Phylogenetic relationships and character evolution in Primula L.: The usefulness of ITS sequence data

ABSTRACT

The main goals of this research were to reconstruct the infrageneric phylogeny of the genus Primula based on both nuclear and chloroplast DNA sequences, and to use the resulting phylogenies to elucidate the evolution of breeding systems, morphological characters, chromosome number, and biogeographic distribution in the genus. In this paper, the results of a pilot study based on the nuclear ribosomal Internal Transcribed Spacer (ITS) region are described. ITS sequences from 21 taxa produced a number of variable characters sufficient to resolve relationships among sections. The resulting phylogeny confirmed the monophyly of sections Auricula and Aleuritia. Sections Armerina, Proliferae, Crystallophlomis, Parryi, and Auricula, with a base chromosome number of x = 11, and sect. Aleuritia, with a base chromosome number of x = 9, formed two well supported clades. The ITS topology also suggested that leaves with revolute vernation, previously believed to be a derived state, might represent the ancestral condition in Primula, with later reversals to the involute condition. Finally, this initial ITS tree provides preliminary support to the proposed role of the widespread, diploid and heterostylous P. mistassinica as having given origin to the polyploid, homostylous P. incana and P. laurentiana.     

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.     

Evolution of biogeographic patterns, ploidy levels, and breeding systems in a diploid-polyploid species complex of Primula

Primula sect. Aleuritia subsect. Aleuritia (Aleuritia) includes diploid, self-incompatible heterostyles and polyploid, self-compatible homostyles, the latter generally occurring at higher latitudes than the former. This study develops a phylogenetic hypothesis for Aleuritia to elucidate the interactions between Pleistocene glacial cycles, biogeographic patterns, ploidy levels and breeding systems. Sequences from five chloroplast DNA loci were analyzed with parsimony to reconstruct a phylogeny, haplotype network, and ancestral states for ploidy levels and breeding systems.The results supported the monophyly of Aleuritia and four major biogeographic lineages: an amphi-Pacific, a South American, an amphi-Atlantic and a European/North American lineage. At least four independent switches to homostyly and five to polyploidy were inferred. An Asian ancestor probably gave origin to an amphi-Pacific clade and to a lineage that diversified on the European and American continents. Switches to homostyly occurred exclusively in polyploid lineages, which mainly occupy previously glaciated areas. The higher success of the autogamous polyploid species at recolonizing habitats freed by glacial retreat might be explained in terms of selection for reproductive assurance.     

SMALL AND UGLY? PHYLOGENETIC ANALYSES OF THE “SELFING SYNDROME” REVEAL COMPLEX EVOLUTIONARY FATES OF MONOMORPHIC PRIMROSE FLOWERS

One of the most common trends in plant evolution, loss of self‐incompatibility and ensuing increases in selfing, is generally assumed to be associated with a suite of phenotypic changes, notably a reduction of floral size, termed the selfing syndrome. We investigate whether floral morphological traits indeed decrease in a deterministic fashion after losses of self‐incompatibility, as traditionally expected, using a phylogeny of 124 primrose species containing nine independent transitions from heterostyly (heteromorphic incompatibility) to homostyly (monomorphic self‐compatibility), a classic system for evolution of selfing. We find similar overall variability of homostylous and heterostylous species, except for diminished herkogamy in homostyles. Bayesian mixed models demonstrate differences between homostylous and heterostylous species in all traits, but net effects across species are small (except herkogamy) and directionality differs among traits. Strongly drift‐like evolutionary trajectories of corolla tube length and corolla diameter inferred by Ornstein–Uhlenbeck models contrast with expected deterministic trajectories toward small floral size. Lineage‐specific population genetic effects associated with evolution of selfing may explain that reductions of floral size represent one of several possible outcomes of floral evolution after loss of heterostyly in primroses. Contrary to the traditional paradigm, selfing syndromes may, but do not necessarily evolve in response to increased selfing.     

Are any primroses (Primula) primitively monomorphic?

Primula (c. 430 species) and relatives (Primulaceae) are paradigmatic to our understanding of distyly. However, the common co-occurrence of distyly and monomorphy in closely related groups within the family has made the interpretation of its evolution difficult.Here, we infer a chloroplast DNA (cpDNA) phylogeny for 207 accessions, including 51% of the species and 95% of the sections of Primula with monomorphic populations, using Bayesian methods. With this tree, we infer the distribution of ancestral states on critical nodes using parsimony and likelihood methods.The inferred cpDNA phylogeny is consistent with prior estimates. The most recent common ancestor (MRCA) of Primula is resolved as distylous using both methods of inference. However, whether the distyly in Primula, Hottonia, and Vitaliana arose once or three independent times is not clear.We conclude that monomorphism in descendants of the MRCA of Primula is derived from distyly in all cases. Thus, scenarios for the evolution of distyly that rely on the persistence of primitive monomorphy (such as in Primula section Sphondylia) require re-evaluation.     

