Indirect effects shape species fitness in coevolved mutualistic networks.

Ecological interactions are one of the main forces that sustain Earth's biodiversity. A major challenge for studies of ecology and evolution is to determine how these interactions affect the fitness of species when we expand from studying isolated, pairwise interactions to include networks of interacting species1-4. In networks, chains of effects caused by a range of species have an indirect effect on other species they do not interact with directly, potentially affecting the fitness outcomes of a variety of ecological interactions (such as mutualism)5-7. Here we apply analytical techniques and numerical simulations to 186 empirical mutualistic networks and show how both direct and indirect effects alter the fitness of species coevolving in these networks. Although the fitness of species usually increased with the number of mutualistic partners, most of the fitness variation across species was driven by indirect effects. We found that these indirect effects prevent coevolving species from adapting to their mutualistic partners and to other sources of selection pressure in the environment, thereby decreasing their fitness. Such decreases are distributed in a predictable way within networks: peripheral species receive more indirect effects and experience higher reductions in fitness than central species. This topological effect was also evident when we analysed an empirical study of an invasion of pollination networks by honeybees. As honeybees became integrated as a central species within networks, they increased the contribution of indirect effects on several other species, reducing their fitness. Our study shows how and why indirect effects can govern the adaptive landscape of species-rich mutualistic assemblages.

The role of the brown bear Ursus arctos as a legitimate megafaunal seed disperser.

Megafaunal frugivores can consume large amounts of fruits whose seeds may be dispersed over long distances, thus, affecting plant regeneration processes and ecosystem functioning. We investigated the role of brown bears (Ursus arctos) as legitimate megafaunal seed dispersers. We assessed the quantity component of seed dispersal by brown bears across its entire distribution based on information about both the relative frequency of occurrence and species composition of fleshy fruits in the diet of brown bears extracted from the literature. We assessed the quality component of seed dispersal based on germination experiments for 11 fleshy-fruited plant species common in temperate and boreal regions and frequently eaten by brown bears. Across its distribution, fleshy fruits, on average, represented 24% of the bear food items and 26% of the total volume consumed. Brown bears consumed seeds from at least 101 fleshy-fruited plant species belonging to 24 families and 42 genera, of which Rubus (Rosaceae) and Vaccinium (Ericaceae) were most commonly eaten. Brown bears inhabiting Mediterranean forests relied the most on fleshy fruits and consumed the largest number of species per study area. Seeds ingested by bears germinated at higher percentages than those from whole fruits, and at similar percentages than manually depulped seeds. We conclude that brown bears are legitimate seed dispersers as they consume large quantities of seeds that remain viable after gut passage. The decline of these megafaunal frugivores may compromise seed dispersal services and plant regeneration processes.

Conflicting selection on Cneorum tricoccon (Rutaceae) seed size caused by native and alien seed dispersers.

The disappearance of native seed dispersers due to anthropogenic activities is often accompanied by the introduction of alien species, which may to some extent replace the ecological service provided by the extinct ones. Yet, little empirical evidence exists demonstrating the evolutionary consequences of such alien "replacement." Here, we document the conflicting selection exerted on seed size by two native lizards (Podarcis lilfordi and P. pityusensis) and an alien mammal species (Martes martes), all acting as legitimate seed dispersers of the Mediterranean relict Cneorum tricoccon. While lizards mostly exerted a negative directional selection on seed diameter, especially P. pityusensis, the much larger pine marten exerted positive selection on seed size. Our findings suggest that this among-disperser variation in the selection regimes, together with the occurrence of spatial variation in the presence of each seed disperser, help to create the geographical variation observed for seed size of C. tricoccon. To our knowledge, this is the first empirical evidence showing opposing selective pressures between native and alien species in the seed dispersal process in an invaded ecosystem.

Honeybees disrupt the structure and functionality of plant-pollinator networks.

