Dioecious Plant Species Research Articles

Differential growth rate, water use efficiency and climate sensitivity between males and females of Ilex aquifolium in north-western Spain.

Dioecious plant species, i.e., those in which male and female functions are housed in different individuals, are particularly vulnerable to global environmental changes. For long-lived plant species, such as trees, long-term studies are imperative to understand how growth patterns and their sensitivity to climate variability differentially affect the sexes. Here, we explore long-term intersexual differences in wood traits, namely radial growth rates, water use efficiency quantified as stable carbon isotope abundance of wood cellulose, and their climate sensitivity in Ilex aquifolium trees growing in a natural population in NW Spain. We found that sex differences in secondary growth rates were variable over time, with males outperforming females in both radial growth rates and water use efficiency in recent decades. Summer water stress significantly reduced the growth of female trees in the following growing season, while the growth of male trees was primarily favoured by cloudy and rainy conditions the previous fall and winter combined with low cloud cover and warm conditions in summer. Sex-dependent lagged correlations between radial growth and water availability were found, with a strong association between tree growth and cumulative water availability in females at 30 months and in males at 10 months. Overall, our results point to greater vulnerability of female tress to increasing drought, which could lead to sex-ratio biases threatening population viability in the future.

Extra-stigmatic pollen germination and pistil elongation: a novel strategy towards reproductive assurance in Dodonaea viscosa subsp. angustifolia

Context Reproductive success in dioecious plant species may be limited by severe pollen limitation owing to their separate sexes and pollination barriers. Aims Dodonaea viscosa subsp. angustifolia (Jacq.) (Sapindaceae) is a dioecious and wind-pollinated species that has a long flowering period. This study sought to determine the relationship between its reproductive behaviour and pollen availability during different flowering phases. Methods Extra-stigmatic pollen germination and reproductive performance during different phases of flowering were investigated and correlated with pistil elongation under natural conditions. Results The species offers whole stigmatic and stylar surfaces for pollen to land and germinate under natural conditions. During pollen-limiting conditions, the length of the pistil increases significantly to enhance pollen capture. Depending on where on the pistil pollen lands, the timing of pollen tube arrival at the ovary varies. Conclusions Dodonaea viscosa subsp. angustifolia is the first reported wild species in the Sapindaceae where pistil elongation is regulated by pollination conditions and extra-stigmatic pollen germination ensures reproduction during phases of differing pollen availability. Our results indicated that the flexibility of female function and reproductive behaviour in Dodonaea viscosa subsp. angustifolia adds to the evolutionary possibilities to overcome pollination constraints. Implications This unique strategy for increasing female fitness through pollen presentation in D. viscosa subsp. angustifolia could be explored in other subspecies of D. viscosa on the Australian subcontinent.

How does plant sex alter microbiota assembly in dioecious plants?

Plant microbiota can greatly impact plant growth, defense, and health in different environments. Thus, it might be evolutionarily beneficial for plants to be able to control processes related to microbiota assembly. Dioecious plant species display sexual dimorphism in morphology, physiology, and immunity. These differences imply that male and female individuals might differently regulate their microbiota, but the role of sex in microbiota assembly has been largely neglected so far. Here, we introduce the mechanism of how sex controls microbiota in plants analogically to the sex regulation of gut microbiota in animals, in particular in humans. We argue that plant sex imposes selective pressure on filtering and constructing microbiota in the rhizosphere, phyllosphere, and endosphere along the soil-plant continuum. Since male plants are more resistant than female plants to environmental stresses, we suggest that a male host forms more stable and resistant plant microbiota that cooperate more effectively with the host to resist stresses. Male and female plants can distinguish whether a plant is of the same or different sex, and males can alleviate stress-caused damage in females. The impact of a male host on microbiota would protect female plants from unfavorable environments.

Drought stress-mediated differences in phyllosphere microbiome and associated pathogen resistance between male and female poplars.

