Evolutionary breakdown of distyly to homostyly is accompanied by reductions of floral scent in Primula oreodoxa

Reproductive traits that function in pollinator attraction may be reduced or lost during evolutionary transitions from outcrossing to selfing. Although floral scent plays an important role in attracting pollinators in outcrossing species, few studies have investigated associations between floral scent variation and intraspecific mating system transitions. The breakdown of distyly to homostyly represents a classic example of a shift from outcrossing to selfing and provides an opportunity to test whether floral fragrances have become reduced and/or changed in composition with increased selfing. Here, we evaluate this hypothesis by quantifying floral volatiles using gas chromatography-mass spectrometry in two distylous and four homostylous populations of Primula oreodoxa Franchet, a perennial herb from SW China. Our analysis revealed significant variation of volatile organic compounds (VOCs) among populations of P. oreodoxa. Although there was no difference in VOCs between floral morphs in distylous populations as predicted, we detected a substantial reduction in VOC emissions and the average number of scent compounds in homostylous compared with distylous populations. A total of 12 compounds, mainly monoterpenoids and sesquiterpenoids, distinguished homostylous and distylous morphs; of these, (E)-β-ocimene was the most important in contributing to the difference in volatiles, with significantly lower emissions in homostyles. Our findings support the hypothesis that the transition from outcrossing to selfing is accompanied by the loss of floral volatiles. The modification to floral fragrances in P. oreodoxa associated with mating system change might occur because high selfing rates in homostylous populations result in relaxed selection for floral attractiveness.     

EVOLUTIONARY HISTORY OF THE MATING SYSTEM IN AMSINCKIA (BORAGINACEAE)

A survey of restriction site variation in the chloroplast genome of the annual plant genus Amsinckia, together with estimation of outcrossing rates, was conducted to analyze the evolutionary history of the mating system. Species, and in some cases populations within species, differ markedly in their mating system. Five taxa are distylous and predominantly outcrossing, or show mixed mating systems, while the remaining taxa are homostylous and predominantly self-fertilizing. Reconstruction of the molecular phylogeny of the group places different distylous and homostylous taxa at four separate branch tips. When distyly is treated as ancestral in the group, or when the loss of distyly is assumed to be more common than its gain, the results of the phylogenetic analysis support the hypothesis that the self-fertilizing taxa are of recent origin from outcrossing relatives. These findings are discussed with respect to theory for the evolution and breakdown of distyly and the probability of extinction of selfing lineages.     

Characterization of 24 microsatellite markers in Primula chungensis (Primulaceae), a distylous homostylous species,using MiSeq sequencing

Primula chungensis is a species with considerable floral and mating-system variation, including distylous (outcrossing), homostylous (selfing) and mixed populations that contain both outcrossing and selfing forms. We isolated 24 microsatellite markers from P.chungensis using Illumina MiSeq sequencing. Polymorphism and genetic diversity were then measured based on a sample of 24 individuals from a natural population in southern Tibet. All loci were polymorphic with the number of alleles per locus ranging from 2 to4. The observed and expected heterozygosity ranged from 0 to 1 and 0.219 to 0.708, respectively. The microsatellite markers we have identified will serve as valuable tools for the investigation of the population genetic structure and phylogeography of P.chungensis and will inform models of the evolutionary history of mating systems in the species.     

Characterization of 24 microsatellite markers in Primula chungensis(Primulaceae),a distylous-homostylous species,using MiSeq sequencing

Primula chungensis is a species with considerable floral and mating-system variation,including distylous(outcrossing),homostylous(selfing) and mixed populations that contain both outcrossing and selfing forms.We isolated 24 microsatellite markers from P.chungensis using Illumina Mi Seq sequencing.Polymorphism and genetic diversity were then measured based on a sample of 24 individuals from a natural population in southern Tibet.All loci were polymorphic with the number of alleles per locus ranging from 2 to 4.The observed and expected heterozygosity ranged from 0 to 1 and 0.219 to 0.708,respectively.The microsatellite markers we have identified will serve as valuable tools for the investigation of the population genetic structure and phylogeography of P.chungensis and will inform models of the evolutionary history of mating systems in the species.     

