Pollinator Specificity Research Articles

Why are the seed cones of conifers so diverse at pollination?

Form and function relationships in plant reproductive structures have long fascinated biologists. Although the intricate associations between specific pollinators and reproductive morphology have been widely explored among animal-pollinated plants, the evolutionary processes underlying the diverse morphologies of wind-pollinated plants remain less well understood. Here we study how this diversity may have arisen by focusing on two conifer species in the pine family that have divergent reproductive cone morphologies at pollination. Standard histology methods, artificial wind pollination assays and phylogenetic analyses were used in this study. A detailed study of cone ontogeny in these species reveals that variation in the rate at which their cone scales mature means that pollination occurs at different stages in their development, and thus in association with different specific morphologies. Pollination experiments nevertheless indicate that both species effectively capture pollen. In wind-pollinated plants, morphological diversity may result from simple variation in development among lineages rather than selective pressures for any major differences in function or performance. This work also illustrates the broader importance of developmental context in understanding plant form and function relationships; because plant reproductive structures perform many different functions over their lifetime, subtle differences in development may dramatically alter the specific morphologies that they use to meet these demands.

Co-evolution of enzyme activities and latex in fig (Ficus carica L.) during fruit maturity process

In this study, enzyme activities and latex production were assessed on twenty fig cultivars of San Pedro, Common, Smyrna and Caprifig-types, throughout different maturity stage (unripe, mid-ripe and ripe) for three times a day (morning, mid-day and late afternoon). Comparison of latex yield revealed negative correlation between the developmental changes of enzymes in the target fig types. The highest latex yield was 1% recorded in the Common-type cultivar Kahli and both San Pedro-type cultivars Bither Abiadh and Bither Akhal, while this yield declined 10-fold in the Smyrna-type cultivar Njali at mid-maturity stage. The protease activity exhibited the highest level of 1,312,652U/ml in cultivar Bidh Beghal (Smyrna-type) at the ripening stage that was correlated with the increase of lipase level by 286.33U/ml at the mid-maturity stage, while its inhibition was recorded by 100% at the unripe stage. The synergic effect between lipase and protease activities was recorded during the receptivity period, which means that enzyme expression of the fig latex depends upon the emitted volatile compounds. The apparent reliance on constituents as attractants and few and specific pollinators may indicate co-evolution.

Relative contribution of reproductive attributes to the density-dependent effects on fruit-set.

Reproductive success of a plant species can be affected by the distribution pattern of its conspecifics in a small population. Besides the low mate availability, the dynamics of breeding system and pollination mechanism may also contribute to low fruit-set in such populations. We examined the relative contribution of these reproductive attributes on fruit-set across the contrasting distribution pattern (denser vs. sparser plots) in two isolated natural populations of a near-threatened tree species, Anogeissus sericea var. nummularia. Although flowers in the species are of generalist type, the narrow stigmatic surface appears to impose a requirement for a specialist pollinator. Pollination in the tree species is mediated only by the flies. The trees exhibit partial selfing and suffer from strong inbreeding depression at the early life-history stages of the selfed progeny. We recorded significant difference between the denser and sparser plots in terms of inflorescence visits per tree, and the number of trees covered in a bout by the pollinators. Moreover, tree density showed a strong positive correlation with fruit-set. Besides the requirement of having proximity among the conspecifics to facilitate pollinator movement, pollen quality also seemed to be a crucial attribute in the reproductive success of the tree species. It is inferred that the mating pattern and fecundity of plants in small and isolated populations are significantly influenced by the extent of sexual incompatibility and magnitude of their dependence on pollinators.

