Target Enrichment and Extensive Population Sampling Help Untangle the Recent, Rapid Radiation of Oenothera Sect. Calylophus.

Oenothera sect. Calylophus is a North American group of 13 recognized taxa in the evening primrose family (Onagraceae) with an evolutionary history that may include independent origins of bee pollination, edaphic endemism, and permanent translocation heterozygosity. Like other groups that radiated relatively recently and rapidly, taxon boundaries within Oenothera sect. Calylophus have remained challenging to circumscribe. In this study, we used target enrichment, flanking noncoding regions, gene tree/species tree methods, tests for gene flow modified for target-enrichment data, and morphometric analysis to reconstruct phylogenetic hypotheses, evaluate current taxon circumscriptions, and examine character evolution in Oenothera sect. Calylophus. Because sect. Calylophus comprises a clade with a relatively restricted geographic range, we were able to extensively sample across the range of geographic, edaphic, and morphological diversity in the group. We found that the combination of exons and flanking noncoding regions led to improved support for species relationships. We reconstructed potential hybrid origins of some accessions and note that if processes such as hybridization are not taken into account, the number of inferred evolutionary transitions may be artificially inflated. We recovered strong evidence for multiple evolutionary origins of bee pollination from ancestral hawkmoth pollination, edaphic specialization on gypsum, and permanent translocation heterozygosity. This study applies newly emerging techniques alongside dense infraspecific sampling and morphological analyses to effectively reconstruct the recalcitrant history of a rapid radiation. [Gypsum endemism; Oenothera sect. Calylophus; Onagraceae; phylogenomics; pollinator shift; recent radiation; target enrichment.].

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

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

Genotype accounts for intraspecific variation in the timing and duration of multiple, sequential life-cycle events in a willow species.

PREMISE: Phenological variation among individuals within populations is common and has a variety of ecological and evolutionary consequences, including forming the basis for population-level responses to environmental change. Although the timing of life-cycle events has genetic underpinnings, whether intraspecific variation in the duration of life-cycle events reflects genetic differences among individuals is poorly understood. METHODS: We used a common garden experiment with 10 genotypes of Salix hookeriana (coastal willow) from northern California, United States to investigate the extent to which genetic variation explains intraspecific variation in the timing and duration of multiple, sequential life-cycle events: flowering, leaf budbreak, leaf expansion, fruiting, and fall leaf coloration. We used seven clones of each genotype, for a total of 70 individual trees. RESULTS: Genotype affected each sequential life-cycle event independently and explained on average 62% of the variation in the timing and duration of vegetative and reproductive life-cycle events. All events were significantly heritable. A single genotype tended to be "early" or "late" across life-cycle events, but for event durations, there was no consistent response within genotypes. CONCLUSIONS: This research demonstrates that genetic variation can be a major component underlying intraspecific variation in the timing and duration of life-cycle events. It is often assumed that the environment affects durations, but we show that genetic factors also play a role. Because the timing and duration of events are independent of one another, our results suggest that the effects of environmental change on one event will not necessarily cascade to subsequent events.

Differential gene expression associated with a floral scent polymorphism in the evening primrose Oenothera harringtonii (Onagraceae).

