Phenotypic selection patterns in a hybrid zone between two Calceolaria species with contrasting pollinators: insights from field surveys and fitness assessments.

Hybrid zones provide natural experimental settings to test hypotheses about species divergence. We concentrated on a hybrid swarm in which oil-collecting bees and flower-pecking birds act as pollinators of two Calceolaria species. We asked whether both pollinators contributed to flower divergence by differentially promoting prezygotic fitness at the phenotypic extremes that represent parentals. We studied pollinator-mediated selection on phenotypic traits critical in plant-pollinator mechanical interaction, namely plant height, reward-to-stigma distance, and flower shape. We utilised the quantity and quality of pollen deposited as fitness measures and distinguished between the contribution of the two pollinator types. Results showed uni- and bivariate disruptive selection for most traits through pollen grains deposited by both pollinators. Bird-mediated fitness favoured low plants with a long reward-to-stigma distance and a straight corolla, while bee-mediated fitness favoured tall plants with a short reward-to-stigma distance and curved corolla. In addition, stabilising selection at one end of the phenotypic range showed a bird-mediated reproductive asymmetry within the swarm. The disruptive pattern was countered, albeit weakly, by hybrids receiving higher-quality pollen on the stigmas. Results suggest that pollinator-mediated selection promotes divergence of integrated flower phenotypes mechanically adjusted either to bees or birds underscoring the importance of pollinator specialisation in diversification.

Honeybee (Apis mellifera L.) pollination enhances the yield and flavor quality of kiwifruit.

Pollination is essential for achieving high yields and enhancing the quality of kiwifruit cultivation, both of which significantly influence growers' interests and consumers' preferences. However, compared to studies on yield, there are fewer studies exploring the impact of pollination methods on the flavor of kiwifruit Actinidia chinensis Planchon. This study examined the effects of bee (Apis mellifera L.) pollination and artificial pollination on the yield and flavor of kiwifruit in the main producing areas of China. Compared with those pollinated artificially, bee-pollinated kiwifruit exhibited a greater fruit set rate, heavier fruit weight, and greater number of seeds. Notably, the number of seeds was positively correlated with fruit weight in bee-pollinated kiwifruit, whereas no such correlation was detected in artificially pollinated fruit. Bee pollination not only enhanced the yield but also improved the flavor of kiwifruit. Specifically, bee-pollinated kiwifruit contained higher levels of sucrose and lower concentrations of glucose and fructose, while the acid content was less affected by pollination methods. Furthermore, significant differences were observed in the volatile organic compound (VOC) levels in kiwifruit subjected to different pollination treatments, with bee-pollinated fruit exhibiting a superior flavor. Our findings provide new insights into the beneficial role of bee pollination in enhancing kiwifruit yield and quality, underscoring the crucial importance of bees in kiwifruit pollination.

Direct tracking of pollen with quantum dots reveals surprising uniformity in dispersal distance across 11 populations of an annual plant.

PREMISE: Pollen movement is a crucial component of dispersal in seed plants. Although pollen dispersal is well studied, methodological constraints have made it challenging to directly track pollen flow within multiple populations across landscapes. We labeled pollen with quantum dots, a new technique that overcomes past limitations, to evaluate the spatial scale of pollen dispersal and its relationship with conspecific density within 11 populations of Clarkia xantiana subsp. xantiana, a bee-pollinated annual plant. METHODS: We used experimental arrays in two years to track pollen movement across distances of 5-35 m within nine populations and across distances of 10-70 m within two additional populations. We tested for distance decay of pollen dispersal, whether conspecific density modulated dispersal distance, and whether dispersal kernels varied among populations across an environmentally complex landscape. RESULTS: Labeled pollen receipt did not decline with distance over 35 m within eight of nine populations or over 70 m within either of two populations. Pollen receipt increased with conspecific density. Overall, dispersal kernels were consistent across populations. CONCLUSIONS: The surprising uniformity in dispersal distance within different populations was likely influenced by low precipitation and plant density in our study years. This suggests that spatiotemporal variation in the abiotic environment substantially influences the extent of gene flow within and among populations.

