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Background and Aims Historical changes in environmental conditions and colonization–extinction dynamics have a direct impact on the genetic structure of plant populations. However, understanding how past environmental conditions influenced the evolution of species with high gene flow is challenging when signals for genetic isolation and adaptation are swamped by gene flow. We investigated the spatial distribution and genetic structure of the widespread terrestrial orchid Epipactis helleborine to identify glacial refugia, characterize postglacial population dynamics and assess its adaptive potential. Methods Ecological niche modelling was used to locate possible glacial refugia and postglacial recolonization opportunities of E. helleborine. A large single-nucleotide polymorphism (SNP) dataset obtained through genotyping by sequencing was used to define population genetic diversity and structure and to identify sources of postglacial gene flow. Outlier analyses were used to elucidate how adaptation to the local environment contributed to population divergence. Key Results The distribution of climatically suitable areas was restricted during the Last Glacial Maximum to the Mediterranean, south-western Europe and small areas in the Alps and Carpathians. Within-population genetic diversity was high in E. helleborine (mean expected heterozygosity, 0.373 ± 0.006; observed heterozygosity, 0.571 ± 0.012; allelic richness, 1.387 ± 0.007). Italy and central Europe are likely to have acted as important genetic sources during postglacial recolonization. Adaptive SNPs were associated with temperature, elevation and precipitation. Conclusions Forests in the Mediterranean and Carpathians are likely to have acted as glacial refugia for Epipactis helleborine. Postglacial migration northwards and to higher elevations resulted in the dispersal and diversification of E. helleborine in central Europe and Italy, and to geographical isolation and divergent adaptation in Greek and Italian populations. Distinguishing adaptive from neutral genetic diversity allowed us to conclude that E. helleborine has a high adaptive potential to climate change and demonstrates that signals of adaptation and historical isolation can be identified even in species with high gene flow.

Premise of research. In hermaphroditic plants, pollen production can enhance fitness through siring success by attracting pollen-seeking pollinators. However, pollen production can also decrease fe...

Hybridization can often lead to the formation of novel taxa which can have traits that resemble either or both parental species. Determining the similarity of hybrid traits to parental taxa is particularly important in plant conservation, as hybrids that form between rare and common taxa may more closely resemble a rare parental species, thereby putting the rare parental taxon at further risk of extinction via increased backcrossing and introgression. We investigated the floral (morphological and chemical) traits and orchid mycorrhizal (OrM) fungal associations of the endangered orchid Orchis patens, its more common sister species O. provincialis, and their natural hybrid O. × fallax in natural sympatric populations. We found that both morphological and chemical floral traits of O. × fallax are shared by the parents but are more similar to O. patens than O. provincialis. OrM fungi were shared among all three taxa, indicating that the availability of OrM fungi should not represent a barrier to establishment of individuals of any of these taxa. These results suggest that O. × fallax may be able to expand its distribution within a similar niche to O. patens. This highlights the importance of quantifying differences between hybrids and parental taxon in species conservation planning.

AIM: Interactions with mycorrhizal fungi are increasingly recognized as an important factor underlying the distribution and abundance of orchid species. However, the geographical distribution of orchid mycorrhizal fungi (OMF) and how their communities vary over large geographical areas are less well understood. Because climatic and environmental similarity may decrease with geographical distance or because some OMF have limited dispersal capabilities, similarities in orchid mycorrhizal communities can be expected to decrease with increasing distances separating orchid populations. However, up till now empirical evidence is largely lacking. LOCATION: Eurasia. TAXA: Gymnadenia conopsea (L.) R. Brown and Epipactis helleborine (L.) Crantz. METHODS: High‐throughput sequencing was used to perform a cross‐continental comparison of OMF that associate with two widespread Eurasian terrestrial orchids, Epipactis helleborine and Gymnadenia conopsea. Both phylogenetic and nonphylogenetic measures of community dissimilarity and their components were calculated and related to geographical distances using Mantel tests. RESULTS: Our results showed that in both orchid species similarity in mycorrhizal communities decreased significantly with geographical distance. Decomposing the contribution of spatial turnover and nestedness to overall dissimilarity showed that the observed dissimilarity was mainly the result of species replacement between regions, and not of species loss. Similarly, a strong relationship was observed between phylogenetic community dissimilarity and geographical distance. Decomposing PCD values into a nonphylogenetic and phylogenetic component showed that orchid populations located closely next to each other were likely to contain the same operational taxonomic units (OTUs), but that the non‐shared taxa came from different phylogenetic clades. Species indicator analyses showed that the majority of OMF OTUs were restricted to particular geographical areas. However, some OTUs occurred in both continents, indicating that some fungi have very wide distributions. MAIN CONCLUSIONS: Overall, these results demonstrate that orchid mycorrhizal communities differ substantially across large geographical areas, but that the distribution of orchids is not necessarily restricted by the distribution of particular OMF. Hence, widespread orchid species can be considered mycorrhizal generalists that are flexible in the OMF with which they associate across large geographical areas. ispartof: JOURNAL OF BIOGEOGRAPHY vol:47 issue:2 pages:421-433 status: published