The honeybee is the primary managed species worldwide for both crop pollination and honey production. Owing to beekeeping activity, its high relative abundance potentially affects the structure and functioning of pollination networks in natural ecosystems. Given that evidences about beekeeping impacts are restricted to observational studies of specific species and theoretical simulations, we still lack experimental data to test for their larger-scale impacts on biodiversity. Here we used a three-year field experiment in a natural ecosystem to compare the effects of pre- and post-establishment stages of beehives on the pollination network structure and plant reproductive success. Our results show that beekeeping reduces the diversity of wild pollinators and interaction links in the pollination networks. It disrupts their hierarchical structural organization causing the loss of interactions by generalist species, and also impairs pollination services by wild pollinators through reducing the reproductive success of those plant species highly visited by honeybees. High-density beekeeping in natural areas appears to have lasting, more serious negative impacts on biodiversity than was previously assumed.

Functional consequences of plant-animal interactions along the mutualism-antagonism gradient.

Plant-animal interactions are pivotal for ecosystem functioning, and usually form complex networks involving multiple species of mutualists as well as antagonists. The costs and benefits of these interactions show a strong context-dependency directly related to individual variation in partner identity and differential strength. Yet understanding the context-dependency and functional consequences of mutualistic and antagonistic interactions on individuals remains a lasting challenge. We use a network approach to characterize the individual, plant-based pollination interaction networks of the Canarian Isoplexis canariensis (Plantaginaceae) with a mixed assemblage of vertebrate mutualists (birds and lizards) and invertebrate antagonists (florivores, nectar larcenists, and predispersal seed predators). We identify and quantify interaction typologies based on the sign (mutualistic vs. antagonistic) and strength (weak vs. strong) of animal-mediated pollination and test the relationship with individual female reproductive success (FRS). In addition, we document pollinator movement patterns among individual plants to infer events of pollen transfer/receipt that define the plant mating networks and test the relationship with FRS. We identify six interaction typologies along a mutualism-antagonism gradient, with two typologies being over-represented involving both mutualists and antagonists and influencing FRS. Plants showing strong mutualistic interactions, but also (weak or strong) interactions with antagonists are relatively better connected in the mating network (i.e., with higher potential to transfer or receive pollen). Thus, mixed flower visitor assemblages with mutualists and antagonists give plants increased their importance in the mating networks, promote outcrossing and increasing both female and male fitness. Our approach helps characterize plant-animal interaction typologies, the context-specificity of diversified mutualisms, and a better forecasting of their functional consequences.

Pollinator shifts drive petal epidermal evolution on the Macaronesian Islands bird-flowered species.

Pollinator shifts are considered to drive floral trait evolution, yet little is still known about the modifications of petal epidermal surface at a biogeographic region scale. Here we investigated how independent shifts from insects to passerine birds in the Macaronesian Islands consistently modified this floral trait (i.e. absence of papillate cells). Using current phylogenies and extensive evidence from field observations, we selected a total of 81 plant species and subspecies for petal microscopy and comparative analysis, including 19 of the 23 insular species pollinated by opportunistic passerine birds (Macaronesian bird-flowered element). Species relying on passerine birds as the most effective pollinators (bird-pollinated) independently evolved at least five times and in all instances associated with a loss of papillate cells, whereas species with a mixed pollination system (birds plus insects and/or other vertebrates) evolved at least five times in Macaronesia and papillate cells were lost in only 25% of these transitions. Our findings suggest that petal micromorphology is a labile trait during pollinator shifts and that papillate cells tend to be absent on those species where pollinators have limited mechanical interaction with flowers, including opportunistic passerine birds that forage by hovering or from the ground.

The signatures of Anthropocene defaunation: cascading effects of the seed dispersal collapse.