Hawkmoth and bee pollinators impact pollen dispersal at the landscape but not local scales in two species of Oenothera.

PREMISE: Animal pollinators play an important role in pollen dispersal. Here, we assessed differences in pollen and seed dispersal and the role of pollinator functional groups with different foraging behaviors in generating patterns of genetic diversity over similar geographic ranges for two closely related taxa. We focused on two members of Oenothera section Calylophus (Onagraceae) that co-occur on gypsum outcrops throughout the northern Chihuahuan Desert but differ in floral phenotype and primary pollinator: Oenothera gayleana (bee) and O. hartwegii subsp. filifolia (hawkmoth). METHODS: We measured breeding system and floral traits and studied gene flow and population differentiation at the local (<13 km; four populations) and landscape (60-440 km; five populations) scales using 10-11 nuclear (pollen dispersal) and three plastid (seed dispersal) microsatellite markers. RESULTS: Both taxa were self-incompatible and floral traits were consistent with expectations for different pollinators. Seed and pollen dispersal patterns were distinctly different for both species. We found no evidence of genetic structure at the local scale but did at the landscape scale; O. gayleana showed greater differentiation and significant isolation by distance than in O. hartwegii subsp. filifolia. The plastid data were consistent with gravity dispersal of seeds and suggest that pollen dispersal is the principal driver of genetic structure in both species. CONCLUSIONS: We demonstrated that pollinator functional groups can impact genetic differentiation in different and predictable ways. Hawkmoths, with larger foraging distances, can maintain gene flow across greater spatial scales than bees.

Osmophores and petal surface traits in Bignonieae species.

This study characterizes the osmophores and corolla traits in 18 species of Bignonieae Dumort., a Bignoniaceae tribe occurring in the Cerrado, a neotropical savanna in Brazil. To detect osmophore distribution, whole, newly opened flowers were immersed in Neutral Red Solution. Samples from the corolla tube and lobes were also fixed and analyzed micromorphologically, anatomically, and histochemically. The osmophores showed six markedly different distribution patterns that were not clearly associated with histological features. In most species, osmophores comprised papillose secretory epidermises and a few layers of subepidermal parenchyma. Starch grains, lipid droplets, and terpenes were detected in osmophores. An ornamented cuticle, cuticular folds, glandular and non-glandular trichomes, raised stomata and epicuticular wax granules are common traits in the species studied and may be useful in determining the taxonomy of the group. We found that 94% of the species visited by bees had papillose epidermises while the single hummingbird-pollinated species presented a flattened epidermis. Variations in osmophore pattern among species visited by bees, including variations within the same plant genus, are novel finding. Additionally, the Bignonieae species visited by bees presented a textured corolla surface, which has been reported as facilitating bee attachment and movement towards the floral resource. Future studies with a greater number of Bignonieae species and more detailed pollinator behavioral assays may help in the interpretation of the variations in corolla traits and functional relationships between flowers and pollinators.

Evolutionary lability in floral ontogeny affects pollination biology in Trimezieae.

PREMISE: There is little direct evidence linking floral development and pollination biology in plants. We characterize both aspects in plain and ornamented flowers of Trimezieae (Iridaceae) to investigate how changes in floral ontogeny may affect their interactions with pollinators through time. METHODS: We examined floral ontogeny in 11 species and documented pollination biology in five species displaying a wide range of floral morphologies. We coded and reconstructed ancestral states of flower types over the tribal phylogeny to estimate the frequency of transition between different floral types. RESULTS: All Trimezieae flowers are similar in early floral development, but ornamented flowers have additional ontogenetic steps compared with plain flowers, indicating heterochrony. Ornamented flowers have a hinge pollination mechanism (newly described here) and attract more pollinator guilds, while plain flowers offer less variety of resources for a shorter time. Although the ornamented condition is plesiomorphic in this clade, shifts to plain flowers have occurred frequently and abruptly during the past 5 million years, with some subsequent reversals. CONCLUSIONS: Heterochrony has resulted in labile morphological changes during flower evolution in Trimezieae. Counterintuitively, species with plain flowers, which are endemic to the campo rupestre, are derived within the tribe and show a higher specialization than the ornamented species, with the former being visited by pollen-collecting bees only.