Floral Trait Variations Among Wild Tobacco Populations Influence the Foraging Behavior of Hawkmoth Pollinators

Most pollinators visit flowers in the search of nectar rewards. However, as the floral nectar can often not be directly detected by pollinators, many flower visitors use secondary metabolites such as odor- or taste-proxies to anticipate nectar quantity and quality. Plants might exploit these sensory inferences of the pollinator to increase their pollination rates without increasing their caloric investment into their floral rewards. Here we investigated the effects of natural variation in certain primary and secondary floral metabolites in three populations of the wild tobacco, Nicotiana attenuata, on the pollination behavior of the hawkmoth Manduca sexta. Although offering the same caloric value per flower, the plants of these populations differ in the compositions and concentrations of sugars within the nectar. Moreover, the flowers of these plants emitted highly contrasting levels of attractive floral volatiles (benzyl acetone), but did not differ in the amounts of defensive nectar metabolites (nicotine). In wind tunnel assays with M. sexta moths, plants from those populations that released the largest amount of benzyl acetone as well as those that had a higher ratio of nectar sucrose were more frequently visited and re-visited by the hawkmoth. High emissions of benzyl acetone additionally correlated with a higher time investment of the moths into individual flowers on each visit, leading to the largest foraging success of the moths on those flowers that were most strongly scented. We propose that it is the variation of flower metabolites and their detection by the pollinator rather than the actual caloric value of the nectar, which determines pollinator visitations to a certain flower population. Hence, plants could potentially create a specialist pollinator community by altering their floral signals, either by producing volatiles that pollinators prefer or by providing nectar sugars that pollinators are most sensitive to, while at the same time keeping the caloric value of their nectar rewards at a constant level. These findings highlight the importance of variations among floral signals and their detection by specific pollinators for plant diversification as well as for the evolution of specific plant-pollinator interactions.

Evolution of nectar spur length in a clade of Linaria reflects changes in cell division rather than in cell expansion.

Background and AimsNectar spurs (tubular outgrowths of a floral organ which contain, or give the appearance of containing, nectar) are hypothesized to be a ‘key innovation’ which can lead to rapid speciation within a lineage, because they are involved in pollinator specificity. Despite the ecological importance of nectar spurs, relatively little is known about their development. We used a comparative approach to investigate variation in nectar spur length in a clade of eight Iberian toadflaxes.MethodsSpur growth was measured at the macroscopic level over time in all eight species, and growth rate and growth duration compared. Evolution of growth rate was reconstructed across the phylogeny. Within the clade we then focused on Linaria becerrae and Linaria clementei, a pair of sister species which have extremely long and short spurs, respectively. Characterization at a micromorphological level was performed across a range of key developmental stages to determine whether the difference in spur length is due to differential cell expansion or cell division.Key ResultsWe detected a significant difference in the evolved growth rates, while developmental timing of both the initiation and the end of spur growth remained similar. Cell number is three times higher in the long spurred L. becerrae compared with L. clementei, whereas cell length is only 1.3 times greater. In addition, overall anisotropy of mature cells is not significantly different between the two species.ConclusionsWe found that changes in cell number and therefore in cell division largely explain evolution of spur length. This contrasts with previous studies in Aquilegia which have found that variation in nectar spur length is due to directed cell expansion (anisotropy) over variable time frames. Our study adds to knowledge about nectar spur development in a comparative context and indicates that different systems may have evolved nectar spurs using disparate mechanisms.

The largest early-diverging angiosperm family is mostly pollinated by ovipositing insects and so are most surviving lineages of early angiosperms.

Insect pollination in basal angiosperms is assumed to mostly involve 'generalized' insects looking for food, but direct observations of ANITA grade (283 species) pollinators are sparse. We present new data for numerous Schisandraceae, the largest ANITA family, from fieldwork, nocturnal filming, electron microscopy, barcoding and molecular clocks to infer pollinator/plant interactions over multiple years at sites throughout China to test the extent of pollinator specificity. Schisandraceae are pollinated by nocturnal gall midges that lay eggs in the flowers and whose larvae then feed on floral exudates. At least three Schisandraceae have shifted to beetle pollination. Pollination by a single midge species predominates, but one species was pollinated by different species at three locations and one by two at the same location. Based on molecular clocks, gall midges and Schisandraceae may have interacted since at least the Early Miocene. Combining these findings with a review of all published ANITA pollination data shows that ovipositing flies are the most common pollinators of living representatives of the ANITA grade. Compared to food reward-based pollination, oviposition-based systems are less wasteful of plant gametes because (i) none are eaten and (ii) female insects with herbivorous larvae reliably visit conspecific flowers.

