Effects of habitat fragmentation on the reproductive success of the nectar-producing orchid Gymnadenia conopsea and the nectarless Orchis mascula

Habitat fragmentation decreases plant population sizes and increases spatial isolation, which hampers the exchange of seeds and pollen between fragmented populations. This may result in decreased population viability. We compared the effects of population size and isolation on the reproductive success of two orchid species, Gymnadenia conopsea (nectar-producing) and Orchis mascula (nectarless) growing in highly fragmented calcareous grassland in southern Belgium. We expected that the nectar-producing species would be more susceptible to the negative reproductive consequences of habitat fragmentation compared to the nectarless species. Nectar production has been associated with increased geitonogamous pollination and, therefore, with lower seed viability. Our results show that seed viability increased with increasing population size in O. mascula, whereas it was always low in G. conopsea, even in large populations. In contrast, percentage fruit set was positively related to population size in the nectar-producing G. conopsea, but no such effect was observed in the nectarless O. mascula, where fruit set was low even in large populations. Population isolation was not related to reproductive success for either species. Our results suggest that even in large populations, where pollinators are expected to be more abundant, increased geitonogamous pollination reduces seed viability in the nectar-producing G. conopsea. In contrast, seed viability in O. mascula seems to benefit from increased pollinator availability in larger populations. For the latter species, however, fruit set remains low, even in large populations, compared to G. conopsea. This may be explained by the relatively low attractiveness of nectarless orchid species for pollinators. Our results indicate that small population size may negatively influence reproductive success in both nectarless and nectar-producing orchids by reducing seed viability and fruit set, respectively.     

Floral isolation is the major reproductive barrier between a pair of rewarding orchid sister species

The crucial role of reproductive isolation in speciation has long been recognized; however, a limited number of studies quantify different isolation barriers and embed reproductive isolation in a phylogenetic context. In this study, we investigate reproductive isolation between the often sympatrically occurring orchid species, Gymnadenia conopsea and G. odoratissima. We examine the phylogenetic relationship between the two species and analyse floral isolation, fruit set and seed viability from interspecies crosses, as well as the ploidy level. Additionally, we quantify interspecies differences in floral signals and morphology. The results suggest that the two species have a sister–species relationship. In terms of reproductive isolation, we found complete floral isolation between the two species, but little to no post‐pollination isolation; the species also mostly had the same ploidy level in the studied populations. We also show clear distinctions in floral signals, as well as in floral size and spur length. We propose that respective adaptation to short‐ vs. long‐tongued pollinators was the driver of speciation in the here studied Gymnadenia species. Our study supports the key role of floral isolation in orchid speciation and shows that floral isolation is not restricted to highly specialized pollination systems, but can also occur between species with less specialized pollination.     

Reproductive system and fitness of Vriesea friburgensis,a self‐sterile bromeliad species

Reproductive biology and plant fertility are directly related to many aspects of plant evolution and conservation biology. Vriesea friburgensis is an epiphytic and terrestrial bromeliad endemic to the Brazilian Atlantic rainforest. Hand‐pollination experiments were used to examine the reproductive system in a wild population of V. friburgensis. Plant fertility was assigned considering flower production, fruit and seed set, seed germination, and pollen viability. Self‐sterility observed from spontaneous selfing and manual self‐pollination treatments may be the consequence of late‐acting self‐incompatibility. Hand‐pollination results indicated no pollen limitation in the population studied. Floral biology features such as a few daily open flowers, nectar production, and sugar concentration corroborate hummingbirds as effective pollinators, although bees were also documented as pollinators. Components of fitness such as high flower, fruit, and seed production together with high seed and pollen viability indicate that this wild population is viable. From a conservation point of view, we highlight that this self‐sterile species depends on pollinator services to maintain its population fitness and viability through cross‐pollination. Currently, pollinators are not limited in this population of V. friburgensis. Conversely, the maintenance and continuous conservation of this community is essential for preserving this plant–pollinator mutualism.     

