The extent of introgression between incipient Clarkia species is determined by temporal environmental variation and mating system.

Introgression is pervasive across the tree of life but varies across taxa, geography, and genomic regions. However, the factors modulating this variation and how they may be affected by global change are not well understood. Here, we used 200 genomes and a 15-y site-specific environmental dataset to investigate the effects of environmental variation and mating system divergence on the magnitude of introgression between a recently diverged outcrosser-selfer pair of annual plants in the genus Clarkia. These sister taxa diverged very recently and subsequently came into secondary sympatry where they form replicated contact zones. Consistent with observations of other outcrosser-selfer pairs, we found that introgression was asymmetric between taxa, with substantially more introgression from the selfer to the outcrosser. This asymmetry was caused by a bias in the direction of initial F1 hybrid formation and subsequent backcrossing. We also found extensive variation in the outcrosser's admixture proportion among contact zones, which was predicted nearly entirely by interannual variance in spring precipitation. Greater fluctuations in spring precipitation resulted in higher admixture proportions, likely mediated by the effects of spring precipitation on the expression of traits that determine premating reproductive isolation. Climate-driven hybridization dynamics may be particularly affected by global change, potentially reshaping species boundaries and adaptation to novel environments.

Isolation by environment and its consequences for range shifts with global change: Landscape genomics of the invasive plant common tansy.

Invasive species are a growing global economic and ecological problem. However, it is not well understood how environmental factors mediate invasive range expansion. In this study, we investigated the recent and rapid range expansion of common tansy across environmental gradients in Minnesota, USA. We densely sampled individuals across the expanding range and performed reduced representation sequencing to generate a dataset of 3071 polymorphic loci for 176 individuals. We used non-spatial and spatially explicit analyses to determine the relative influences of geographic distance and environmental variation on patterns of genomic variation. We found no evidence for isolation by distance but strong evidence for isolation by environment, indicating that environmental factors may have modulated patterns of range expansion. Land use classification and soils were particularly important variables related to population structure although they operated on different spatial scales; land use classification was related to broad-scale patterns and soils were related to fine-scale patterns. All analyses indicated a distinctive genetic cluster in the most recently invaded portion of the range. Individuals from the far northwestern range margin were separated from the remainder of the range by reduced migration, which was associated with environmental resistance. This portion of the range was invaded primarily in the last 15 years. Ecological niche models also indicated that this cluster was associated with the expansion of the niche. While invasion is often assumed to be primarily influenced by dispersal limitation, our results suggest that ongoing invasion and range shifts with climate change may be strongly affected by environmental heterogeneity.

Population histories of variable reproductive success and low winter precipitation correlate with risk-averse seed germination in a mediterranean-climate winter annual.

PREMISE: Seed germination involves risk; post-germination conditions might not allow survival and reproduction. Variable, stressful environments favor seeds with germination that avoids risk (e.g., germination in conditions predicting success), spreads risk (e.g., dormancy), or escapes risk (e.g., rapid germination). Germination studies often investigate trait correlations with climate features linked to variation in post-germination reproductive success. Rarely are long-term records of population reproductive success available. METHODS: Supported by demographic and climate monitoring, we analyzed germination in the California winter-annual Clarkia xantiana subsp. xantiana. Sowing seeds of 10 populations across controlled levels of water potential and temperature, we estimated temperature-specific base water potential for 20% germination, germination time weighted by water potential above base (hydrotime), and a dormancy index (frequency of viable, ungerminated seeds). Mixed-effects models analyzed responses to (1) temperature, (2) discrete variation in reproductive success (presence or absence of years with zero seed production by a population), and (3) climate covariates, mean winter precipitation and coefficient of variation (CV) of spring precipitation. For six populations, records enabled analysis with a continuous metric of variable reproduction, the CV of per-capita reproductive success. RESULTS: Populations with more variable reproductive success had higher base water potential and dormancy. Higher base water potential and faster germination occurred at warmer experimental temperatures and in seeds of populations with wetter winters. CONCLUSIONS: Geographic variation in seed germination in this species suggests local adaptation to demographic risk and rainfall. High base water potential and dormancy may concentrate germination in years likely to allow reproduction, while spreading risk among years.

Bet Hedging Is Not Sufficient to Explain Germination Patterns of a Winter Annual Plant.

