The relationship between cell division and elongation during development of the nectar-yielding petal spur in Centranthus ruber (Valerianaceae)

Background and Aims Floral spurs are hollow, tubular outgrowths that typically conceal nectar. By their involvement in specialized pollinator interactions, spurs have ecological and evolutionary significance, often leading to speciation. Despite their importance and diversity in shape and size among angiosperm taxa, detailed investigations of the mechanism of spur development have been conducted only recently.Methods Initiation and growth of the nectar-yielding petal spur of Centranthus ruber ‘Snowcloud’ was investigated throughout seven stages, based on bud size and developmental events. The determination of the frequency of cell division, quantified for the first time in spurs, was conducted by confocal microscopy following 4'',6-diamidino-2-phenylindole (DAPI) staining of mitotic figures. Moreover, using scanning electron microscospy of the outer petal spur surface unobstructed by trichomes, morphometry of epidermal cells was determined throughout development in order to understand the ontogeny of this elongate, hollow tube.Key Results Spur growth from the corolla base initially included diffuse cell divisions identified among epidermal cells as the spur progressed through its early stages. However, cell divisions clearly diminished before a petal spur attained 30 % of its final length of 4·5 mm. Thereafter until anthesis, elongation of individual cells was primarily responsible for the spur’s own extension. Consequently, a prolonged period of anisotropy, wherein epidermal cells elongated almost uniformly in all regions along the petal spur’s longitudinal axis, contributed principally to the spur’s mature length.Conclusions This research demonstrates that anisotropic growth of epidermal cells – in the same orientation as spur elongation – chiefly explains petal spur extension in C. ruber. Representing the inaugural investigation of the cellular basis for spur ontogeny within the Euasterids II clade, this study complements the patterns in Aquilegia species (order Ranunculales, Eudicots) and Linaria vulgaris (order Lamiales, Euasterids I), thereby suggesting the existence of a common underlying mechanism for petal spur ontogeny in disparate dicot lineages.     

Genetic diversity and evolutionary history of four closely related Aquilegia species revealed by 10 nuclear gene fragments

The genus Aquilegia is emerging as the new model system for plant development, ecology, and evolution studies. Previous research showed that pollinator shift might drive the diversification of North American Aquilegia species, and natural selection on the length of petal nectar spur might play a crucial role. In this genus, A. ecalcarata Maxim. is the only taxon that has lost nectar spurs. Previous phylogenetic results indicated that A. ecalcarata, A. yabeana Kitag., A. oxysepala var. kansuensis Bruhl., and A. rockii Munz comprised a monophyletic group. However, their pattern of genetic diversity remains unknown. In addition, little is known about the evolutionary relationship among the four species on the population level. We carried out a population genetics study with 21 representative populations based on 10 single-copy nuclear gene fragments and found that: (i) A. yabeana conserved the highest genetic diversity (both π_sil and θ_sil) and A. oxysepala var. kansuensis had the lowest level; (ii) A. ecalcarata split into two groups, with one population clustered with A. rockii and the other five populations clustered with A. oxysepala var. kansuensis; and (iii) the allele frequency spectrum showed an excess of low frequency alleles in all four species, implying that they may undergo the mutation-drift equilibrium. Our findings provide the first investigation of genetic diversity and evolutionary relationships in A. yabeana, A. oxysepala var. kansuensis, A. rockii, and A. ecalcarata. They lay the foundation for future evolutionary studies, such as speciation mediated by pollinators.     

Effects of fire frequency on oak litter decomposition and nitrogen dynamics

Rapid speciation within some plant families has been attributed to the evolution of floral spurs and to the effect of spur length on plant reproductive success. The flowers of Impatiens capensis (jewelweed) possess a long, curved spur in which nectar is produced and stored. Spur length and curvature varies among plants within one population. Here I document that spur shape is variable in natural populations, variation within plants is less than variation among plants, and spur shape is correlated with components of female and male reproductive success. The apparent natural selection is weakly directional in 1 of 2 years, with greatest seed production and pollen removal occurring in flowers with the greatest spur curvature. Bee pollinator visit length is longest at flowers with highly curved spurs, and they leave less nectar in these spurs than in flowers with straighter spurs. Spur angle evolution may be limited, at least in part, by opposing selection by nectar-robbers who prefer to visit flowers with greater spur curvature. Other factors that might contribute to the maintenance of spur angle variation are temporal variation in the strength of selection and potential genetic correlations of spur shape with other traits under selection.     