Phyllosphere-associated microbes play a crucial role in plant-pathogen interactions while their composition and diversity are strongly influenced by drought stress. As dioecious plant species exhibited secondary dimorphism between the two sexes in response to drought stress, whether such difference will lead to sex-specific differences in phyllosphere microbiome and associated pathogen resistance between male and female conspecifics is still unknown. In this study, we subjected female and male full siblings of a dioecious poplar species to a short period of drought treatment followed by artificial infection of a leaf pathogenic fungus. Our results showed that male plants grew better than females with or without drought stress. Female control plants had more leaf lesion area than males after pathogen infection, whereas drought stress reversed such a difference. Further correlation and in vitro toxicity tests suggested that drought-mediated sexual differences in pathogen resistance between the two plant sexes could be attributed to the shifts in structure and function of phyllosphere-associated microbiome rather than the amount of leaf main defensive chemicals contained in plant leaves. Supportively, the microbiome analysis through high-throughput sequencing indicated that female phyllosphere enriched a higher abundance of ecologically beneficial microbes that serve as biological plant protectants, while males harbored abundant phytopathogens under drought-stressed conditions. The results could provide potential implications for the selection of suitable poplar sex to plants in drought or semi-drought habitats.

Male and female poplars exhibited sex-specific differences in metabolic and transcriptional responses to two levels of water deficit

Dioecious plant species exhibit sexual dimorphism in morphology and physiology under water deficit conditions, but the metabolic and molecular mechanisms underlying such differences are barely explored. In this study, we integrated metabolome and transcriptome analyses of female and male full-sibs of Populus deltoides that featured different drought tolerance in response to moderate and severe water shortage conditions. The results showed that male poplars had higher chlorophyll content and photosynthetic capacity, which result in a better growth performance than female poplars under drought conditions. The metabolome profiling revealed that both the quantification and qualification of differentially expressed metabolites in male poplars were higher than those in females. The transcriptome analysis revealed that the KEGG pathway was enriched in plant hormone signal transduction and carotenoid biosynthesis pathway in both male and female poplars under water deficiency. Moreover, the number of differentially expressed genes (DEGs) that were altered by plant sex was much lower than that by drought treatment. Simultaneously, most of these DEGs were up-regulated in male poplars, especially the key genes that involve in ABA synthetic pathway. The results revealed the metabolic and molecular mechanisms underlying the better ability of drought resistance observed in male poplar plants.

Pollination ecology of the early-spring-blooming dioecious shrub Eurya japonica (Pentaphylacaceae)

While the majority of angiosperm plants have hermaphrodite flowers, where a single pollinator visit can achieve both pollen removal and deposition, 5%–6% of angiosperms, including zoophilous species, are dioecious, necessitating pollen transport from male plants to females. As such, dioecy is considered less adaptive under low-temperature conditions that tend to restrict or lower potential pollinator activity. Interestingly, however, a number of dioecious, zoophilous plants such as Eurya japonica Thunb. (Pentaphylacaceae) bloom in low-temperature seasons. This study made a quantitative assessment of flower visitors/pollinators of E. japonica blooming in early spring in central Japan. A total of 15 families in 4 orders of insects were recorded on both pistillate and staminate flowers of E. japonica, of which Diptera (Empididae and Chironomidae, in particular) and Hymenoptera were predominant regardless of site/year. Hymenopteran visitors were more active at higher temperatures, while dipterans were observed even at low temperatures. The fruiting rate and the number of seeds were greater at the site where dipteran visitors were more abundant. The present study suggests that the system of multi-taxa, Diptera-dominated pollination may play an important role in the reproductive ecology of dioecious plant species, in particular those flowering at low temperatures.

Sexual differentiation in dioecious cucurbits - A molecular perspective

Understanding the evolution and genetics of sex differences is significant in both pure and applied research. Sex determination in the family Cucurbitaceae is complex owing to the existence of multiple sexual phenotypes and evolutionary changes between different systems as well as variability in chromosome numbers and structures. Dioecy (or the separate sexed condition) is widespread in angiosperms and the recent availability of molecular markers, next-generation based genome sequencing and small RNA profiling had advanced understanding of the evolution and genetics of sexual dimorphism in several dioecious plant species. Many cucurbits including some economically important species are dioecious. In this review, we summarize knowledge about the molecular mechanisms associated with sexual differentiation in dioecious cucurbits including sexlinked regions and genes identified using molecular markers. We also discuss the roles of small regulatory molecules, phytohormones and transposon variation affecting sex phenotypes in dioecious cucurbits for commercial exploitation.