Specificity and influence of climatic factors on the pollination of south-eastern Brazilian Sinningia

Ornithophily is a widespread pollination syndrome that has evolved independently several times along the evolution of plants. It is reported in sixty-five plant families, including Gesneriaceae. One of the most diverse genera within gesneriads is Sinningia, which comprises species pollinated by bees, hummingbirds and bats. However, little is known about the reproduction biology of either species or on pollinator specificity in this genus. Based on investigations on flowering phenology, flower morpho-anatomy, volume and concentration of floral nectar, pollinators, and breeding system, this paper reports the reproduction and the pollinator specificity of Sinningia douglasii and Sinningia araneosa in natural areas in Brazil. Both species offer nectar as a resource and are exclusively pollinated by a single species of hermit hummingbird. Nectar is produced by glands around the ovary. Pollen is deposited between the base of the beak and the head feathers. Both species are self-compatible and pollinator-dependent. In both Sinningia species, the combination of protandry, pollinator specificity, due to long corolla tubes that exclude the visitation of short-billed hummingbirds. The presence of particular flower features influence the occurrence of pollination by specific pollen vectors, such as hermit hummingbirds, in deterrence of illegitimate visitors (i.e. territorial hummingbirds), protection against vigorous foraging, and accurate placement of pollen on legitimate pollinators. In addition, the foraging of Phaethornis on flowers of Sinningia is independent of relative humidity and air temperature, whereas the main factor influencing hummingbird visits is the luminosity.

Species-specific male pollinators found for three native New Zealand greenhood orchids (Pterostylis spp.) suggest pollination by sexual deception

Many orchids achieve pollination by deceptive means. Sexually deceptive orchids are pollinated by male insects, which are lured to flowers that mimic the sex pheromones and/or appearance of their female conspecifics. This specialised pollination strategy was recently confirmed for the first time in a Pterostylis species in Australia. We investigated whether this pollination strategy may also be operating in Pterostylis species in New Zealand where generalised plant–insect pollination strategies are most commonly documented. The breeding systems of Pterostylis oliveri Petrie and Pterostylis irsoniana Hatch were investigated in the field with pollination treatments. Sticky traps were set up over flowering P. oliveri, P. irsoniana and Pterostylis venosa Colenso to catch potential pollinators of the flowers. Insects caught carrying orchid pollinia were identified, and the pollinia were identified to plant species with nuclear rDNA internal transcribed spacer (nrDNA ITS) sequences. Both P. oliveri and P. irsoniana were found to be self-compatible, but dependent on insects for pollination. Pollinia from each of the three Pterostylis spp. were found to be carried species-specifically by male fungus gnats (Diptera: Mycetophilidae): only Mycetophila latifascia fungus gnats carried the pollinia of P. oliveri, Morganiella fusca gnats carried the pollinia of P. irsoniana, and Tetragoneura sp. carried the pollinia of P. venosa. The pollinator specificity indicates that each of the male fungus gnat species was attracted to the flowers of a specific Pterostylis orchid. This strongly suggests that each of the orchid species emit a specific floral volatile, most probably resembling the sex pheromones of the female conspecifics, to lure their male pollinators. These are the first documented cases of highly specialised sexually deceptive pollination in New Zealand orchids, which were thought to be predominantly self-pollinating.