New populations of endangered plants may establish outside of protected areas if a suitable habitat becomes available. However, it is unclear whether such populations are genetically uniform, as a result of a founder effect, or whether they contain genetic variation resulting from continuous gene flow from source populations, and subsequent recruitment after establishment. We addressed this by examining haplotype variation in the endangered orchid, Anacamptis robusta, which has formed a new population outside of its protected area within the past 20 years. To assess population growth, the number of A. robusta flowering plants were counted every year for 22 years in both the new population and populations within the reserve. Haplotype diversity and fine-scale structure were examined with spatial autocorrelation analysis, both in the new population and representative populations from the protected area. The number of flowering individuals in the new population increased from 9 to 2,277 between 2003 and 2012, whereas within the reserve flowering population sizes varied between years. Seventeen of 23 haplotypes detected for A. robusta were detected in the new population, with seven of these occurring more frequently in the new population than reserve populations. In the new population, there was strong fine-scale spatial structure of haplotypes, similar to patterns found in large populations from the protected area, suggesting multiple colonization events and subsequent local recruitment. This highlights that ongoing demographic and genetic monitoring of plant populations is vital to improve our understanding of population colonization and the conservation of narrow endemics.

Pollinator taxa may vary in their contribution to plant outcrossing rates and hence differ in their influence on plant fitness. In plants that have ornithophilous floral traits (large, deep-tubed orange-red flowers with dilute nectar), it is expected that birds should be the most effective agents of outcrossing. However, bees often pollinate plants with ornithophilous floral traits and may contribute to their outcrossing. To assess the contribution of bees to outcrossing rates and inbreeding in the rare self-compatible treelet Aloe thraskii, we compared outcrossing rates (t) and adult inbreeding coefficients (F), in bird-excluded and open-pollinated plants. We then tested whether outcrossing rates, and inbreeding depression (δ) estimated using Ritland’s genetic marker-based approach, varied according to bee and bird visitation rates, plant height, flowering population size, population density, and isolation distance across eight populations. Outcrossing rates did not differ between progeny from open-pollinated and bird-excluded inflorescences. Outcrossing rates increased with bee visitation, but not bird visitation. Adult inbreeding coefficients were low (F = 0–0.1), and inbreeding depression estimates were high (δ = 0.59–1). Estimates of inbreeding depression did not vary according to population aggregation. These results highlight the contribution to outcrossing that insect pollinators can make to outcrossing rates of rare plant species that are seemingly adapted for bird pollination.

Aim: Biodiversity is known to be unevenly distributed along latitudinal gradients. While this pattern has been observed for many different organisms, it is unclear whether the distributions of ecologically important belowground mutualists, such as orchid mycorrhizal fungi (OMF), also vary according to latitude.Location: Europe. Taxon: Spiranthes spiralis (L. Chevall) Orchidaceae. Methods: We sampled 37 populations over a >3,000 km latitudinal gradient of the European orchid Spiranthes spiralis to test whether the diversity and community com-position of OMF are influenced by latitude. For this, we sequenced the broad spectrum ITS3/4OF region to identify OMF and quantified environmental variables that may influence their occurrence. To assess whether seed germination was affected by suitable OMF in different regions, we performed a reciprocal translocation germina-tion experiment at the northern, central and southern part of its range. Results: In total, 75 OMF taxa were associated with S. spiralis, the majority belonging to the Ceratobasidiaceae and Thelephoraceae. Both taxonomic and phylogenetic OMF diversity decreased with increasing latitude. Three Ceratobasidiaceae taxa were widespread, but the number of sequences of two of these taxa decreased with increasing latitude. Reciprocal translocation germination experiments revealed that S. spiralis seeds germinated in each region regardless of seed provenance, but germination rates were highest in the southern part of its range where mycorrhizal diversity was highest. Main conclusions: OMF diversity decreased with increasing latitude, indicating that communities of OMF may be shaped by latitudinal gradients. These results highlight the need to understand the factors that determine the distribution of belowground mutualists, which are essential to predict how plant distributions will shift under future environmental change.