Autonomous self‐pollination in the nectarless orchid Pogonia minor

The pollination biology of the nectarless orchid Pogonia minor was investigated in central Japan. The investigation revealed that the solitary flowers failed to attract pollinators, while high rates of fruit set were observed in the natural population. Comparable levels of fruit set were obtained in bagged, artificial self‐pollinated and artificial cross‐pollinated plants, indicating that the species is not pollinator‐limited for fruit set under natural conditions. Autonomous self‐pollination in P. minor resulted from a reduced rostellum, which allowed contact between the pollinia and the stigma. Self‐pollination is thought to be an adaptive response that provides reproductive assurance under conditions of pollinator limitation. Since pollen limitation is generally known to be frequent among deceptive orchids, strong pollen limitation is probably a driving force in the autonomous self‐pollination mechanism in the nectarless orchid P. minor.     

Low legitimate pollen flow in distylic Turnera hermannioides (Passifloraceae) and its consequences on fruit and seed set

Turnera hermannioides is a ruderal distylic subshrub, native to NE-Brazil. In the Catimbau National Park, situated within the semi-arid Caatinga region, we studied the pollination ecology of this species, emphasizing (1) effective pollinators; (2) characteristics of short- and long-styled flowers; (3) intra- and intermorph pollen flow; and (4) fruit and seed set. Short and long-styled morphs differ in pollen size and ornamentation, stigmatic surface, style and stamen length and nectar production. The flowers are obligate intermorphic outcrossers and depend on animals for pollination. The flowers of T. hermannioides attracted insect visitors of 25 species, among them butterflies, beetles, but mainly bees. Polylectic bees, such as Apis mellifera, stingless bees, and solitary Callonychium brasiliense were the most frequent visitors and the principal pollinators. The frequency of visits, however, was very low (on average 1.9 visits/flower/day), resulting in a low and unbalanced legitimate pollen flow, i.e. a much lower number of pollen grains from short-styled flowers reaching stigmas of long-styles than vice versa, which in turn compromised fruit and seed set. Surprisingly, Protomeliturga catimbaui and P. turnerae, both oligolectic on flowers of the Turneraceae-clade of the Passifloraceae s.l., were very rare flower visitors and did not contribute significantly to the pollination of Turnera hermannioides. We discuss how the low visitation rate and the inefficiency of pollinators (i.e. inefficiency in promoting legitimate pollination) compromise pollen flow and, consequently, fruit and seed set in this species.     

Self-incompatibility, Inbreeding Depression and Crossing Potential in Five BrazilianPleurothallis (Orchidaceae) Species

Intra- and interspecific experimental pollinations were madeto determine the mating systems and the interspecific crossingpotential in individuals from 24 populations of five Pleurothallisspecies. Pleurothallis johannensis, P. ochreata and P. teresshow weak or partial self-incompatibility while P. adamantinensisand P.fabiobarrosii are more strictly self-incompatible. Wefound no differences in fruit set between intra- and interpopulationintraspecific crosses, and there was no correlation betweenfruit set and genetic variability or inbreeding in these species.All species are interfertile and showed no correlation betweengenetic similarity and crossing potential. We scored fruitsfor seed viability and observed a strong inbreeding depressionin all populations; however, there was no difference in seedviability among intrapopulation, interpopulation intraspecificand interspecific cross-pollinations. These species are pollinatedby flies with a behavioural pattern that facilitates self-pollination.Self-incompatibility and inbreeding depression are apparentlyimportant in the maintenance of the unusually high levels ofgenetic variability found in these species. As generally observedin other orchid species, barriers to hybridization between thesePleurothallis species are at the level of pollination. Copyright2001 Annals of Botany Company Pleurothallis, Orchidaceae, mating system, self-incompatibility, inbreeding depression, fruit set, seed set, polyembryony, interspecific compatibility     

Abnormalities in sexual development and pollinator limitation in Michelia coriacea (Magnoliaceae), a critically endangered endemic to Southeast Yunnan,China

Michelia coriacea (Magnoliacae) is a critically endangered tree, endemic to Southeast Yunnan province, China. Most of the individuals in the extant populations normally bear flowers, but fruit set and fertile seed production rates were only 6.7% and 0.2%, respectively. To identify possible causes of reproductive barriers, the following studies were carried out: examination of stamen and pistil development; experiments using controlled pollination; observations of behavior and frequency of floral visitors in both cultivated and natural populations. The results revealed that about 60% of pollen was abnormal and approx. 70% of ovules had delayed development. Hand pollination could effectively enhance the fruit set (F_4,25=35.139, P<0.0001) and seed set per fruit (F_4,25=85.022, P<0.0001). Both cultivated and wild M. coriacea had an extremely low frequency of floral visitors. Some beetles, a few species of Andrenidae and some Bombus sp. are likely to be the effective pollinators. The fruit set and seed set per fruit from controlled self-pollination and cross-pollination were significantly different (P<0.05), and thus it is inferred, that inbreeding depression may be a contributing factor in the very low seed production. It appears that low seed set in M. coriacea is due to a combination of factors: abnormalities in pollen and ovules, low number of effective pollinators, and inbreeding depression.     