VARIATION AND EVOLUTION OF BREEDING SYSTEMS IN THE TURNERA ULMIFOLIA L. COMPLEX (TURNERACEAE)

The evolutionary and functional relationships among breeding systems and floral morphology were investigated in the Turnera ulmifolia complex. Predictions of a model of breeding system evolution among distylous and homostylous varieties were tested. Chromosome counts of 73 accessions revealed an association between breeding system and chromosome number. Diploid and tetraploid populations of five taxonomic varieties are distylous and self-incompatible, whereas hexaploid populations of three varieties are homostylous and self-compatible. The latter occur at different margins of the geographical range of the complex. Crossing studies and analyses of pollen and ovule fertility in F₁'s revealed that the three homostylous varieties are intersterile. To test the prediction that, homostylous varieties are long homostyles that have originated by crossing over within the distyly supergene, a crossing program was undertaken among distylous and homostylous plants. Residual incompatibility was observed in styles and pollen of each homostylous variety with patterns consistent with predictions of the cross-over model. The intersterility of hexaploid varieties suggests that long homostyly has arisen on at least three occasions in the complex by recombination within the supergene controlling distyly. Deviation from expected compatibility behavior occurs in populations of var. angustifolia that have the longest styles. These phenotypes displayed the greatest separation between anthers and stigmas (herkogamy) and set little seed in crosses with long- or short-styled plants. This suggests that they are derived from long homostyles with shorter length styles. It is proposed that selection for increased outcrossing has favored the evolution of herkogamy in long homostyles. Estimates of outcrossing rate in a distylous population using allozyme markers confirmed that dimorphic incompatibility enforces complete outcrossing. Significant genetic variation for floral traits likely to influence the mating system, such as stigma-anther separation, occurs within and among homostylous populations of var. angustifolia on Jamaica. Estimates of the mating system of families from a population with varying degrees of stigma-anther separation, using five isozyme loci, were heterogeneous and ranged from t = 0.04–0.79. Families exhibiting the largest mean stigma-anther separation have higher outcrossing rates than those with little separation.     

Limited phenological and pollinator-mediated isolation among selfing and outcrossing Arabidopsis lyrata populations

Transitions from outcrossing to selfing have been a frequent evolutionary shift in plants and clearly play a role in species divergence. However, many questions remain about the initial mechanistic basis of reproductive isolation during the evolution of selfing. For instance, how important are pre-zygotic pre-pollination mechanisms (e.g. changes in phenology and pollinator visitation) in maintaining reproductive isolation between newly arisen selfing populations and their outcrossing ancestors? To test whether changes in phenology and pollinator visitation isolate selfing populations of Arabidopsis lyrata from outcrossing populations, we conducted a common garden experiment with plants from selfing and outcrossing populations as well as their between-population hybrids. Specifically, we asked whether there was isolation between outcrossing and selfing plants and their between-population hybrids through differences in (1) the timing or intensity of flowering; and/or (2) pollinator visitation. We found that phenology largely overlapped between plants from outcrossing and selfing populations. There were also no differences in pollinator preference related to mating system. Additionally, pollinators preferred to visit flowers on the same plant rather than exploring nearby plants, creating a large opportunity for self-fertilization. Overall, this suggests that pre-zygotic pre-pollination mechanisms do not strongly reproductively isolate plants from selfing and outcrossing populations of Arabidopsis lyrata.     

Pedigree-Free Estimates of Heritability in the Wild: Promising Prospects for Selfing Populations

Estimating the genetic variance available for traits informs us about a population’s ability to evolve in response to novel selective challenges. In selfing species, theory predicts a loss of genetic diversity that could lead to an evolutionary dead-end, but empirical support remains scarce. Genetic variability in a trait is estimated by correlating the phenotypic resemblance with the proportion of the genome that two relatives share identical by descent (‘realized relatedness’). The latter is traditionally predicted from pedigrees (Φ_A: expected value) but can also be estimated using molecular markers (average number of alleles shared). Nevertheless, evolutionary biologists, unlike animal breeders, remain cautious about using marker-based relatedness coefficients to study complex phenotypic traits in populations. In this paper, we review published results comparing five different pedigree-free methods and use simulations to test individual-based models (hereafter called animal models) using marker-based relatedness coefficients, with a special focus on the influence of mating systems. Our literature review confirms that Ritland’s regression method is unreliable, but suggests that animal models with marker-based estimates of relatedness and genomic selection are promising and that more testing is required. Our simulations show that using molecular markers instead of pedigrees in animal models seriously worsens the estimation of heritability in outcrossing populations, unless a very large number of loci is available. In selfing populations the results are less biased. More generally, populations with high identity disequilibrium (consanguineous or bottlenecked populations) could be propitious for using marker-based animal models, but are also more likely to deviate from the standard assumptions of quantitative genetics models (non-additive variance).     

