Mating system, morphological and genetic evidence endorse clonality as an essential reproductive mode in Daphnopsis filipedunculata (Thymelaeaceae), a dioecious and endemic species from the Amazon.

Background and Aims: Clonality is characterized by the formation of independent individuals of the same genotype that are capable of reproducing and propagating vegetatively. Although clonality is an important mechanism that facilitates the persistence of a population, its extensive use can lead to negative impacts on sexual reproduction due to trade-offs in the investment of resources. Therefore, studies on the sexual reproduction of species that exhibit clonality can provide information about resilience to environmental changes, information about fecundity, the risk of the absence of pollinators and the ability to persist in unfavourable conditions and to successfully occupy new areas. Here, we investigated the role of clonal propagation and sexual reproduction in Daphnopsis filipedunculata (Thymelaeaceae), a dioecious species distributed only in Serra dos Carajás. Methods: We evaluated the extent of clonality in this species using molecular tools and anatomical analyses of the underground system responsible for developing new ramets. Furthermore, we analysed the sexual system and its contribution to reproductive success through morphometric analyses of floral types and pollination experiments in the field. Key Results: Overall, we found that clonal propagation plays an important role in maintaining the population of D. filipedunculata. Specifically, we demonstrated that this species presents functional male and female plants, indicating that D. filipedunculata is an obligate xenogamous species but has low reproductive success. We also showed that clonal vegetative propagation is the main form of asexual reproduction in this species, with roots responsible for clonal growth. Finally, our results indicated that this species presents an intermediate phalanx-guerrilla clonal architecture. Conclusions: Our study provides the first insights into sexual reproduction and clonal propagation in D. filipedunculata and can inform management practices, conservation and the restoration of endemic species.

Germinación, crecimiento vegetativo y morfología floral de Dasylirion cedrosanum del matorral rosetófilo de Coahuila / Germination, vegetative growth and floral morphology of Dasylirion cedrosanum from the rosetophyllous scrub of Coahuila

RESUMEN El sotol (Dasylirion cedrosanum Trel.), especie endémica del noreste de México, presenta interés económico y social por su aprovechamiento comercial. El objetivo del presente trabajo fue caracterizar la germinación, crecimiento vegetativo y morfología floral de D. cedrosanum del matorral rosetófilo de Coahuila. Se sometieron 3 lotes de 100 semillas a un proceso de imbibición y se registró el porcentaje de germinación. La caracterización de la germinación se realizó a las plántulas cultivadas en laboratorio, mientras que la morfología vegetativa se determinó en plantas adultas tomadas de un jardín botánico, con fecha de siembra conocida y a las cuales no se les dio un manejo de cultivo. La descripción de la morfología floral se realizó en plantas silvestres del cañón de San Lorenzo, Saltillo, Coahuila, México. La primera fase de la imbibición duró 48 h e incrementó 52 % el peso; la segunda o de latencia duró 48 h alcanzando un 55 %; y la tercera concluyó a las 120 h con un incremento total del 60 % de peso. Las semillas emitieron el hipocótilo a partir del quinto día. En el décimo día se obtuvo 98 % de germinación. Entre las características morfológicas distintivas de la especie, las inflorescencias, conocidas como "escapos florales", registraron una altura promedio de 3.39 m. Las flores fueron actinomorfas y unisexuales. La flor estaminada presentó perianto sin diferenciación de tépalos y sépalos. Las flores pistiladas mostraron pedicelos y receptáculo corto. El polen fue elíptico, de abertura monosulcada, con dimensión de 15 μm x 30 μm. Los frutos fueron secos e indehiscentes, conocidos como samara. Las semillas fueron turbinadas de contorno trigonal. El análisis de las características de germinación y morfología vegetativa y floral del sotol permitieron comprender las adaptaciones que esta planta ha desarrollado para crecer en los ambientes semiáridos del país.

