Abstract Staminodes (sterile stamens) are widely interpreted as enhancing pollination through pollinator attraction and efficiency, yet their contribution to seed production remains uncertain in species with petaloid staminodes, short anthesis, and arid habitats where opportunities for pollinator attraction may be restricted. We tested how petaloid staminodes influence pollinator behaviour and seed set in Mentzelia integra (Loasaceae), an arid-adapted species with short-lived, evening-blooming flowers, using paired-choice field experiments near St. George, UT, USA. We compared paired intact and staminode-removed flowers for visitation rate, first landing, visit duration, flower constancy, and seed set. Flowers with intact staminodes received more bee visits and first landings, whereas visit duration did not differ between treatments in Perdita, the only taxon analysed for this metric, suggesting a primary role in attraction rather than handling efficiency. Previous flower type influenced subsequent landing choice in Megachile and Perdita, but not in Anthophora. Only Perdita showed a reciprocal transition pattern consistent with flower constancy, with subsequent landings tending to remain within the same treatment. Flowers with intact staminodes had higher seed set than flowers without staminodes. Overall, petaloid staminodes in M. integra enhance seed set through attraction-mediated pathways, highlighting the evolutionary co-option of sterile staminal structures as floral display traits.
Abstract Temporal colour change within individual flowers serves as a dynamic visual signal that maintains and even enhances pollinator attraction throughout the floral lifespan. This study investigates variation in floral traits associated with pollinator attraction in Nicotiana mutabilis during the age-related transition of flower colour from white to magenta. Using greenhouse-grown plants, we assessed stigma receptivity, nectar volume, sugar concentration, and energy content, and characterized floral pigments, pollen, and corolla volatile compounds, and corolla shape and size, across colour phases of N. mutabilis. The transition from white to magenta consistently occurred under controlled common-garden conditions, as documented in nature, typically 3 to 8 days post-anthesis. Stigma remained receptive in both flower colour phases. Nectar production increased progressively after anthesis, concomitant with the transition from white to magenta. Magenta flowers exhibited higher nectar volumes, sugar concentrations, and total energetic content, which rose markedly over the first 96 hours post-anthesis. Magenta flowers contained 54% more anthocyanins in the 400–600 nm range, whereas flavonol levels in the UV-A/B range were similar between colour phases, indicating a shared UV-absorbent phenotype likely attractive to hawkmoths in both phases. The scent compounds were similar in the floral colour stages and in night and day emissions. The composition of volatile organic compounds differs between the corolla and pollen, indicating that these floral parts may serve distinct olfactory functions in pollinator attraction. Corolla shape did not differ between floral colour phases; however, tube size was larger in the magenta phase. Notably, our results showed that N. mutabilis maintains its ability to attract pollinators and continues to offer floral resources in both colour phases. Nectar volume, sugar concentration per flower, floral morphology, and the composition of floral pigments, pollen, and corolla scent are probably attractive to multiple pollinator groups.
Abstract Botanical inventories across Europe increasingly reveal rapid and often underestimated change, driven by ongoing introductions and improved detection of overlooked taxa. Cryptic invasions—where non-native or unrecognized lineages remain hidden due to morphological similarity or taxonomic complexity—contribute substantially to this underestimation, particularly among small, short-lived ruderal plants. We combined flow cytometry, chromosome counting, morphometric analyses, and nuclear ITS sequence data to investigate diversity within the genus Holosteum in central Europe. Sampling in Czechia, Hungary, and Slovakia revealed the coexistence of two distinct cytotypes: diploids (2n = 20) corresponding to H. umbellatum s.s., and an as yet unknown tetraploid cytotype (2n = 40) not assignable to any described taxon. Morphometric analyses showed clear differentiation, with leaf shape, pedicel indumentum, and stamen number providing the most reliable diagnostic characters. ITS data indicate the tetraploid is an allotetraploid sharing one ribotype with diploid H. umbellatum and another of unknown origin. Based on combined morphological and molecular evidence, we describe the tetraploids as representing a new species, Holosteum cerastioides sp. nov. It is currently known from salt-affected roadside habitats and is apparently undergoing rapid expansion. Our results highlight the importance of integrative taxonomy for detecting cryptic diversity and overlooked components of changing European floras.