AbstractBet hedging consists of life history strategies that buffer against environmental variability by trading off immediate and long-term fitness. Delayed germination in annual plants is a classic example of bet hedging and is often invoked to explain low germination fractions. We examined whether bet hedging explains low and variable germination fractions among 20 populations of the winter annual plant Clarkia xantiana ssp. xantiana that experience substantial variation in reproductive success among years. Leveraging 15 years of demographic monitoring and 3 years of field germination experiments, we assessed the fitness consequences of seed banks and compared optimal germination fractions from a density-independent bet-hedging model to observed germination fractions. We did not find consistent evidence of bet hedging or the expected trade-off between arithmetic and geometric mean fitness, although delayed germination increased long-term fitness in 7 of 20 populations. Optimal germination fractions were two to five times higher than observed germination fractions, and among-population variation in germination fractions was not correlated with risks across the life cycle. Our comprehensive test suggests that bet hedging is not sufficient to explain the observed germination patterns. Understanding variation in germination strategies will likely require integrating bet hedging with complementary forces shaping the evolution of delayed germination.

Rapid evolution during climate change: demographic and genetic constraints on adaptation to severe drought.

Populations often vary in their evolutionary responses to a shared environmental perturbation. A key hurdle in building more predictive models of rapid evolution is understanding this variation-why do some populations and traits evolve while others do not? We combined long-term demographic and environmental data, estimates of quantitative genetic variance components, a resurrection experiment and individual-based evolutionary simulations to gain mechanistic insights into contrasting evolutionary responses to a severe multi-year drought. We examined five traits in two populations of a native California plant, Clarkia xantiana, at three time points over 7 years. Earlier flowering phenology evolved in only one of the two populations, though both populations experienced similar drought severity and demographic declines and were estimated to have considerable additive genetic variance for flowering phenology. Pairing demographic and experimental data with evolutionary simulations suggested that while seed banks in both populations probably constrained evolutionary responses, a stronger seed bank in the non-evolving population resulted in evolutionary stasis. Gene flow through time via germ banks may be an important, underappreciated control on rapid evolution in response to extreme environmental perturbations.

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.

Pleiotropy facilitates local adaptation to distant optima in common ragweed (Ambrosia artemisiifolia).

Pleiotropy, the control of multiple phenotypes by a single locus, is expected to slow the rate of adaptation by increasing the chance that beneficial alleles also have deleterious effects. However, a prediction arising from classical theory of quantitative trait evolution states that pleiotropic alleles may have a selective advantage when phenotypes are distant from their selective optima. We examine the role of pleiotropy in regulating adaptive differentiation among populations of common ragweed (Ambrosia artemisiifolia); a species that has recently expanded its North American range due to human-mediated habitat change. We employ a phenotype-free approach by using connectivity in gene networks as a proxy for pleiotropy. First, we identify loci bearing footprints of local adaptation, and then use genotype-expression mapping and co-expression networks to infer the connectivity of the genes. Our results indicate that the putatively adaptive loci are highly pleiotropic, as they are more likely than expected to affect the expression of other genes, and they reside in central positions within the gene networks. We propose that the conditionally advantageous alleles at these loci avoid the cost of pleiotropy by having large phenotypic effects that are beneficial when populations are far from their selective optima. We further use evolutionary simulations to show that these patterns are in agreement with a model where populations face novel selective pressures, as expected during a range expansion. Overall, our results suggest that highly connected genes may be targets of positive selection during environmental change, even though they likely experience strong purifying selection in stable selective environments.

Of mutualism and migration: will interactions with novel ericoid mycorrhizal communities help or hinder northward Rhododendron range shifts?

Rapid climate change imperils many small-ranged endemic species as the climate envelopes of their native ranges shift poleward. In addition to abiotic changes, biotic interactions are expected to play a critical role in plant species' responses. Below-ground interactions are of particular interest given increasing evidence of microbial effects on plant performance and the prevalence of mycorrhizal mutualisms. We used greenhouse mesocosm experiments to investigate how natural northward migration/assisted colonization of Rhododendron catawbiense, a small-ranged endemic eastern U.S. shrub, might be influenced by novel below-ground biotic interactions from soils north of its native range, particularly with ericoid mycorrhizal fungi (ERM). We compared germination, leaf size, survival, and ERM colonization rates of endemic R. catawbiense and widespread R. maximum when sown on different soil inoculum treatments: a sterilized control; a non-ERM biotic control; ERM communities from northern R. maximum populations; and ERM communities collected from the native range of R. catawbiense. Germination rates for both species when inoculated with congeners' novel soils were significantly higher than when inoculated with conspecific soils, or non-mycorrhizal controls. Mortality rates were unaffected by treatment, suggesting that the unexpected reciprocal effect of each species' increased establishment in association with heterospecific ERM could have lasting demographic effects. Our results suggest that seedling establishment of R. catawbiense in northern regions outside its native range could be facilitated by the presence of extant congeners like R. maximum and their associated soil microbiota. These findings have direct relevance to the potential for successful poleward migration or future assisted colonization efforts.

