Interspecific pollen competition as a reproductive barrier between sympatric species of Helianthus (Asteraceae)

Artificial crosses between Helianthus annuus and H. petiolaris using 1:9, 1:1, and 9:1 mixtures of intraspecific: interspecific pollen were conducted to determine the role of interspecific pollen competition as a reproductive barrier in Helianthus. Of 1,245 achenes analyzed from the pollen competition experiments, only 49 were hybrids. The number of hybrids observed was significantly less than expectations for all three pollen mixtures, regardless of the identity of maternal parent (P < 0.01). Stigma age and pollen ratio had no significant impact on hybrid frequency. However, hybrids were significantly more frequent with H. annuus than with H. petiolaris as the maternal parent (P < 0.01). Analysis of pollen tube growth rates revealed no differences in the rate of growth of intraspecific vs. interspecific pollen. Likewise, pollinations with either intraspecific or interspecific pollen or with different pollen ratios did not affect the percentage of filled achenes. Thus, the mechanism responsible for selective fertilization by intraspecific pollen in mixed pollen loads remains unclear. Nonetheless, these findings suggest that interspecific pollen competition plays an important role in controlling the formation of hybrids between H. annuus and H. petiolaris and may partially account for patterns or differential cytoplasmic vs. nuclear introgression in Helianthus.     

Inbreeding and outbreeding effects on pollen fitness and zygote survival in Silene nutans (Caryophyllaceae)

In plants, selfing and outcrossing may be affected by maternal mate choice and competition among pollen and zygotes. To evaluate this in Silene nutans, we pollinated plants with mixtures of (1) self‐ and outcross pollen and (2) pollen from within a population and from another population. Pollen fitness and zygote survival was estimated from the zygote survival and paternity of seeds. Self pollen had a lower fitness than outcross pollen, and selfed zygotes were less likely, or as likely, to develop into seeds. Hybrid zygotes survived as frequently or more than local zygotes, and pollen from one of the populations fertilized most ovules in both populations. Our results thus indicate strong maternal discrimination against selfing, whereas the success of outbreeding seems mostly affected by divergent pollen performance. The implications for the evolution of maternal mate choice are discussed.     

Mating Systems of Three Tropical Dry Forest Tree Species1

Polymorphic allozyme loci were used to estimate outcrossing rates for three tree species from a disturbed dry forest in southern Costa Rica. Estimates of the multilocus outcrossing rates of Cedrela odorata and Jacaranda copaia were 0.969 and 0.982, respectively, and suggest that these species may be self-incompatible. The subcanopy tree Stemmadenia donnell-smithii also demonstrated little self-fertilization based on an estimated outcrossing rate of 0.896. Significant heterogeneity in pollen allele frequencies among maternal trees was detected for at least two enzyme loci for each species. A test of correlated mating between progeny of S. donnell-smithii revealed that all seeds within a fruit were singly sired. In addition, the low estimates of biparental inbreeding and significant differences in pollen and ovule allele frequencies for this species suggest that gene flow into the sampled forest fragment may occur. The implications of deforestation on the mating systems of these tropical tree taxa are discussed.     

Are hybrid species more fit than ancestral parent species in the current hybrid species habitats?

Hybrid speciation is thought to be facilitated by escape of early generation hybrids into new habitats, subsequent environmental selection and adaptation. Here, we ask whether two homoploid hybrid plant species (Helianthus anomalus, H. deserticola) diverged sufficiently from their ancestral parent species (H. annuus, H. petiolaris) during hybrid speciation so that they are more fit than the parent species in hybrid species habitats. Hybrid and parental species were reciprocally transplanted into hybrid and parental habitats. Helianthus anomalus was more fit than parental species in the H. anomalus actively moving desert dune habitat. The abilities to tolerate burial and excavation and to obtain nutrients appear to be important for success in the H. anomalus habitat. In contrast, H. deserticola failed to outperform the parental species in the H. deserticola stabilized desert dune habitat, and several possible explanations are discussed. The home site advantage of H. anomalus is consistent with environmental selection having been a mechanism for adaptive divergence and hybrid speciation and supports the use of H. anomalus as a valuable system for further assessment of environmental selection and adaptive traits.     