BACKGROUND: Plant volatiles play an important role in both plant-pollinator and plant-herbivore interactions. Intraspecific polymorphisms in volatile production are ubiquitous, but studies that explore underlying differential gene expression are rare. Oenothera harringtonii populations are polymorphic in floral emission of the monoterpene (R)-(-)-linalool; some plants emit (R)-(-)-linalool (linalool+ plants) while others do not (linalool- plants). However, the genes associated with differential production of this floral volatile in Oenothera are unknown. We used RNA-Seq to broadly characterize differential gene expression involved in (R)-(-)-linalool biosynthesis. To identify genes that may be associated with the polymorphism for this trait, we used RNA-Seq to compare gene expression in six different Oenothera harringtonii tissues from each of three linalool+ and linalool- plants. RESULTS: Three clusters of differentially expressed genes were enriched for terpene synthase activity: two were characterized by tissue-specific upregulation and one by upregulation only in plants with flowers that produce (R)-(-)-linalool. A molecular phylogeny of all terpene synthases identified two putative (R)-(-)-linalool synthase transcripts in Oenothera harringtonii, a single allele of which is found exclusively in linalool+ plants. CONCLUSIONS: By using a naturally occurring polymorphism and comparing different tissues, we were able to identify candidate genes putatively involved in the biosynthesis of (R)-(-)-linalool. Expression of these genes in linalool- plants, while low, suggests a regulatory polymorphism, rather than a population-specific loss-of-function allele. Additional terpene biosynthesis-related genes that are up-regulated in plants that emit (R)-(-)-linalool may be associated with herbivore defense, suggesting a potential economy of scale between plant reproduction and defense.

Evolution of selfing syndrome and its influence on genetic diversity and inbreeding: A range-wide study in Oenothera primiveris.

PREMISE: To avoid inbreeding depression, plants have evolved diverse breeding systems to favor outcrossing, such as self-incompatibility. However, changes in biotic and abiotic conditions can result in selective pressures that lead to a breakdown in self-incompatibility. The shift to increased selfing is commonly associated with reduced floral features, lower attractiveness to pollinators, and increased inbreeding. We tested the hypothesis that the loss of self-incompatibility, a shift to self-fertilization (autogamy), and concomitant evolution of the selfing syndrome (reduction in floral traits associated with cross-fertilization) will lead to increased inbreeding and population differentiation in Oenothera primiveris. Across its range, this species exhibits a shift in its breeding system and floral traits from a self-incompatible population with large flowers to self-compatible populations with smaller flowers. METHODS: We conducted a breeding system assessment, evaluated floral traits in the field and under controlled conditions, and measured population genetic parameters using RADseq data. RESULTS: Our results reveal a bimodal transition to the selfing syndrome from the west to the east of the range of O. primiveris. This shift includes variation in the breeding system and the mating system, a reduction in floral traits (flower diameter, herkogamy, and scent production), a shift to greater autogamy, reduced genetic diversity, and increased inbreeding. CONCLUSIONS: The observed variation highlights the importance of range-wide studies to understand breeding system variation and the evolution of the selfing syndrome within populations and species.

Incomplete reproductive isolation and low genetic differentiation despite floral divergence across varying geographic scales in Castilleja.

PREMISE: Divergence depends on the strength of selection and frequency of gene flow between taxa, while reproductive isolation relies on mating barriers and geographic distance. Less is known about how these processes interact at early stages of speciation. Here, we compared population-level differentiation in floral phenotype and genetic sequence variation among recently diverged Castilleja to explore patterns of diversification under different scenarios of reproductive isolation. METHODS: Using target enrichment enabled by the Angiosperms353 probe set, we assessed genetic distance among 50 populations of four Castilleja species. We investigated whether patterns of genetic divergence are explained by floral trait variation or geographic distance in two focal groups: the widespread C. sessiliflora and the more restricted C. purpurea species complex. RESULTS: We document that C. sessiliflora and the C. purpurea complex are characterized by high diversity in floral color across varying geographic scales. Despite phenotypic divergence, groups were not well supported in phylogenetic analyses, and little genetic differentiation was found across targeted Angiosperms353 loci. Nonetheless, a principal coordinate analysis of single nucleotide polymorphisms revealed differentiation within C. sessiliflora across floral morphs and geography and less differentiation among species of the C. purpurea complex. CONCLUSIONS: Patterns of genetic distance in C. sessiliflora suggest species cohesion maintained over long distances despite variation in floral traits. In the C. purpurea complex, divergence in floral color across narrow geographic clines may be driven by recent selection on floral color. These contrasting patterns of floral and genetic differentiation reveal that divergence can arise via multiple eco-evolutionary paths.