Ecogeographical isolation is a fundamental prezygotic barrier to reproduction and a step towards diversification in flowering plants. However, whether ecogeographical isolation acts as a reproductive barrier between species and thus as a mechanism for species divergence is unclear and is expected to change as species distributions shift under climate change. Using a Maxent framework, we quantified the extent of ecogeographical isolation of nine closely related species of the European plant genus Pulmonaria, which lack premating barriers and often form hybrids. We investigated ecogeographical isolation under both current conditions and four climate change scenarios; two unrelated future climate models, each with two predictions of the potential trajectories of climate change. Compared with current climates, under these future climate scenarios, there will be an increase in mean annual temperature, but not in mean annual precipitation in the range of Pulmonaria throughout Europe. This will result in a range contraction in three of the nine Pulmonaria species, a range expansion in two species, and no clear effect of climate change on the ranges of four species. Climate change will result in an overall increase in ecogeographical isolation between these species in future. In particular, species that have low ecogeographical isolation under current conditions are predicted to increase the most in their ecogeographical isolation under climate change. Synthesis. These results highlight that climate change can strongly impact on a major premating barrier of closely related species, therefore potentially affect the evolutionary trajectory of plants in the future. ispartof: JOURNAL OF ECOLOGY vol:107 issue:1 pages:167-177 status: published

Nursery pollination is an unusual plant–insect interaction in which an insect is both pollinator and seed predator. Depending on the abundance of the nursery pollinator and of other pollinators, this interaction can range from mutualism to parasitism and it is thus likely to vary geographically. We investigated this mechanism in the widespread species Silene latifolia in a Mediterranean environment that is likely to offer a rich pollinator community to the plant, thus decreasing the dependence on the nursery pollinator. Surprisingly, we found that although generalist pollinators contribute significantly to plant fitness, the nursery pollinator is still the more efficient., Plants involved in specialized pollinator interactions, such as nursery pollination, may experience trade-offs in their female fitness, as the larvae of their pollinators may also consume seeds produced by the flowers they pollinate. These interactions could potentially shift between mutualism and parasitism, depending on the presence and abundance of both the nursery pollinator and of other pollinators. We investigated the fitness trade-off in a Mediterranean plant (Silene latifolia), which has a specialist nocturnal nursery pollinator moth (Hadena bicruris) and is also visited by several diurnal pollinators. We estimated the pollination rates and fecundity of S. latifolia in both natural and experimental populations in the Mediterranean. We estimated natural pollination rates in different flowering times and with presence/absence of the H. bicruis moth. Then by exposing plants to each pollinator group either during the day or at night, we quantified the contribution of other diurnal pollinators and the specialized nocturnal nursery pollinator to plant female fitness. We found no difference in plant fruit set mediated by diurnal versus nocturnal pollinators, indicating that non-specialist pollinators contribute to plant female fitness. However, in both natural and experimental populations, H. bicruris was the most efficient pollinator in terms of seeds produced per fruit. These results suggest that the female fitness costs generated by nursery pollination can be overcome through higher fertilization rates relative to predation rates, even in the presence of co-pollinators. Quantifying such interactions is important for our understanding of the selective pressures that promote highly specialized mutualisms, such as nursery pollination, in the Mediterranean region, a centre of diversification of the carnation family.

Plant species that are effective colonisers of transient habitats are expected to have a capacity for uniparental reproduction and show flexibility in pollination systems. Such traits may enable populations to be established from a small number of founding individuals without these populations succumbing to reductions in fecundity arising from pollinator limitation. We tested these predictions for Aloe thraskii (Xanthorrhoeaceae), a succulent treelet that colonises shifting coastal dunes and has both bird and bee pollinators. We performed hand-pollination experiments, and selectively excluded bird visitors to determine differences in pollinator effectiveness. We measured pollinator visitation rates and fecundity in populations varying in their size, density and isolation distance. Controlled hand-pollinations revealed that unlike most other Aloe species, A. thraskii is self-compatible and thus capable of uniparental reproduction. The species does however depend on pollinators and is visited by various bird species as well as by bees. Fruit and seed set are not affected by selective exclusion of birds, thus indicating that bees are effective pollinators. Bird visitation rates increased with increasing plant height and population size, while bee visitation rates increased with increasing population size and density. We found that seed set per flower was lower in large populations than in small populations. These results suggest that establishment of populations of A. thraskii from a small number of individuals is unlikely to be limited by the fecundity of individual plants.