ABSTRACT Sotol (Dasylirion cedrosanum Trel.), endemic species of northeastern Mexico, has attracted economic and social interest due to its commercial potencial. The objective of this work was to characterize the germination, vegetative growth and floral morphology of D. cedrosanum from the rosetophyllous scrub of Coahuila. Three batches of 100 seeds were submitted to an imbibition process and the germination percentage was recorded. The characterization of the germination was carried out on the seedlings grown in the laboratory, while the vegetative morphology was determined in adult plants taken from a botanical garden, with a known planting date and which were not given a crop management. The description of the floral morphology was carried out on wild plants from the San Lorenzo canyon, near the municipality of Saltillo, Coahuila, Mexico. The imbibition showed the first phase of absorption within 48 h, with an increase of 52 % in weight. The third phase began at 96 h with an increase of 55 % in weight. The seeds emitted the hypocotyl from the fifth day. On the tenth day 98 % germination was obtained. Among the distinctive morphological characteristics of the species, the inflorescences, known as "floral scapes", recorded an average height of 3.39 m. The flowers were actinomorphic and unisexual. The staminate flower presented perianth without differentiation of tepals and sepals, while the pistillate flowers showed pedicels and a short receptacle. The pollen was elliptical, with a monosulcate opening, with a dimension of 15 μm x 30 μm. The fruits were dry and indehiscent, known as samara. The seeds were turbinated with a trigonal contour. The germination and characteristics of the sotol corresponded to adaptations that allow it to grow in the semi-arid environment of this region of the country.

Looking for non-hermaphrodite cacti: multidisciplinary studies in Gymnocalycium bruchii endemic to central Argentina.

KEY MESSAGE: Through a multidisciplinary study we found that Gymnocalycium bruchii, an endemic cactus from central Argentina, acts as a dioecious species, which is the first record in this genus. Cactaceae species are typically hermaphroditic; however, about 2% have other different reproductive systems. These non-hermaphroditic species may develop sexual dimorphism in flowers or other reproductive, vegetative or ecological traits, besides a specific breeding system and floral ontogeny. Therefore, multidisciplinary research is necessary to fully understand reproduction in those species. For this purpose, we studied Gymnocalicium bruchii, a globose cactus endemic to central Argentina that is presumably dioecious or gynodioecious. We made observations in two natural and two cultivated populations. We made morphological observations of plants and flowers, and performed quantitative analyses to determine the sex ratio, size of plants and flowers, flower production, fruiting, among other variables. We performed hand-pollination, self-fertilization and free-pollination tests to determine the breeding system. Finally, we studied the anatomy and ontogeny of the reproductive organs using permanent histological slides of flower morphs at different stages. Our results confirm that Gymnocalicium bruchii is a dioecious species. Female flowers have atrophied anthers and a functional gynoecium that produces fruits and seeds. Male flowers are bigger and have a functional androecium but a sterile gynoecium. In the cultivated population, the sex ratio was 1/1, whereas the number of male individuals was higher in both natural populations. Pollination tests corroborated dioecy. Ontogenetic studies revealed that in female flowers the anthers collapse before microspore maturation, while in male flowers the gynoecium shows normal development of the ovary, style, stigma, and ovules; however, the latter are never fertilized.

Flower structure, anatomy, and sexuality of Chrysophyllum gonocarpum (Sapotaceae).

The Sapotaceae are a significant component of the humid forests of the Neotropics and have many species of economic interest. Chrysophyllum gonocarpum is one of them and its edible fruits have currently acquired a high commercial value. Since there are no studies that cover its floral anatomy and elucidate its sexual system, the objective of the present study is to describe these aspects based on field observations and a detailed anatomical analysis of their flowers. Conventional techniques of plant anatomy are implemented. The results indicate that the species presents cryptic dioecy, showing specimens with morphologically and functionally pistillate flowers (with reduced staminodes), and trees with morphologically hermaphrodite and functionally staminate flowers. In addition, data on floral nectaries and laticiferous are provided.

Strongly unbalanced gender attractiveness in a dioecious mass flowering tropical tree pollinated by stingless bees.