Applying the zoo model to conservation of threatened exceptional plant species.

Maintaining a living plant collection is the most common method of ex situ conservation for plant species that cannot be seed banked (i.e., exceptional species). Viability of living collections, and their value for future conservation efforts, can be limited without coordinated efforts to track and manage individuals across institutions. Using a pedigree-focused approach, the zoological community has established an inter-institutional infrastructure to support long-term viability of captive animal populations. We assessed the ability of this coordinated metacollection infrastructure to support the conservation of 4 plant species curated in living collections at multiple botanic gardens around the world. Limitations in current practices include the inability to compile, share, and analyze plant collections data at the individual level, as well as difficulty in tracking original provenance of ex situ material. The coordinated metacollection framework used by zoos can be adopted by the botanical community to improve conservation outcomes by minimizing the loss of genetic diversity in collections. We suggest actions to improve ex situ conservation of exceptional plant species, including developing a central database to aggregate data and track unique individuals of priority threatened species among institutions and adapting a pedigree-based population management tool that incorporates life-history aspects unique to plants. If approached collaboratively across regional, national, and global scales, these actions could transform ex situ conservation of threatened plant species.

Aplicación del Modelo Zoológico a la Conservación de Especies Excepcionales de Plantas Amenazadas Resumen El mantenimiento de una colección de plantas vivas es el método más común para de conservación ex situ para especies de plantas que no pueden almacenarse en bancos de semillas (i. e., especies excepcionales). La viabilidad de las colecciones vivientes, junto con el valor que representan para los futuros esfuerzo de conservación, puede estar limitada si no existen esfuerzos coordinados para rastrear y manejar a los individuos entre las instituciones. Mediante una estrategia enfocada en el linaje, la comunidad de zoológicos ha establecido una infraestructura interinstitucional que respalda la viabilidad a largo plazo de las poblaciones de animales en cautiverio. Evaluamos la habilidad de esta infraestructura coordinada de metacolecciones para apoyar en la conservación de cuatro especies de plantas curadas en colecciones vivientes en varios jardines botánicos de todo el mundo. Las limitaciones de las prácticas contemporáneas incluyen la incapacidad de recopilar, compartir y analizar los datos de las colecciones de plantas a nivel individual, así como la dificultad de rastrear la procedencia original del material ex situ. El marco de trabajo de metacolecciones coordinadas que utilizan los zoológicos puede ser adoptado por la comunidad botánica para mejorar los resultados de conservación al minimizar la pérdida de la diversidad genética que ocurre en las colecciones. Sugerimos acciones que aumenten la conservación ex situ de las especies excepcionales de plantas. Estas acciones incluyen el desarrollo de una base de datos central para acumular datos y rastrear entre las instituciones a los individuos únicos de las especies amenazadas prioritarias y la adaptación de una herramienta de manejo poblacional basada en el linaje que incorpore los aspectos únicos de la historia de vida de las plantas. Si estas acciones se plantean colaborativamente a escala regional, nacional y global, podrían transformar la conservación ex situ de las especies amenazadas de plantas.

Pollinator identity and spatial isolation influence multiple paternity in an annual plant.

The occurrence and extent of multiple paternity is an important component of variation in plant mating dynamics. However, links between pollinator activity and multiple paternity are generally lacking, especially for plant species that attract functionally diverse floral visitors. In this study, we separated the influence of two functionally distinct floral visitors (hawkmoths and solitary bees) and characterized their impacts on multiple paternity in a self-incompatible, annual forb, Oenothera harringtonii (Onagraceae). We also situated pollinator-mediated effects in a spatial context by linking variation in multiple paternity to variation in plant spatial isolation. We documented pronounced differences in the number of paternal sires as function of pollinator identity: on average, the primary pollinator (hawkmoths) facilitated mating with nearly twice as many pollen donors relative to the secondary pollinator (solitary bees). This effect was consistent for both isolated and nonisolated individuals, but spatial isolation imposed pronounced reductions on multiple paternity regardless of pollinator identity. Considering that pollinator abundance and pollen dispersal distance did not vary significantly with pollinator identity, we attribute variation in realized mating dynamics primarily to differences in pollinator morphology and behaviour as opposed to pollinator abundance or mating incompatibility arising from underlying spatial genetic structure. Our findings demonstrate that functionally distinct pollinators can have strongly divergent effects on polyandry in plants and further suggest that both pollinator identity and spatial heterogeneity have important roles in plant mating dynamics.