Breeding system variation, genetics and evolution in the Turneraceae

We review the genetics and evolution of breeding systems in the Turneraceae. Distyly occurs in seven of 10 genera and 81% of species. The remaining species are homostylous. Polyploid evolution has been significant in Turnera. Approximately 60% of species are polyploid ranging from diploid through decaploid. No relationship between breeding system and polyploidy is evident. The genetics of distyly involves a one-locus two-allele system (S and s). Evidence from crosses with homostylous species and mutants is consistent with the possibility that a "Primula-type" supergene underlies distyly but does not prove this to be the case. A polygalacturonase, and an alpha-dioxygenase specific to the transmitting tissue of short-styled plants both exhibit morph-limited expression in concert with predictions from an evolutionary model. The function of the proteins in distyly, if any, is unknown. We have begun constructing a fine-scale genetic map of Turnera. Two genetic markers lie within 0.2 cm of the distyly locus. This should provide a starting point for positional cloning of the distyly locus and reveal the genetic architecture and molecular basis of distyly.     

On the occurrence of unilateral incompatibility in Primula section Aleuritia Duby and the origin of Primula scotica Hook

The occurrence of unilateral incompatibility was tested for a distylous heteromorphic system, using crosses between a self-sterile and three self-fertile species in Primula L. section Aleuritia Duby. The crosses showed non-reciprocity but not in the same direction as would be predicted in the case of unilateral incompatibility. Pollen from the self-compatible (s-c) species was not always inhibited on the style of the self-incompatible (s-i) species, and cross-fertility between s-i and s-c crosses more resembled that between different s-c species, which was also non-reciprocal. Cytoplasmic-nuclear DNA interactions and the possibility of embryo-endosperm imbalance could both explain these results. In crosses between Primula farinosa L. (s-i diploid) and P. scotica Hook. (s-c hexaploid), heterostylous pin tetraploid offspring were produced. This result is discussed in relation to the genome of P. scotica and the possibility that pin morph plants may occur in wild P scotica populations.     

The evolution of apomixis in angiosperms: A reappraisal

Abstract

Apomixis, the asexual reproduction via seed, has long been regarded a blind alley of evolution. This hypothesis was based on the assumption that apomixis is an irreversible, phylogenetically derived trait that would rapidly lead to extinction of the respective lineages. However, recent updates of the taxonomic distribution of apomixis in angiosperms suggest an alternative evolutionary scenario. Apomixis is taxonomically scattered and occurs in both early and late branching lineages, with several reversals from apomixis to obligate sex along phylogeny. Genetic control of apomixis is based on altered expression patterns of the same genes that control sexual development; epigenetic changes following polyploidization and/or hybridization may trigger shifts from sexuality to apomixis. Mendelian inheritance confirms the facultative nature and possible reversibility of apomixis to sexual reproduction. Apomixis, therefore, could represent a transition period in the evolution of polyploid complexes, with polyspory in paleopolyploids being a remnant of lost apomixis. In neopolyploids, apomixis helps to overcome sterility and allows for geographical range expansions of agamic polyploid complexes. The facultative nature of apomixis allows for reversal to sexuality and further speciation of paleopolyploid lineages. Thus, apomixis may facilitate diversification of polyploid complexes and evolution in angiosperms.     

Pollen flow in a population of Primula vulgaris Huds.

A British population of Primula vulgaris was visited four times between 1971 and 1976 for study of the nature and functioning of distyly. The stainability and production of pollen from pins exceeded those of thrums. About 20% of pin pollen and 48% of thrum pollen was removed from anthers by insect visitors. Pins and thrums were present in a 1:1 ratio in 1971 but pins outnumbered thrums in 1976. Thrums produced more seeds per flower than pins though the number of ovules produced by each was similar. Two methods for collecting stigmas from open flowers and analyzing them for pollen loans produced somewhat different results. For pin stigmas, the pollen load consisted of 2–23% thrum pollen; for thrum stigmas, the pollen load consisted of 0–71% pin pollen with most stigmas having less than 50% pin pollen. In general, intermorph pollen flow is less than would be expected if pollen flow were random. It is probable that most intramorph pollen on stigmas is a result of self- or geitonogamous pollination. The extensive literature concerning the natural pollinators of the primrose is reviewed. Although Darwin's hypothesis concerning the functional significance of distyly in promoting intermorph pollination was never quantified, the pollen flow patterns observed in P. vulgaris are unexpectedly deviant and are similar to those patterns observed in several unrelated heterostylous species in other plant families.     