Microbes, mutualism, and range margins: testing the fitness consequences of soil microbial communities across and beyond a native plant's range.

Interactions between plants and soil fungi and bacteria are ubiquitous and have large effects on individual plant fitness. However, the degree to which spatial variation in soil microbial communities modulates plant species' distributions remains largely untested. Using the California native plant Clarkia xantiana ssp. xantiana we paired glasshouse and field reciprocal transplants of plant populations and soils to test whether plant-microbe interactions affect the plant's geographic range limit and whether there is local adaptation between plants and soil microbe communities. In the field and glasshouse, one of the two range interior inocula had a positive effect on plant fitness. In the field, this benefit was especially pronounced at the range edge and beyond, suggesting possible mutualist limitation. In the glasshouse, soil inocula from beyond-range tended to increase plant growth, suggesting microbial enemy release beyond the range margin. Amplicon sequencing revealed stark variation in microbial communities across the range boundary. Plants dispersing beyond their range limit are likely to encounter novel microbial communities. In C. x. xantiana, our results suggest that range expansion may be facilitated by fewer pathogens, but could also be hindered by a lack of mutualists. Both negative and positive plant-microbe interactions will likely affect contemporary range shifts.

Consequences of ignoring dispersal variation in network models for landscape connectivity.

Habitat loss and fragmentation can negatively influence population persistence and biodiversity, but the effects can be mitigated if species successfully disperse between isolated habitat patches. Network models are the primary tool for quantifying landscape connectivity, yet in practice, an overly simplistic view of species dispersal is applied. These models often ignore individual variation in dispersal ability under the assumption that all individuals move the same fixed distance with equal probability. We developed a modeling approach to address this problem. We incorporated dispersal kernels into network models to determine how individual variation in dispersal alters understanding of landscape-level connectivity and implemented our approach on a fragmented grassland landscape in Minnesota. Ignoring dispersal variation consistently overestimated a population's robustness to local extinctions and underestimated its robustness to local habitat loss. Furthermore, a simplified view of dispersal underestimated the amount of habitat substructure for small populations but overestimated habitat substructure for large populations. Our results demonstrate that considering biologically realistic dispersal alters understanding of landscape connectivity in ecological theory and conservation practice.

Consecuencias de la Omisión de la Variación en la Dispersión en los Modelos de Redes para la Conectividad de Paisajes Resumen La pérdida y la fragmentación del hábitat pueden influir negativamente la persistencia de poblaciones y biodiversidad. Sin embargo, estos efectos pueden ser mitigados si las especies tienen una dispersión exitosa entre los fragmentos aislados de hábitat. Los modelos de redes son la herramienta principal para la cuantificación de la conectividad del paisaje, no obstante en la práctica, se tiende a usar una visión excesivamente simplista de la dispersión de especies. Es común que estos modelos ignoren la variación que existe entre individuos en sus habilidades de dispersión y que asuman que todos los individuos se pueden mover la misma distancia y con la misma probabilidad. En este estudio, desarrollamos una estrategia de modelaje para (minimizar o aminorar) estas limitaciones incorporando kernels de dispersión dentro de los modelos de redes para determinar cómo la variación individual de la dispersión altera el entendimiento de la conectividad a nivel de paisaje. Como un ejemplo, implementamos esta estrategia en un paisaje de pastizal fragmentado en Minnesota. Omitir la variación en la dispersión generó una sobreestimación sistemática de la robustez de la población ante las extinciones locales y una subestimación de la robustez ante la pérdida local del hábitat. Además, una visión simplificada de la dispersión subestimó la complejidad de hábitat para las poblaciones pequeñas, sin emgargo sobreestimó la complejidad para las poblaciones grandes. Nuestros resultados demuestran que incorporar parámetros que describan una dispersión biológica realista tiene implicaciones importantes en la teoría de conectividad de paisajes e implementación de practicas de conservación.