Size Variation of the Nonrecombining Region on the Mating-Type Chromosomes in the Fungal Podospora anserina Species Complex

Sex chromosomes often carry large nonrecombining regions that can extend progressively over time, generating evolutionary strata of sequence divergence. However, some sex chromosomes display an incomplete suppression of recombination. Large genomic regions without recombination and evolutionary strata have also been documented around fungal mating-type loci, but have been studied in only a few fungal systems. In the model fungus Podospora anserina (Ascomycota, Sordariomycetes), the reference S strain lacks recombination across a 0.8-Mb region around the mating-type locus. The lack of recombination in this region ensures that nuclei of opposite mating types are packaged into a single ascospore (pseudohomothallic lifecycle). We found evidence for a lack of recombination around the mating-type locus in the genomes of ten P. anserina strains and six closely related pseudohomothallic Podospora species. Importantly, the size of the nonrecombining region differed between strains and species, as indicated by the heterozygosity levels around the mating-type locus and experimental selfing. The nonrecombining region is probably labile and polymorphic, differing in size and precise location within and between species, resulting in occasional, but infrequent, recombination at a given base pair. This view is also supported by the low divergence between mating types, and the lack of strong linkage disequilibrium, chromosomal rearrangements, transspecific polymorphism and genomic degeneration. We found a pattern suggestive of evolutionary strata in P. pseudocomata. The observed heterozygosity levels indicate low but nonnull outcrossing rates in nature in these pseudohomothallic fungi. This study adds to our understanding of mating-type chromosome evolution and its relationship to mating systems.     

Postglacial ecotype formation under outcrossing and self‐fertilization in Arabidopsis lyrata

The formation of ecotypes has been invoked as an important driver of postglacial biodiversity, because many species colonized heterogeneous habitats and experienced divergent selection. Ecotype formation has been predominantly studied in outcrossing taxa, while far less attention has been paid to the implications of mating system shifts. Here, we addressed whether substrate‐related ecotypes exist in selfing and outcrossing populations of Arabidopsis lyrata subsp. lyrata and whether the genomic footprint differs between mating systems. The North American subspecies colonized both rocky and sandy habitats during postglacial range expansion and shifted the mating system from predominantly outcrossing to predominantly selfing in a number of regions. We performed an association study on pooled whole‐genome sequence data of 20 selfing or outcrossing populations, which suggested genes involved in adaptation to substrate. Motivated by enriched gene ontology terms, we compared root growth between plants from the two substrates in a common environment and found that plants originating from sand grew roots faster and produced more side roots, independent of mating system. Furthermore, single nucleotide polymorphisms associated with substrate‐related ecotypes were more clustered among selfing populations. Our study provides evidence for substrate‐related ecotypes in A. lyrata and divergence in the genomic footprint between mating systems. The latter is the likely result of selfing populations having experienced divergent selection on larger genomic regions due to higher genome‐wide linkage disequilibrium.     

The Evolution of Selfing Is Accompanied by Reduced Efficacy of Selection and Purging of Deleterious Mutations

The transition from outcrossing to selfing is predicted to reduce the genome-wide efficacy of selection because of the lower effective population size (N_e) that accompanies this change in mating system. However, strongly recessive deleterious mutations exposed in the homozygous backgrounds of selfers should be under strong purifying selection. Here, we examine estimates of the distribution of fitness effects (DFE) and changes in the magnitude of effective selection coefficients (N_es) acting on mutations during the transition from outcrossing to selfing. Using forward simulations, we investigated the ability of a DFE inference approach to detect the joint influence of mating system and the dominance of deleterious mutations on selection efficacy. We investigated predictions from our simulations in the annual plant Eichhornia paniculata, in which selfing has evolved from outcrossing on multiple occasions. We used range-wide sampling to generate population genomic datasets and identified nonsynonymous and synonymous polymorphisms segregating in outcrossing and selfing populations. We found that the transition to selfing was accompanied by a change in the DFE, with a larger fraction of effectively neutral sites (N_es < 1), a result consistent with the effects of reduced N_e in selfers. Moreover, an increased proportion of sites in selfers were under strong purifying selection (N_es > 100), and simulations suggest that this is due to the exposure of recessive deleterious mutations. We conclude that the transition to selfing has been accompanied by the genome-wide influences of reduced N_e and strong purifying selection against deleterious recessive mutations, an example of purging at the molecular level.     