Mast fruiting in a hawkmoth-pollinated orchid Habenaria glaucifolia: an 8-year survey

Aims The hypothesis of predator satiation has been proposed to explain mast fruiting in various flowering plants. It considers that the simultaneous production of large numbers of seeds by a plant population reduces the risk of seed predation for each individual. Orchids produce huge numbers of seeds per fruit and rarely experience seed predation. It remains unclear which factors may affect fluctuating fruit production in orchids, which generally suffer a widespread pollen limitation. To explore the temporal pattern of fruiting and potential factors related to fluctuation in fruit production, we investigated reproductive success of a long-spurred orchid (Habenaria glaucifolia) in an alpine meadow with thousands of individuals over 8 years.Methods To estimate reproductive success, pollinator observation was conducted by day and at night, and pollinia removal and receipt were recorded in the field population for 8 years. To examine whether fruit set and seed set are pollen limited, we conducted supplementary pollination experiments and compared fruit set, seed set and pollinia movement of open-pollinated flowers from 2011 to 2013. We measured lengths of spurs and pollinator proboscises, and nectar volume and concentration, to identify potential pollinators.Important findings Hawkmoths were seen to be effective pollinators for H. glaucifolia in 3 years, whereas in the remaining 5 years no pollinators were observed, and consequently pollinia were rarely transferred. Numerous pollinia movements were observed in 2012, 2013 and 2014 (pollinia removal: 48, 59 and 85%; pollinia receipt 51, 70 and 80%), and correspondingly fruit set was significantly higher in 2012 and 2013 (59 and 46%) than in 2011 (25%). It was fruit set, rather than seed set, that was pollen limited in this orchid in the 3 years, in that supplementary pollination increased fruit set but did not increase seed set per fruit compared to natural. Three species of hawkmoths had proboscis lengths that matched the spur length of H. glaucifolia. Fruit set in this long spurred orchid depends on the activity of long-tongued hawkmoths, resulting in significant temporal variation in fruit production. Mast fruiting in this alpine orchid could be attributed to a 'sit and wait' strategy, awaiting an abundance of effective pollinators.     

Pollination biology and breeding systems in the terrestrial orchid Bletilla striata

The levels of fruit set in different habitats (three inland populations vs. three coastal populations) and the breeding system in the terrestrial orchid Bletilla striata were examined in two groups of populations in Haenam-gun, Jollanam-do Province in southern Korea. In total inland populations (mean fruit set=13.3%) showed about eight-fold higher fruit set per plant than the coastal populations (1.6%), probably due to mild climatic conditions in the inland populations during the flowering period. Honey bee workers Apis mellifera were observed in the inland populations, but no visitors were found in the coastal populations. However, a great increase in fruit set was achieved by hand-pollination: the percentage of fruit set through artificial self-pollination and geitonogamous pollination were close to 90%, indicating that B. striata is self-compatible. Agamospermy and spontaneous autogamy (automatic self-pollination) were not detected in this study. These indicate that pollinia vectors are essential for achieving fruit set in natural populations of B. striata. Except in one inland population examined in 2001, there was no significant correlation between the patch size per population and the average percentage of fruit set, which might reflect deception-pollination of B. striata and/or paucity of pollinators. Overall levels of fruit sets within and among patches in each population were homogeneous during a two-year period.     

Effects of distance from models on the fitness of floral mimics

• 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.

     

Does floral trait variability enhance reproductive success in deceptive orchids?