Context Dependence of Local Adaptation to Abiotic and Biotic Environments: A Quantitative and Qualitative Synthesis.

Understanding how spatially variable selection shapes adaptation is an area of long-standing interest in evolutionary ecology. Recent meta-analyses have quantified the extent of local adaptation, but the relative importance of abiotic and biotic factors in driving population divergence remains poorly understood. To address this gap, we combined a quantitative meta-analysis and a qualitative metasynthesis to (1) quantify the magnitude of local adaptation to abiotic and biotic factors and (2) characterize major themes that influence the motivation and design of experiments that seek to test for local adaptation. Using local-foreign contrasts as a metric of local adaptation (or maladaptation), we found that local adaptation was greater in the presence than in the absence of a biotic interactor, especially for plants. We also found that biotic environments had stronger effects on fitness than abiotic environments when ignoring whether those environments were local versus foreign. Finally, biotic effects were stronger at low latitudes, and abiotic effects were stronger at high latitudes. Our qualitative analysis revealed that the lens through which local adaptation has been examined differs for abiotic and biotic factors. It also revealed biases in the design and implementation of experiments that make quantitative results challenging to interpret and provided directions for future research.

The opportunity for outcrossing varies across the geographic range of the primarily selfing Clarkia xantiana ssp. parviflora.

PREMISE: The timing of self-fertilization has potentially important consequences for the trajectory of mating system evolution, the opportunity for outcrossing, and the maintenance of genetic variation in populations. For primarily selfing taxa, it remains poorly understood as to how floral variation influences the opportunity for outcrossing and whether those attributes vary among populations across geographic ranges. METHODS: We examined variation in floral traits (herkogamy, protandry, flower size, stigma stage at anthesis, timing of stigma receptivity) in seven populations of Clarkia xantiana ssp. parviflora, a primarily selfing taxon, spanning from the western to eastern margins of its distribution. We also performed experimental emasculations and pollinations (followed by stigma severing) to quantify the extent of opportunities for outcrossing across flower development. RESULTS: There was marked among-population variation in all floral traits, particularly between far eastern and western populations. Emasculation experiments showed that the eastern populations had minimal autonomous selfing, but western populations had high rates of selfing within 24 h after anthesis. Population variation in autofertility was significantly predicted by floral trait variation, especially protandry and petal size. CONCLUSIONS: Greater protandry both extended the time over which outcrossing could potentially occur and reduced the probability of autonomous selfing, suggesting that there may be a tradeoff that results in fitness loss when pollinator visits are not common. The east-west pattern of differentiation in some floral traits parallels that of postglacial range expansion, suggesting that selection on the mating system may have been strong in the process of range expansion.

Biotic Interactions Contribute to the Geographic Range Limit of an Annual Plant: Herbivory and Phenology Mediate Fitness beyond a Range Margin.

Species' geographic distributions have already shifted during the Anthropocene. However, we often do not know what aspects of the environment drive range dynamics, much less which traits mediate organisms' responses to these environmental gradients. Most studies focus on possible climatic limits to species' distributions and have ignored the role of biotic interactions, despite theoretical support for their importance in setting distributional limits. We used field experiments and simulations to estimate contributions of mammalian herbivory to a range boundary in the Californian annual plant Clarkia xantiana ssp. xantiana. A steep gradient of increasing probability of herbivory occurred across the boundary, and a reanalysis of prior transplant experiments revealed that herbivory drove severalfold declines in lifetime fitness at and beyond the boundary. Simulations showed that populations could potentially persist beyond the range margin in the absence of herbivory. Using data from a narrowly sympatric subspecies, Clarkia xantiana parviflora, we also showed that delayed phenology is strongly associated with C. xantiana ssp. xantiana's susceptibility to herbivory and low fitness beyond its border. Overall, our results provide some of the most comprehensive evidence to date of how the interplay of demography, traits, and spatial gradients in species interactions can produce a geographic range limit, and they lend empirical support to recent developments in range limits theory.

Does adaptation to historical climate shape plant responses to future rainfall patterns? A rainfall manipulation experiment with common ragweed.