Differential reward in "male" versus "female" pollen of functionally dioecious Solanum.

PREMISE: Five to six percent of angiosperm species exhibit a dioecious sexual system, with unisexual "male" or "female" flowers borne on separate plants. The consequent need for inter-individual pollen exchange is a special challenge for taxa where pollen is the sole pollinator reward. Dioecious Australian Solanum assure visits from pollen-foraging bees via production of inaperturate pollen in functionally female (morphologically bisexual) flowers. Biochemical composition of pollen from Australian Solanum has not been assessed nor compared to porate pollen from staminate flowers to reveal whether these flowers differ in their pollinator reward potential. METHODS: Porate pollen from male flowers and inaperturate pollen from functionally female flowers of two functionally dioecious Australian species were compared for protein and amino acid content. We also assessed pollen from bisexual and staminate flowers of a closely related andromonoecious species, in which all pollen is porate, as a comparison across co-occurring sexual systems. RESULTS: In both functionally dioecious species, porate pollen grains from staminate flowers had significantly higher levels of proteins and amino acids than inaperturate pollen grains from functionally female flowers. Levels of proteins and amino acids were highest in bisexual and staminate flowers of the andromonoecious species. CONCLUSIONS: Higher levels of proteins and amino acids in porate pollen of "male" flowers in our functionally dioecious Solanum species suggests a greater nutritive reward for bees foraging on "male" plants than for those foraging on functionally "female" plants. Greater reward in porate pollen (including andromonoecious species) may be connected to the potential to generate a pollen tube.

Darwin's vexing contrivance: a new hypothesis for why some flowers have two kinds of anther.

Heteranthery, the presence of two or more anther types in the same flower, is taxonomically widespread among bee-pollinated angiosperms, yet has puzzled botanists since Darwin. We test two competing hypotheses for its evolution: the long-standing 'division of labour' hypothesis, which posits that some anthers are specialized as food rewards for bees whereas others are specialized for surreptitious pollination, and our new hypothesis that heteranthery is a way to gradually release pollen that maximizes pollen delivery. We examine the evolution of heteranthery and associated traits across the genus Clarkia (Onagraceae) and study plant-pollinator interactions in two heterantherous Clarkia species. Across species, heteranthery is associated with bee pollination, delayed dehiscence and colour crypsis of one anther whorl, and movement of that anther whorl upon dehiscence. Our mechanistic studies in heterantherous species show that bees notice, forage on and export pollen from each anther whorl when it is dehiscing, and that heteranthery promotes pollen export. We find no support for division of labour, but multifarious evidence that heteranthery is a mechanism for gradual pollen presentation that probably evolved through indirect male-male competition for siring success.

Gradual replacement of wild bees by honeybees in flowers of the Mediterranean Basin over the last 50 years.

Evidence for pollinator declines largely originates from mid-latitude regions in North America and Europe. Geographical heterogeneity in pollinator trends combined with geographical biases in pollinator studies can produce distorted extrapolations and limit understanding of pollinator responses to environmental changes. In contrast with the declines experienced in some well-investigated European and North American regions, honeybees seem to have increased recently in some areas of the Mediterranean Basin. Because honeybees can have negative impacts on wild bees, it was hypothesized that a biome-wide alteration in bee pollinator assemblages may be underway in the Mediterranean Basin involving a reduction in the relative number of wild bees. This hypothesis was tested using published quantitative data on bee pollinators of wild and cultivated plants from studies conducted between 1963 and 2017 in 13 countries from the European, African and Asian shores of the Mediterranean Sea. The density of honeybee colonies increased exponentially and wild bees were gradually replaced by honeybees in flowers of wild and cultivated plants. The proportion of wild bees at flowers was four times greater than that of honeybees at the beginning of the period, the proportions of both groups becoming roughly similar 50 years later. The Mediterranean Basin is a world biodiversity hotspot for wild bees and wild bee-pollinated plants, and the ubiquitous rise of honeybees to dominance as pollinators could in the long run undermine the diversity of plants and wild bees in the region.

A pollinator shift explains floral divergence in an orchid species complex in South Africa.