Hermaphroditism is the most common se xual system in plants, occurring in approximately 72% of angiosperms ( de Jong and Klinkhamer, 2006 ). Despite the success of hermaphroditism in angiosperms, hermaphrodites still need to overcome compro-mises in both the quality and quantity of seed production owing to the close spatial proximity of male and female reproductive organs ( Barrett and Harder, 1996 ). As a result, hermaphroditic plants have evolved various forms of self-incompatibility (SI) and they often separate male and female reproductive function in space and time to reduce selfi ng and promote outcrossing ( de Nettancourt 1997 ; de Jong and Klinkhamer, 2006 ). Despite this, there is still potential for interference between sex functions. The most widely studied form of reproductive interference is pollen discounting (a form of female interference with male function) which occurs in both self-compatible and self-incompatible taxa ( Kohn and Barrett, 1994 ; Harder and Barrett, 1995 ; Eckert, 2000 ; Fishman, 2000 ). A much less well-studied phenomenon is ovule discounting, a form of male interference with female function where ovules penetrated by self-pollen tubes fail to develop, thus reducing opportunities for seed production ( Aizen and Harder, 2007 ). Ovule discounting can be diffi cult to quantify in the fi eld ( Aizen and Harder, 2007 ) and has often been considered to arise solely from early acting inbreeding depression ( Eckert et al., 2010 ). However, it could also arise from ovarian (late-acting) self-incompatibility in which ovules are penetrated by tubes from self-pollen, but then fail to develop ( Seavey and Bawa, 1986 ). Compared with pollen discounting, ovule discounting has sel-dom been quantifi ed in natural populations ( Waser and Price, 1991 ; V aughton and Ramsey, 2010 ). In plants prone to ovule dis-counting, experimental removal of self-pollen by emasculation should increase seed set ( Vaughton and Ramsey, 2010 ), but the outcomes of emasculation can be complex because it can nega-tively affect the rate of pollinator visitation if pollen is used as a reward ( Duffy and Johnson, 2011 ). In this study we consider whether experimental emascula-tion affects seed production in the perennial hermaphrodite, Kniphofi a linearifolia Baker (Xanthorrhoeaceae, formerly Aspho- delaceae). Studies of related species in this lineage have shown that ovules penetrated by self-pollen tubes uniformly fail to de-velop seeds, which suggests either late-acting self-incompatibility or strong early inbreeding depression ( Naaborgh and Willemse, 1991 ; Hargreaves et al., 2012 ; S. D. Johnson, unpublished data). As

Rewardless plants can attract pollinators by mimicking floral traits of rewarding heterospecific plants. This should result in the pollination success of floral mimics being dependent on the relative abundance of their models, as pollinator abundance and conditioning on model signals should be higher in the vicinity of the models. However, the attraction of pollinators to signals of the models may be partially innate, such that spatial isolation of mimics from model species may not strongly affect pollination success of mimics. We tested whether pollination rates and fruit set of the rewardless orchid Disa pulchra were influenced by proximity and abundance of its rewarding model species, Watsonia lepida. Pollination success of the orchid increased with proximity to the model species, while fruit set of the orchid increased with local abundance of the model species. Orchids that were experimentally translocated outside the model population experienced reduced pollinaria removal and increased pollinator-mediated self-pollination. These results confirm predictions that the pollination success of floral mimics should be dependent on the proximity and abundance of model taxa, and thus highlight the importance of ecological facilitation among species involved in mimicry systems.

Most orchid mycorrhizal fungi (OMF) are free-living decomposers that do not depend on orchids for establishment or survival. Despite their importance for completion of the orchid life cycle, the geographical distribution of OMF remains poorly understood. Here we review studies that examined the relationship between orchids and their mycorrhizal associates and place them in a biogeographical context. We first describe the diversity of OMF and then examine the patterns of OMF distributions according to geographical locations, biomes, and islands. OMF have been found across all major biogeographic realms and biomes. While the distribution of the major mycorrhizal fungal families that associate with orchids appears to be entirely cosmopolitan, we know little about how the distribution of particular OMF shapes the distribution of the orchids that rely on them. In the face of ongoing rapid anthropogenic influence on the natural environment, more information is needed to show how OMF communities vary across large geographical scales and how this affects the distribution of orchids. In particular, we need to identify the environmental variables (e.g., rainfall, geological substrate) that drive spatial variation in OMF communities. This information will be crucial for setting up future orchid conservation and restoration projects.