Anthropogenic activity is driving population declines and extinctions of large-bodied, fruit-eating animals worldwide. Loss of these frugivores is expected to trigger negative cascading effects on plant populations if remnant species fail to replace the seed dispersal services provided by the extinct frugivores. A collapse of seed dispersal may not only affect plant demography (i.e., lack of recruitment), but should also supress gene flow via seed dispersal. Yet little empirical data still exist demonstrating the genetic consequences of defaunation for animal-dispersed plant species. Here, we first document a significant reduction of seed dispersal distances along a gradient of human-driven defaunation, with increasing loss of large- and medium-bodied frugivores. We then show that local plant neighbourhoods have higher genetic similarity, and smaller effective population sizes when large seed dispersers become extinct (i.e., only small frugivores remain) or are even partially downgraded (i.e., medium-sized frugivores providing less efficient seed dispersal). Our results demonstrate that preservation of large frugivores is crucial to maintain functional seed dispersal services and their associated genetic imprints, a central conservation target. Early signals of reduced dispersal distances that accompany the Anthropogenic defaunation forecast multiple, cascading effects on plant populations.

Hotspots of damage by antagonists shape the spatial structure of plant-pollinator interactions.

The balance between mutualistic and antagonistic plant-animal interactions and their spatial variation results in a highly dynamic mosaic of reproductive success within plant populations. Yet, the ecological drivers of this small-scale heterogeneity of interaction patterns and their outcomes remain virtually unexplored. We analyzed spatial structure in the frequency and intensity of interactions that vertebrate pollinators (birds and lizards) and invertebrate antagonists (florivores, nectar larcenists, and seed predators) had when interacting with the insular plant Isoplexis canariensis, and their effect on plant fitness. Spatially autocorrelated variation in plant reproductive success (fruit and viable seed set) emerged from the combined action of mutualists and antagonists, rather than reflecting the spatial pattern of any specific animal group. However, the influence of antagonists on plant fitness was stronger primarily due to the florivores' action on earlier reproductive stages, consuming and damaging floral structures before the arrival of pollinators. Our results indicate that the early action of antagonists creates hotspots of increased plant damage, where the effects of later acting mutualists are not translated into increased reproductive benefits. We foresee the potential for antagonists to shape the intra-population mosaics of plant fitness in situations where antagonists outnumber mutualists, when their interactions occur before those of mutualists, and when mutualists can detect and avoid damaged plants while foraging. Severely damaged plants in antagonistic hotspots might be excluded from the mating network and render a limited production of viable seeds, reducing both the growth rate of the plant population and the effective population size.

Birds see the true colours of fruits to live off the fat of the land.

Communication is a characteristic of life, but its reliability and basic definition are hotly debated. Theory predicts that trade among mutualists requires high reliability. Here, we show that moderate reliability already allows mutualists to optimize their rewards. The colours of Mediterranean fleshy-fruits indicate lipid rewards (but not other nutrients) to avian seed dispersers on regional and local scales. On the regional scale, fruits with high lipid content were significantly darker and less chromatic than congeners with lower lipid content. On the local scale, two warbler species (Sylvia atricapilla and Sylvia borin) selected fruit colours that were less chromatic, and thereby maximized their intake of lipids-a critical resource during migration and wintering. Crucially, birds were able to maximize lipid rewards with moderate reliability from visual fruit colours (r(2) = 0.44-0.60). We suggest that mutualisms require only that any association between the quality and sensory aspects of signallers is learned through multiple, repeated interactions. Because these conditions are often fulfilled, also in social communication systems, we contend that selection on reliability is less intense than hitherto assumed. This may contribute to explaining the extraordinary diversity of signals, including that of plant reproductive displays.

Adaptation of flower and fruit colours to multiple, distinct mutualists.

Communication in plant-animal mutualisms frequently involves multiple perceivers. A fundamental uncertainty is whether and how species adapt to communicate with groups of mutualists having distinct sensory abilities. We quantified the colour conspicuousness of flowers and fruits originating from one European and two South American plant communities, using visual models of pollinators (bee and fly) and seed dispersers (bird, primate and marten). We show that flowers are more conspicuous than fruits to pollinators, and the reverse to seed dispersers. In addition, flowers are more conspicuous to pollinators than to seed dispersers and the reverse for fruits. Thus, despite marked differences in the visual systems of mutualists, flower and fruit colours have evolved to attract multiple, distinct mutualists but not unintended perceivers. We show that this adaptation is facilitated by a limited correlation between flower and fruit colours, and by the fact that colour signals as coded at the photoreceptor level are more similar within than between functional groups (pollinators and seed dispersers). Overall, these results provide the first quantitative demonstration that flower and fruit colours are adaptations allowing plants to communicate simultaneously with distinct groups of mutualists.