Unusually high intra-specific floral trait variability has often been described within deceptive orchid populations, as opposed to rewarding ones. Such variability is traditionally thought to have consequences on reproduction in this orchid group, i.e. phenotypically variable deceptive species may have a reproductive success advantage compared to those with a constant floral display. The proposed reason for this hypothetic pattern is that floral trait variability decreases pollinator avoidance learning in dealing with nectarless flowers, hence increasing their visitation rate. However, despite an intuitive and appealing hypothesis and a possible mechanism to explain it, the often-cited higher reproductive success induced by floral trait variability still remains unsupported.Here, we review the literature and consider eight studies that have experimentally or correlatively tested this hypothesis in deceptive orchids. In all these experiments, we have found no difference in average reproductive success between populations with high versus low flower trait variability, either in scent variable or colour polymorphic species. We discuss possible explanations for the lack of this pattern including the incapability of pollinators in perceiving the variability, the scarce relevance of polymorphic traits in the choice of species to forage on, or a different pollinator behaviour than the one proposed. We suggest that the high phenotypic variability is not likely to enhance deceptive orchids’ reproductive success, but is more likely to be a consequence of relaxed selection by pollinators. Nonetheless, information regarding orchid pollination strategy or pollinator cognitive abilities is often superficial, hence calling researchers for additional investigations that can contribute to a better understanding of this debated and yet unsupported hypothesis.     

Experimental evidence for specialized bird pollination in the endangered South African orchid Satyrium rhodanthum and analysis of associated floral traits

The bird pollination syndrome is characterized by red, unscented flowers with dilute nectar in long nectar tubes. However, the extent to which plants with such traits actually depend on birds for seed production is seldom determined experimentally, and traits such as colour and scent production are often assessed only subjectively. We documented bird pollination and quantified floral traits in the critically endangered Satyrium rhodanthum (Orchidaceae) from mistbelt grasslands in the summer‐rainfall region of South Africa. Direct observations and motion trigger camera footage revealed amethyst sunbirds as the only pollinators, despite the presence of other potential pollinators. Experimental exclusion of sunbirds significantly reduced pollination and fruit set to near zero. Pollination success in naturally pollinated plants was close to 100% in one year, and fruit set varied from 23 to 64% in other years. Pollen transfer efficiency was 5.8%, which is lower than in related insect‐pollinated species, probably due to a tendency of birds to wipe pollinaria from their beak. Flowers of S. rhodanthum only reflect light in the red range of the spectrum, and they produce only a few aliphatic and monoterpene scent compounds at comparatively low emission rates. Nectar volume and sugar concentration varied between 2.7 and 3.7 μL and 23.7 and 25.9%, respectively. We conclude that S. rhodanthum is highly specialized for pollination by sunbirds. Colour, scent and nectar characteristics differ from insect‐pollinated Satyrium species and are consistent with those expected for bird‐pollinated flowers, and may contribute to lack of visitation by other potential long‐tongued pollinators. Habitat loss probably underlies the critically endangered conservation status of S. rhodanthum, but the specialization for pollination by a single bird species means that reproduction in this orchid is vulnerable to losses in surrounding communities of plants that subsidize the energetic requirements of sunbirds. © 2014 The Linnean Society of London, Botanical Journal of the Linnean Society, 2014, 177 , 141–150.     

Pollination and seed set of an obligatory outcrossing plant in an urban–peri-urban gradient

Urban land use strongly affects pollinators and plant pollination due to fragmentation and isolation of natural plant populations. On the other hand, urban land use can potentially be beneficial to pollinators through the presence of urban gardens and parks which are usually designed to ensure abundant flowering. However, little is known about the effects of urban land use on bees and the pollination services they provide. In an urban–peri-urban gradient around Leuven, Belgium we studied flower visitation rates to, and seed set of Trifolium repens (white clover) in public lawns. T. repens is an obligatory outcrossing plant and is therefore sensitive to reductions in pollinator services. We related our results to both local and regional variables using Boosted Regression Trees. The two variables that best explained the variability in visitation rates and seed set were the amount of green areas (gardens, parks, grasslands) in the surroundings and the abundance of T. repens in the lawns. Surprisingly, an increasing amount of green areas in the surroundings had a negative effect on both flower visitation rates to, and seed set of T. repens. Flower visitation rates by bumblebees responded positively to urban land use resulting in higher visitation rates and increased seed set in the more urban sites. This may point either to increased abundance of bumblebees in more urban sites or to a concentration effect of bumblebees in our urban study sites due to a lack of alternative forage resources. Responses will likely differ for other bee and plant species, but this shows that at least for T. repens, pollination is not compromised by urban land use.     