Land-use change has no detectable effect on reproduction of a disturbance-adapted, hawkmoth-pollinated plant species.

PREMISE OF THE STUDY: Land-use change is cited as a primary driver of global biodiversity loss, with myriad consequences for species, populations, and ecosystems. However, few studies have examined its impact on species interactions, particularly pollination. Furthermore, when the effects of land-use change on pollination have been studied, the focus has largely been on species pollinated by diurnal pollinators, namely, bees and butterflies. Here, we focus on Oenothera harringtonii, a night-flowering, disturbance-adapted species that has experienced a range-wide gradient of land-use change. We tested the hypothesis that the negative impacts of land-use change are mitigated by long-distance pollination. METHODS: Our study included both temporal (4 yr) and spatial (19 populations range-wide, and 1, 2, and 5 km from the population center) data, providing a comprehensive understanding of the role of land-use change on pollination biology and reproduction. KEY RESULTS: We first confirmed that O. harringtonii is self-incompatible and reliant on pollinators for reproduction. We then showed that hawkmoths (primarily Hyles lineata) are highly reliable and effective pollinators in both space and time. Unlike other studies, we did not detect an effect of population size, increased isolation, or a reduction in suitable habitat in areas with evidence of land-use change on pollination (visitation, pollen removal and deposition). Furthermore, the proportion of suitable habitat and other fragmentation metrics examined were not associated with population size or density in this plant species. CONCLUSIONS: We conclude that nocturnal pollination of Oenothera harringtonii via hawkmoths is robust to the negative impacts of land-use change.

Local topography shapes fine-scale spatial genetic structure in the Arkansas Valley evening primrose, Oenothera harringtonii (Onagraceae).

Identifying factors that shape the spatial distribution of genetic variation is crucial to understanding many population- and landscape-level processes. In this study, we explore fine-scale spatial genetic structure in Oenothera harringtonii (Onagraceae), an insect-pollinated, gravity-dispersed herb endemic to the grasslands of south-central and southeastern Colorado, USA. We genotyped 315 individuals with 11 microsatellite markers and utilized a combination of spatial autocorrelation analyses and landscape genetic models to relate life history traits and landscape features to dispersal processes. Spatial genetic structure was consistent with theoretical expectations of isolation by distance, but this pattern was weak (Sp = 0.00374). Anisotropic analyses indicated that spatial genetic structure was markedly directional, in this case consistent with increased dispersal along prominent slopes. Landscape genetic models subsequently confirmed that spatial genetic variation was significantly influenced by local topographic heterogeneity, specifically that geographic distance, elevation and aspect were important predictors of spatial genetic structure. Among these variables, geographic distance was ~68% more important than elevation in describing spatial genetic variation, and elevation was ~42% more important than aspect after removing the effect of geographic distance. From these results, we infer a mechanism of hydrochorous seed dispersal along major drainages aided by seasonal monsoon rains. Our findings suggest that landscape features may shape microevolutionary processes at much finer spatial scales than typically considered, and stress the importance of considering how particular dispersal vectors are influenced by their environmental context.

Conservation genomics assessment of Tharp's bluestar (Amsonia tharpii) with comparisons to widespread (A. longilora) and narrowly endemic (A. fugatei) congeners.