Characterization of microsatellite loci in Primula modesta Bisset et Moore (Primulaceae)

Eleven microsatellite loci were isolated from alpine plant Primula modesta. Enriched repeat libraries were constructed and screened by colony hybridization. Forty‐eight polymerase chain reaction (PCR) primer pairs were designed, of which 11 pairs producted successful amplification. A total of 31–35 adult individuals were genotyped for allelic diversity. The polymorphism ranged from three to 14 alleles, and the heterozygosity ranged from 0.161 to 0.828. No linkage disequilibrium was found between these 11 loci.     

Functional heterostyly in Tylosema esculentum (Caesalpinioideae)

Tylosema esculentum is a long-lived perennial species endemic to arid areas of southern Africa. Its potential as a crop species has long been recognized as a result of the high oil and protein content of its seeds. The reproductive biology and breeding systems of the species were investigated in wild and experimental populations growing in Botswana. Field observations confirmed that the species is heterostylous with the pistil and anthers exhibiting reciprocal heights in the two morphs, although pollen size and sculpturing do not vary. The wet, nonpapillate stigma characteristic of the species is the first to be reported in the Caesalpinioideae. In vivo and in vitro diallel crossing experiments demonstrated that a diallelic self-incompatability system exists in T. esculentum. The major site of pollen tube inhibition in the intramorph crosses was found to be in the style. This is the first report of functional heterostyly in the Fabaceae and of a confirmed self-incompatibility system in the Caesalpinioideae. Three separate lines of evidence, the monitoring of fruit development in open-pollinated plants, fruit set in diallel crossing experiments, and observations made in wild populations, demonstrated that fruit set and, by implication, seed set, are very low in this species. Floral abscission was a major limitation to the production of mature pods but there were also significant losses at other developmental stages of fruit production. The results suggest that low seed set may be an adaptation of the species to an environment in which rainfall is scarce.     

Species Biology of Primula sieboldii for the Conservation of its Lowland-habitat Population: I. Inter-clonal Variations in the Flowering Phenology, Pollen Load and Female Fertility Components

Abstract Intra-population variations of several characteristics related to fertility were assessed for a nature-reserve population of Primula sieboldii in the flood plain of the Arakawa River. Although there were large intra- and inter-clonal variations of fruit and seed sets as well as other female fertility components in the 13 clones examined, female fertility was generally low, except in a homomorph clone. Pollinator availability was assessed by continuous monitoring of insect visits with 8 mm film. No insect visits were recorded during the monitoring of total 68 flowers over 16 hours in the nature-reserve population. In contrast, a high pollinator visitation rate was recorded in a reference population of P. sieboldii on the flank of Mt. Yatugatake. Thus, the low rates of seed production in the nature reserve was due to a shortage of compatible pollens as a result of the limitation of pollinator availability. Stigmas of flowers in various clones occasionally were loaded with pollen grains whose sizes were not significantly different from those of anther pollens of the same flowers. However, pollen germination on stigmas was observed only in the flowers of a homo-morph clone whose flowers have pin-type stigmas and anthers with compatible thrum-type pollens at the same height. Exceptionally high fertility of the homo-morph clone would be associated with its inbreeding habit.     

珍稀植物景东报春的地理分布和生态生物学特性

Primula interjacens Chen, an endemic and rare species to China, is restrictively distributed in Mt. Wuliangshan of Yunnan Province. Only three populations are found. The species has been fallen into endangered condition. Its present state of distribution, biological and ecological characteristics are investigated. In addition, the factors causing this species endangered and conservation strategies are analyzed briefly.     

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.     

The dimorphic heterostyly inAconogonon campanulatum (Polygonaceae)

Heterostyly is clearly confirmed inAconogonon campanulatum. This distylous species is dimorphic for tepals, styles, stigma surface, stamens, pollen grain size, and pollen sexine ornamentation. The floral shape is campanulate and thrum flowers are slightly larger than pin flowers. Small solitary bees were observed as flower visitors and probably effected pollination. The possible evolution of dioecy via heterostyly within the genusAconogonon is discussed.