Abstract In the genus Luehea, a taxonomic complex comprising the species L. grandiflora, L. macrophylla, L. rufescens, L. speciosa, and L. tomentella has been historically recognized, but its classification remains unresolved. We carried out a study that combined morphometric and molecular analyses to clarify the taxonomic relationships among these taxa. These analyses proved useful in circumscribing the studied species within the complex and in identifying the most diagnostic morphological characters that can be used to distinguish the accepted species. In light of the results obtained, we propose treating this complex as comprising two distinct species, L. macrophylla and L. speciosa, whereas L. grandiflora, L. rufescens, and L. tomentella are treated as new synonyms of L. speciosa.
Speciation has long been a central topic in evolutionary biology, with reproductive isolation (RI) playing a fundamental role in this process by limiting gene flow between diverging populations. However, the relative contributions and temporal sequence of establishment of different reproductive barriers remain poorly understood, particularly in plants. In this study, we quantify RI between Actaea purpurea and A. japonica-two species exhibiting significant differences in floral morphology and pollinator preferences-with particular emphasis on pollinator isolation. We measured five reproductive barriers: ecogeographic isolation, phenological isolation, pollinator isolation, pollen-stigma incompatibility, and post-zygotic isolation, using hybridization experiments and ecological niche modelling with varying grid size. Pollinator isolation was assessed by establishing artificial sympatric populations. Our results demonstrate strong total RI between the two species (>0.99), supporting their status as distinct biological species with minimal gene flow. Pre-zygotic barriers contributed more significantly to RI than post-zygotic isolation. Ecogeographic isolation was the largest contributor to total RI, followed by pollinator isolation. This study underscores the importance of both pollinator isolation and ecogeographic factors in speciation and species maintenance in Actaea, and suggests that pre-zygotic reproductive barriers may arise earlier and play a dominant role in speciation: a pattern that may be widespread in flowering plants.
Abstract Pollination is a major driver of floral diversity and reproductive success in angiosperms, but comprehensive field-based data are still limited for many plant families. We conducted a systematic review and meta-analysis of 128 studies on Solanaceae, compiling 249 plant taxa with records of planta-pollinators in 25 genera and 154 species. We extracted data on floral traits, including colour, symmetry, reward type, and the presence of floral spots, and analysed their associations with primary and secondary pollinator groups using generalized linear models. Bees were the most frequent pollinators, particularly attracted to zygomorphic flowers offering both pollen and nectar, whereas hawkmoths preferred pale flowers and hummingbirds favoured flowers without spots. Floral traits strongly influenced pollinator visitation and species richness, although patterns varied among pollinator groups. Our results highlight gaps in taxonomic and geographic coverage and provide a synthesis that can guide future research on floral evolution, ecological specialization, and conservation strategies in Solanaceae.
Understanding fungal diversity in plants is essential for microbial ecology but is often limited by the time, geographical range, and labour required for field sampling. Herbarium specimens offer a promising alternative for large-scale, long-term, or historical studies of fungal communities. This study explores the feasibility of using herbarium collections to investigate leaf-associated fungi by examining how preservation methods, storage environments, and potential contamination might influence fungal communities. We analysed fungal communities in the plant specimens from three major herbaria in Taiwan using high-throughput sequencing. Our results indicate that fungal diversity and community structure remain stable across fresh leaves, dried leaves, and newly prepared herbarium specimens, suggesting that drying and short-term preservation have limited impact on fungal communities. However, prolonged storage may drive community shifts. Furthermore, specimens across different herbaria contain similar fungal communities, reflecting that herbarium environments do not affect fungal communities. Additionally, minimal overlap between airborne fungal species in the herbarium and those detected in the specimens implies limited contamination from ambient fungi. These results demonstrate that herbarium specimens are valuable resources for studying fungal communities across time and space, but caution is needed when interpreting data from historical specimens, even those stored for just a few years.