In dioecious, bee-pollinated tree species, male and female flowers offer different resources. It is unclear how this unbalanced quantity and quality of floral resources affects flower visits and pollen transfer to female flowers. We asked, what characteristics of flowering and dynamics of resource production by trees favour flights of bees between male and female trees. We quantified the floral resources produced by individual flowers and entire trees of Myracrodruon urundeuva (Anacardiaceae), measured pollen flow to female flowers, fruit set of naturally pollinated flowers and determined the effective pollinators. Crown volume of male trees was four-fold, flower number 15-fold and nectar volume 60 times higher than in female trees. While ~70% of male flowers opened in the morning, ~70% of female flowers opened in the afternoon. Fruit set was 27%. Stingless bee species were the main pollinators, while honeybees were common only on male flowers. Strongly unbalanced production of floral resources, high potential lifespan of female flowers and anticipated opening of male flowers favour pollinator movement and pollen deposition on female flowers, albeit low (0.003% of pollen that reached stigmas), but sufficient to produce thousands of fruits per tree. Besides being an astonishing pollen and nectar source for numerous social bee species, only a few of them were effective pollinators. Our quantitative approach to floral resource production of each gender provides new insights, such as the proportion of resources allocated to each gender and the corresponding behaviour of flower visitors, for understanding the reproductive strategy of dioecious tropical mass-flowering trees.

Distance Dependent Contribution of Ants to Pollination but Not Defense in a Dioecious, Ambophilous Gymnosperm.

Dioecious plants are obligate outcrossers with separate male and female individuals, which can result in decreased seed set with increasing distance between the sexes. Wind pollination is a common correlate of dioecy, yet combined wind and insect pollination (ambophily) could be advantageous in compensating for decreased pollen flow to isolated females. Dioecious, ambophilous gymnosperms Ephedra (Gnetales) secrete pollination drops (PDs) in female cones that capture airborne pollen and attract ants that feed on them. Plant sugary secretions commonly reward ants in exchange for indirect plant defense against herbivores, and more rarely for pollination. We conducted field experiments to investigate whether ants are pollinators and/or plant defenders of South American Ephedra triandra, and whether their contribution to seed set and seed cone protection varies with distance between female and male plants. We quantified pollen flow in the wind and assessed the effectiveness of ants as pollinators by investigating their relative contribution to seed set, and their visitation rate in female plants at increasing distance from the nearest male. Ants accounted for most insect visits to female cones of E. triandra, where they consumed PDs, and pollen load was larger on bigger ants without reduction in pollen viability. While wind pollination was the main contributor to seed set overall, the relative contribution of ants was distance dependent. Ant contribution to seed set was not significant at shorter distances, yet at the farthest distance from the nearest male (23 m), where 20 times less pollen reached females, ants enhanced seed set by 30% compared to plants depending solely on wind pollination. We found no evidence that ants contribute to plant defense by preventing seed cone damage. Our results suggest that, despite their short-range movements, ants can offset pollen limitation in isolated females of wind-pollinated plants with separate sexes. We propose that ants enhance plant reproductive success via targeted delivery of airborne pollen, through frequent contact with ovule tips while consuming PDs. Our study constitutes the first experimental quantification of distance-dependent contribution of ants to pollination and provides a working hypothesis for ambophily in other dioecious plants lacking pollinator reward in male plants.

In situ radiation explains the frequency of dioecious palms on islands.