Life-history characteristics and climate correlates of dioecious plant species in central southern Australia

Context The proportion of dioecious species can vary considerably among climates and habitats. However, studies often involve isolated communities or large diverse areas and fail to capture how proportions vary across diverse landscapes.Aims To identify (1) life-history associations of terrestrial dioecious plant species in central southern Australia, (2) whether proportion of dioecy varies spatially across central southern Australia, and (3) whether proportion of dioecy is correlated with life-history and/or climate factors.Methods Species growth form, pollination mechanisms and seed-dispersal features were extracted from herbarium databases to determine potential dioecy-linked traits. Distribution data for native terrestrial species in 66 Interim Biogeographical Regionalisation of Australia subregions were extracted from the Australasian Virtual Herbarium to calculate the proportion of total native species richness that are dioecious. Climate data for each subregion were also obtained from Terrestrial Ecology Research Network databases to investigate relationships among climate, life-history traits and dioecy.Key results Woodiness, abiotic pollination and endozoochory were more prevalent in dioecious than non-dioecious taxa. Proportion of dioecy ranged from 1.7% to 8.5% among subregions and correlated negatively with annual temperature range, January to March rainfall and precipitation seasonality and with average annual daily mean, minimum, maximum and average annual minimum temperature. The highest-ranked models of dioecy incorporated the additive effects of the relative proportion of woody species and either annual temperature ranges, January to March rainfall or average annual daily maximum temperature.Conclusions Dioecy was associated with woodiness, abiotic pollination and endozoochory, in line with studies of other flora, with the model of stable temperature range and woodiness being the highest-ranked model of dioecy.Implications Areas with higher proportions of dioecy can be targeted for future investigations into dioecious plant ecology to aid conservation and ecosystem management.

Sex-based metabolic and microbiota differences in roots and rhizosphere soils of dioecious papaya (Carica papaya L.).

Dioecious plant species have a high genetic variation that is important for coping with or adapting to environmental stress through natural selection. Intensive studies have reported dimorphism morphism in morphology, physiology, as well as biotic and abiotic stress responses in dioecious plants. Here, we demonstrated the dimorphism of metabolic profile and the preference of some microorganisms in the roots and rhizosphere soils of male and female papaya. The metabolic composition of roots were significantly different between the males and females. Some sex hormones occurred in the differential metabolites in roots and rhizosphere soils. For example, testosterone was up-regulated in male papaya roots and rhizosphere soils, whereas norgestrel was up-regulated in the female papaya roots, indicating a possible balance in papaya roots to control the sexual differentiation. Plant hormones such as BRs, JAs, SA and GAs were also detected among the differential metabolites in the roots and rhizosphere soils of dioecious papaya. In addition, some metabolites that have medicinal values, such as ecliptasaponin A, crocin, berberine and sapindoside A were also expressed differentially between the two sexes. Numerous differential metabolites from the papaya roots were secreted in the soil, resulting in the differences in microbial community structure in the roots and rhizosphere soils. Some nitrogen-fixing bacteria such as Allorhizobium-Neorhizobium-Pararhizobium-Rhizobium, Brevundimonas and Microvirga were enriched in the male papaya roots or rhizosphere soils. While Candidatus Solibacter and Tumebacillus, which utilize organic matters, were enriched in the roots or rhizosphere soils of the female papaya. Some differences in the fungi abundance were also observed in both male and female papaya roots. These findings uncovered the effect of sex types on the metabolic and microbiota differences in roots and rhizosphere soils in papaya and will lead to investigations of underlining genomic and molecular mechanisms.

New Evidence Suggests No Sex Bias in Herbivory or Plant Defense.