We conducted a genus-wide survey of leaf micromorphology in Veratrum (Melanthiaceae) to assess the taxonomic significance of epidermal traits. Light and scanning electron microscopy were applied to 98 herbarium and field specimens representing 18 taxa from two recognized sections of the genus, Veratrum and Fuscoveratrum. Principal coordinates analysis of quantitative and qualitative micromorphological traits, including stomatal features, was used to assess their taxonomic value. The analyses recovered two well-defined leaf epidermis micromorphological types that are congruent with the current infrageneric classification: pseudostem-forming taxa of section Veratrum show abundant unicellular trichomes, elongated guard cells (60.22 +/- 7.39 & micro;m), and sparse styloids, whereas glabrous-leaved section Fuscoveratrum combines shorter guard cells (48.91 +/- 4.85 & micro;m) with dense raphides. For species-level diagnosis, V. nigrum shows a high density of styloids, whereas V. stamineum lacks trichomes and exhibits raphide dominance. Observed associations between habitat conditions and foliar micromorphological traits suggest a potential relationship with soil moisture, although direct functional evidence is lacking. Overall, leaf-surface characters provide reliable markers for sectional delimitation and contribute to species diagnosis in Veratrum, offering a framework for integrating micromorphological evidence with cytogenetic and phylogenomic data.
The Amazon and the Atlantic Forest harbour the greatest diversity of herbaceous bamboos (Poaceae, Bambusoideae, Olyreae) in the Americas, especially the region comprising southern Bahia and northern Espirito Santo states, which exhibits a high degree of endemism for this group. Over the past three decades, molecular systematics has significantly advanced the resolution of taxonomic uncertainties within the Olyreae, including a revised circumscription of subtribes Buergersiochloinae and Olyrinae, as well as the recognition of Olyra as non-monophyletic. To further elucidate these relationships, we sequenced four plastid (ndhF, rpl32-trnL, trnD-trnT, and trnS-trnG) and one nuclear (ITS1-5.8S-ITS2) genome regions for Olyra and other members of the tribe Olyreae. Our phylogenetic analyses support the segregation of two monospecific lineages distinct from Olyra as novel genera: Pataxoa and Tupinambarum. Both genera are formally described and illustrated here, and their relationships with sister taxa are compared. Proposed International Union for Conservation of Nature assessments are included for both genera. Our results indicate that these new genera are more closely related to Lithachne and Arberella than to Olyra. Furthermore, the name O. filiformis is lectotypified in accordance with the International Code of Nomenclature. Both new genera have been named to honour the indigenous peoples of the regions where these bamboos occur. These discoveries improve knowledge about the biodiversity and evolutionary history of bamboos in the Atlantic Forest and also recognize the contribution of indigenous peoples to conserving the biodiversity of the region.
Abstract Taxonomy within Hymenophyllum subgenus Mecodium (Hymenophyllaceae) has long presented significant challenges due to high morphological similarity among species. In this study, we conduct a taxonomic revision by combining a literature review, morphological examination of herbarium specimens, field investigations, and phylogenetic analyses based on several chloroplast markers (rbcL, accD, rps4, rbcL-accD, and rps4-trnS). Our revision yields an updated checklist of Chinese Mecodium, comprising 11 species and one variety. Integrating four Hymenophyllum-associated fossils, we performed divergence time estimation using both CladeDate and tip-dating methods. We infer for the subgenus Mecodium an origin in the Palaeogene, and a single arrival in Continental Asia dated from the Eocene. Major diversification events in Asia appear to coincide with the Himalayan orogeny and Miocene climatic shifts. Biogeographical reconstruction indicates multiple dispersal routes, including transoceanic movement from East Asia to North America and southeastward expansion from the Himalayan region within Asia. These findings offer new insights into the evolutionary history of this taxonomically complex fern group.