BACKGROUND AND AIMS: Floral diversification driven by shifts between pollinators has been one of the key explanations for the radiation of angiosperms. According to the Grant-Stebbins model of pollinator-driven speciation, these shifts result in morphologically distinct 'ecotypes' which may eventually become recognizable as species. The current circumscription of the food-deceptive southern African orchid Eulophia parviflora encompasses a highly variable monophyletic species complex. In this study, two forms were identified within this complex that differ in distribution, floral morphology, scent chemistry and phenology, and a test was made of whether these differences represent adaptations for different pollinators. METHODS AND RESULTS: Multivariate analysis of floral and vegetative traits revealed that there are at least two discrete morphological forms in the species complex. Field observations revealed that each form is pollinated by a different insect species, and thus represent distinct ecotypes. The early-flowering coastal form which has long spurs and floral scent dominated by sesquiterpene compounds is pollinated exclusively by the long-tongued bee Amegilla fallax (Apidae, Anthophorinae), while the late-flowering inland form with short spurs and floral scent dominated by benzenoid compounds is pollinated exclusively by the beetle Cyrtothyrea marginalis (Cetoniinae; Scarabaeidae). Choice experiments in a Y-maze olfactometer showed that beetles are preferentially attracted to the scent of the short-spurred form. A spur-shortening experiment showed that long spurs are required for effective pollination of the bee-pollinated form. Although it was initially thought likely that divergence occurred across a geographical pollinator gradient, plants of the long-spurred form were effectively pollinated when transplanted to an inland locality outside the natural coastal range of this form. Thus, the underlying geographical basis for the evolution of ecotypes in the E. parviflora complex remains uncertain, although early flowering in the long-spurred form to exploit the emergence of naïve bees may restrict this form to coastal areas where there is no frost that would damage flower buds. Later flowering of the short-spurred form coincides closely with the emergence of the pollinating beetles following winter frosts. CONCLUSIONS: This study identifies a shift between bee and beetle pollination as the main driver of floral divergence in an orchid species complex. Floral scent and spur length appear to be key traits in mediating this evolutionary transition.

Flies as pollinators of melittophilous Salvia species (Lamiaceae).

UNLABELLED: • PREMISE OF THE STUDY: Floral adaptation to a functional pollinator group does not necessarily mean close specialization to a few pollinator species. For the more than 950 species of Salvia, only bee and bird pollinations are known. Restriction to these pollinators is mainly due to the specific flower construction (lever mechanism). Nevertheless, it has been repeatedly suggested that Salvia flowers might also be pollinated by flies. Are flies able to handle the lever mechanism? Are they functionally equivalent pollinators? In this study, we compared and quantified pollen transfer by bees and flies to test whether flies are true pollinators in Salvia.• METHODS: We identified pollinators using field observations and photos. Video documentation of the visitation rate and the site of pollen placement on the pollinator body, morphometric measurements, quantification of pollen placement, pollen load, handling time, and stigma contact ratio were analyzed.• KEY RESULTS: Field investigations revealed that 19 insect species pollinated S. virgata and four pollinated S. verticillata, including 16 bee species from seven genera of the Apidae and three fly species from three genera of the Nemestrinidae and Tabanidae.• CONCLUSIONS: Flies have been found to be pollinators in primarily bee-pollinated Salvia species. This result demonstrates the potential of a given "melittoid" flower construction to broaden the range of pollinators to guarantee successful pollination and seed production. Though bees, particularly Bombus terrestris, were more efficient than flies, the study shows that flies significantly contribute to pollen transfer in Salvia.

Darwin's inflorescence syndrome is indeed associated with bee pollination.