Insights Gained From 50 Years of Studying the Evolution of Self-Compatibility in <Emphasis Type="Italic">Leavenworthia</Emphasis> (Brassicaceae)

The shift from outcrossing to selfing is one of the most common evolutionary trends in plants, and there is intense interest in why this is so. The genus Leavenworthia has been the focus of research on this question for half of a century, with particular attention paid to the evolution of self-compatibility from self-incompatibility. In this review, we discuss the last 50 years of research concerning this evolutionary transition in Leavenworthia. Selfing appears to have evolved independently at minimum three times within this genus of eight species. Work on the ecological basis of mating system evolution in Leavenworthia has clarified that selection among individuals is likely a major force behind the recurrent evolution of selfing. Although inadequate pollination is appreciated as a factor favoring selfing, definitive ecological mechanisms that act to favor selfing are still not known and future work on the efficacy of pollinating bees and the effects of climate change is needed. Recent research has likely identified the SRK ortholog at the S-locus controlling self-incompatibility in Leavenworthia alabamica. Analyses of S-locus variation have revealed substantial S-allele diversity in outcrossing populations, with the recurrent fixation of mutations at the S-locus permitting the parallel evolution of selfing in this species. Although we appreciate some of the factors that may explain the evolution of selfing in this group, there is less known about the mechanisms underlying the widespread maintenance of outcrossing at the population and species levels. Studies in Leavenworthia have revealed that genetic diversity is lost over the long-term within selfing populations and leads to elevated population subdivision, but work is needed to determine why these genetic consequences of selfing cause lineages to become evolutionary dead ends.     

Global Population Genetic Structure of Caenorhabditis remanei Reveals Incipient Speciation

Mating system transitions dramatically alter the evolutionary trajectories of genomes that can be revealed by contrasts of species with disparate modes of reproduction. For such transitions in Caenorhabditis nematodes, some major causes of genome variation in selfing species have been discerned. And yet, we have only limited understanding of species-wide population genetic processes for their outcrossing relatives, which represent the reproductive state of the progenitors of selfing species. Multilocus-multipopulation sequence polymorphism data provide a powerful means to uncover the historical demography and evolutionary processes that shape genomes. Here we survey nucleotide polymorphism across the X chromosome for three populations of the outcrossing nematode Caenorhabditis remanei and demonstrate its divergence from a fourth population describing a closely related new species from China, C. sp. 23. We find high genetic variation globally and within each local population sample. Despite geographic barriers and moderate genetic differentiation between Europe and North America, considerable gene flow connects C. remanei populations. We discovered C. sp. 23 while investigating C. remanei, observing strong genetic differentiation characteristic of reproductive isolation that was confirmed by substantial F(2) hybrid breakdown in interspecific crosses. That C. sp. 23 represents a distinct biological species provides a cautionary example of how standard practice can fail for mating tests of species identity in this group. This species pair permits full application of divergence population genetic methods to obligately outcrossing species of Caenorhabditis and also presents a new focus for interrogation of the genetics and evolution of speciation with the Caenorhabditis model system.     

Mating Systems of Psychotria tenuinervis (Rubiaceae): Distance from Anthropogenic and Natural Edges of Atlantic Forest Fragment

The aim of this study was to determine and compare the mating systems among Psychotria tenuinervis populations at anthropogenic edges, natural edges, and the forest interior using allozyme electrophoresis of naturally pollinated progeny arrays. P. tenuinervis showed low outcrossing rates, varying from 37% to 50% of the mating attributable to outcrossing and 50% to 63% attributable to self-fertilization, in the three habitats. The forest interior had the highest outcrossing rate (t _m = 0.50 and t _s = 0.43) among the three habitats. However, there were no differences in either multilocus or single-locus rates among the three habitats, indicating that the contribution of biparental inbreeding to the apparent selfing rate in these populations was very low. The multilocus (t _m) and single-locus (t _s) outcrossing rates for the P. tenuinervis in the sample plots within each habitat showed great heterogeneity. In conclusion, edge creation seems not to influence its mating systems. Additionally, although P. tenuinervis is a distylous species, the population’s inbreeding can be attributed almost entirely to self-fertilization.     

