of crop production in mast species do not track crop-year climate, but instead are regulated by climate cues in prior-years. Whether the pattern of year-to-year seed mass variation is coupled in time with mast seeding, maintaining seed mass-number trade-offs, and coherently driven by similar climate cues as other seed traits (e.g. seed germination) remains unknown. Using ca. 6,000 long-term seed inventory data over the years 1955-2015 in conifers, this retrospective study revealed the temporal patterns of mast species seed mass and its associated trait, seed germination. To pinpoint their ecological drivers, pairwise correlation analysis was performed between each trait and seasonal climates in crop year and four prior-years. Using climate variables key to each trait, regression models were constructed to project trait values. Findings showed minor seed mass variation among years, which rejects the generality of seed mass-number trade-offs in many plant species. This result reasonably arises as the economies of scale (compensating benefits) theory are often used to account for mast seeding but not for seed mass. Moreover, final germination fraction also varied little over time, but exhibited an increasing tendency. In addition, we found that temperature-based climate variables drive seed mass, number, and germination variation, but these variables in different seasons of crop year or prior-years did not have equal influences on trait variability. Finally, regression models showed that the number of frost-free days and evapotranspiration are crucial to the three traits and climate in autumn is a critical season, followed by summer and winter. This study holds considerable promise for explaining reproductive strategies of taxonomic groups with mast seeding characteristics in allocating reproductive resources to different life-history traits using ecological signals.

BackgroundMelastomes are well known for their striking diversity in stamen morphologies mostly adapted to buzz pollination by bees. The variously modified connective appendages and heteranthery in the family have fascinated botanists for more than two centuries and a variety of functions associated with pollination have been discovered for these staminal traits over the years. The repeated evolutionary shifts in these traits have been linked to pollinator shifts, likely contributing to diversification in the family. The evolutionary lability of staminal traits, especially the connective morphology, led us to hypothesize that these traits might be controlled by relatively simple genetic mechanisms and we here take the first steps to test this hypothesis by using a comparative transcriptomics approach with Arthrostemma ciliatum as our model. We also tested the functional significance of heteranthery and whether the classical division of labour hypothesis holds true for this species by comparing the number, size and viability of pollen in the two stamen types. ResultsStaminal development of this species was studied and suitable stages for transcriptome comparisons were identified. Differential expression analyses between the morphologically distinct stamen whorls at four developmental stages showed the differential expression of several transcripts involved in stamen development/elongation. Pollen comparisons between the two whorls showed that the antepetalous/inner whorl stamens have a significantly higher number of pollen and higher germination rates while the antesepalous/outer whorl stamens have significantly larger pollen. ConclusionsWe identified Jasmonate and Gibberellin signalling pathway genes (JAZ, GID1, DELLA and ARF homologs), EPF/EPFL family genes, autophagy related genes (VPE homologs) and S Locus ELF homologs as putative candidates involved in causing staminal dimorphism in A. ciliatum. Our results indicate that, for the heterantherous morph of this species, the shorter stamens (antepetalous/inner whorl) have both pollinating and feeding functions contradicting the division of labour theory. We also report the possible existence of heterostyly in A. ciliatum as an outbreeding mechanism.

Premise of StudyThe parentage of a widespread member of the carnivorous sundew genus Drosera, the allopolyploid Drosera anglica, remains uncertain despite over 100 years of morphological, cytological, and, more recently, molecular study. MethodsUsing transcriptomic and genomic data from 12 species Drosera sect. Drosera including four D. anglica populations and a disjunct Idaho population of D. intermedia, we assembled genes in HybPiper and phased sequences in HybPhaser. We estimated heterozygosity and generated flow cytometry data to assess ploidy levels. We estimated species relationships with phylogenetic and pairwise genetic distance methods. Additionally, we assembled rbcL and ITS reads to compare to previous data. Key ResultsSequences from phased subgenomes highly supported D. anglica as sister to D. rotundifolia and D. linearis, differing from previous analyses based on chromosome pairing and Sanger sequencing with limited taxon sampling. Both ITS and rbcL sequences of D. anglica were the most similar to D. linearis. Drosera anglica is intermediate between both parents in leaf shape and microhabitat; however, across D. sect. Drosera, neither leaf shape nor biogeographic distribution were reliable indicators of phylogenetic relationships. Despite a range-wide sampling, we did not find evidence for multiple origins of D. anglica. Additionally, we confirmed that the Idaho population previously identified as D. intermedia is D. anglica. ConclusionsDrosera anglica arose from allopolyploidy between D. linearis (the chloroplast donor) and D. rotundifolia. Our study demonstrates the importance of taxon sampling and careful examining complex phylogenomic data, and presents an exemplar of analyzing allopolyploid relationships in plant lineages.

O_LIPlant mating systems and pollen limitation often vary along elevational gradients, yet empirical evidence remains mixed and rarely comes from multi-species experimental studies in tropical mountain ecosystems. C_LIO_LIWe tested how elevation affects natural seed set, pollen limitation, and selfing capacity, and whether these patterns are associated with variation in flower visitation, in Afromontane grasslands on Mount Cameroon. We conducted a hand-pollination experiment on seven zoogamous plant species at four elevations above the timberline (2,300-3,800 m), applying autonomous selfing, geitonogamous selfing, outcrossing, and open-pollination treatments to 1,776 flowers and quantifying seed set. In parallel, we quantified pollinator visitation on unmanipulated plants to estimate visitation frequency, morphospecies richness, and functional-group richness. C_LIO_LINatural reproductive success of the studied plants exhibited a pronounced mid-elevation peak and declined sharply towards the summit, where pollen limitation increased strongly. At the highest elevation, some species produced few or no seeds even under outcross-pollination, indicating physiological reproductive constraints. C_LIO_LIIndices of autonomous selfing and geitonogamy varied among species, with no consistent elevational patterns. Despite detected partial self-compatibility, summit populations did not exhibit expected shifts towards selfing, suggesting limited reproductive assurance under high-elevation conditions. C_LIO_LIPollinator visitation frequency and diversity declined at the highest elevation. Natural seed set was positively associated with visitation frequency and mildly negatively associated with morphospecies richness, whereas pollen limitation and selfing indices showed no clear relationships with visitation metrics, consistent with the influence of additional physiological and developmental constraints on reproduction. C_LIO_LISynthesis. Our study shows that in isolated Afrotropical montane grasslands, plant reproductive success at high elevations is jointly constrained by declining pollination service and abiotic limitations, which is not compensated by increased selfing. This suggests that successful plant reproduction near upper vegetation limits depends on both sufficient pollination service and favourable physiological conditions. Under ongoing climate change, upslope range shifts of plants may therefore not guarantee reproductive success if plant and pollinator responses to warming are asynchronous or if a part of extreme high-elevation conditions remain limiting. These findings advance understanding of how biotic interactions and abiotic constraints together shape plant reproduction along tropical elevational gradients. C_LI