KEY MESSAGE: A relationship between vertical acropetal inflorescences with protandrous flowers and bee pollination was hypothesized by Darwin back in 1877. Here we provide empirical evidence supporting this association across the angiosperms. Plant reproduction is not only determined by flower traits but also by the arrangement of flowers within inflorescences. Based on his observations of the orchid Spiranthes autumnalis, Darwin proposed in 1877 that bee-pollinated plants presenting protandrous flowers on vertical acropetal inflorescences, where proximal flowers open first, can exploit the stereotypical foraging behavior of their pollinators (i.e., upward movement through the inflorescence) to promote pollen exportation and reduce self-pollination. In these inflorescences, male-phase flowers lie spatially above female-phase flowers. To examine this untested hypothesis, we compiled literature information from 718 angiosperms species and evaluated the association between vertical acropetal inflorescences with protandrous flowers and bee pollination within a phylogenetic comparative framework. Results reveal that this type of inflorescence is indeed more common in species pollinated by bees. Moreover, this association does not seem to be weakened by the presence of alternative self-pollination avoidance mechanisms, like self-incompatibility, suggesting that this inflorescence type benefits mainly male rather than female fitness. Other inflorescence types placing male-phase flowers above female-phase flowers, e.g., vertical basipetal inflorescences with protogynous flowers, do not provide strong evidence of a differential association with pollination by bees. Female-biased nectar production in vertical acropetal inflorescences with protandrous flowers may reinforce the behavior of bees to fly upwards, rendering Darwin's configuration more adaptive than other inflorescence configurations.

To each their own! Nectar plasticity within a flower mediates distinct ecological interactions.

Nuptial and extranuptial nectaries are involved in interactions with different animal functional groups. Nectar traits involved in pollination mutualisms are well known. However, we know little about those traits involved in other mutualisms, such as ant-plant interactions, especially when both types of nectaries are in the same plant organ, the flower. Here we investigated if when two types of nectaries are exploited by distinct functional groups of floral visitors, even being within the same plant organ, the nectar secreted presents distinct features that fit animal requirements. We compared nectar secretion dynamics, floral visitors and nectar chemical composition of both nuptial and extranuptial nectaries in natural populations of the liana Amphilophium mansoanum (Bignoniaceae). For that we characterized nectar sugar, amino acid and specialized metabolite composition by high-performance liquid chromatography. Nuptial nectaries were visited by three medium- and large-sized bee species and extranuptial nectaries were visited mainly by ants, but also by cockroaches, wasps and flies. Nuptial and extranuptial nectar differed regarding volume, concentration, milligrams of sugars per flower and secretion dynamics. Nuptial nectar was sucrose-dominated, with high amounts of γ-aminobutyric acid and ß-aminobutyric acid and with theophylline-like alkaloid, which were all exclusive of nuptial nectar. Whereas extranuptial nectar was hexose-rich, had a richer and less variable amino acid chemical profile, with high amounts of serine and alanine amino acids and with higher amounts of the specialized metabolite tyramine. The nectar traits from nuptial and extranuptial nectaries differ in energy amount and nutritional value, as well as in neuroactive specialized metabolites. These differences seem to match floral visitors' requirements, since they exclusively consume one of the two nectar types and may be exerting selective pressures on the composition of the respective resources of interest.

Temporal Trends in Pollination Deficits and Its Potential Impacts on Chinese Agriculture.

Worldwide, there is increasing evidence that shows a decline in pollinators, limiting crop pollination and production. However, it is unclear to what extent Chinese agriculture could be impacted by pollinator deficits. Data for 84 major crops in China between 1961 and 2018 were analyzed for the temporal trends in crop area and production, agricultural economic contribution of pollination, crop yield deficits, and honey bee pollination demand. We found a rapid increase in agricultural dependence on insect pollinators: both the cultivated area and total production of pollinator-dependent crops increased faster than those of pollinator-independent crops during 1961-2018. The total economic value of pollination amounted to US$ 106.08 billion in 2010, representing 19.12% of the total production value of Chinese agriculture, approximately twice the 9.5% value estimated for global agriculture. Crops with higher pollinator dependence showed greater mean growth in cultivated area than those with lower dependence, but lower mean growth of crop production and yield. Crop yield growth was also more unstable with increasing pollinator dependence. The minimum pollination demand for honey bee colonies was about three times the stock of honey bee colonies available in 2018. Furthermore, we found a decline in crop yield deficit with the increase in honey bee colony pollination service capacity. We considered that the shortage of pollinators resulted in the yield deficits for pollinator-dependent crops. Future increase in the area of pollinator-dependent crops will increase the need for more pollinators, suggesting the importance of implementing measures to protect pollinators to ensure a better-secured future for agricultural production in China.