Pollinators of the Rocky Mountain columbine: temporal variation, functional groups and associations with floral traits

Background and Aims

Pollinators together with other biotic and some abiotic factors can select for floral traits. However, variation in pollinator abundance over time and space can weaken such selection. In the present study, the variation in pollinator abundance over time and space was examined in populations of the Rocky Mountain columbine. The variation in three floral traits is described and correlations between pollinator type, functional pollinator groups or altitude and floral traits are examined.

Methods

Pollinator observations took place in six Aquilegia coerulea populations over 1–4 years and spur length, flower colour and sepal length were measured in 12 populations. Pollinator abundance, measured as visits per flower per hour, was compared among populations and years. Pollinators were grouped into two functional groups: pollen or nectar collectors. The following associations were examined: annual presence of hawkmoths and whiter flowers with longer spurs; the presence of Sphinx vashti and longer spurs; and higher altitudes and whiter flowers. The study looked at whether an increase in the proportion of hawkmoths in a population was associated with whiter and larger flowers with longer spurs.

Key Results

The abundance of different pollinator groups varied over time and space. Floral traits varied among populations. Higher altitude was correlated with bluer flowers. Whiter flowers were associated with the annual presence of hawkmoths. Populations visited by Sphinx vashti had longer spurs than populations visited only by Hyles lineata. Populations with greater percentage of nectar-collecting pollinators did not have whiter, larger flowers with longer spurs.

Conclusions

Despite the large variation in pollinator abundance over time and space, one species of bumble-bee or hawkmoth tended to predominate in each population each year. Future studies of Aquilegia coerulea should examine the specific influences of pollinators and the environment on flower colour and of hawkmoth species on spur length.Key words: Aquilegia coerulea, columbine, pollinator abundance, bumble-bee, hawkmoth, flower colour, spur length, functional pollinator group, altitude, floral trait     

Continuous petal variation in Epimedium tianmenshanense (Berberidaceae), a taxon endemic to western Hunan,China

In the protologue, Epimedium tianmenshanense (Berberidaceae), a species endemic to western Hunan, China, was described as ‘flower small, 0.2–0.4 cm diam., inner sepals white, petals as long as inner sepals or a little shorter than the latter, spur very short, ca 5.0 mm’. However, both morphological characteristics and molecular evidence suggest that E. tianmenshanense is closely related to E. baojingense, a taxon with a long spur, thus suggesting that the size of the floral parts is not as reliable as previously believed. When investigating the variability of E. tianmenshanense in more detail, in the field as well as in cultivation, we found that the petals were are highly variable in morphology (both shape and size), being cucullate, subulate, short to long spurred, and with various transitions. The flowers size varied from small to large accordingly. The flowers with cucullate and subulate petals, which were a little shorter than the inner sepals or almost as long as the latter, were small (about 0.8 cm in diameter). The flowers with long spurs, which were much longer than the inner sepals, were also large (about 2.5–3.5 cm in diameter). Finally, the flowers with short spurs, which were a little longer than the inner sepals, were medium-sized (about 1.0–1.2 cm in diameter). In addition, the color of inner sepals was revised from ‘white, occasionally light mulberry-purple’ to yellowish green or yellowish white. Epimedium tianmenshanense is a perfect example of natural petal evolution, which could be used for further taxonomic and evolutionary studies. The reason for the variation and the taxonomic treatment of the species still need further study.     

基于同质园栽培的21种淫羊藿属植物花部特征的分类学意义

以采自中国多地并同质园栽培于吉首大学种质资源圃的21种淫羊藿为研究材料,通过观测这些物种的花梗长、花直径、内萼片长宽、蜜距长、雌雄蕊长、花药长、花柱长、胚珠数、花粉大小及花粉量等16个花部特征数量性状和花序类型、有无蜜距、花瓣颜色、内萼片颜色、内萼片与花瓣相对长度及花粉颜色6个质量性状,并对其分类学意义进行探讨。结果表明,(1)21种淫羊藿内轮萼片花瓣状,4枚花瓣除无距淫羊藿外均特化成蜜距,但花部形态种间存在显著差异,尤其体现于花直径、内萼片长宽、蜜距长、雌蕊长、胚珠数、花粉量、花粉直径等特征。(2)主成分分析显示花直径、内萼片长、花药长、胚珠数和花粉直径等性状信息负荷量较大,在该属分类中起主要作用。(3)根据花部特征聚类分析,物种间区分良好,21物种可分为4支:第1支花小,花瓣短于内轮萼片,囊状距或兜状距;第2支花较大,花瓣远长于内轮萼片,长距状;第3支花小,花瓣没有特化形成蜜距;第4支花较小,花丝伸长明显,可达10 mm,花蜜距呈钻状。(4)依据所观察到的花部形态特征编写了淫羊藿属21种植物花部特征分种检索表。研究认为,基于同质园栽培下的淫羊藿属物种的花部特征具有分类学意义,可以为该属分类提供一定依据。     