It is commonly assumed that the geographical distributions of plants are governed mainly by abiotic variables. However, interactions with other organisms, such as pollinators, also have the potential to influence plant distributions. To investigate this, we developed niche models for 32 plant taxa that have specialized pollination systems and which are native to a biodiversity hotspot (South Africa). We found that the distributions of these taxa are best explained by a combination of biotic (pollinators) and abiotic factors, rather than by abiotic factors alone. For approximately 66% of these plant taxa, pollinator distributions were the factor that provided the best predictor of their niches. Furthermore, co-occurrence of these plants and their pollinators was generally not explained solely by shared abiotic niches, and thus probably reflects broad-scale positive ecological interactions. These results are consistent with the proposal that pollinator distributions may constrain plant distributions and highlight the general potential for species distributions to be shaped by positive interactions with other species. This has important consequences for predicting how distributions of species might change in the face of loss of their key mutualists.

The pollen of hermaphrodite plants is often utilised by flower-visiting animals. While pollen production has obvious benefits for plant male fitness, its consequences for plant female fitness, especially in self-incompatible hermaphrodite species, are less certain. Pollen production could either enhance seed production though increased pollinator attraction, or reduce it if ovules are discounted by deposition of self pollen, as can occur in species with late-acting self-incompatibility. To test the effects of pollen reward provision on female fitness, we artificially emasculated flowers in two populations of the succulent Aloe maculata (Asphodelaceae), which has a late-acting self-incompatibility system, over the course of its flowering period. Flowers of this species are visited by sunbirds (for nectar) and native bees (for pollen and nectar). We measured floral visitation rates, floral rejection rates, pollen deposition on stigmas and fruit and seed set in both emasculated and non-emasculated plants. We found that flowers of emasculated plants suffered reduced visitation and increased rejection (arrival without visitation) by bees, but not by sunbirds; had fewer pollen grains deposited on stigmas and showed an overall decrease in fruit set and seed set. Rates of seed abortion were, however, greatly reduced in emasculated flowers. This study shows that pollen rewards can be important for seed set, even in self-incompatible plants, which have been assumed to rely on nectar rewards for pollinator attraction. Seed abortion was, however, increased by pollen production, a result that highlights the complexity of selection on pollen production in hermaphrodite flowers.

Genetic diversity can play a key role in the survival of endangered species. Both the number and size of populations of a species can affect genetic diversity and may be reduced by fragmentation of natural habitats. Here we examine the effects of a low number of populations and small population sizes on the population genetics of the nationally endangered and red-listed orchid Pseudorchis albida. To determine genetic variability within and among populations of P. albida, we employed AFLP and four plastid microsatellite (SSR) markers. In addition, we compared the SSR data from Irish P. albida to the closely related P. straminea. AFLP markers showed that P. albida has a high level of polymorphism and that significant differentiation occurred both within and among populations, with increasing population genetic isolation through distance. However, SSR mark ers revealed no variation, but did show that all Irish individuals had the same alleles as P. straminea rather than P. albida. Further work is required to assess the taxonomic status of P. albida/straminea in Western Europe. Irish populations of P. albida s.l. require immediate conservation attention due to their decreasing number and low numbers of individuals within populations.