Land-use change and habitat fragmentation are threats to biodiversity. The decrease in available habitat, increase in isolation, and mating within populations can lead to elevated inbreeding, lower genetic diversity, and poor fitness. Here we investigate the genetics of two rare and threatened plant species, Amsonia tharpii and A. fugatei, and we compare them to a widespread congener A. longiflora. We also report the first phylogenetic study of the genus Amsonia (Apocynaceae), including 10 of the 17 taxa and multiple sampling locations, to understand species relationships. We used a double digest restriction-site associated DNA sequencing (ddRADseq) approach to investigate the genetic diversity and gene flow of each species and to create a maximum likelihood phylogeny. The ddRADseq data was mapped to a reference genome to separate out the chloroplast and nuclear markers for population genetic analysis. Our results show that genetic diversity and inbreeding were low across all three species. The chloroplast and nuclear dataset in A. tharpii were highly structured, whereas they showed no structure for A. fugatei, while A. longiflora lacked structure for nuclear data but not chloroplast. Phylogenetic results revealed that A. tharpii is distinct and sister to A. fugatei, and together they are distantly related to A. longiflora. Our results demonstrated that evolutionary history and contemporary ecological processes largely influences genetic diversity within Amsonia. Interestingly, we show that in A. tharpii there was significant structure despite being pollinated by large, bodied hawkmoths that are known to be able to carry pollen long distances, suggesting that other factors are contributing to the structure observed among A. tharpii populations. Conservation efforts should focus on protecting all of the A. tharpii populations, as they contain unique genetic diversity, and a protection plan for A. fugatei needs to be established due to its limited distribution.

Adaptive generalization in pollination systems: Hawkmoths increase fitness to long-tubed flowers, but secondary pollinators remain important.

Selection on floral traits by animal pollinators is important in the evolution of flowering plants, yet whether floral divergence requires specialized pollination remains uncertain. Longer floral tubes, a trait associated with long-tongued pollinators, can also exclude other pollinators from accessing rewards, a potential mechanism for specialization. Across most of its range, Castilleja sessiliflora displays much longer corollas than most Castilleja species, though tube length varies geographically and correlates partially with hawkmoth visitation. To assess whether long corolla tubes reflect adaptation to hawkmoth pollinators, we performed a day/night pollinator exclusion experiment in nine natural populations that varied in corolla length across the range of C. sessiliflora and short-tubed members of the parapatric C. purpurea complex. We compared the fitness contributions of nocturnal and diurnal visitors, revealing that long-tubed populations visited predominantly by hawkmoths experienced greater fruit set at night, in contrast with short-tubed populations or those visited mainly by diurnal pollinators. Next, leveraging a range-wide multiyear dataset of pollinator visitation to these species, we identify that hawkmoth visitation is associated with increased fitness in long-tubed populations overall, and that long tubes are associated with less diverse visitor assemblages. Thus, long corollas represent an adaptation to hawkmoth pollination at the exclusion of diverse pollinators. Nonetheless, while hawkmoths were scarce in the northern range, secondary diurnal pollinators contributed to fruit set across the range, providing reproductive assurance despite possible trait mismatch. This study illustrates adaptive generalization in pollination systems and that floral divergence may proceed along a continuum of generalized and specialized pollinator interactions.

A phylogeny of the evening primrose family (Onagraceae) using a target enrichment approach with 303 nuclear loci.