AbstractDioecious plants can exhibit sexual dimorphism across a suite of plant traits, including susceptibility to herbivory and secondary chemistry. One hypothesis is that because of greater costs of reproduction in females, males should grow faster and invest less in defense, resulting in male-biased herbivory. Indeed, a series of articles and a prominent meta-analysis have established male-biased herbivory as a rule. However, more recent reviews have raised questions about how general the pattern is, citing the low breadth of taxon sampling. The literature on this topic has not been formally quantified by meta-analysis in over 15 years. Here, we report the results of a meta-analysis of studies that measured sex bias in either herbivory and/or secondary defense in 71 dioecious plant species. We added 58 observations of herbivory and 41 of secondary chemistry to the original. We control for nonindependence of effects from the same study and taxonomic group to address critiques of earlier studies. For secondary chemistry, we found no support for any consistent difference between male and female plants. For herbivory, results are directionally similar to earlier reports, although not statistically significant once we accounted for taxonomic group and study. We also found that earlier studies reported stronger male bias than more recent studies. We discuss our results in light of the decline effect, where the magnitude of a described effect declines as the number of observations increases, and consider whether the data sets exhibit signs of evidence of the type(s) of biases that can result in declining effect sizes over time.

Drought stress drives sex-specific differences in plant resistance against herbivores between male and female poplars through changes in transcriptional and metabolic profiles

Drought stress poses adverse influence on plant growth and further alters plant-herbivore interactions. Such effect is enhanced as drought occurrence is reported to increase due to global warming. Although dioecious plant species have shown sex-specific response to drought stress through the changes in growth performance and stress tolerance, whether such changes will drive sex-specific differences in defense against herbivores between male and female plant conspecifics is barely studied. In the current study, female and male poplar full-siblings were submitted to moderate (75 % field water capacity) and severe drought (50 % field water capacity) stresses, followed by herbivore growth and feeding bioassays to test the effect of plant gender on herbivore growth and feeding performance of two specialist and two generalist leaf herbivores. The results showed that although the growth of both plant sexes was inhibited by the two drought levels, male plants performed better than female conspecifics. In the paired-choice bioassays, the specialist herbivores preferred female plants while the generalist herbivores fed more on the male plants without drought stress. Both the moderate and severe drought stresses reversed such preferences. In the triple-choice bioassays, the specialist herbivores preferred female control plants while the generalist herbivores fed more on female plants under severe drought. In addition, the specialist herbivores fed on female plants from severe drought stress grew the worst while the generalist herbivores gained the highest fresh weight. The transcriptomic and metabolomic profiling revealed that female plant leaves contained higher levels of flavonoids than males under control condition while severe drought stress remarkably reduced the levels of defensive metabolites such as flavonoids, isoflavonoids, neoflavonoids and alkaloids in female but not in male plant leaves.

The differences in cocoon and silk qualities among sex-related mulberry and silkworm feeding groups.

Sexual dimorphism is seen in many dioecious plant and animal species, which may influence their trophic interactions. The differences in trophic interactions derived from sexual dimorphism in plants may influence herbivorous performance and population dynamics. Both silkworm (Bombyx mori L.) and mulberry (Morus alba L.) usually exhibit sexual dimorphism. However, few studies have been conducted on the effect of sex-related silkworm and mulberry pairings on cocoon and silk qualities, which are crucial in sericulture. Here, we compared the differences in cocoon and silk qualities under four feeding combinations (FS-FL: female silkworm fed with leaves from female mulberry trees; MS-FL: male silkworm fed with leaves from female mulberry trees; FS-ML: female silkworm fed with leaves from male mulberry trees; MS-ML: male silkworm fed with leaves from male mulberry trees). The results showed that silkworms exhibited male-biased herbivory with more male mulberry leaves digested. The FS-ML group had higher silk weight and silk ratio of fresh cocoons than the FS-FL group, and the MS-ML group had lower coarse points than the MS-FL group. Compared with groups FS-ML and FS-FL, both MS-FL and MS-ML had smaller cocoons with longer silk lengths and a higher silk ratio of the fresh cocoons. In addition, the Entropy Weight-TOPSIS method showed the cocoon quality rank as FS-ML > FS-FL > MS-FL > MS-ML, whereas silk quality rank was MS-ML > FS-FL > FS-ML > MS-FL. These results indicate that the quality of cocoon and silk is related to the interaction of silkworm and mulberry at the sex level. Furthermore, female silkworms fed with female and male tree leaves have a higher total yield in cocoon production, while male silkworms fed with male tree leaves produced higher silk quality.