Plastid microsatellite loci developed for Cephalanthera longifolia were used to examine the level of genetic variation within and between populations of the three widespread Cephalanthera species (C. damasonium, C. longifolia and C. rubra). The most detailed sampling was in C. longifolia (42 localities from Ireland to China; 147 individuals). Eight haplotypes were detected. One was detected in the vast majority of individuals and occurred from Ireland to Iran. Three others were only found in Europe (Ireland to Italy, England to Italy and Austria to Croatia). Two were only found in the Middle East and two only in Asia. In C. damasonium, 21 individuals from 10 populations (England to Turkey) were sampled. Only one haplotype was detected. In C. rubra, 34 individuals from eight populations (England to Turkey) were sampled. Although it was not possible to amplify all loci for all samples of this species, nine haplotypes were detected. Short alleles for the trnS-trnG region found in two populations of C. rubra were characterized by sequencing and were caused by deletions of 26 and 30 base pairs. At this level of sampling, it appears that C. rubra shows the greatest genetic variability. Cephalanthera longifolia, C. rubra and C. damasonium have previously been characterized as outbreeding, outbreeding with facultative vegetative reproduction and inbreeding, respectively. Patterns of genetic variation here are discussed in the light of these reproductive system differences. The primers used in these three species of Cephalanthera were also demonstrated to amplify these loci in another five species (C. austiniae, C. calcarata, C. epipactoides, C. falcata and C. yunnanensis). Although it is sometimes treated as a synonym of C. damasonium, the single sample of C. yunnanensis from China had a markedly different haplotype from that found in C. damasonium. All three loci were successfully amplified in two achlorophyllous, myco-heterotrophic species, C. austinae and C. calcarata. © 2010 The Linnean Society of London, Botanical Journal of the Linnean Society, 2010, 163, 181–193.

We demonstrate that invasive populations of Rhododendron ponticum in Ireland are genetically distinct from ancestral populations in Spain and produce flowers which have wider floral tubes. Although Irish populations are spreading and Spanish ones are declining, we found low genetic diversity among individual plants within populations in both regions, and limited between-population gene flow. Wider floral tubes may have evolved in response to novel pollinators in Ireland. Few studies examine invasive species in both their introduced and native habitats, but this approach is needed to understand invasive species evolution and ecology., Populations of introduced species in their new environments are expected to differ from native populations, due to processes such as genetic drift, founder effects and local adaptation, which can often result in rapid phenotypic change. Such processes can also lead to changes in the genetic structure of these populations. This study investigated the populations of Rhododendron ponticum in its introduced range in Ireland, where it is severely invasive, to determine both genetic and flower width diversity and differentiation. We compared six introduced Irish populations with two populations from R. ponticum's native range in Spain, using amplified fragment length polymorphism and simple sequence repeat genetic markers. We measured flower width, a trait that may affect pollinator visitation, from four Irish and four Spanish populations by measuring both the width at the corolla tip and tube base (nectar holder width). With both genetic markers, populations were differentiated between Ireland and Spain and from each other in both countries. However, populations displayed low genetic diversity (mean Nei's genetic diversity = 0.22), with the largest proportion (76–93 %) of genetic variation contained within, rather than between, populations. Although corolla width was highly variable between individuals within populations, tube width was significantly wider (>0.5 mm) in introduced, compared with native, populations. Our results show that the same species can have genetically distinct populations in both invasive and native regions, and that differences in floral width may occur, possibly in response to ecological sorting processes or local adaptation to pollinator communities.

Hermaphroditism can lead to both intra- and intersexual conflict between male and female gender functions. However, the effect that such gender conflicts have on pollination efficiency has seldom been investigated. By artificially reducing the number of available male gametes on an individual, we quantified whether male interference with pollination efficiency occurs in the self-compatible, moth-pollinated orchid Satyrium longicauda. We partially emasculated S. longicauda inflorescences and compared pollination success and fecundity in these plants to intact controls. Pollen in both groups of plants was colour-labelled so that its dispersal by pollinators could be tracked directly in the field. Intact flowers on partially emasculated inflorescences exported more pollen and received more cross-pollen and less self-pollen than those on intact inflorescences. Proportion of fruit set per plant was similar between the two treatments; however, fruits on partially emasculated plants had proportionally more viable seeds than those on intact controls. These results provide empirical evidence that male interference with pollination efficiency can occur in a hermaphroditic plant and that such interference can compromise fecundity. The most likely mechanism for such male interference is competition for placement on the proboscis of hawkmoth pollinators. Consequently, male competition for siring success may influence the evolution of sexual systems in hermaphroditic pollinator-dependent plants.