BACKGROUND: The evening primrose family (Onagraceae) includes 664 species (803 taxa) with a center of diversity in the Americas, especially western North America. Ongoing research in Onagraceae includes exploring striking variation in floral morphology, scent composition, and breeding system, as well as the role of these traits in driving diversity among plants and their interacting pollinators and herbivores. However, these efforts are limited by the lack of a comprehensive, well-resolved phylogeny. Previous phylogenetic studies based on a few loci strongly support the monophyly of the family and the sister relationship of the two largest tribes but fail to resolve several key relationships. RESULTS: We used a target enrichment approach to reconstruct the phylogeny of Onagraceae using 303 highly conserved, low-copy nuclear loci. We present a phylogeny for Onagraceae with 169 individuals representing 152 taxa sampled across the family, including extensive sampling within the largest tribe, Onagreae. Deep splits within the family are strongly supported, whereas relationships among closely related genera and species are characterized by extensive conflict among individual gene trees. CONCLUSIONS: This phylogenetic resource will augment current research projects focused throughout the family in genomics, ecology, coevolutionary dynamics, biogeography, and the evolution of characters driving diversification in the family.

Microsatellite primers in Oenothera harringtonii (Onagraceae), an annual endemic to the shortgrass prairie of Colorado.

PREMISE OF THE STUDY: Microsatellite markers were developed in the annual herb, Oenothera harringtonii, to investigate patterns of genetic diversity, gene flow, and parentage within and among populations of this Colorado endemic. METHODS AND RESULTS: Ten polymorphic loci were identified in O. harringtonii and tested in four populations sampled across the range of the species. These loci contained trinucleotide repeats with 7-29 alleles per locus. Nine of the 10 loci also amplified in O. caespitosa subsp. macroglottis, O. caespitosa subsp. marginata, and O. caespitosa subsp. navajoensis. In addition, we optimized three markers developed for O. biennis and provide reports of their effectiveness in all four taxa. CONCLUSIONS: These results indicate the utility of these markers in O. harringtonii for future studies of genetic structure, gene flow, and parentage as well as their applicability in other members of the O. caespitosa species complex.

Supporting long-term sustainability of ex situ collections using a pedigree-based population management approach.

Premise: Living collections maintained for generations are at risk of diversity loss, inbreeding, and adaptation to cultivation. To address these concerns, the zoo community uses pedigrees to track individuals and implement crosses that maximize founder contributions and minimize inbreeding. Using a pedigree management approach, we demonstrate how conducting strategic crosses can minimize genetic issues that have arisen under current practices. Methods: We performed crosses between collections and compared progeny fitness, including plant performance and reproductive health. We genotyped the progeny and parental accessions to measure changes in diversity and relatedness within and between accessions. Results: The mean relatedness values among individuals within each accession suggest they are full siblings, demonstrating that there was high inbreeding and low diversity within accessions, although less so among accessions. Progeny from the wider crosses had increased genetic diversity and were larger and more fertile, while self-pollinated accessions were smaller and less fertile. Discussion: Institutions that hold exceptional species should consider how diversity is maintained within their collections. Implementing a pedigree-based approach to managing plant reproduction ex situ will slow the inevitable loss of genetic diversity and, in turn, result in healthier collections.

Influences of landscape and pollinators on population genetic structure: examples from three Penstemon (Plantaginaceae) species in the Great Basin.

PREMISE OF THE STUDY: Despite rapid growth in the field of landscape genetics, our understanding of how landscape features interact with life history traits to influence population genetic structure in plant species remains limited. Here, we identify population genetic divergence in three species of Penstemon (Plantaginaceae) similarly distributed throughout the Great Basin region of the western United States but with different pollination syndromes (bee and hummingbird). The Great Basin's mountainous landscape provides an ideal setting to compare the interaction of landscape and dispersal ability in isolating populations of different species. METHODS: We used eight highly polymorphic microsatellite loci to identify neutral population genetic structure between populations within and among mountain ranges for eight populations of P. deustus, 10 populations of P. pachyphyllus, and 10 populations of P. rostriflorus. We applied traditional population genetics approaches as well as spatial and landscape genetics approaches to infer genetic structure and discontinuities among populations. KEY RESULTS: All three species had significant genetic structure and exhibited isolation by distance, ranging from high structure and low inferred gene flow in the bee-pollinated species P. deustus (F(ST) = 0.1330, R(ST) = 0.4076, seven genetic clusters identified) and P. pachyphyllus (F(ST) = 0.1896, R(ST) = 0.2531, four genetic clusters identified) to much lower structure and higher inferred gene flow in the hummingbird-pollinated P. rostriflorus (F(ST) = 0.0638, R(ST) = 0.1116, three genetic clusters identified). CONCLUSIONS: These three Penstemon species have significant yet strikingly different patterns of population genetic structure, findings consistent with different interactions between landscape features and the dispersal capabilities of their pollinators.