BACKGROUND AND AIMS: Dioecy has evolved up to 5000 times in angiosperms, despite the potentially high intrinsic costs to unisexuality. Dioecy prevents inbreeding, which is especially relevant on isolated islands when gene pools are small. Dioecy is also associated with certain dispersal traits, such as fruit size and type. However, the influence of dioecy on other life history traits and island distribution remains poorly understood. Here, we test the effect of dioecy on palm (Arecaceae) speciation rates, fruit size and frequency on islands. METHODS: We used phylogenetic comparative methods to estimate the ancestral state of the sexual system and its impact on speciation rates and fruit size. Frequency of sexual systems, effect of insularity on the probability of being dioecious, and phylogenetic clustering of island dioecious vs. mainland species were inferred. Lastly, we determined the interplay of insularity and sexual system on speciation rates. KEY RESULTS: Palms repeatedly evolved different sexual systems (dioecy, monoecy and polygamy) from a hermaphrodite origin. Differences in speciation rates and fruit size among the different sexual systems were not identified. An effect of islands on the probability of the palms being dioecious was also not found. However, we found a high frequency and phylogenetic clustering of dioecious palms on islands, which were not correlated with higher speciation rates. CONCLUSIONS: The high frequency and phylogenetic clustering may be the result of in situ radiation and suggest an 'island effect' for dioecious palms, which was not explained by differential speciation rates. This island effect also cannot be attributed to long-distance dispersal due to the lack of fruit size difference among sexual systems, and particularly because palm dispersal to islands is highly constrained by the interaction between the sizes of fruit and frugivores. Taken together, we suggest that trait flexibility in sexual system evolution and the in situ radiation of dioecious lineages are the underlying causes of the outstanding distribution of palms on islands.

Pleistocene aridification underlies the evolutionary history of the Caribbean endemic, insular, giant Consolea (Opuntioideae).

PREMISE: The Caribbean islands are in the top five biodiversity hotspots on the planet; however, the biogeographic history of the seasonally dry tropical forest (SDTF) there is poorly studied. Consolea consists of nine species of dioecious, hummingbird-pollinated tree cacti endemic to the West Indies, which form a conspicuous element of the SDTF. Several species are threatened by anthropogenic disturbance, disease, sea-level rise, and invasive species and are of conservation concern. However, no comprehensive phylogeny yet exists for the clade. METHODS: We reconstructed the phylogeny of Consolea, sampling all species using plastomic data to determine relationships, understand the evolution of key morphological characters, and test their biogeographic history. We estimated divergence times to determine the role climate change may have played in shaping the current diversity of the clade. RESULTS: Consolea appears to have evolved very recently during the latter part of the Pleistocene on Cuba/Hispaniola likely from a South American ancestor and, from there, moved into the Bahamas, Jamaica, Puerto Rico, Florida, and the Lesser Antilles. The tree growth form is a synapomorphy of Consolea and likely aided in the establishment and diversification of the clade. CONCLUSIONS: Pleistocene aridification associated with glaciation likely played a role in shaping the current diversity of Consolea, and insular gigantism may have been a key innovation leading to the success of these species to invade the often-dense SDTF. This in-situ Caribbean radiation provides a window into the generation of species diversity and the complexity of the SDTF community within the Antilles.

Isolation and characterization of microsatellite loci in Astroniumgraveolens (Anacardiaceae) and cross amplification in related species.

Microsatellite markers are a useful genetic tool to answer ecological and conservation genetics questions. Microsatellite primers were developed and characterized to evaluate forest fragmentation effects on genetic structure, diversity and gene flow patterns in the dioecious tropical tree Astroniumgraveolens (Anacardiaceae). Using genomic library enrichment, sixteen microsatellite loci were developed for A.graveolens. Polymorphism was evaluated in 80 individuals from four localities in Mexico. The overall number of alleles detected was 63, average alleles per locus 3.9 with a range from one to 11 per locus. Cross amplification trails on related species in the Anacardiaceae family: Spondiaspurpurea and Amphipterygiumadstringens, achieved successful amplification for all primers. Microsatellite markers described here are the first to be characterized for A.graveolens. These genetic markers will be a useful tool to assess the genetic consequences of habitat fragmentation and selective logging on A.graveolens populations. Cross amplification success on S.purpureaandA.adstringens suggest that they may be used for population genetic studies in other species within the family.

Comparative development of staminate and pistillate flowers in the dioecious cactus Opuntia robusta.