Abstract Insect pollination has been important in angiosperm diversification, but alternative reproductive strategies also occur. Wind pollination is widespread in Cyperaceae, Juncaceae, and Poaceae; however, ambophily or strict entomophily may have evolved secondarily in some lineages. We conducted a systematic review to synthesize historical evidence of biotic pollination in Cyperaceae and mapped the phylogenetic distribution of genera with confirmed or putative ambophily. We also propose a theoretical gradient of floral attributes to infer pollination modes in the family. Direct evidence of biotic pollination (from field experiments) is documented in three genera (Cyperus, Eleocharis, and Rhynchospora) and indirect evidence, based mainly on floral visitor records, occurs in 10 genera: Bolboschoenus, Carex, Diplasia, Eleocharis, Ficinia, Hypolytrum, Krenakia, Lagenocarpus, Mapania, and Schoenoplectiella. One case of hydrophily is known in Eleocharis. Species with putative biotic pollination are mostly tropical, and ambophily appears to have evolved independently at least 12 times. A trait gradient seems to span from strictly anemophilous species with inconspicuous, widely spaced spikelets to entomophilous taxa with compact, colourful inflorescences and pollenkitt production, with ambophilous forms in intermediate zones. Thus, Cyperaceae provide a valuable model for research on reproductive evolution.
Fungus gnats have been identified as pollinators of the species-rich plant genus Arisaema, but the specificity of the interaction between Arisaema species and flower visitors remains incompletely known. Oviposition site mimicry (mushroom mimicry) has traditionally been postulated as the mechanism of pollinator attraction in Arisaema, but recently it has been suggested that some species may employ sexual mimicry. While oviposition site mimicry is expected to be associated with even or female-biased sex ratios of flower visitors and a potentially diverse flower visitor assemblage, sexual mimicry has been inferred in cases with strongly male-biased sex ratios and can be expected to be associated with a limited number of flower visitor species. We examined the species composition and sex ratios of fungus gnat visitors in female inflorescences of 26 Arisaema species sampled at 17 sites across China. The fungus gnat assemblage differed between sympatric Arisaema species, but also varied among conspecific populations at different sites, and among years. Most Arisaema species interacted with taxonomically diverse sets of fungus gnat visitors, including species from both Mycetophilidae and Sciaridae and multiple genera, consistent with attraction based on oviposition site mimicry to which many fungus gnat species respond. Three cases of strongly male-biased sex ratios were documented. In one Arisaema population, a strongly male-biased sex ratio combined with dominance by a single flower visitor species was consistent with predictions of sexual mimicry. The results indicate that differences in pollinator assemblages are likely to contribute to reproductive isolation among many sympatric Arisaema species, and prompt further experimental studies of pollinator efficiencies and the mechanisms of flower-visitor attraction in this genus.
Historical herbaria allow previous flora to be reconstructed and to understand long-term environmental changes. By using Ulisse Aldrovandi's herbarium, one of the oldest of Europe, we attempted to reconstruct historical plant communities at the habitat level, selecting the ancient wetland system of Valle Padusa (SE Po Plain, northern Italy) as a case study, which has now disappeared due to land reclamation and development. We analysed 85 specimens (belonging to 54 species) collected by Aldrovandi in this area, clearly attributed to freshwater sites; after which we made a list of potential habitats (as defined by Directive 92/43/EEC and regional classifications of vegetation under conservation interest), on the basis of the presence of physiognomic and companion species (e.g. species generally acknowledged as frequent in target vegetation) and abundance observations from Aldrovandi's manuscripts. The landscape shown is characterized by a complex mosaic of water bodies, riparian environments and marshes: 17 habitats have been identified, out of which 12 are recognized by the Directive; among them 3150, 3260, 3290, 7210* plus regional vegetation types Mc (Magnocaricion) and Pa (Phragmites australis thickets), which definitely existed. Many of the collected species are now either regionally extinct, critically endangered, or protected, underlining a dramatic decline in inland water plant diversity primarily driven by wetland destruction. Our study confirms the potential of using herbaria for reconstructing past landscapes, especially when different disciplines are integrated.