Genetic mapping of sexually dimorphic volatile and non-volatile floral secondary chemistry of a dioecious willow.

Secondary chemistry often differs between sexes in dioecious plant species, a pattern attributed to its possible role in the evolution and/or maintenance of dioecy. We used GC-MS to measure floral volatiles emitted from, and LC-MS to quantitate non-volatile secondary compounds contained in, female and male Salix purpurea willow catkins from an F2 family. Using the abundance of these chemicals, we then performed quantitative trait locus (QTL) mapping to locate them on the genome, identified biosynthetic candidate genes in the QTL intervals, and examined expression patterns of candidate genes using RNA-seq. Male flowers emitted more total terpenoids than females, but females produced more benzenoids. Male tissue contained greater amounts of phenolic glycosides, but females had more chalcones and flavonoids. A flavonoid pigment and a spermidine derivative were found only in males. Male catkins were almost twice the mass of females. Forty-two QTL were mapped for 25 chemical traits and catkin mass across 16 of the 19 S. purpurea chromosomes. Several candidate genes were identified, including a chalcone isomerase associated with seven compounds. A better understanding of the genetic basis of the sexually dimorphic chemistry of a dioecious species may shed light on how chemically mediated ecological interactions may have helped in the evolution and maintenance of dioecy.

Transcriptome Analysis and Identification of a Female-Specific SSR Marker in Pistacia chinensis Based on Illumina Paired-End RNA Sequencing.

Pistacia chinensis Bunge (P. chinensis), a dioecious plant species, has been widely found in China. The female P. chinensis plants are more important than male plants in agricultural production, as their seeds can serve as an ideal feedstock for biodiesel. However, the sex of P. chinensis plants is hard to distinguish during the seedling stage due to the scarcity of available transcriptomic and genomic information. In this work, Illumina paired-end RNA sequencing assay was conducted to unravel the transcriptomic profiles of female and male P. chinensis flower buds. In total, 50,925,088 and 51,470,578 clean reads were obtained from the female and male cDNA libraries, respectively. After quality checks and de novo assembly, a total of 83,370 unigenes with a mean length of 1.3 kb were screened. Overall, 64,539 unigenes (77.48%) could be matched in at least one of the NR, NT, Swiss-Prot, COG, KEGG, and GO databases, 71 of which were putatively related to the floral development of P. chinensis. Additionally, 21,662 simple sequence repeat (SSR) motifs were identified in 17,028 unigenes of P. chinensis, and the mononucleotide motif was the most dominant type of repeats (52.59%) in P. chinensis, followed by dinucleotide (22.29%), trinucleotide (20.15%). The most abundant repeats were AG/CT (13.97%), followed by AAC/GTT (6.75%) and AT/TA (6.10%). Based on these SSR, 983 EST-SSR primers were designed, 151 of which were randomly chosen for validation. Of these validated EST-SSR markers, 25 SSR markers were found to be polymorphic between male and female plants. One SSR marker, namelyPCSSR55, displayed excellent specificity in female plants, which could clearly distinguish between male and female P. chinensis. Altogether, our findings not only reveal that the EST-SSR marker is extremely effective in distinguishing between male and female P. chinensis but also provide a solid framework for sex determination of plant seedlings.

Environmental variation in sex ratios and sexual dimorphism in three wind‐pollinated dioecious plant species

Variation in plant sex ratios is often attributable to sex‐specific mortality in heterogeneous environments that differentially limit male and female plant reproduction. Yet sexual dimorphism and plastic responses to environmental heterogeneity are common and may co‐vary with variation in sex ratios. Here, we show that the sex ratio and the degree of sexual dimorphism for a number of plant traits varied along climatic and elevation gradients in three wind‐pollinated dioecious species, Rumex lunaria, Urtica dioica and Salix helvetica. Some of the observed sex‐specific responses to climatic variation are consistent with greater sensitivity of females to water scarcity, but most responses rather point to the greater sensitivity of males to ecological stress, consistent with larger male reproductive effort, as has been commonly reported for wind‐pollinated plants. In contrast, we found no evidence for variation in either sex ratios or sexual dimorphism expected under sexual selection. Interestingly, sex ratios and sexual dimorphism varied both along distinct and the same ecological axes of variation, suggesting that the evolution of sexual dimorphism in the measured traits was not sufficient to prevent sex‐specific mortality.