Disturbance regimes, gap-demanding trees and seed mass related to tree height in warm temperate rain forests worldwide.

For tropical lowland rain forests, Denslow (1987) hypothesized that in areas with large-scale disturbances tree species with a high demand for light make up a larger proportion of the flora; results of tests have been inconsistent. There has been no test for warm temperate rain forests (WTRFs), but they offer a promising testing ground because they differ widely in the extent of disturbance. WTRF is dominated by microphylls sensu Raunkiaer and has a simpler structure and range of physiognomy than tropical or subtropical rain forests. It occurs in six parts of the world: eastern Asia, New Zealand, Chile, South Africa, SE Australia and the Azores. On the Azores it has been mostly destroyed, so we studied instead the subtropical montane rain forest (STMRF) on the Canary Islands which also represents a relict of the kind of WTRF that once stretched across southern Eurasia. We sought to find whether in these six regions the proportion of tree species needing canopy gaps for establishment reflects the frequency and/or extent of canopy disturbance by wind, landslide, volcanic eruptions (lava flow and ash fall), flood or fire. We used standard floras and ecological accounts to draw up lists of core tree species commonly reaching 5 m height. We excluded species which are very rare, very localized in distribution, or confined to special habitats, e.g. coastal forests or rocky sites. We used published accounts and our own experience to classify species into three groups: (1) needing canopy gaps for establishment; (2) needing either light shade throughout or a canopy gap relatively soon (a few months or years) after establishment; and (3) variously more shade-tolerant. Group 1 species were divided according the kind of canopy opening needed: tree-fall gap, landslide, lava flow, flood or fire. Only some of the significant differences in proportion of Group 1 species were consistent with differences in the extent of disturbance; even in some of those cases other factors seem likely to have had a major determining influence during evolution. We also sought to determine whether the species that are at least 'short-term persistent' in the soil seed bank (lasting 2-4 years) are all species needing canopy gaps for establishment. The answer was negative; large numbers of seeds of some shade-tolerants accumulate in the soil, and these species are able to benefit from soil disturbance in deep shade. We found a significant and strong positive relationship in Japan between mean seed mass and mature tree height, a weak positive relationship in New Zealand and no relationship in any of the other four regions. When comparing the seed mass values of Group 1 and Group 3 species we obtained different answers depending on whether or not we confined ourselves to taxonomically controlled contrasts. In only two of the four regions with an appreciable number of species in Group 1 is the mean seed mass of such species significantly lower than that of Group 3 species when taxonomic relatedness is ignored.

Quantity and quality components of effectiveness in insular pollinator assemblages.

Ecologically isolated habitats (e.g., oceanic islands) favor the appearance of small assemblages of pollinators, generally characterized by highly contrasted life modes (e.g., birds, lizards), and opportunistic nectar-feeding behavior. Different life modes should promote a low functional equivalence among pollinators, while opportunistic nectar feeding would lead to reduced and unpredictable pollination effectiveness (PE) compared to more specialized nectarivores. Dissecting the quantity (QNC) and quality (QLC) components of PE, we studied the opportunistic bird-lizard pollinator assemblage of Isoplexis canariensis from the Canary Islands to experimentally evaluate these potential characteristics. Birds and lizards showed different positions in the PE landscape, highlighting their low functional equivalence. Birds were more efficient than lizards due to higher visitation frequency (QNC). Adult lizards differed from juveniles in effecting a higher production of viable seeds (QLC). The disparate life modes of birds and lizards resulted in ample intra- and inter-specific PE variance. The main sources of PE variance were visitation frequency (both lizards and birds), number of flowers probed (lizards) and proportion of viable seeds resulting from a single visit (birds). The non-coincident locations of birds and lizards on the PE landscape indicate potential constraints for effectiveness. Variations in pollinator abundance can result in major effectiveness shifts only if QLC is relatively high, while changes in QLC would increase PE substantially only at high QNC. The low functional equivalence of impoverished, highly contrasted pollinator assemblages may be an early diagnostic signal for pollinator extinction potentially driving the collapse of mutualistic services.