Asymmetric reproductive interference: The consequences of cross‐pollination on reproductive success in sexual–apomictic populations of Potentilla puberula (Rosaceae)

Apomixis evolves from a sexual background and usually is linked to polyploidization. Pseudogamous gametophytic apomicts, which require a fertilization to initiate seed development, of various ploidy levels frequently co‐occur with their lower‐ploid sexual ancestors, but the stability of such mixed populations is affected by reproductive interferences mediated by cross‐pollination. Thereby, reproductive success of crosses depends on the difference in ploidy levels of mating partners, that is, on tolerance of deviation from the balanced ratio of maternal versus paternal genomes. Quality of pollen can further affect reproductive success in intercytotype pollinations. Cross‐fertilization, however, can be avoided by selfing which may be induced upon pollination with mixtures of self‐ and cross‐pollen (i.e., mentor effects). We tested for reproductive compatibility of naturally co‐occurring tetraploid sexuals and penta‐ to octoploid apomicts in the rosaceous species Potentilla puberula by means of controlled crosses. We estimated the role of selfing as a crossing barrier and effects of self‐ and cross‐pollen quality as well as maternal: paternal genomic ratios in the endosperm on reproductive success. Cross‐fertilization of sexuals by apomicts was not blocked by selfing, and seed set was reduced in hetero‐ compared to homoploid crosses. Thereby, seed set was negatively related to deviations from balanced parental genomic ratios in the endosperm. In contrast, seed set in the apomictic cytotypes was not reduced in hetero‐ compared to homoploid crosses. Thus, apomictic cytotypes either avoided intercytotype cross‐fertilization through selfing, tolerated intercytotype cross‐fertilizations without negative effects on reproductive success, or even benefitted from higher pollen quality in intercytotype pollinations. Our experiment provides evidence for asymmetric reproductive interference, in favor of the apomicts, with significantly reduced seed set of sexuals in cytologically mixed populations, whereas seed set in apomicts was not affected. Incompleteness of crossing barriers further indicated at least partial losses of a parental genomic endosperm balance requirement.     

Individual and Temporal Variation in Outcrossing Rates and Pollen Flow Patterns in Ceiba pentandra (Malvaceae: Bombacoidea)

Ceiba pentandra is a tropical tree with high rates of selfing in some populations. In mixed‐mating species, variation in selfing is due to changes in adult density or variability of incompatibility systems. The effect of spatial isolation and phenology on selfing rates and pollen flow distances was analyzed using microsatellites in a fragmented population of Ceiba pentandra, in southern Costa Rica. Adult trees within a heterogeneous landscape were classified as grouped or isolated. We compared selfing rates at the individual level, between isolation conditions and 2 yr (2007, 2009), which differed in the number of flowering individuals. Mixed mating was estimated in both years (t_m = 0.624–0.759). Trees mated predominantly by outcrossing, while only a few trees reproduced through selfing. Spatial isolation did not significantly affect outcrossing rates. The progeny of grouped trees was mostly sired by near‐neighbors (<1 km) and by long‐distance pollen flow events in isolated trees. A reduction in the number of flowering individuals in 2009 reduced near‐neighbor matings, increased selfing in grouped trees, and decreased the number of unsampled sires in the progeny. Comparing selfing rates on individuals that flowered in both reproductive periods suggests a flexible mating system. Variation in self‐fertilization rates in this population appears to depend on variation of individual traits, such as genetic variability in self‐incompatibility genes, but it is independent of landscape heterogeneity. In contrast, pollen flow distances depend on local tree density as bats concentrate their foraging between near individuals to maximize energy efficiency.     