Pollinator shifts, contingent evolution, and evolutionary constraint drive floral disparity in Salvia (Lamiaceae): Evidence from morphometrics and phylogenetic comparative methods.

Switches in pollinators have been argued to be key drivers of floral evolution in angiosperms. However, few studies have tested the relationship between floral shape evolution and switches in pollination in large clades. In concert with a dated phylogeny, we present a morphometric analysis of corolla, anther connective, and style shape across 44% of nearly 1000 species of Salvia (Lamiaceae) and test four hypotheses of floral evolution. We demonstrate that floral morphospace of New World (NW) Salvia is largely distinct from that of Old World (OW) Salvia and that these differences are pollinator driven; shifts in floral morphology sometimes mirror shifts in pollinators; anther connectives (key constituents of the Salvia staminal lever) and styles co-evolved from curved to linear shapes following shifts from bee to bird pollination; and morphological differences between NW and OW bee flowers are partly the legacy of constraints imposed by an earlier shift to bird pollination in the NW. The distinctive staminal lever in Salvia is a morphologically diverse structure that has evolved in concert with both the corolla and style, under different pollinator pressures, and in contingent fashion.

Economic Valuation of Bee Pollination Services for Passion Fruit (Malpighiales: Passifloraceae) Cultivation on Smallholding Farms in São Paulo, Brazil, Using the Avoided Cost Method.

This paper estimates the economic value of ecosystem services provided by Brazilian native bee, Xylocopa spp. Latreille (Hymenoptera: Apidae), pollination on a scale relevant to individual smallholder farmers that produce yellow passion fruit (Passiflora edulis Sims). The study areas are located in the vicinity of Pedro de Toledo and Itariri (Sao Paulo State-Brazil), in the Atlantic Forest region. The local economy is based on family farms, small stores, and ecotourism. The value was obtained using the ecological economics Avoided Cost Method, also known as replacement cost. Farms from this region informally hire temporary day laborers to supplement natural pollination with manual pollination of passion fruit flowers, so the cost of contracting temporary laborers was used to estimate the economic value of bee pollination. The value of pollination services was estimated at US$ 2,583.00 per hectare over 2 yr of P. edulis farming. Our estimates based on passion fruit farmer surveys and ecological valuation over 2 yr suggest that manual pollination accounts for 44-48% of production costs and results in a loss of ~58% of profits when wild bee pollination services are not available and manual pollination is required. We suggest that smallholder farmers follow the suggestions of previous studies and conserve adequate forest habitat for bee nesting and foraging, plan pesticide use around flowering and pollination, and supplement bee populations to maximize the benefit of the pollination ecosystem service and profits.

Floral micromorphology and nectar composition of the early evolutionary lineage Utricularia (subgenus Polypompholyx, Lentibulariaceae).

Utricularia (Lentibulariaceae) is a genus comprising around 240 species of herbaceous, carnivorous plants. Utricularia is usually viewed as an insect-pollinated genus, with the exception of a few bird-pollinated species. The bladderworts Utricularia multifida and U. tenella are interesting species because they represent an early evolutionary Utricularia branch and have some unusual morphological characters in their traps and calyx. Thus, our aims were to (i) determine whether the nectar sugar concentrations and composition in U. multifida and U. tenella are similar to those of other Utricularia species from the subgenera Polypompholyx and Utricularia, (ii) compare the nectary structure of U. multifida and U. tenella with those of other Utricularia species, and (iii) determine whether U. multifida and U. tenella use some of their floral trichomes as an alternative food reward for pollinators. We used light microscopy, histochemistry, and scanning and transmission electron microscopy to address those aims. The concentration and composition of nectar sugars were analysed using high-performance liquid chromatography. In all of the examined species, the floral nectary consisted of a spur bearing glandular trichomes. The spur produced and stored the nectar. We detected hexose-dominated (fructose + glucose) nectar in U. multifida and U. tenella as well as in U. violacea. In both U. multifida and U. tenella, there were trichomes that blocked the entrance into the throat and spur. Because these trichomes were rich in chromoplasts and contained lipid droplets, they may form an additional visual attractant. Bearing in mind the phylogenetic hypothesis for the genus, we suggest that an early ancestor of Utricularia had a nectariferous spur flower with a lower lip that formed a wide landing platform for bee pollinators.