Reproductive trade-offs maintain bract color polymorphism in Scarlet Indian paintbrush (Castilleja coccinea).

Populations of scarlet Indian paintbrush (Castilleja coccinea) in the Midwestern United States exhibit a bract color polymorphism, with each population having predominantly yellow or scarlet bracts. We investigated a possible mechanism for this maintenance of bract color polymorphism in C. coccinea by conducting hand-pollination experiments in two nearby populations, one predominantly yellow and one predominantly scarlet. The hand-pollination treatments were either self-pollination or cross pollination using pollen from within and between populations. Both color morphs were used as pollen donors for the within and between crosses. We found that both color morphs of C. coccinea were self-compatible. When the scarlet morph was the maternal plant it had higher seed set. When pollinators were excluded, the yellow morph outperformed the scarlet morph in fruit set and seed set. The apparent trade-offs between a higher reproductive output in the scarlet morph and a reproductive assurance advantage in the yellow morph may explain the maintenance of the polymorphism in C. coccinea. While many previous studies have provided evidence for pollinator preference playing a role in floral color polymorphism, the results of the current study indicate that reproductive assurance, which would be important for fluctuations in pollinator abundance or colonizing new areas, may act as a selective agent to maintain such polymorphisms.

Characterization of microsatellite loci in Brighamia insignis and transferability to other genera in the Hawai'ian lobelioid group.

PREMISE: Microsatellite markers were developed to measure genetic diversity and relatedness of ex situ collections of Brighamia insignis (Campanulaceae). METHODS AND RESULTS: Potential microsatellite markers were identified from two sources; 28 were developed for B. insignis and an additional 12 markers from a previously published study of Lobelia villosa. Primer pairs were tested on 30 individuals of B. insignis and 24 individuals of B. rockii to provide measures of genetic diversity and inbreeding. We assessed cross-species amplification in an additional 13 taxa that represented all six genera within the Hawai'ian lobelioid group to determine the broader applicability of the markers. CONCLUSIONS: Results indicate that these primers will provide useful estimates of genetic diversity and relatedness of ex situ collections of both Brighamia species. In addition, we have also demonstrated the widespread applicability of these markers for use in population genetic studies of several species within the Hawai'ian lobelioid group.

Microsatellites for Oenothera gayleana and O. hartwegii subsp. filifolia (Onagraceae), and their utility in section Calylophus.

PREMISE OF THE STUDY: Eleven nuclear and four plastid microsatellite markers were screened for two gypsum endemic species, Oenothera gayleana and O. hartwegii subsp. filifolia, and tested for cross-amplification in the remaining 11 taxa within Oenothera sect. Calylophus (Onagraceae). METHODS AND RESULTS: Microsatellite markers were tested in two to three populations spanning the ranges of both O. gayleana and O. hartwegii subsp. filifolia. The nuclear microsatellite loci consisted of both di- and trinucleotide repeats with one to 17 alleles per population. Several loci showed significant deviation from Hardy-Weinberg equilibrium, which may be evidence of chromosomal rings. The plastid microsatellite markers identified one to seven haplotypes per population. The transferability of these markers was confirmed in all 11 taxa within Oenothera sect. Calylophus. CONCLUSIONS: The microsatellite loci characterized here are the first developed and tested in Oenothera sect. Calylophus. These markers will be used to assess whether pollinator foraging distance influences population genetic parameters in predictable ways.