Jatropha curcas L. (Euphorbiaceae) exhibits a mixed mating system, high correlated mating and apomixis

The hierarchical mating system among and within fruits of Jatropha curcas was investigated in a base population using five microsatellite loci, employing mixed mating and correlated mating models. Open-pollinated fruits were collected from 15 randomly selected seed trees, sampling seven fruits per tree for a total of 21 seeds from each tree. We detected multilocus genotypes identical to the mother tree in 13 % of offspring, implying the occurrence of apomixis in J. curcas. The presumed apomictic individuals were excluded from the analysis of the remaining results. Evidence of substantial selfing was provided by the average multilocus outcrossing rate (t _m?=?0.683), showing that the species exhibits a mixed mating system. The outcrossing rate showed a large variation among seed trees, ranging from 0.21 to 1.0, indicating that the species is not self-incompatible. Significant differences were detected between the multilocus and the single locus outcrossing rates (t _m???t _s?=?0.347) that suggested mating among related individuals, possibly because of the presence of individuals from the same progeny (sibs) in the base population. The multilocus paternity correlation was extremely high for the population (r _p(m)?=?0.999), indicating that the progenies were manly composed of full-sibs. As a consequence of selfing and a high paternity correlation, the co-ancestry coefficient within the progeny was higher (Θ?=?0.369) than expected for panmictic populations. Our results indicated that J. curcas produces seeds asexually by apomixis and sexually by a mixed mating system, combining selfing and outcrossing.     

Broad geographic covariation between floral traits and the mating system in Camissoniopsis cheiranthifolia (Onagraceae): multiple stable mixed mating systems across the species' range?

Background and Aims

Plants vary widely in the extent to which seeds are produced via self-fertilization vs. outcrossing, and evolutionary change in the mating system is thought to be accompanied by genetic differentiation in a syndrome of floral traits. We quantified the pattern of variation and covariation in floral traits and the proportion of seeds outcrossed (t) to better understand the evolutionary processes involved in mating system differentiation among and within populations of the short-lived Pacific coastal dune endemic Camissoniopsis cheiranthifolia across its geographic range in western North America.

Methods

We quantified corolla width and herkogamy, two traits expected to influence the mating system, for 48 populations sampled in the field and for a sub-sample of 29 populations grown from seed in a glasshouse. We also measured several other floral traits for 9–19 populations, estimated t for 16 populations using seven allozyme polymorphisms, and measured the strength of self-incompatibility for nine populations.

Key Results

Floral morphology and self-incompatibility varied widely but non-randomly, such that populations could be assigned to three phenotypically and geographically divergent groups. Populations spanned the full range of outcrossing (t = 0·001–0·992), which covaried with corolla width, herkogamy and floral life span. Outcrossing also correlated with floral morphology within two populations that exhibited exceptional floral variation.

Conclusions

Populations of C. cheiranthifolia seem to have differentiated into three modal mating systems: (1) predominant outcrossing associated with self-incompatibility and large flowers; (2) moderate selfing associated with large but self-compatible flowers; and (3) higher but not complete selfing associated with small, autogamous, self-compatible flowers. The transition to complete selfing has not occurred even though the species appears to possess the required genetic capacity. We hypothesize that outcrossing populations in this species have evolved to different stable states of mixed mating.     

Speciation and Introgression between Mimulus nasutus and Mimulus guttatus

Mimulus guttatus and M. nasutus are an evolutionary and ecological model sister species pair differentiated by ecology, mating system, and partial reproductive isolation. Despite extensive research on this system, the history of divergence and differentiation in this sister pair is unclear. We present and analyze a population genomic data set which shows that M. nasutus budded from a central Californian M. guttatus population within the last 200 to 500 thousand years. In this time, the M. nasutus genome has accrued genomic signatures of the transition to predominant selfing, including an elevated proportion of nonsynonymous variants, an accumulation of premature stop codons, and extended levels of linkage disequilibrium. Despite clear biological differentiation, we document genomic signatures of ongoing, bidirectional introgression. We observe a negative relationship between the recombination rate and divergence between M. nasutus and sympatric M. guttatus samples, suggesting that selection acts against M. nasutus ancestry in M. guttatus.     