Climate change is affecting both the volume and distribution of precipitation, which in turn is expected to affect the growth and reproduction of plant populations. The near ubiquity of local adaptation suggests that adaptive differentiation may have important consequences for how populations are affected by and respond to changing precipitation. Here, we manipulated rainfall in a common garden to examine how differentiation among populations of common ragweed, Ambrosia artemisiifolia (Asteraceae) affects responses to water availability expected under climate change. We collected seeds from 26 populations along gradients of historical rainfall and used event-based rainout shelters and watering additions to simulate drier summer conditions and more extreme rainfall events, respectively. Ambrosia artemisiifolia had higher fitness on average under reduced rainfall, suggesting it may spread and become more abundant in areas projected to become hotter and drier during the summer months. We also found strong evidence for phenotypic and fitness clines across both latitude and longitude, and that phenological responses and fitness effects of altered rainfall depended on seed source or historical climate. The effect of rainfall treatment on female fitness was highest in western and mid longitudes, but there was little effect on eastern populations. Across latitude, the effect of rainfall treatment on male fitness was highest in southern populations. These phenology and fitness clines suggest that adaptive differentiation across the species' range has the potential to shape future responses of A. artemisiifolia populations to climate change, particularly altered patterns of rainfall.

Little plant, big city: a test of adaptation to urban environments in common ragweed (Ambrosia artemisiifolia).

A full understanding of how cities shape adaptation requires characterizing genetically-based phenotypic and fitness differences between urban and rural populations under field conditions. We used a reciprocal transplant experiment with the native plant common ragweed, (Ambrosia artemisiifolia), and found that urban and rural populations have diverged in flowering time, a trait that strongly affects fitness. Although urban populations flowered earlier than rural populations, plants growing in urban field sites flowered later than plants in rural field sites. This counter-gradient variation is consistent adaptive divergence between urban and rural populations. Also consistent with local adaptation, both urban and rural genotypes experienced stronger net selection in the foreign than in the local habitat, but this pattern was not significant for male fitness. Despite the evidence for local adaptation, rural populations had higher lifetime fitness at all sites, suggesting that selection has been stronger or more uniform in rural than urban populations. We also found that inter-population differences in both flowering time and fitness tended to be greater among urban than rural populations, which is consistent with greater drift or spatial variation in selection within urban environments. In summary, our results are consistent with adaptive divergence of urban and rural populations, but also suggest there may be greater environmental heterogeneity in urban environments which also affects evolution in urban landscapes.

Global biogeography of mating system variation in seed plants.

Latitudinal gradients in biotic interactions have been suggested as causes of global patterns of biodiversity and phenotypic variation. Plant biologists have long speculated that outcrossing mating systems are more common at low than high latitudes owing to a greater predictability of plant-pollinator interactions in the tropics; however, these ideas have not previously been tested. Here, we present the first global biogeographic analysis of plant mating systems based on 624 published studies from 492 taxa. We found a weak decline in outcrossing rate towards higher latitudes and among some biomes, but no biogeographic patterns in the frequency of self-incompatibility. Incorporating life history and growth form into biogeographic analyses reduced or eliminated the importance of latitude and biome in predicting outcrossing or self-incompatibility. Our results suggest that biogeographic patterns in mating system are more likely a reflection of the frequency of life forms across latitudes rather than the strength of plant-pollinator interactions.

Mating System Evolution under Strong Pollen Limitation: Evidence of Disruptive Selection through Male and Female Fitness in Clarkia xantiana.

Selection on floral traits in hermaphroditic plants is determined by both male and female reproductive success. However, predictions regarding floral trait and mating system evolution are often based solely on female fitness. Selection via male fitness has the potential to affect the outcomes of floral evolution. In this study, we used paternity analysis to assess individual selfing rates and selection on floral traits via male and female fitness in an experimental population of Clarkia xantiana where pollen limitation of seed set was strong. We detected selection through both female and male fitness with reinforcing or noninterfering patterns of selection through the two sex functions. For female fitness, selection favored reduced herkogamy and protandry, traits that promote increased autonomous selfing. For male fitness, selection on petal area was disruptive, with higher trait values conferring greater pollinator attraction and outcross siring success and smaller trait values leading to higher selfed siring success. Combining both female and male fitness, selection on petal area and protandry was disruptive because intermediate phenotypes were less successful as both males and females. Finally, functional relationships among male and female fertility components indicated that selfing resulted in seed discounting and pollen discounting. Under these functional relationships, the evolutionarily stable selfing rate can be intermediate or predominantly selfing or outcrossing, depending on the segregating load of deleterious mutations.