Competition, trait–mediated facilitation, and the structure of plant–pollinator communities

In plant–pollinator communities many pollinators are potential generalists and their preferences for certain plants can change quickly in response to changes in plant and pollinator densities. These changes in preferences affect coexistence within pollinator guilds as well as within plant guilds. Using a mathematical model, we study how adaptations of pollinator preferences influence population dynamics of a two-plant–two-pollinator community interaction module. Adaptation leads to coexistence between generalist and specialist pollinators, and produces complex plant population dynamics, involving alternative stable states and discrete transitions in the plant community. Pollinator adaptation also leads to plant–plant apparent facilitation that is mediated by changes in pollinator preferences. We show that adaptive pollinator behavior reduces niche overlap and leads to coexistence by specialization on different plants. Thus, this article documents how adaptive pollinator preferences for plants change the structure and coexistence of plant–pollinator communities.

Genetic Mapping Reveals an Anthocyanin Biosynthesis Pathway Gene Potentially Influencing Evolutionary Divergence between Two Subspecies of Scarlet Gilia (Ipomopsis aggregata).

Immense floral trait variation has likely arisen as an adaptation to attract pollinators. Different pollinator syndromes-suites of floral traits that attract specific pollinator functional groups-are repeatedly observed across closely related taxa or divergent populations. The observation of these trait syndromes suggests that pollinators use floral cues to signal the underlying nectar reward, and that complex trait combinations may persist and evolve through genetic correlations. Here, we explore pollinator preferences and the genetic architecture of floral divergence using an extensive genetic mapping study in the hybrid zone of two Ipomopsis aggregata subspecies that exhibit a hummingbird and a hawkmoth pollinator syndrome. We found that natural selection acts on several floral traits, and that hummingbirds and hawkmoths exhibited flower color preferences as predicted by their respective pollinator syndromes. Our quantitative trait loci (QTL) analyses revealed 46 loci affecting floral features, many of which colocalize across the genome. Two of these QTL have large effects explaining >15% of the phenotypic variance. The strongest QTL was associated with flower color and localized to a SNP in the anthocyanin biosynthesis pathway gene, dihydroflavonol-4-reductase (DFR). Further analysis revealed strong associations between DFR SNP variants, gene expression, and flower color across populations from the hybrid zone. Hence, DFR may be a target of pollinator-mediated selection in the hybrid zone of these two subspecies. Together, our findings suggest that hummingbirds and hawkmoths exhibit contrasting flower color preferences, which may drive the divergence of several floral traits through correlated trait evolution.

Changes in floral biology and inbreeding depression in native and invaded regions of Datura stramonium.

Plant populations invading new environments might compromise their fitness contribution to the next generation, because of the lack of native specialist pollinators and/or potential mates. Thus, changes in plant mating system and traits linked to it are expected in populations colonising new environments where selection would favour selfing and floral traits that maximise reproductive output. To test this, we studied native (Mexico) and non-native (Spain) populations of the obligate sexual reproducing annual weed Datura stramonium. Flower size, herkogamy, total number of seeds per plant, number of visits by and type of pollinators, and inbreeding depression were assessed in native and non-native populations. Finally, we measured phenotypic selection on corolla size and herkogamy in each population. Flower size and herkogamy showed wide and similar variation in both ranges. However, the largest average flower size was found in one non-native population whereas the highest average positive herkogamy was detected in one native population. On average, flowers in the native range received more visits by pollinators. Hawkmoths were the main visitors in the native populations while only bees were observed visiting flowers in Spain's populations. Only in the native range was inbreeding depression detected. Selection to reduce herkogamy was found only in one native population. Absence of both inbreeding depression and selection on floral traits suggest a change in mating system of D.stramonium in a new range where generalist pollinators may be promoting high reproductive success. Selection against deleterious alleles might explain the reduction of inbreeding depression, promoting the evolution of selfing.

Multiple factors contribute to reproductive isolation between two co-existing Habenaria species (Orchidaceae).