The birds and the seeds: opportunistic avian nectarivores enhance reproduction in an endemic montane aloe

Opportunistic bird pollination has become more evident in studies that confirm distinct differences in floral adaptations that attract opportunistic, rather than specialist, bird pollinators. Pollination syndrome studies investigating the effectiveness of different pollinator guilds on reproduction seldom do so by measuring seed viability. We studied pollination in Aloe peglerae, a high altitude endemic succulent of the Magaliesberg mountain range, previously thought to be largely sunbird (specialist) pollinated. Using field observation and pollinator exclusion treatments, i.e. (i) open to all visitors, (ii) bird excluded, and (iii) all visitors excluded, we established that birds contributed significantly more to fruit (2.3–5.6 times) and seed (1.3–1.4 times) set than insect or self-pollination, respectively. Overall, pollination by opportunistic avian nectarivores amplified seed production per aloe ~7 and 10 times compared to insects and self-pollination, respectively. One of three opportunistic nectarivores, the Cape Rock-Thrush (Monticola rupestris), played a significant role in pollination, contributing ~60 % of all probes at inflorescences. The difference in reproductive output between insect visited and visitor excluded flowers was not significant and suggests possible self-pollination in A. peglerae which is particularly unusual in Aloe species. Breeding system experiments would help clarify this. In assessing the effectiveness of pollinator guild on seed viability, we found no differences in percentage seed viability, seed germination or seedling emergence between exclusion treatments. Seed viability and germination were low and variable; however, ~19 % seedling emergence was observed across the treatments. Practically, the net effect of bird pollination may result in 8–12 times more potential seedlings compared to insect and self-pollination respectively. These findings highlight the importance of pollination by opportunistic avian nectarivores in Aloe.     

Autogamy in Boronia falcifolia: consequences for inbreeding and persistence in fragmented coastal heaths

Coastal heath is becoming increasingly fragmented, resulting in reduced pollinator abundance affecting species viability. The reproductive ecology and inbreeding of a common coastal heath species, Boronia falcifolia (Rutaceae), was investigated. Pollination observations and experiments were carried out at one site, and inbreeding was assessed using genetic markers at eight populations. Four pollination treatments were assessed: cross-pollination, self-pollination, autogamous pollination and open (insect) pollination. Boronia falcifolia was found to be highly self-compatible, frequently setting seeds from autogamous pollination. There were no significant differences between the autogamous, self and cross pollen treatments for any measures of reproductive success (pollen tube growth, fruit set, swollen locules and seed production). Insect activity resulted in fewer seeds per fruit than hand cross pollination, although pollinator visits were few. All populations studied were highly inbred. Autogamous self-pollination is widespread in B. falcifolia and this combined with its ability to vegetatively regenerate enhances its potential for survival when pollinators are few.     

What We Think We Know vs. What We Need to Know About Orchid Pollination and Conservation: <Emphasis Type="Italic">Cypripedium</Emphasis> L. as a Model Lineage

While Darwin (1862, 1877) showed that reproductive success in orchid populations depended on adaptive floral morphology coupled with pollinator visitation a more recent review of the literature (Tremblay et al., 2005) confirmed that many out-breeding species are pollinator-limited because most orchid species showing low fecundity also lack rewards. The absence of rewards depresses both pollinator fidelity and the frequency of pollinator visits to an orchid population even though orchid flowers that lack rewards retain the same interlocking floral structures for precise pollinia removal and deposition found in related species that offer rewards. Using the genus, Cypripedium, as a model lineage of non-rewarding flowers this study also shows that the correlation between low fruit set in a Cypripedium sp. and its specific pollinator(s) is insufficient to predict specific frequencies of low fecundity. Annual rates of fruit set often vary broadly between populations of the same species and within the same population over several seasons. We speculate that fruit-set rates also decline when orchid demography and additional biotic and abiotic factors interrupt rates of pollinator activity (pre-zygotic) and fertilization/fruit maturation (post-zygotic). We suggest that that traditional field studies on pollination ecology and breeding systems be combined with data sets recording genetic variation and orchid flower demography in relation to seasonal variation in climate. We also propose that the same information be collected in regard to genetic variation, demography and phenology of populations of known orchid pollinators and co-blooming angiosperm species native to orchid habitats.     