Abstract Floral oil is a highly specialized reward secreted by dense areas of glandular trichomes known as elaiophores. Tigridieae (Iridaceae), a predominantly South American tribe represented by two main clades, A and B, exhibits diverse floral rewards, including oil, a rare resource restricted to few specialized lineages. To evaluate whether elaiophores influenced diversification in Cypella and closely related genera (Clade A), we generated a dated phylogeny, clarified the definition of elaiophores in Tigridieae, and explored diversification patterns. A well-sampled, time-calibrated phylogeny was inferred using nine DNA regions, three nuclear and six plastidial, for 61 taxa, including 32 newly generated sequences and representatives of all genera of Tigridieae Clade A. Divergence times were estimated in BEAST, diversification rates were analysed using RevBayes and Bayesian analysis of macroevolutionary mixtures. Trait-dependent diversification was tested using state-dependent speciation and extinction models. We recovered six well-supported major clades and revealed that the most species-rich genera, Calydorea, Cypella, and Herbertia, were not monophyletic. Our findings support a broader view of floral evolution driven by diffuse selective pressures within complex pollination systems, suggesting that oil-based rewards in Tigridieae Clade A evolved through shifts from specialized to more generalized adaptive strategies. These findings provide valuable insights into reward evolution in angiosperms.
Apomixis is an asexual mode of reproduction through seeds and nearly all gametophytic apomictic species are polyploid. Psidium cattleyanum, a polyploid species with several described ploidy levels, exhibits apomixis and requires effective pollination for fruiting (pseudogamy). The aim of this study was to determine the frequency of apomixis and sexuality in heptaploid and octoploid accessions of P. cattleyanum, and to analyse maternal and paternal contributions to endosperm formation. Flow cytometry was used on mature seeds obtained from hand-pollinated crosses between two octoploid and two heptaploid accessions to determine the ploidy levels of embryo and endosperm. Among the 492 seeds analysed, embryo: endosperm ploidy ratios ranged from the expected 2:5 (64.2%, pseudogamous apomixis) and 2:3 (4.7%, sexuality) to unexpected values including 1:3 (0.4%), 2:6 (2.4%), 2:7 (0.2%), 3:5 (8.8%), 2:3:5 (18.5%), and 2:4:6 (0.8%), associated with polyhaploid parthenogenesis, pseudogamous apomixis involving reduced and unreduced gametophytes, and female gametophytes with two or three polar nuclei. Embryos with both higher and lower ploidy than the maternal plant were detected, and the implications of these findings for wild populations are discussed.
The distribution of dichogamy in Acanthaceae is unknown, yet studies in the genus Justicia from Africa described the occurrence of protandry with staminal movement. A review of the genus suggests African species show protandry with staminal movement but New World species do not. In this study, we test this hypothesis. We also propose that hummingbird pollination led to the loss of this form of dichogamy. We inferred a time-calibrated molecular phylogeny of Acanthaceae and scored species within tribe Justicieae for protandry with staminal movement and pollination syndrome. We used ancestral state reconstruction to determine how many times staminal movement evolved in Justicieae, and tested for correlated evolution between changes in staminal movement and changes in pollination syndrome and geography. We scored a total of 333 species in tribe Justicieae, of which 58 showed protandry with staminal movement and 275 did not. Mapping staminal movement onto the phylogeny revealed multiple origins of this type of protandry, and significantly a loss in the common ancestor of New World 'justicioids' with a single regain in the Neotropics. Protandry with staminal movement is common in Old World Justicieae but confirmed to be almost absent in the New World. As Neotropical Justicia are nested in an Old World clade, it is clear they lost protandry with staminal movement and regained it once in the New World. Insect foraging behaviour probably drove the evolution of protandry with staminal movement and hummingbird foraging behaviour probably drove the loss of protandry with staminal movement in this clade.This summary, translated into Spanish, is available in the Supporting Information section (Appendix S1).