KEY MESSAGE: PCD role in unisexual flowers. The developmental processes underlying the transition from hermaphroditism to unisexuality are key to understanding variation and evolution of floral structure and function. A detailed examination of the cytological and histological patterns involved in pollen and ovule development of staminate and pistillate flowers in the dioecious Opuntia robusta was undertaken, and the potential involvement of programmed cell death in the abortion of the sex whorls was explored. Flowers initiated development as hermaphrodites and became functionally unisexual by anthesis. Female individuals have pistillate flowers with a conspicuous stigma, functional ovary, collapsed stamens and no pollen grains. Male individuals have staminate flowers, with large yellow anthers, abundant pollen grains, underdeveloped stigma, style and an ovary that rarely produced ovules. In pistillate flowers, anther abortion resulted from the premature degradation of the tapetum by PCD, followed by irregular deposition of callose wall around the microsporocytes, and finally by microspore degradation. In staminate flowers, the stigma could support pollen germination; however, the ovaries were reduced, with evidence of placental arrest and ovule abortion through PCD, when ovules were present. We demonstrate that PCD is recruited in both pistillate and staminate flower development; however, it occurs at different times of floral development. This study contributes to the understanding of the nature of the O. robusta breeding system and identifies developmental landmarks that contribute to sexual determination in Cactaceae.

Completing the jigsaw: the first record of the female plant of Daphnopsisfilipedunculata (Thymelaeaceae), an endemic species from the Brazilian Amazon.

The results of intensive fieldwork in the National Forest of Carajás (FLONA Carajás) led to the discovery of pistillate plants of Daphnopsisfilipedunculata, an endemic species from the Serra dos Carajás, previously known only from staminate individuals. These newly discovered populations add valuable missing information related to pistillate buds, mature flowers and fruits.

Best in the company of nearby males: female success in the threatened cycad, Zamia portoricensis.

Variation in plant reproductive success is affected by ecological conditions including the proximity of potential mates. We address the hypothesis that spatial distribution of sexes affects female reproductive success (RS) in the dioecious cycad, Zamia portoricensis. Are the frequencies of males, operational sex ratios, and distances to the nearest mate associated with RS in females? We studied the spatial distribution of sexes in two populations in Puerto Rico and compared RS of target females with the number of males and operational sex ratios. Population structure suggests regular successful recruitment. Adults, males, and females were randomly distributed with respect to one another. Reproductive success of females was highly variable, but was higher in neighborhoods with more males than females and generally decreased with increasing distance to the nearest male, becoming statistically significant beyond 190 cm. This possible mate-finding Allee effect indicates that pollinator movement among plants may be limited for this mutually dependent plant-pollinator interaction. Yet being close to male plants is a matter of chance, perhaps a factor generating the high intra-population genetic diversity in Z. portoricensis.

Female fruit production depends on female flower production and crown size rather than male density in a continuous population of a tropical dioecious tree (Virola surinamensis).

PREMISE OF THE STUDY: Factors related to pollen and resource limitation were evaluated to predict female fruit production in a tropical dioecious tree. Pollen limitation via variation in the male density at local scales is expected to limit female reproduction success in dioecious plants. METHODOLOGY: We modeled the roles of local male density, female crown size, crown illumination, and female flower production on female fruit initiation and mature fruit production in a continuous population (62 ha plot) of a tropical dioecious tree (Virola surinamensis). In addition, we used microsatellites to describe the scale of effective pollen flow, the male effective population size, and the spatial genetic structure within/between progenies and males. KEY RESULTS: The local male density was not related to female fruit initiation or mature fruit production. Female floral production had a positive effect on fruit initiation. The female crown size was positively related to fruit maturation. Seeds from the same female and seeds from different but spatially proximal females were generally half-siblings; however, proximal females showed greater variation. Proximal male-female adult pairs were not significantly more genetically related than distant pairs. The probability of paternity was negatively affected by the distance between seeds and males; most effective pollen dispersal events (∼85%) occurred from males located less than 150 m from females. The number of males siring progenies was greater than the number of males found at local scales. CONCLUSIONS: Female fecundity in this continuous population of Virola surinamensis is not limited by the availability of pollen from proximal males. Rather, resource allocation to floral production may ultimately determine female reproductive success.

Reproductive biology of Syzygiella rubricaulis (Nees) Steph. (Adelanthaceae, Marchantiophyta), a liverwort disjunctly distributed in high-altitude Neotropical mountains.