Evolution of a hybrid zone of two willow species (Salix L.) in the European Alps analyzed by RAD-seq and morphometrics

Natural hybridization of plants can result in many outcomes with several evolutionary consequences, such as hybrid speciation and introgression. Natural hybrid zones can arise in mountain systems as a result of fluctuating climate during the exchange of glacial and interglacial periods, where species retract and expand their territories, resulting in secondary contacts. Willows are a large genus of woody plants with an immense capability of interspecific crossing. In this study, the sympatric area of two diploid sister species, S. foetida and S. waldsteiniana in the eastern European Alps, was investigated to study the genomic structure of populations within and outside their contact zone and to analyze congruence of morphological phenotypes with genetic data. Eleven populations of the two species were sampled across the Alps and examined using phylogenetic network and population genetic structure analyses of RAD Seq data and morphometric analyses of leaves. The results showed that a homoploid hybrid zone between the two species was established within their sympatric area. Patterns of genetic admixture in homoploid hybrids indicated introgression with asymmetric backcrossing to not only one of the parental species but also one hybrid population forming a separate lineage. The lack of F1 hybrids indicated a long-term persistence of the hybrid populations. Insignificant isolation by distance suggests that gene flow can act over large geographical scales. Morphometric characteristics of hybrids supported the molecular data and clearly separated populations of the parental species, but showed intermediacy in the hybrid zone populations with a bias toward S. waldsteiniana. The homoploid hybrid zone might have been established via secondary contact hybridization, and its establishment was fostered by the low genetic divergence of parental species and a lack of strong intrinsic crossing barriers. Incomplete ecological separation and the ability of long-distance dispersal of willows could have contributed to the spatial expansion of the hybrid zone.     

FEMALE FREQUENCIES IN GYNODIOECIOUS POPULATIONS CORRELATED WITH SELFING RATES IN HERMAPHRODITES

Gynodioecious populations consist of separate hermaphroditic and female individuals. Females are at a selective disadvantage because they contribute genes to the next generation only through ovules, while hermaphrodites contribute genes through ovules and pollen. For females to be maintained in populations they must have some compensating selective advantage. The outcrossing hypothesis postulates that females are maintained because their progeny result from obligate outcrossing, whereas some of the progeny of hermaphrodites result from self-fertilization and are less fit because of inbreeding depression. If correct, the frequency of females should be positively correlated with selfing rates of hermaphrodites in populations. We found a strong positive correlation between female frequency and selfing rates of hermaphrodites (r = 0.91, P < 0.01) in eight gynodioecious populations of Hawaiian species of Bidens. Our results confirm that the obligate outcrossing of females is a major factor maintaining females in gynodioecious populations. However, the observed selfing rates are insufficient by themselves to account for the frequency of females in these populations.     

Small reductions in corolla size and pollen: ovule ratio,but no changes in flower shape in selfing populations of the North American <Emphasis Type="Italic">Arabidopsis lyrata</Emphasis>

The shift from outcrossing to selfing is often accompanied by striking changes in floral morphology towards a “selfing syndrome”, which is characterized by flowers with reduction in size, pollen: ovule (P/O) ratio, and herkogamy. This study aims to test whether such changes have occurred in the North American Arabidopsis lyrata, which is of particular interest because of the relatively recent transitions to selfing in this system. Flower size, flower shape, herkogamy levels, P/O ratio, and floral integration of six self-incompatible (outcrossing) and six self-compatible (selfing) populations of A. lyrata were measured in a common environment using conventional and geometric morphometrics methods. Although selfers had on average 9.2% smaller corollas, 8.4% longer pistils, and 21.5% lower P/O ratios than outcrossers, there were no differences in shape, floral integration, and herkogamy between outcrossing and selfing populations. Moreover, most variation in floral traits was explained by population genetic background rather than by mating system. We conclude that selfing populations in A. lyrata have not evolved a selfing syndrome.     

Mating system in Mexican populations of the annual herb Solanum rostratum Dunal (Solanaceae)