With 58 species, the pantropical papilionoid legume genus Macropsychanthus (Leguminosae: Diocleae) has long posed taxonomic challenges. In this study, we reconstructed its evolutionary history and revised its infrageneric classification using a comprehensive phylogenetic framework based on our newly produced plastome and nuclear DNA data from 59 taxa, including 51 species of Macropsychanthus. Our analyses confirm the monophyly of the genus and its division into the subgenera Macropsychanthus and Platylobium, each characterized by both molecular and morphological traits. Within subg. Macropsychanthus, we identify six well-supported clades, here proposed as sections, each supported by synapomorphies and biogeographic patterns. Divergence dating places the origin of the genus in the Amazon during the Miocene (similar to 12 Mya), followed by rapid diversification and subsequent long-distance dispersals into South-east Asia and Africa, representing rare cases of amphi-Pacific rainforest disjunctions. Overall, this study resolves long-standing taxonomic uncertainties, establishes a stable infrageneric classification, and provides novel insights into the timing, geography, and drivers of diversification in Macropsychanthus. The genus emerges as a valuable model for investigating biome shifts, long-distance dispersal, and morphological convergence in tropical lianas.
Understanding the evolutionary history of taxonomically complex plant genera remains a central challenge in systematics, especially in species-rich tropical lineages shaped by intricate patterns of morphological variation and biogeographical diversification. Machaerium is one such lineage from among the largest woody genera within neotropical Papilionoideae (Leguminosae). It comprises nearly 130 species predominantly distributed across the American tropics, and showing exceptional diversity in Brazil, particularly in the rainforest biome of Amazonia and the Atlantic Forest, as well as in the seasonally dry vegetation of the Caatinga and the fire-prone savannas of the Cerrado. Molecular phylogenetic assessments have placed Machaerium within the pantropical Dalbergioid clade. However, despite these advances, infrageneric relationships within Machaerium are still poorly resolved. In this study, we address this gap by conducting maximum likelihood and Bayesian phylogenetic analyses of Machaerium using both plastid matK and nuclear ITS 5.8S DNA markers. Although our Sanger-based sequencing approach did not fully resolve the evolutionary relationships within Machaerium, it unequivocally supported the monophyly of the genus with the inclusion of Steinbachiella, which is deeply nested within Machaerium. Considering this well-supported phylogenetic placement, we propose the necessary nomenclatural changes to reflect the revised circumscription of the genus. A robust sister-group relationship between Machaerium and Ctenodon was consistently recovered in all combined analyses, further reinforcing their close evolutionary affinity within the Dalbergioid clade. The results of our study do not support the monophyly of all traditionally recognized sections (Machaerium sect. Machaerium, M. sect. Acutifolia, M. sect. Lineata, M. sect. Oblonga, and M. sect. Reticulata). However, despite the lack of strong infrageneric structure, we identified diagnostic morphological characters for a subset of the species previously assigned to sect. Lineata, such as the presence of spinescent stipules and a cladodromous pattern of secondary venation in the leaflets. Our findings underscore the need for a reassessment of infrageneric classification in Machaerium using high-throughput genomic sequencing integrated with morphological traits to advance the systematics and understanding of the evolutionary history in this ecologically diverse neotropical genus.
Hybridization is common among plant species and presents many challenges for conservation. One emerging issue is unrecognized hybrids being mistaken for rare species and receiving conservation management that may not be warranted. Here, we used genome-wide single nucleotide polymorphism (SNP) markers to test hybridization hypotheses for three threatened species in Western Australia: Eucalyptus crispata, E. leprophloia, and E. pruiniramis. All three were suspected hybrids arising from one shared parent, E. accedens, and three other species, E. arachnaea subsp. arachnaea, E. zopherophloia, and E. loxophleba subsp. loxophleba, respectively. Results confirmed hybrid origins for E. crispata and E. leprophloia as hypothesized, although an additional hybrid combination was detected within E. leprophloia. In all combinations, clonality and hybrid class testing determined that most populations exist as singleton F1 individuals scattered across the landscape, with few backcrosses. Consequently, these taxa do not represent stabilized hybrid species but transient hybrids resulting from rare reproductive events between their parental taxa. Here, we recommend taxonomic revision as nothotaxa and re-assessment of their conservation status. In contrast, E. pruiniramis showed no signal of hybrid origin, supporting its continued recognition as a valid, conservation-listed species. These varied outcomes highlight the value of using molecular data for clarifying hybrid status to inform appropriate conservation actions and resourcing.