Syzygiella rubricaulis is a dioecious leafy liverwort disjunctly distributed and restricted to high-altitude mountains in the Neotropics and the Azores. This study is part of a larger project examining the phylogeography of S. rubricaulis in the Neotropics, and our main goals were to understand its reproductive biology, where sex expression occurs, if vegetative propagules are frequently found, how the sexes are distributed in populations, how frequently sporophytes are formed and what environmental conditions influence sexual expression. S. rubricaulis patches are mostly female, but all patches also contain non sex-expressing shoots. Out of 42 patches examined, 29 (69%) were sex-expressing: 25 were unisexual (21 female and four male) and four of mixed sex (two male-biased and two unbiased). At shoot level, out of 4200 shoots 18% were female and 7% male; among sex-expressing shoots, 73% were female, representing a sex ratio of 0.8 (female-biased). We encountered a total of 33 sporophytes in six patches (in Brazil, Venezuela and Ecuador). Leaf regenerants were found in one patch in Mexico. Low rates of sporophytes were likely related to low frequencies of male shoots and large distances between the sexes. As 25% of S. rubricaulis shoots expressed sex (occasionally producing sporophytes), we suggest that short-distance (and rarely long-distance) spore dispersal events occur in mountainous areas on a short-term basis. On a long-term basis, however, these events likely contribute to dynamic exchanges among populations in the Neotropics.

Correlated polymorphism in cytotype and sexual system within a monophyletic species, Lycium californicum.

BACKGROUND AND AIMS: Polyploidy has important effects on reproductive systems in plants and has been implicated in the evolution of dimorphic sexual systems. In particular, higher ploidy is associated with gender dimorphism across Lycium species (Solanaceae) and across populations within the species Lycium californicum. Previous research on the association of cytotype and sexual system within L. californicum sampled a limited portion of the species range, and did not investigate evolutionary transitions between sexual systems. Lycium californicum occurs in arid regions on offshore islands and mainland regions in the south-western United States and Mexico, motivating a more comprehensive analysis of intraspecific variation in sexual system and cytotype across the full range of this species. METHODS: Sexual system (dimorphic vs. cosexual) was determined for 34 populations across the geographical range of L. californicum using field observations of pollen production, and was confirmed using morphological measurements and among-plant correlations of primary sexual traits. Ploidy was inferred using flow cytometry in 28 populations. DNA sequence data from four plastid and two nuclear regions were used to reconstruct relationships among populations and to map transitions in sexual system and ploidy. KEY RESULTS: Lycium californicum is monophyletic, ancestrally diploid and cosexual, and the association of gender dimorphism and polyploidy appears to have two evolutionary origins in this species. Compared with cosexual populations, dimorphic populations had bimodal anther size distributions, negative correlations between male and female floral traits, and larger coefficients of variation for primary sexual traits. Flow cytometry confirmed tetraploidy in dimorphic populations, whereas cosexual populations were diploid. CONCLUSIONS: Tetraploidy and gender dimorphism are perfectly correlated in L. californicum, and the distribution of tetraploid-dimorphic populations is restricted to populations in Arizona and the Baja California peninsula. The analysis suggests that tetraploidy and dimorphism likely established in Baja California and may have evolved multiple times.

Pollination biology in the dioecious orchid Catasetum uncatum: How does floral scent influence the behaviour of pollinators?