Traditionally, annual colonising species are expected to have high rates of self‐fertilisation, although recent theoretical and empirical studies have shown that cross‐fertilisation can be selected for under heterogeneous pollination environments. Solanum rostratum is a self‐compatible annual herb that colonises disturbed habitats. Despite the lack of physiological mechanisms to prevent self‐fertilisation, pollen transfer between individuals is expected to be favoured because of its complex floral morphology. In previous studies of S. rostratum it has been shown that anther dimorphism within flowers results in precise pollen placement on the pollinator's body, and the presence of mirror‐image floral morphs within plants promotes outcrossing in experimental arrays. However, the mating system of natural populations of S. rostratum has never been assessed, and thus whether it is predominantly selfing or outcrossing remains unknown. We hypothesise that floral and inflorescence morphology of S. rostratum should facilitate cross‐fertilisation, making it a predominantly outcrossing despite its lack of a self‐incompatibility system. To test this hypothesis, we estimated outcrossing rates by genotyping 700 individuals at 13 microsatellite loci, sampled from four populations across a 690‐km transect in the species' native range. We found that populations had mean outcrossing rates of 0.70 ± 0.03, with multiple sires contributing to paternity of each progeny array (average effective number of sires = 8.97 ± 0.57). This indicates that natural populations S. rostratum have relatively high levels of outcrossing, probably facilitated by its floral and inflorescence morphology. We speculate that partial selfing in this species may be an unavoidable consequence of displaying multiple flowers at the same time (geitonogamy), as well as the result of self‐pollen transfer by illegitimate visitors.     

Persistence of sunflower crop traits and fitness in Helianthus petiolaris populations

Transgenic plants have increased interest in the study of crop gene introgression in wild populations. Genes (or transgenes) conferring adaptive advantages persist in introgressed populations, enhancing competitiveness of wild or weedy plants. This represents an ecological risk that could increase problems of weed control. Introgression of cultivar alleles into wild plant populations via crop–wild hybridisations is primarily governed by their fitness effect. To evaluate this, we studied the second generation of seven wild–crop interspecific hybrids between weedy Helianthus petiolaris and cultivated sunflower, H. annuus var. macrocarpus. The second generation comprised open‐pollinated progeny and backcrosses to the wild parent, mimicking crosses that occur in natural situations. We compared a number of morphological, life history and fitness traits. Multivariate analysis showed that the parental species H. annuus and H. petiolaris differed in a number of morphological traits, while the second hybrid generation between them was intermediate. Sunflower crop introgression lowered fitness of interspecific hybrids, but fitness parameters tended to recover in the following generation. Relative frequency of wild/weedy and introgressed plants was estimated through four generations, based on male and female parent fitness. In spite of several negative selection coefficients observed in the second generation, introgressed plants could be detected in stands of <100 weedy H. petiolaris populations. The rapid recovery of fecundity parameters leads to prediction that any trait conferring an ecological advantage will diffuse into the wild or weedy population, even if F1 hybrids have low fitness.     

山莨菪(茄科)的传粉生物学

茄科的多数种类具有自交不亲和的特点, 主要通过异花传粉结实; 但是, 一些物种或者物种内的部分种群或者个体却高度自交亲合, 转变为自交的繁育系统。该科植物山莨菪(Anisodus tanguticus)主要分布在青藏高原, 开花较早, 比其他晚开花的植物种类更加缺少有效的异花传粉昆虫。我们选择了位于不同海拔高度的2个种群进行比较研究, 主要目的是检验该物种的繁育系统是否在极端环境下由于传粉者的缺乏而发生了部分改变。研究发现,山莨菪的花不完全雌性先熟, 柱头和花药间的平均距离随着花开放时间的延长而不断缩小, 但两者在多数花的单花花期结束时并没有发生接触。因此, 山莨菪花主要表现为适应异花传粉的雌雄异位特征。然而, 少数花 (4.9％)的柱头和花药发生接触, 为“自动自交”的传粉解除了空间隔离。2个种群的多数个体存在自交不亲和机制, 应具有异花传粉的繁育系统; 但是部分个体具有明显的自交亲和能力, 为自交提供了生理基础。高海拔种群的传粉昆虫主要是厕蝇(Fannia sp.), 它们在不同植株间的活动能够保证异花传粉结实; 同时该种群的部分个体存在“自动自交”。低海拔种群的主要访花昆虫是蚂蚁, 它们在花内的活动导致花粉在同一朵花内传递, 而引起“协助自交”; 而异花传粉昆虫厕蝇的访花频率则较高海拔种群低。两个种群的结实均由于异花传粉者不足而受到传粉限制。因此两种不同类型的自交机制为该早期开花植物异花访花昆虫的不足提供了一定程度上的繁殖补偿。     

Evolution of the Selfing Syndrome in Arabis alpina (Brassicaceae)

Introduction

The transition from cross-fertilisation (outcrossing) to self-fertilisation (selfing) frequently coincides with changes towards a floral morphology that optimises self-pollination, the selfing syndrome. Population genetic studies have reported the existence of both outcrossing and selfing populations in Arabis alpina (Brassicaceae), which is an emerging model species for studying the molecular basis of perenniality and local adaptation. It is unknown whether its selfing populations have evolved a selfing syndrome.