Long-term demographic consequences of a seed dispersal disruption.

The loss or decline of vertebrate frugivores can limit the regeneration of plants that depend on them. However, empirical evidence is showing that this is still very scarce, as functionally equivalent species may contribute to maintain the mutualistic interaction. Here, we investigated the long-term consequences of the extinction of frugivorous lizards on the population persistence of a Mediterranean relict shrub Cneorum tricoccon (Cneoraceae). We examined the demographic parameters among 26 insular and mainland populations, which encompass the entire plant distributional range, comparing populations with lizards with those in which these are extinct, but in which alien mammals currently act as seed dispersers. Plant recruitment was found to be higher on island populations with lizards than on those with mammals, and the long-term effects of the native disperser's loss were found in all vital phases of plant regeneration. The study thus gives evidence of the cascading effects of human-induced changes in ecosystems, showing how the disruption of native ecological processes can lead to species regression and, in the long term, even to local extinctions.

Consequences of plant-pollinator and floral-herbivore interactions on the reproductive success of the Canary Islands endemic Canarina canariensis (Campanulaceae).

PREMISE OF THE STUDY: Pollination is a critical phase for plant reproduction, but ecological and evolutionary outcomes by pollinators may be counteracted by floral herbivores. These interacting assemblages may also be altered (directly or indirectly) by introduced species, especially on oceanic islands. In this study, we analyzed the effects of opportunistic nectar-feeding passerine birds and native (semi-slugs) and introduced (rats) floral herbivores on the reproductive success of Canarina canariensis. • METHODS: Manual pollination experiments were conducted to determine plant breeding system and pollen limitation. We also identified floral visitors and their visitation frequencies by censuses. Bird pollination effectiveness was evaluated by selective exclosures. The intensity of floral herbivory by native vs. introduced herbivores and its effect on plant fitness was estimated in different areas within the Canary island of Tenerife. • KEY RESULTS: Canarina canariensis had a very low spontaneous selfing ability and high pollen limitation, despite being self-compatible. Birds increased fruit set and the percentage of viable seeds per fruit, while florivores, the principal floral visitors, reduced them. Semi-slugs mainly consumed male reproductive organs, while rats preferred female. There was a strong within-island spatial variation in the herbivory intensity. • CONCLUSIONS: Opportunistic nectar-feeding birds increase the production of viable seeds in C. canariensis, but their beneficial effects are counteracted by the high incidence of floral herbivory. Because native semi-slugs damaged anthers more frequently than did introduced rats, these florivores may differ in their effects on male and female plant reproductive success.

Isolation and characterization of 20 microsatellite loci for laurel species (Laurus, Lauraceae).

PREMISE OF THE STUDY: Microsatellite primers were developed for the evergreen tree Laurus to investigate population genetic structure and patterns of gene flow via animal-dispersed pollen and seeds. METHODS AND RESULTS: Twenty polymorphic nuclear microsatellite markers were developed using CA, GA, AAC, and ATG n-enriched genomic libraries. Given the tetraploidy of the sampled populations, we analyzed our data both as dominant loci and as codominant genotypic data to calculate allele frequencies and genetic diversity. A total of 196 and 222 alleles were found in 37 Mediterranean (L. nobilis) and 26 Macaronesian islands (L. azorica) individuals, respectively. CONCLUSIONS: Levels of polymorphism of the reported markers are adequate for studies of diversity and parentage in natural populations of this Tertiary relict tree.

Geographic patterns in fruit colour diversity: do leaves constrain the colour of fleshy fruits?