Increased anthropogenic influence is expected to reduce the size of natural populations, which may decrease individual fecundity and long-term survival. However, it is unclear whether populations of a species that occur in a variety of habitats will suffer decreased fecundity similarly. Certain habitats, such as those in agricultural landscapes that used to undergo traditional management, may be altered more than others and therefore individuals within such populations may be more prone to decreased fecundity. This is particularly important to quantify in Mediterranean landscapes, where habitat change is a major threat to biodiversity. We examined the fecundity of 88 populations of the orchid Spiranthes spiralis, across multiple habitats on the Greek island of Lesvos. In each population we recorded both the habitat and geological substrate. At the end of the flowering season, population size was quantified in each population by counting the number of fruiting plants and the percentage fruit set was determined. To test whether seed set was reduced due to pollination limitation we performed a pollen supplementation experiment. Large populations had significantly lower fruit set than small populations, and this effect was most pronounced in olive groves. Pollen supplementation showed that S. spiralis may suffer reduced seed set due to pollen limitation. The results of this study show that larger olive grove populations may be pollinator-limited and there may be two reasons for this: first, olive groves are less favourable sites for honeybees – the major and more efficient pollinators of S. spiralis on Lesvos; second, such populations have been suffering severe management changes in recent years (e.g. chemical treatments, chopping) and are most vulnerable to reductions in reproductive output.

Summary Factors underlying the Allee effect are still heavily debated in ecology. For plants that rely on pollinators for seed production, decreases in conspecific aggregation may reduce attractiveness to floral visitors and lead to an Allee effect. However, floral visitors often differ in their pollination effectiveness; hence, the likelihood of an Allee effect in plant fecundity may depend on how various flower visitors respond to plant aggregation. We tested for Allee effects on fecundity of individuals across two years in the self-incompatible perennial, Kniphofia linearifolia Baker (Xanthorrhoeaceae), which has two distinct types of pollinator, birds and native bees. For this, we used three measures of aggregation; population size, density and isolation. We made replicated pollinator observations in populations of various aggregations and quantified fecundity in these populations. To determine the differences in pollinator effectiveness and assess their contribution to fecundity, we selectively excluded bird visitors from K. linearifolia in these populations. We found that population size, but not density or isolation distance, was associated with increased bird abundance and seed set in one of the two years of the study. Bird visitation rate increased with increased plant aggregation within populations. Fruit set and seed set per flower were positively related to bird visitation rate. The difference in seed set per flower between bird-excluded and unmanipulated plants increased with increasing population size. Although birds were much less frequent visitors than bees (on average 2.1 visits plant−1 h−1 compared to 57.5 visits plant-1 h−1), selective exclusion experiments indicated that birds are consistently the more effective pollinators of this species, and therefore most likely to influence fecundity. Synthesis. In this system, characterised by an Allee effect on plant fecundity, birds were the most effective pollinators, responded positively to plant aggregation and were associated with increased fecundity. Therefore, the responses of effective pollinators to plant aggregation may be a factor that underlies Allee effects on plant fecundity.

Variation in within-population floral density can affect interactions between plants and pollinators, resulting in variable pollen export for plants. We investigated the effects of conspecific and heterospecific floral densities on pollination success both of two related, self-compatible, nectar-rewarding orchid species in Ireland, Spiranthesromanzoffiana (rare and listed as endangered) and its congener, S. spiralis (more abundant and not of conservation concern). Floral densities, insect visitation rates, and orchid pollen transport were recorded in multiple quadrats in four populations of both orchid species over their flowering season. We found that conspecific and heterospecific co-flowering plant density affected pollination in both orchid species. For S. romanzoffiana, higher heterospecific density increased pollen removal. For S. spiralis, higher conspecific visitation increased pollen removal and increased heterospecific density decreased pollen deposition. In addition, increased conspecific density increased pollen deposition in both species. This study shows that plants may interact to facilitate or compete for different components of the pollination process, namely; pollinator attraction, pollen removal and deposition. Such interactions have immediate consequences for endangered plant species, as increases in both conspecific and heterospecific coflowering density may ameliorate the negative effects of rarity on pollination, hence overall reproductive success.

Genetic diversity can play a key role in the survival of endangered species. Both the number and size of populations of a species can affect genetic diversity and may be reduced by fragmentation of natural habitats. Here we examine the effects of a low number of populations and small population sizes on the population genetics of the nationally endangered and red-listed orchid Pseudorchis albida. To determine genetic variability within and among populations of P. albida, we employed AFLP and four plastid microsatellite (SSR) markers. In addition, we compared the SSR data from Irish P. albida to the closely related P. straminea. AFLP markers showed that P. albida has a high level of polymorphism and that significant differentiation occurred both within and among populations, with increasing population genetic isolation through distance. However, SSR markers revealed no variation, but did show that all Irish individuals had the same alleles as P. straminea rather than P. albida. Further work is required to assess the taxonomic status of P. albida/straminea in Western Europe. Irish populations of P. albida s.l. require immediate conservation attention due to their decreasing number and low numbers of individuals within populations.