Who helps whom? Pollination strategy of <Emphasis Type="Italic">Iris tuberosa</Emphasis> and its relationship with a sexually deceptive orchid

Reproductive success of plants may be affected by interactions with co-flowering species either negatively, through competition for pollinators, or positively, by means of a magnet species effect and floral mimicry. In this study, potential interactions between Iris tuberosa, a rewarding species, and Ophrys fusca, a sexually deceptive orchid, were explored in four populations in southern Italy. In each population plots showing different ratios of the examined species were arranged in the field, and in each plot the number of pollinators and fruit set were assessed. In addition, flower size and floral hydrocarbons produced by the two species were analysed. Morphological and scent data pointed out that flower size and aliphatic compounds did not differ significantly between the two species. Interestingly, both species shared tricosane and 11-nonacosene, electrophysiologically active compounds in the shared dominant pollinator Adrena. We have found that fruit production and number of pollinators in I. tuberosa varied significantly among plots, while percentage of capsules and number of pollinators of O. fusca captured showed no significant differences across plots. These results suggested, that the presence of O. fusca contributes differentially to pollinator attraction, and thus, to total reproductive success of I. tuberosa, according to a different ratio of aggregation. These findings suggest that I. tuberosa profits from the greater abundance of insects attracted by the presence of orchid specimens, and that a sexually deceptive orchid may be a magnet species in pollination strategy.     

Pollination of invasive Eichhornia crassipes (Pontederiaceae) by the introduced honeybee (Apis mellifera L.) in South China

Heterostyly functions as an outcrossing mechanism facilitating accurate pollen transfer from anthers to stigmas of particular heights as a result of the behavior of specialist pollinators. However, heterostylous plants are also visited by generalist pollinators, which may affect the plant–pollinator mutualism. Eichhornia crassipes is a tristylous invasive species, with only the mid- and long-styled morphs (M and L) found in China. We recorded flower-visiting insects in Zhuhai, Zhongshan and Nanning in South China. We hand-pollinated the two morphs to determine their compatibility. In addition, by allowing controlled insect pollination in artificial isoplethically monomorphic and bimorphic populations, we undertook a detailed analysis of pollen deposition between the floral morphs, and fruit and seed set. Ranked by relative abundance, the flower-visiting insects were: Apis mellifera, A. cerana, Lasioglossum sp. and Eristalis arvorum. Hand pollination showed that both the M and L morphs were self-compatible, but the former was probably more so than the latter. Intra-morph pollen transfer by A. mellifera within a population was significantly greater than legitimate pollen transfer between populations, suggesting that the pollen exchange between populations was limited. Seed set of the L morph was significantly greater than that of the M morph in monomorphic populations, indicating intra-morph pollen deposition in the former was higher than in the latter. The results showed that A. mellifera was the major pollinator in South China and able to pollinate E. crassipes legitimately and to promote its fruit and seed set, even though high levels of intra-morph pollination occurred.     

Reduced fecundity in large populations of a Mediterranean orchid – Evidence for pollinator limitation

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.     

Floral morph composition and pollen limitation in the seed set of <Emphasis Type="Italic">Nymphoides indica</Emphasis> populations

We investigated the floral morph composition and seed set of the populations in a heterostylous aquatic plant, Nymphoides indica O. Kuntze (Menyanthaceae). The bias of floral morph ratio in the populations was negatively correlated with the seed set. In the populations with biased floral morph ratio, the pollen on the stigma of the common morphs were dominated by pollen of the same morph, resulting in reduced seed set. The addition of pollen of the opposite morph on the stigmas of the common morph resulted in an increase of fruit and seed set rates. The intermorph pollination rate and the fruit and seed set rates of Nymphoides indica were highest when two floral morphs were close in the populations. Various insect species including Lepidoptera, Diptera and Hymenoptera visited the flowers. However, geitonogamous pollination was common because of the short flying distance of the pollinators. These results indicate that the seed set of Nymphoides indica is limited by the shortage of compatible pollens and that the success of intermorph pollination is influenced by spatial distribution of the two floral morphs. As Nymphoides indica is endangered, conservation of the populations with high fecundity is proposed by us.