Consequences of multiple mating‐system shifts for population and range‐wide genetic structure in a coastal dune plant

Evolutionary transitions from outcrossing to selfing can strongly affect the genetic diversity and structure of species at multiple spatial scales. We investigated the genetic consequences of mating‐system shifts in the North American, Pacific coast dune endemic plant Camissoniopsis cheiranthifolia (Onagraceae) by assaying variation at 13 nuclear (n) and six chloroplast (cp) microsatellite (SSR) loci for 38 populations across the species range. As predicted from the expected reduction in effective population size (N_e) caused by selfing, small‐flowered, predominantly selfing (SF) populations had much lower nSSR diversity (but not cpSSR) than large‐flowered, predominantly outcrossing (LF) populations. The reduction in nSSR diversity was greater than expected from the effects of selfing on N_e alone, but could not be accounted for by indirect effects of selfing on population density. Although selfing should reduce gene flow, SF populations were not more genetically differentiated than LF populations. We detected five clusters of nSSR genotypes and three groups of cpSSR haplotypes across the species range consisting of parapatric groups of populations that usually (but not always) differed in mating system, suggesting that selfing may often initiate ecogeographic isolation. However, lineage‐wide genetic variation was not lower for selfing clusters, failing to support the hypothesis that selection for reproductive assurance spurred the evolution of selfing in this species. Within three populations where LF and SF plants coexist, we detected genetic differentiation among diverged floral phenotypes suggesting that reproductive isolation (probably postzygotic) may help maintain the striking mating‐system differentiation observed across the range of this species.     

Wild sorghum from different eco-geographic regions of Kenya display a mixed mating system

Knowledge of mating systems is required in order to understand the genetic composition and evolutionary potential of plant populations. Outcrossing in a population may co-vary with the ecological and historical factors influencing it. However, literature on the outcrossing rate is limited in terms of wild sorghum species coverage and eco-geographic reference. This study investigated the outcrossing rates in wild sorghum populations from different ecological conditions of Kenya. Twelve wild sorghum populations were collected in four sorghum growing regions. Twenty-four individuals per population were genotyped using six polymorphic simple sequence repeat (SSR) markers to compute their indirect equilibrium estimates of outcrossing rate as well as population structure. In addition, the 12 populations were planted in a field in a randomised block design with five replications. Their progeny (250 individuals per population) were genotyped with the six SSR markers to estimate multi-locus outcrossing rates. Equilibrium estimates of outcrossing rates ranged from 7.0 to 75.0%, while multi-locus outcrossing rates (t _m) ranged from 8.9 to 70.0% with a mean of 49.7%, indicating that wild sorghum exhibits a mixed mating system. The wide range of estimated outcrossing rates in wild sorghum populations indicate that environmental conditions may exist under which fitness is favoured by outcrossing and others under which selfing is more advantageous. The genetic structure of the populations studied is concordant with that expected for a species displaying mixed mating system.     

Do selfing species have greater niche breadth? Support from ecological niche modeling

We explore the relationship between plant mating system (selfing or outcrossing) and niche breadth to gain new insights into processes that drive species distributions. Using a comparative approach with highly selfing versus highly outcrossing sister species, we test the extent to which: (1) species pairs have evolved significant niche divergence and less niche overlap, (2) selfers have wider niche breadths than outcrossers or vice versa, and (3) niches of selfers and outcrossers are defined by significant differences in environmental variables. We applied predictive ecological niche modeling approaches to estimate and contrast niche divergence, overlap and breadth, and to identify key environmental variables associated with each species’ niche for seven sister species with divergent mating systems. Data from 4862 geo-referenced herbarium occurrence records were compiled for 14 species in Collinsia and Tonella (Plantaginaceae) and 19 environmental variables associated with each record. We found sister species display significant niche divergence, though not as a function of divergence time, and overall, selfers have significantly wider niche breadths compared to their outcrossing sisters. Our results suggest that a selfing mating system likely contributes to the greater capacity to reach, reproduce, establish, and adapt to new habitats, which increases niche breadth of selfers.