We tested for geographic patterns in fruit colour diversity. Fruit colours are thought to promote detection by seed dispersers. Because seed dispersers differ in their spectral sensitivities, we predicted that fruit colour diversity would be higher in regions with higher seed disperser diversity (i.e. the tropics). We collected reflectance data on 232 fruiting plant species and their natural backgrounds in seven localities in Europe, North and South America, and analysed fruit colour diversity according to the visual system of birds-the primary consumer types of these fruits. We found no evidence that fruit colours are either more conspicuous or more diverse in tropical areas characterised by higher seed disperser diversity. Instead, fruit colour diversity was lowest in central Brazil, suggesting that fruit colours may be more diverse in temperate regions. Although we found little evidence for geographic variation in fruit hues, the spectral properties of fruits were positively associated with the spectral properties of backgrounds. This result implies that fruit colours may be influenced by selection on the reflectance properties of leaves, thus constraining the evolution of fruit colour. Overall, the results suggest that fruit colours in the tropics are neither more diverse nor more conspicuous than temperate fruits, and that fruit colours may be influenced by correlated selection on leaf reflectance properties.

Isolation and characterization of 13 microsatellite loci for Neochamaelea pulverulenta (Cneoraceae).

We report 13 polymorphic nuclear microsatellite loci for Neochamaelea pulverulenta (Cneoraceae) using an enriched-library approach. Although this plant species is tetraploid, expected patterns for tetrasomic segregation were completely absent, and all loci analysed showed a diploid pattern of inheritance. We detected a total of 102 alleles in 57 individuals genotyped (mean number of alleles per locus was 7.85). The values of observed and expected heterozygosities ranged from 0.193 to 0.737 and 0.425 to 0.812 respectively. Disomic segregation, levels of polymorphism and the exclusionary power of the developed markers render them readily applicable for parentage assignment of dispersed seeds, and for analyses of spatial genetic structure and population connectivity.

Opportunistic nectar-feeding birds are effective pollinators of bird-flowers from Canary Islands: experimental evidence from Isoplexis canariensis (Scrophulariaceae).

Insular floras, characterized by simple pollination networks, sometimes include novel mutualistic agents such as nonspecialist nectarivores. In this study we confirmed the effective pollination of Isoplexis canariensis by opportunistic nectar-feeding birds in Tenerife, Canary Islands, Spain. This plant is among the ornithophilous species of the Canarian flora that lack past and present specialist nectarivorous birds. Experimental hand pollinations revealed self-compatibility, but cross-pollinated flowers produced a greater percentage of viable seeds than self-pollinated ones. Flowers were visited by five species of birds (Phylloscopus canariensis, Parus caeruleus, Sylvia melanocephala, Serinus canarius, and Fringilla coelebs) and by the endemic lizard (Gallotia galloti, Lacertidae). Insect pollination was absent, and the few insect visitors acted as nectar thieves or secondary nectar robbers. Birds represented 93.1% of total visits, with the Canarian Chiffchaff, Ph. canariensis, being the most frequent visitor. Flowers visited by birds set more, larger, and heavier fruit than flowers from which birds were excluded. Bird visitation also enhanced seed viability. These results demonstrate the active role of these opportunistic birds as effective pollinators of this Canarian bird-flower species. Further, the results reveal the need to consider the effect of these birds on the evolution of ornithophilous floral traits in absence of specialist nectarivores.

Body size in ecological networks.

Body size determines a host of species traits that can affect the structure and dynamics of food webs, and other ecological networks, across multiple scales of organization. Measuring body size provides a relatively simple means of encapsulating and condensing a large amount of the biological information embedded within an ecological network. Recently, important advances have been made by incorporating body size into theoretical models that explore food web stability, the patterning of energy fluxes, and responses to perturbations. Because metabolic constraints underpin body-size scaling relationships, metabolic theory offers a potentially useful new framework within which to develop novel models to describe the structure and functioning of ecological networks and to assess the probable consequences of biodiversity change.