The probability and time of extinction of taxa is often inferred from statistical analyses of historical records. Many of these analyses require the exclusion of multiple records within a unit of time (i.e., a month or a year). Nevertheless, spatially explicit, temporally aggregated data may be useful for identifying clusters of sightings (i.e., sighting clusters) in space and time. Identification of sighting clusters highlights changes in the historical recording of endangered taxa. I used two methods to identify sighting clusters in historical records: the Ederer-Myers-Mantel (EMM) test and the space-time permutation scan (STPS). I applied these methods to the spatially explicit sighting records of three species of orchids that are listed as endangered in the Republic of Ireland under the Wildlife Act (1976): Cephalanthera longifolia, Hammarbya paludosa, and Pseudorchis albida. Results with the EMM test were strongly affected by the choice of the time interval, and thus the number of temporal samples, used to examine the records. For example, sightings of P. albida clustered when the records were partitioned into 20-year temporal samples, but not when they were partitioned into 22-year temporal samples. Because the statistical power of EMM was low, it will not be useful when data are sparse. Nevertheless, the STPS identified regions that contained sighting clusters because it uses a flexible scanning window (defined by cylinders of varying size that move over the study area and evaluate the likelihood of clustering) to detect them, and it identified regions with high and regions with low rates of orchid sightings. The STPS analyses can be used to detect sighting clusters of endangered species that may be related to regions of extirpation and may assist in the categorization of threat status.

Fragmentation of natural habitats can increase numbers of rare species. Conservation of rare species requires experts and resources, which may be lacking for many species. In the absence of regular surveys and expert knowledge, historical sighting records can provide data on the distribution of a species. Numerous models have been developed recently to make inferences regarding the threat status of a taxon on the basis of variation in trends of sightings over time. We applied 5 such models to national and regional (county) data on 3 red-listed orchid species (Cephalanthera longifolia, Hammarbya paludosa, and Pseudorchis albida) and 1 species that has recently come to the attention of conservation authorities (Neotinea maculata) in the Republic of Ireland. In addition, we used an optimal linear estimate to calculate the time of extinction for each species overall and within each county. To account for bias in recording effort over time, we used rarefaction analysis. On the basis of sighting records, we inferred that these species are not threatened with extinction and, although there have been declines, there is no clear geographical pattern of decline in any species. Most counties where these orchid species occurred had a low number of sightings; hence, we were cautious in our interpretation of output from statistical models. We suggest the main drivers of decline in these species in Ireland are modification of habitats for increased agricultural production and lack of appropriate management. Our results show that the application of probabilistic models can be used even when sighting data are scarce, provided multiple models are used simultaneously and rarefaction is used to account for bias in recording effort among species over time. These models could be used frequently when making an initial conservation assessment of species in a region, particularly if there is a relatively constant recording rate and some knowledge of the underlying recording process. Regional-scale analyses, such as ours, complement World Conservation Union criteria for assessment of the extinct category and are useful for highlighting areas of under recording and focusing conservation efforts of rare and endangered species.

Density can affect attraction of pollinators, with rare plants receiving fewer pollinating visits compared with more common co-flowering species. However, if a locally rare species is very attractive in terms of the rewards it offers pollinators, it may be preferentially visited. Spiranthes romanzoffiana is a nectar rewarding, geographically rare, endangered orchid species which forms small populations in Ireland, co-flowering with more common, florally rewarding species. We examined visitation rates to S. romanzoffiana and two nectar rewarding co-flowering species (Mentha aquatica and Prunella vulgaris) in the west of Ireland. These three plant species were visited by three bee species (Bombus pascuorum, B. hortorum and Apis mellifera). B. pascuorum was the most common visitor, while A. mellifera was least common. Our results suggest that individual S. romanzoffiana inflorescences compete intraspecifically for visitation from pollinators at high densities. The relationship between visitation to S. romanzoffiana and total floral density appeared to be positive, suggesting interspecific facilitation of pollinator visitation at high densities. Nectar standing crop varied through the season, among species and between open and bagged flowers. Nectar standing crop was not correlated with visitation in S. romanzoffiana. Despite relatively high visitation, S. romanzoffiana produced no mature fruit during this flowering season. The lack of fruit maturation in this species may be a major factor causing its rarity in Europe.