Reproductive isolation is a key feature that forms barriers to gene flow between distinct plants. In orchids, prezygotic reproductive isolation has been considered to be strong, because their associations with highly specific pollinators. In this study, the reproductive ecology and reproductive isolation of two sympatric Habenaria species, H. davidii and H. fordii, was investigated by floral phenology and morphology, hand-pollination experiments and visitor observation in southwest China. The two species were dependent on insects for pollination and completely self-compatible. A number of factors have been identified to limit gene flow between the two species and achieved full reproductive isolation. Ecogeographic isolation was a weak barrier. H. fordii and H. davidii had completely overlapped flowering periods, and floral morphology plays an important role in floral isolation. The two species shared the same hawkmoth pollinator, Cechenena lineosa, but the pollinaria of the two orchids were attached on different body parts of pollinators. Prezygotic isolation was not complete, but the interspecific pollination treatments of each species resulted in no seed sets, indicating that unlike many other orchid species, in which the postzygotic reproductive isolation is very weak or complete absence, the post-zygotic isolation strongly acted in the stage of seed production between two species. The results illustrate the reproductive isolation between two species involves multiple plant life-history stages and a variety of reproductive barriers can contribute to overall isolation.

How do steppe plants follow their optimal environmental conditions or persist under suboptimal conditions? The differing strategies of annuals and perennials.

For a species to be able to respond to environmental change, it must either succeed in following its optimal environmental conditions or in persisting under suboptimal conditions, but we know very little about what controls these capacities. We parameterized species distribution models (SDMs) for 135 plant species from the Algerian steppes. We interpreted low false‐positive rates as reflecting a high capacity to follow optimal environmental conditions and high false‐negative rates as a high capacity to persist under suboptimal environmental conditions. We also measured functional traits in the field and built a unique plant trait database for the North‐African steppe. For both perennial and annual species, we explored how these two capacities can be explained by species traits and whether relevant trait values reflect species strategies or biases in SDMs. We found low false‐positive rates in species with small seeds, flowers attracting specialist pollinators, and specialized distributions (among annuals and perennials), low root:shoot ratios, wide root‐systems, and large leaves (perennials only) (R 2 = .52–58). We found high false‐negative rates in species with marginal environmental distribution (among annuals and perennials), small seeds, relatively deep roots, and specialized distributions (annuals) or large leaves, wide root‐systems, and monocarpic life cycle (perennials) (R 2 = .38 for annuals and 0.65 for perennials). Overall, relevant traits are rarely indicative of the possible biases of SDMs, but rather reflect the species' reproductive strategy, dispersal ability, stress tolerance, and pollination strategies. Our results suggest that wide undirected dispersal in annual species and efficient resource acquisition in perennial species favor both capacities, whereas short life spans in perennial species favor persistence in suboptimal environmental conditions and flowers attracting specialist pollinators in perennial and annual species favor following optimal environmental conditions. Species that neither follow nor persist will be at risk under future environmental change.

The Biosynthesis of Unusual Floral Volatiles and Blends Involved in Orchid Pollination by Deception: Current Progress and Future Prospects.

Flowers have evolved diverse strategies to attract animal pollinators, with visual and olfactory floral cues often crucial for pollinator attraction. While most plants provide reward (e.g., nectar, pollen) in return for the service of pollination, 1000s of plant species, particularly in the orchid family, offer no apparent reward. Instead, they exploit their often specific pollinators (one or few) by mimicking signals of female insects, food source, and oviposition sites, among others. A full understanding of how these deceptive pollination strategies evolve and persist remains an open question. Nonetheless, there is growing evidence that unique blends that often contain unusual compounds in floral volatile constituents are often employed to secure pollination by deception. Thus, the ability of plants to rapidly evolve new pathways for synthesizing floral volatiles may hold the key to the widespread evolution of deceptive pollination. Yet, until now the biosynthesis of these volatile compounds has been largely neglected. While elucidating the biosynthesis in non-model systems is challenging, nonetheless, these cases may also offer untapped potential for biosynthetic breakthroughs given that some of the compounds can be exclusive or dominant components of the floral scent and production is often tissue-specific. In this perspective article, we first highlight the chemical diversity underpinning some of the more widespread deceptive orchid pollination strategies. Next, we explore the potential metabolic pathways and biosynthetic steps that might be involved. Finally, we offer recommendations to accelerate the discovery of the biochemical pathways in these challenging but intriguing systems.