Methods

Using macro-photography, microscopy and automated cell counting, we compared floral syndromes (size, herkogamy, pollen and ovule numbers) between three outcrossing populations from the Apuan Alps and three selfing populations from the Western and Central Alps (Maritime Alps and Dolomites). In addition, we genotyped the plants for 12 microsatellite loci to confirm previous measures of diversity and inbreeding coefficients based on allozymes, and performed Bayesian clustering.

Results and Discussion

Plants from the three selfing populations had markedly smaller flowers, less herkogamy and lower pollen production than plants from the three outcrossing populations, whereas pistil length and ovule number have remained constant. Compared to allozymes, microsatellite variation was higher, but revealed similar patterns of low diversity and high Fis in selfing populations. Bayesian clustering revealed two clusters. The first cluster contained the three outcrossing populations from the Apuan Alps, the second contained the three selfing populations from the Maritime Alps and Dolomites.

Conclusion

We conclude that in comparison to three outcrossing populations, three populations with high selfing rates are characterised by a flower morphology that is closer to the selfing syndrome. The presence of outcrossing and selfing floral syndromes within a single species will facilitate unravelling the genetic basis of the selfing syndrome, and addressing which selective forces drive its evolution.     

Pollen pool heterogeneity in jack pine (Pinus banksiana Lamb.): a problem for estimating outcrossing rates?

Summary Pollen pool heterogeneity, which violates an assumption of the mixed-mating model, is a major potential problem in measuring plant mating systems. In this study, isozyme markers were used to examine pollen pool heterogeneity in two natural populations of jack pine, Pinus banksiana Lamb., in northwestern Ontario, Canada. Population multilocus estimates of outcrossing rate ranged from 0.83 to 0.95 and differed significantly between populations. Single-tree multilocus outcrossing rates were found to be homogeneous among trees in both populations. Computer simulation studies indicated that a consanguineous pollen pool (pollen gametes related to the mother tree) was capable of biasing population outcrossing estimates downward. Random pollen pool heterogeneity (uncorrelated with maternal genotypes) did not appear to affect population outcrossing estimates in the simulations. Heterogeneity G-tests and Spearman rank tests showed that pollen pool heterogeneity existed in the two natural populations examined; however, it did not have a major effect on population outcrossing estimates, since the consanguineous pollen pool detected was probably a relatively minor component of the outcross pollen pool in both populations. In addition, heterogeneity G-tests were found to be not sensitive in detecting pollen pool heterogeneity caused by consanguineous pollen pool.     

Outcrossing rates and their relationship to phenology in Triticum dicoccoides

Summary Outcrossing rates (t) were estimated in natural and garden populations of wild emmer wheat, Triticum dicoccoides. The estimated t from the natural populations was 0 with the 0.95 upper confidence limit ranging from 0.102 to 0.925 depending on the genetic variation within the population. Outcrossing rates for two genotype classes of this species grown in a common garden were 0.0077 and 0.0018. The difference in Outcrossing rates between the two genotypes is ascribed to phenological differences and hence different available pollen pools. Phenological displacement is discussed as a possible bias in the estimation of outcrossing rates in both predominantly selfing and outcrossing plants.     