Evidence for passerine bird pollination in Rhododendron species.

When insect activity is limited at low temperature, birds may be comparatively more important pollinators than insects for flowering plants. It has been thought that many large-flowered Rhododendron species are pollinated by local birds in the Himalayan regions because most of these species flower in spring at high elevation with cool atmospheric temperature. However, experimental evidence for the role of bird pollination in this hyperdiverse genus remains scarce. To examine the role of birds and insects in pollination, we observed floral visitors to 15 Rhododendron species with different floral sizes and abundant flowering individuals in the eastern Himalayas, Southwest China. To examine the role of birds and insects in female reproductive success in each species, cages were used to exclude birds but not insects from visiting flowers and net bags were used to exclude all floral visitors. Inflorescences where visitation was excluded did not produce fruits in any of the Rhododendron species, indicating that sexual reproduction in these species depended on pollinator visitation. Bird visits were generally less frequent than bee visits in the studied species. However, in the nine species on which bird visitors were observed, fruit and/or seed set were greatly reduced in inflorescences caged to exclude birds but not bees, compared to open-pollinated inflorescences. In the other six species on which bird visitation was not observed, fruit and seed set did not differ significantly between caged and open inflorescences except in one species (R. wardii). Manipulations to achieve selective exclusion of visitors demonstrated that birds could be effective pollinators for 10 out of 15 studied Rhododendron species in the eastern Himalayas. Floral characteristics of these Rhododendron species and weather conditions might favour the evolution of bird pollination systems in the East Himalayas.

Pollination effects on antioxidant content of Perilla frutescens seeds analysed by NMR spectroscopy.

The effects of Perilla frutescens pollination on the content of seed antioxidants were analysed by agronomical and pollination trials, comparing seeds produced from bagged plants in 2013 (A) to prevent access to pollinating insects, and seeds from open-pollinated plants in 2013 (B) and 2015 (C). The seeds of open-pollinated plants were significantly more numerous and heavier than those of self-pollinated plants. 1H NMR seed analysis showed a higher presence of phenolic compounds in open-pollinated seeds, mainly rosmarinic acid and flavonoids, apigenin and luteolin. Flavonoids were present in the glucosylated form in seeds (A) and (C), and in the aglycone form in seeds from (B) plants. Saturated and unsaturated fatty acids (palmitic, linoleic and linolenic) were more abundant in seeds from self-pollinated flowers. Pollination performed almost exclusively by the honeybee notably increased the antioxidant content in perilla seeds and gave rise to a reduction in the fatty acid content.

Bees, birds and yellow flowers: pollinator-dependent convergent evolution of UV patterns.

Colour is one of the most obvious advertisements of flowers, and occurs in a huge diversity among the angiosperms. Flower colour is responsible for attraction from a distance, whereas contrasting colour patterns within flowers aid orientation of flower visitors after approaching the flowers. Due to the striking differences in colour vision systems and neural processing across animal taxa, flower colours evoke specific behavioural responses by different flower visitors. We tested whether and how yellow flowers differ in their spectral reflectance depending on the main pollinator. We focused on bees and birds and examined whether the presence or absence of the widespread UV reflectance pattern of yellow flowers predicts the main pollinator. Most bee-pollinated flowers displayed a pattern with UV-absorbing centres and UV-reflecting peripheries, whereas the majority of bird-pollinated flowers are entirely UV- absorbing. In choice experiments we found that bees did not show consistent preferences for any colour or pattern types. However, all tested bee species made their first antennal contact preferably at the UV-absorbing area of the artificial flower, irrespective of its spatial position within the flower. The appearance of UV patterns within flowers is the main difference in spectral reflectance between yellow bee- and bird-pollinated flowers, and affects the foraging behaviour of flower visitors. The results support the hypothesis that flower colours and the visual capabilities of their efficient pollinators are adapted to each other.