Catasetum is a neotropical orchid genus that comprises about 160 dioecious species with a remarkable sexual dimorphism in floral morphology. Flowers of Catasetum produce perfumes as rewards, which are collected only by male euglossine bees. Currently, floral scents are known to be involved in the selective attraction of specific euglossine species. However, sexual dimorphism in floral scent and its eventual role in the pollination of Catasetum species have never been investigated. Here, we have investigated the pollination of Catasetum uncatum and asked: (1) Is floral scent a sexual dimorphic trait? (2) Does pollinarium removal/deposition affect scent emission? (3) Does sexual dimorphism in floral scent and changed scent emission have implications with regard to the behaviour of the pollinators? The frequency and behaviour of floral visitors were observed in non-manipulated flowers (both flower sexes) and in manipulated flowers (pistillate only) in which pollinaria were deposited. Scents of staminate and pistillate flowers (both manipulated and non-manipulated) were collected by using dynamic headspace methods and analysed chemically. Electrophysiological analyses were performed to detect compounds triggering antennal depolarisation in the euglossine species. C. uncatum is pollinated mainly by males of Euglossa nanomelanotricha. Pollinators were more frequent in pistillate than in staminate inflorescences. Bees approaching staminate flowers frequently flew away without visiting them, a behavioural pattern not observed in pistillate flowers. In the chemical analyses, we recorded 99 compounds, 31 of which triggered antennal depolarisation in pollinators. Multivariate analyses with the electrophysiological-active compounds did not detect differences between the scent composition of staminate and pistillate flowers. Pollinarium removal or deposition resulted in diminished scent emission within 24h in staminate and pistillate flowers, respectively. Surprisingly, bees discriminated pollinated from non-pollinated pistillate flowers as early as 2h after pollination. The rapid loss in the attractiveness of flowers following pollinarium removal/deposition can be interpreted as a strategy to direct pollinators to non-pollinated flowers. We have found no evidence that euglossine males discriminate staminate from pistillate flowers by means of floral scent. Instead, we speculate that bees use visual cues, such as sex dimorphic traits, to discriminate flowers of different sexes. Together, our results provide interesting insights into the evolution of floral signals in gender-dimorphic species and into its significance in plant reproductive biology.

The ecology, evolution, and biogeography of dioecy in the genus Solanum: with paradigms from the strong dioecy in Solanum polygamum, to the unsuspected and cryptic dioecy in Solanum conocarpum.

UNLABELLED: • PREMISE OF THE STUDY: Island plants are over-exploited and "under-explored." Understanding the reproductive biology of plants, especially rare species, is fundamental to clarifying their evolution, estimating potential for change, and for creating effective conservation plans. Clarification of sexual systems like dioecy, and unusual manifestations of it in specific studies within Solanum, helps elucidate evolutionary patterns and genetic and ecological control of sex expression.• METHODS: Studies of reproductive systems of two Caribbean endemics, S. polygamum and S. conocarpum, combined multifaceted analyses of field populations and of multiple generations of greenhouse plants.• KEY RESULTS: The dioecy in both species is, like that in other solanums, largely cryptic, although the gender of S. polygamum flowers is obvious. The rare S. conocarpum is recognized as dioecious; floral gender is not obvious. Variation in sex expression facilitated experiments and promoted hypotheses on control and significance of morphological features and sex expression.• CONCLUSIONS: Confirmed dioecy in at least 15 solanums is distributed across the genus, with perhaps 6 independent origins, and with crypticity in the form of morphologically hermaphroditic, but functionally unisexual, flowers characterizing all species. Dioecy is not more strongly associated with islands. Inaperturate pollen in pistillate flowers characterizes almost all, but not the two dioecious species studied herein. Dioecy in both species indicates leakiness (rare hermaphroditic flowers on male plants) that helps explain island colonization and radiation. Leakiness allowed confirmation-usually impossible for dioecious species-of self-compatibility for S. polygamum, and thus support for the hypothesis that dioecy evolved to promote outcrossing.

Tree size and its relationship with flowering phenology and reproductive output in Wild Nutmeg trees.

Reproductive strategies, sexual selection, and their relationship with the phenotype of individuals are topics widely studied in animals, but this information is less abundant for plants. Variability in flowering phenology among individuals has direct impact on their fitness, but how reproductive phenology is affected by the size of the individuals needs further study. We quantified the flowering intensity, length, and reproductive synchronization of two sympatric dioecious Wild Nutmeg tree species (Virola, Myristicaceae) in the Brazilian Atlantic forest, and analyzed its relationships with tree size. Two distinct strategies in flowering timing and intensity were found between species (annual versus biennial flowering), and among individuals in the annual flowering species (extended versus peak flowering). Only for the annual flowering species the reproductive output is related to tree size and large trees present proportionally higher flower coverage, and lower synchronization than smaller ones. Flowering is massive and highly synchronized in the biennial species. Sex ratios are not different from 1:1 in the two species, and in the two segregated reproductive subgroups in the biennial flowering species. The biennial flowering at individual level is a novelty among reproductive patterns in plants, separating the population in two reproductive subgroups. A proportional increase in the reproductive output with size exists only for the annual flowering species. A biennial flowering can allow resource storage favouring massive flowering for all the individuals diluting their relationship with size.