Mating system and the hybridization between self-compatible Phlox cuspidata and self-incompatible Phlox drummondii

Mating system can impact the frequency of hybridization and therefore the maintenance of species diversity. I evaluate the effects of weak self-incompatibility (SI) in Phlox cuspidata and SI in Phlox drummondii on mating success within species and on hybridization dynamics between species under controlled conditions. The effects of SI on hybridization frequency were assessed by manipulating the relatedness of conspecific pollen and the relative timing of pollen deposition in mixed-donor interspecific pollinations. Selfing as opposed to outcrossing increased hybridization by 16% in P. cuspidata maternal plants and by 48% in P. drummondii maternal plants because self pollen did not compete as well against heterospecific pollen. The relative timing of conspecific versus heterospecific pollen deposition also impacted hybridization. In self-compatible P. cuspidata, the deposition of self pollen 5 h earlier than heterospecific pollen decreased hybridization by 28%. In self-incompatible P. drummondii, a 5 h delay in the deposition of compatible conspecific pollen increased hybridization by 32%. In this hybrid system, early self-pollination can decrease hybridization (but increase inbreeding) by P. cuspidata maternal plants, and SI may increase hybridization by P. drummondii maternal plants.     

THE POPULATION GENETICS OF SPOROPHYTIC SELF‐INCOMPATIBILITY IN THREE HYBRIDIZING SENECIO (ASTERACEAE) SPECIES WITH CONTRASTING POPULATION HISTORIES

Hybridization generates evolutionary novelty and spreads adaptive variation. By promoting outcrossing, plant self‐incompatibility (SI) systems also favor interspecific hybridization because the S locus is under strong negative frequency‐dependent balancing selection. This study investigates the SI mating systems of three hybridizing Senecio species with contrasting population histories. Senecio aethnensis and S. chrysanthemifolius native to Sicily, form a hybrid zone at intermediate altitudes on Mount Etna, and their neo‐homoploid hybrid species, S. squalidus, has colonized disturbed urban habitats in the UK during the last 150 years. We show that all three species express sporophytic SI (SSI), where pollen incompatibility is controlled by the diploid parental genome, and that SSI is inherited and functions normally in hybrids. Large‐scale crossing studies of wild sampled populations allowed direct comparison of SSI between species and found that the main impacts of colonization in S. squalidus compared to Sicilian Senecio was a reduced number of S alleles, increased S allele frequencies, and increased interpopulation S allele sharing. In general, many S alleles were shared between species and the S locus showed reduced intra‐ and interspecific population genetic structure compared to molecular genetic markers, indicative of enhanced effective gene flow due to balancing selection.     

MOLECULAR TESTS OF THE HYPOTHESIZED HYBRID ORIGIN OF TWO DIPLOID HELIANTHUS SPECIES (ASTERACEAE)

Enzyme electrophoresis and restriction-fragment analysis of chloroplast DNA (cpDNA) and nuclear ribosomal DNA (rDNA) were used to test the hypothesis that both Helianthus neglectus and H. paradoxus are stabilized hybrid derivatives of H. annuus and H. petiolaris. The four species are annuals, diploid outcrossers, and have the same chromosome number. Helianthus annuus and H. petiolaris had the same allele in highest frequency for 16 of the 18 isozyme loci examined and had different majority alleles for only 6-Pgd3 and Pgi2. The two species had divergent rDNAs that could be distinguished by seven diagnostic restriction site mutations and three length mutations, and their cpDNAs could be differentiated by three diagnostic restriction site mutations. The alleles observed in H. neglectus were not a combination of those observed in H. annuus and H. petiolaris. Although H. neglectus had only one unique allele, it possessed none of the three alleles specific to H. annuus. In contrast, it had four of the seven alleles specific to H. petiolaris. Furthermore, H. neglectus had the same rDNA type as H. petiolaris and had the same cpDNA as that found in two populations of H. petiolaris ssp. fallax. These data allowed us to speculate that H. neglectus may be a recent derivative of H. petiolaris ssp. fallax, rather than a stabilized hybrid derivative as originally proposed. In contrast, H. paradoxus combined the alleles of H. annuus and H. petiolaris and had no unique alleles. At Adh2, H. paradoxus was monomorphic for an allele found only in H. petiolaris ssp. fallax, whereas at 6-Pgd3 and Pgi2, it was monomorphic for high frequency H. annuus alleles. Furthermore, H. paradoxus combined the rDNA repeat types of both proposed parents and had the chloroplast genome of H. annuus. These data provide compelling evidence that H. paradoxus, in contrast to H. neglectus, was derived via hybridization.