Plant sex affects the structure of plant-pollinator networks in a subtropical forest.

Although it has long been recognized that the diversified sexual systems of plants could influence community patterns and pollination specialization, plant sex is not usually incorporated to quantify plant-pollinator networks. In this study, we observed 1776 visitations corresponding to 84 pollinator species and 28 plant species (19 sexually monomorphic plants and 9 dioecious plants) in a subtropical forest, China. We constructed three networks by, respectively, combining visitations to dioecious female and male plants at the species level, separating them, and retaining the shared visitations between them. When the shared visitations between male and female plants were considered, the modularity was increased and the nestedness was decreased with a significantly low robustness for the plant community. Only in this network, most dioecious and hermaphroditic plants were associated with different pollinator groups and separated to different modules. The results also showed that dioecious plants were more generalized and more likely to be module hubs in sex-combined network and sex-separated network but not in sex-shared network. Only in the sex-separated network, pollinators in dioecious modules were less selective than in hermaphroditic modules. Our study shows incorporating the different visitations between plant sexes could affect the analysis of key network structure properties and the description of pollination niche. To better understand niche partitioning and stability of plant-pollinator communities, it is necessary to compare pollination networks considering plant sexual diversity.

Importance of birds versus insects as pollinators of the African shrub Syncolostemon densiflorus (Lamiaceae)

Flowers visited by a broad assemblage of animals may nevertheless be considered to have a specialized pollination system if pollen deposition is effected mainly by a subset of these visitors. The importance of a particular animal group for pollination derives mainly from the relative rate of floral visitation and per-visit effectiveness for pollen deposition. We assessed the relative importance of birds and insects for pollination of the African shrub Syncolostemon densiflorus (Lamiaceae). Flower visitors were observed to evaluate their visitation rates. Virgin flowers were exposed to single visits by nine animal visitor groups to evaluate their effectiveness at depositing pollen on stigmas. We selectively excluded birds and larger insects from flowers to assess the contribution of smaller visitors to pollen deposition and seed set. Sunbirds and insects visited flowers at a similar frequency, but on a per-visit basis, sunbirds deposited more pollen on stigmas than did any of the insect groups, including honeybees, long-proboscid flies, butterflies and day-flying hawkmoths. Selective exclusion of birds and larger insects resulted in a large decline in pollen deposition on stigmas and seed production. These results show that S. densiflorus is pollinated mainly by sunbirds, despite its attraction of a broad assemblage of insect flower visitors. These findings highlight the importance of per-visit pollination effectiveness and selective exclusion for understanding the ecological importance of different flower visitors.

Floral glands in asclepiads: structure, diversity and evolution

ABSTRACT Species of Apocynaceae stand out among angiosperms in having very complex flowers, especially those of asclepiads, which belong to the most derived subfamily (Asclepiadoideae). These flowers are known to represent the highest degree of floral synorganization of the eudicots, and are comparable only to orchids. This morphological complexity may also be understood by observing their glands. Asclepiads have several protective and nuptial secretory structures. Their highly specific and specialized pollination systems are associated with the great diversity of glands found in their flowers. This review gathers data regarding all types of floral glands described for asclepiads and adds three new types (glandular trichome, secretory idioblast and obturator), for a total of 13 types of glands. Some of the species reported here may have dozens of glands of up to 11 types on a single flower, corresponding to the largest diversity of glands recorded to date for a single structure.

Landscape effects on pollination networks in Mediterranean gypsum islands.