Mating system divergence affects the distribution of sequence diversity within and among populations of recently diverged subspecies of Clarkia xantiana (Onagraceae).

PREMISE OF THE STUDY: The population biology of outcrossing and self-fertilizing taxa is thought to differ because of the advantage that selfers have in colonizing unoccupied sites where mates and pollinators may be limiting (Baker's Law). This reduced tendency for outcrossers to colonize new sites, along with their greater dependence on pollinators to disperse pollen, has the potential to differently influence the genetic diversity and structure of outcrossing and selfing populations. METHODS: We conducted a comparative population genetic study of two sister outcrossing and selfing subspecies of Clarkia xantiana that have very recently diverged. We used DNA sequence variation (>40 kb from eight nuclear loci) from large samples of individuals from 14 populations to assess geographic patterns of genetic diversity and make inferences about the demographic and colonization histories of each subspecies. KEY RESULTS: We show that sequence variation is strongly reduced across all selfing populations. The demographic history of selfing populations exhibits recent colonization bottlenecks, whereas such bottlenecks are rarely observed for the outcrosser. The greater effect of genetic drift in the selfer has resulted in strong population genetic structure, but with no pattern of isolation by distance. By contrast, the stronger effect of gene flow in the outcrosser has resulted in considerably less structure, but a significant pattern of isolation by distance. CONCLUSIONS: Taken together, our results suggest that selfing populations are not at migration-drift equilibrium, are affected by strong episodes of genetic drift during colonization, and experience little or no subsequent gene flow from other populations after those founder events.

Floral and mating system divergence in secondary sympatry: testing an alternative hypothesis to reinforcement in Clarkia.

BACKGROUND AND AIMS: Reproductive character displacement (RCD) is often an important signature of reinforcement when partially cross-compatible taxa meet in secondary sympatry. In this study, floral evolution is examined during the Holocene range expansion of Clarkia xantiana subsp. parviflora from eastern Pleistocene refugia to a western zone of sympatry with its sister taxon, subsp. xantiana. Floral divergence between the two taxa is greater in sympatry than allopatry. The goal was to test an alternative hypothesis to reinforcement - that floral divergence of sympatric genotypes is simply a by-product of adaptation to pollination environments that differ between the allopatric and sympatric portions of the subspecies' range. METHODS: Floral trait data from two common garden studies were used to examine floral divergence between sympatric and allopatric regions and among phylogeographically defined lineages. In natural populations of C. x. parviflora, the magnitude of pollen limitation and reproductive assurance were quantified across its west-to-east range. Potted sympatric and allopatric genotypes were also reciprocally translocated between geographical regions to distinguish between the effects of floral phenotype versus contrasting pollinator environments on reproductive ecology. KEY RESULTS: Sympatric populations are considerably smaller flowered with reduced herkogamy. Pollen limitation and the reproductive assurance value of selfing are greater in sympatric than in allopatric populations. Most significantly, reciprocal translocation experiments showed these differences in reproductive ecology cannot be attributed to contrasting pollinator environments between the sympatric and allopatric regions, but instead reflect the effects of flower size on pollinator attraction. CONCLUSIONS: Floral evolution occurred during the westward range expansion of parviflora, particularly in the zone of sympatry with xantiana. No evidence was found that strongly reduced flower size in sympatric parviflora (and RCD between parviflora and xantiana) is due to adaptation to limited pollinator availability. Rather, floral divergence appears to have been driven by other factors, such as interactions with congenerics in secondary sympatry.

The value of long-term ecological research for evolutionary insights.

Scientists must have an integrative understanding of ecology and evolution across spatial and temporal scales to predict how species will respond to global change. Although comprehensively investigating these processes in nature is challenging, the infrastructure and data from long-term ecological research networks can support cross-disciplinary investigations. We propose using these networks to advance our understanding of fundamental evolutionary processes and responses to global change. For ecologists, we outline how long-term ecological experiments can be expanded for evolutionary inquiry, and for evolutionary biologists, we illustrate how observed long-term ecological patterns may motivate new evolutionary questions. We advocate for collaborative, multi-site investigations and discuss barriers to conducting evolutionary work at network sites. Ultimately, these networks offer valuable information and opportunities to improve predictions of species' responses to global change.