Evolution of the sex-determining region in Ginkgo biloba.

Sex chromosomes or sex-determining regions (SDR) have been discovered in many dioecious plant species, including the iconic ‘living fossil' Ginkgo biloba, though the location and size of the SDR in G. biloba remain contradictory. Here we resolve these controversies and analyse the evolution of the SDR in this species. Based on transcriptome sequencing data from four genetic crosses we reconstruct male- and female-specific genetic maps and locate the SDR to the middle of chromosome 2. Integration of the genetic maps with the genome sequence reveals that recombination in and around the SDR is suppressed in a region of about 50 Mb in both males and females. However, occasional recombination does occur except a small, less than 5 Mb long region that does not recombine in males. Based on synonymous divergence between homologous X- and Y-linked genes in this region, we infer that the Ginkgo SDR is fairly old—at least of Cretaceous origin. The analysis of substitution rates and gene expression reveals only slight Y-degeneration. These results are consistent with findings in other dioecious plants with homomorphic sex chromosomes, where the SDR is typically small and evolves in a region with pre-existing reduced recombination, surrounded by long actively recombining pseudoautosomal regions.This article is part of the theme issue ‘Sex determination and sex chromosome evolution in land plants’.

Molecular sexual determinants in Pistacia genus by KASP assay.

Pistacia is a genus of dioecious plant species whose trees can take 4-5years to reach the economically valuable fruit-bearing stage. The fruits have great importance as raw material in the food, healthcare, and baking industries. For that reason, the identification of individual plants in the early juvenile period for the pollination and positioning of trees is crucial for growers. The objective of this study is to develop markers for each Pistacia species that can help in screening the sex of plant seedlings before they reach the reproductive stage, without waiting for morphological characteristics to appear. Within this context, by using the power of the kompetitive allele-specific PCR (KASP) assay technology as a marker screening system, we successfully discriminated seven out of eight Pistacia species: P. atlantica, P. integerrima, P. khinjuk, P. mutica, P. terebinthus, P. vera, and P. lentiscus. We used a high-throughput DNA sequence read archive (SRA) to assemble a reference genome that was employed in our studies as a de novo bioinformatics method. Four genomic regions from SRA and three single-nucleotide polymorphism (SNP) positions from Kafkas et al. BMC Genomics 16:98, 2015) were selected and sequenced with collected plant material from predominantly the Antepfıstıgı Research Institute Collection Garden, and eight species were aligned intraspecifically for SNP mining. In total, 12 SNP markers were converted to KASP markers, and 5 of them (SNP-PIS-133396, SNP-PIS-167992, P-ATL-91951-565, P-INT-91951-256, P-KHI-91951-115) showed clear allelic discrimination between male and female plants. SNP-PIS-167992 and P-ATL-91951-565 were identified as the best marker assays because they showed allelic frequency differences for all individuals and for both homozygous and heterozygous characters. These markers could be the most comprehensive ones for the whole genus because they showed discriminative power for several species. This study is the first one to use the KASP assay for sex discrimination in Pistacia species, and it can be regarded as a precursor study for sex discrimination by KASP for plants in general.

Chromosome-scale haplotype-phased genome assemblies of the male and female lines of wild asparagus (Asparagus kiusianus), a dioecious plant species.