Programmed cell death promotes male sterility in the functional dioecious Opuntia stenopetala (Cactaceae).

BACKGROUND AND AIMS: The sexual separation in dioecious species has interested biologists for decades; however, the cellular mechanism leading to unisexuality has been poorly understood. In this study, the cellular changes that lead to male sterility in the functionally dioecious cactus, Opuntia stenopetala, are described. METHODS: The spatial and temporal patterns of programmed cell death (PCD) were determined in the anthers of male and female flowers using scanning electron microscopy analysis and histological observations, focusing attention on the transition from bisexual to unisexual development. In addition, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling assays were used as an indicator of DNA fragmentation to corroborate PCD. KEY RESULTS: PCD was detected in anthers of both female and male flowers, but their patterns differed in time and space. Functionally male individuals developed viable pollen, and normal development involved PCD on each layer of the anther wall, which occurred progressively from the inner (tapetum) to the outer layer (epidermis). Conversely, functional female individuals aborted anthers by premature and displaced PCD. In anthers of female flowers, the first signs of PCD, such as a nucleus with irregular shape, fragmented and condensed chromatin, high vacuolization and condensed cytoplasm, occurred at the microspore mother cell stage. Later these features were observed simultaneously in all anther wall layers, connective tissue and filament. Neither pollen formation nor anther dehiscence was detected in female flowers of O. stenopetala due to total anther disruption. CONCLUSIONS: Temporal and spatial changes in the patterns of PCD are responsible for male sterility of female flowers in O. stenopetala. Male fertility requires the co-ordination of different events, which, when altered, can lead to male sterility and to functionally unisexual individuals. PCD could be a widespread mechanism in the determination of functionally dioecious species.

Reproductive biology of Hechtia schottii, a dioecious Bromeliaceae, in Mexico

Hechtia schottii is a terrestrial, rosetofilous, dioecious, polycarpic succulent herb, that grows mainly in shrubby associations, and less frequently, in secondary low caducifolious forests, both on calcareous soils or limestone outcrops in Yucatan and Campeche States, Mexico. We studied phenology, floral and pollination biology, and breeding system at Calcehtok, Yucatan, during two flowering seasons. Plants bloom mainly during the dry season (November-April) and disperse seeds during the rainy season (May-October). Both floral morphs have diurnal anthesis; pollen is removed ca. 1 h after anthesis starts and both floral morphs are visited by several insect species, especially bees, but results suggest that the introduced honey bee, Apis mellifera, is the pollinator. Controlled crossings show that the species is functionally dioecious and requires to be serviced by pollinators based on fruit setting only in unassisted cross pollination crosses.

Hechtia schottii es una hierba terrestre, suculenta, rosetófila, dioica y policárpica, que crece en asociaciones arbustivas y selva baja caducifolia secundaria, ambos en suelos calcáreos o limosos. Estudiamos la fenología, la biología floral, reproductiva y de la polinización en una población en Calcehtok, Yucatán, México, durante dos estaciones de floración. Las plantas florecen principalmente en la época de secas (noviembre-abril) y la dispersión de semillas es durante la estación de lluvias (mayo-octubre). Ambas formas (morphs) florales tienen antesis diurna; el polen es removido ca. 1 h después del comienzo de la antesis y ambas formas florales son visitadas por varias especies de insectos, especialmente abejas, pero los resultados sugieren que la abeja introducida, Apis mellifera, es el polinizador. Cruces controlados muestran que la especie es funcionalmente dioica y que requiere de un polinizador, ya que solo produce frutos por polinización no asistida entre formas.