Habitat fragmentation is a major driver of global change that has operated historically on Mediterranean ecosystems. However, more needs to be understood about how fragmentation influences ecological interactions, particularly pollination. Gypsum outcrops are historically fragmented Mediterranean habitats and settings for the evolution of many endangered soil-specialist plants with narrow ranges. In this study, we aimed to determine how fragmentation (area and connectivity) affects: (i) pollinator community composition and (ii) structural properties of pollination networks; and whether there are differences in the effects of fragmentation on: (iii) the number of interactions and visits among pollinator functional groups; and (iv) the number of interactions and specialisation degree between soil-specialist and soil-generalist plants. We characterised the degree of fragmentation and the pollination network structures in 12 gypsum habitat fragments embedded in a cropland matrix during two consecutive years. We found significant relationships between fragmentation and network structure. The effects of fragmentation differed among pollinator functional groups, but not between soil-specialist and soil-generalist plants, in terms of number of interactions. However, the relatively higher pollinator specialisation of soil-specialist plants suggested greater dependence on pollinators. Inter-annual variations in the network structures demonstrated the importance of temporal replication. The observed patterns related to the landscape structure and pollination at both the network and species levels provide insights into the key ecological processes in gypsum islands. These findings may help to identify the potential drivers of species persistence, especially for endangered soil-specialist plants with narrow ranges in a changing scenario with exacerbated habitat fragmentation.

Resolving Recent Plant Radiations: Power and Robustness of Genotyping-by-Sequencing.

Disentangling species boundaries and phylogenetic relationships within recent evolutionary radiations is a challenge due to the poor morphological differentiation and low genetic divergence between species, frequently accompanied by phenotypic convergence, interspecific gene flow and incomplete lineage sorting. Here we employed a genotyping-by-sequencing (GBS) approach, in combination with morphometric analyses, to investigate a small western Mediterranean clade in the flowering plant genus Linaria that radiated in the Quaternary. After confirming the morphological and genetic distinctness of eight species, we evaluated the relative performances of concatenation and coalescent methods to resolve phylogenetic relationships. Specifically, we focused on assessing the robustness of both approaches to variations in the parameter used to estimate sequence homology (clustering threshold). Concatenation analyses suffered from strong systematic bias, as revealed by the high statistical support for multiple alternative topologies depending on clustering threshold values. By contrast, topologies produced by two coalescent-based methods (NJ$_{\mathrm{st}}$, SVDquartets) were robust to variations in the clustering threshold. Reticulate evolution may partly explain incongruences between NJ$_{\mathrm{st}}$, SVDquartets and concatenated trees. Integration of morphometric and coalescent-based phylogenetic results revealed (i) extensive morphological divergence associated with recent splits between geographically close or sympatric sister species and (ii) morphological convergence in geographically disjunct species. These patterns are particularly true for floral traits related to pollinator specialization, including nectar spur length, tube width and corolla color, suggesting pollinator-driven diversification. Given its relatively simple and inexpensive implementation, GBS is a promising technique for the phylogenetic and systematic study of recent radiations, but care must be taken to evaluate the robustness of results to variation of data assembly parameters.

The nectar spur is not only a simple specialization for long-proboscid pollinators.

Since the time of Darwin, biologists have considered the floral nectar spur to be an adaptation representing a high degree of plant specialization. Nevertheless, some researchers suggest that nature is more complex and that even morphologically specialized plants attract a wide spectrum of visitors. We observed visitors on Impatiens burtonii (Balsaminaceae) and measured the depth of the proboscis insertion into the spur, the distance of the nectar surface from the spur entrance and the visitor's effectiveness. The hoverfly Melanostoma sp., with the shortest proboscis, was most active early in the morning and fed on pollen and nectar near the spur entrance. The honeybee Apis mellifera and the hoverfly Rhingia mecyana were the most frequent visitors before and after noon, respectively. Although R.mecyana, the only visitor able to reach the end of the spur, was the most frequent, it did not deposit the largest number of pollen grains per visit. Nectar spurs may function as complex structures allowing pollination by both short- and long-proboscid visitors and separating their spatial and temporal niches. Spurred plants should be considered as more generalized and exposed to more diverse selection pressures than previously believed.