Asparagus kiusianus is a disease-resistant dioecious plant species and a wild relative of garden asparagus (Asparagus officinalis). To enhance A. kiusianus genomic resources, advance plant science, and facilitate asparagus breeding, we determined the genome sequences of the male and female lines of A. kiusianus. Genome sequence reads obtained with a linked-read technology were assembled into four haplotype-phased contig sequences (∼1.6 Gb each) for the male and female lines. The contig sequences were aligned onto the chromosome sequences of garden asparagus to construct pseudomolecule sequences. Approximately 55,000 potential protein-encoding genes were predicted in each genome assembly, and ∼70% of the genome sequence was annotated as repetitive. Comparative analysis of the genomes of the two species revealed structural and sequence variants between the two species as well as between the male and female lines of each species. Genes with high sequence similarity with the male-specific sex determinant gene in A. officinalis, MSE1/AoMYB35/AspTDF1, were presented in the genomes of the male line but absent from the female genome assemblies. Overall, the genome sequence assemblies, gene sequences, and structural and sequence variants determined in this study will reveal the genetic mechanisms underlying sexual differentiation in plants, and will accelerate disease-resistance breeding in garden asparagus.

The distribution of dioecious plant pollination, sex ratio and seed dispersal research by climate type in Australia

Dioecious-plant life-history characteristics and reproduction vary greatly among species, and have been linked to certain climate and habitat types, particularly tropical and island habitats. Australia is a large, predominantly arid continent, although it also contains mediterranean, temperate and tropical climates. Dioecious plants occupy each of these climate types, with their respective habitats and evolutionary pressures. As a result, dioecious species demonstrate great variation in key reproductive characteristics (pollination mechanism, sex ratio, and seed dispersal), which require understanding so as to conserve dioecious species effectively among climate types. Therefore, to understand dioecious-plant ecology across Australia, research must include all climate types, and investigate reproductive traits in each to better focus conservation strategies. An external factor which could prevent equal study effort across climate types would be geoclimatic preference of study location or biases in researcher interest, causing gaps in practical understanding and less effective dioecious-plant conservation management in understudied systems. To assess the prevalence and nature of dioecious plant research in Australia, we conducted a systematic mapping and scoping review to answer the following questions: (1) what climate zones have been captured by dioecious-plant research in Australia, focusing on pollination, sex ratio and seed dispersal; and (2) what is the distribution of dioecious-plant pollination, sex ratio and seed dispersal research by climate type; of the 43 studies that met the inclusion criteria, 27 (62.8%) studies were conducted in temperate climates, 8 (18.6%) in dry climates, 6 (13.9%) in tropical climates, and 5 (11.6%) in continental climates. Of the three focal areas, pollination studies (n = 24, 75% studies of entomophily) were most common, whereas sex ratio (n = 17, 41% studies of sex ratio over more than one population) and seed dispersal studies (n = 13, 54% of studies investigated abiotic dispersal) were less frequently researched. Considering the climate classification distribution in Australia, studies of dioecious species in dry climates are less prevalent than those in temperate climates. Of the focal areas of research, particularly under-investigated areas were anemophily, biotic seed dispersal, and study of female-biased species. Further studies addressing these gaps would be beneficial to inform conservation initiatives for Australian dioecious plant species. Additionally, it would be informative to investigate whether similar patterns are found in dioecious plant research in other countries with diverse climate types.

Effects of sexual dimorphism on pollinator behaviour in a dioecious species

Floral traits often display sexual dimorphism in insect‐pollinated dioecious plant species, with male individuals typically being showier than females. While this strategy is theorized to be optimal when pollinators are abundant, it might represent a risk when they become scarce, because the disproportionately high number of visits on the most attractive sex, males, might preclude efficient pollen transfer from males to females. Here, the effect of sexual dimorphism on pollination efficiency was assessed in experimental arrays of dioecious Silene dioica that were exposed to one frequent visitor of the species, Bombus terrestris, and that differed in the magnitude of sexual dimorphism for either flower number or flower size. We found that flower number dimorphism negatively affected the number of visits on female plants, on female flowers and on the number of female flowers visited after a male flower. However, flower number dimorphism had no effect on the number of pollen grains deposited per stigma, presumably because the decrease in the number of visits to female flowers was compensated by a higher number of pollen grains deposited per visit. Flower size dimorphism did not affect visitation patterns and pollen transfer in our experimental arrays. These results highlight the need to jointly study visitation patterns and pollen receipt on stigmas when exploring the effect on floral phenotypes on pollination efficiency.