Increases in leaf nitrogen concentration and leaf area did not enhance spur survival and return bloom in almonds (<Emphasis Type="Italic">Prunus dulcis</Emphasis> [Mill.] DA Webb)

Mature almond trees bear fruit mainly on short shoots called spurs, with only a small percentage of fruit produced laterally on long 1-year-old shoots. As a result, maintenance of large numbers of healthy spurs per tree is critical for fruit production. However, spurs that bear fruit have lower leaf area, leaf nitrogen content, and CO₂ assimilation rate than non-fruiting spurs. This has been correlated with reduced percentages of spur survival and return bloom the following season. Thus, we hypothesized that spur leaf area, and ultimately spur health could be enhanced through application of foliar sprays and soil nitrogen treatments that would enhance leaf nitrogen content and spur leaf area. To test our hypothesis, we selected almond trees exhibiting significant yield differences as a consequence of differential soil rates of nitrogen fertilization (N rate) for three prior years (140, 224, and 392 kg/ha). In each tree, three spur types [non-fruiting spurs (F0); spurs with one fruit (F1); spurs with two fruit (F2)] were selected on the east side of the canopy and tracked for one complete season (2011–2012). Four foliar treatments (nutrient replacement, nutrient replacement with biostimulant, nitrogen, and non-spray) were directly applied to individual spurs in each N rate in the spring of 2011 and characteristic such as leaf nitrogen, and fruit quality were recorded throughout the season. In winter of 2012, spur survival and return bloom were addressed through individual visual inspection of the tagged spurs. In this experiment, soil and foliar N treatments effectively increased spur leaf area, fruit, and leaf nitrogen concentration. In the high N treatment, the leaf nitrogen values exceeded the critical nitrogen concentration established for almond trees and the critical leaf area for spur survival and blooming thresholds established by past research in this area. However, none of these positive changes in leaf N or leaf area improved spur survival and/or return boom of any spur type. Indeed, survival and hull + shell weight tended to be lower in the population of fruiting spurs with the highest leaf area and leaf nitrogen concentration and the return bloom probabilities were always lower in fruiting spurs than in non-fruiting spurs, independent of the nitrogen rate. These results and the relationship between nitrogen rate and spur survival are discussed.     

Petal-specific subfunctionalization of an APETALA3 paralog in the Ranunculales and its implications for petal evolution

? The petals of the lower eudicot family Ranunculaceae are thought to have been derived many times independently from stamens. However, investigation of the genetic basis of their identity has suggested an alternative hypothesis: that they share a commonly inherited petal identity program. This theory is based on the fact that an ancient paralogous lineage of APETALA3 (AP3) in the Ranunculaceae appears to have a conserved, petal-specific expression pattern. ? Here, we have used a combination of approaches, including RNAi, comparative gene expression and molecular evolutionary studies, to understand the function of this petal-specific AP3 lineage. ? Functional analysis of the Aquilegia locus AqAP3-3 has demonstrated that the paralog is required for petal identity with little contribution to the identity of the other floral organs. Expanded expression studies and analyses of molecular evolutionary patterns provide further evidence that orthologs of AqAP3-3 are primarily expressed in petals and are under higher purifying selection across the family than the other AP3 paralogs. ? Taken together, these findings suggest that the AqAP3-3 lineage underwent progressive subfunctionalization within the order Ranunculales, ultimately yielding a specific role in petal identity that has probably been conserved, in stark contrast with the multiple independent origins predicted by botanical theories.     

Modelling three-dimensional flow over spur-and-groove morphology

Spur-and-groove (SAG) morphology characterizes the fore reef of many coral reefs worldwide. Although the existence and geometrical properties of SAG have been well documented, an understanding of the hydrodynamics over them is limited. Here, the three-dimensional flow patterns over SAG formations, and a sensitivity of those patterns to waves, currents, and SAG geometry were characterized using the physics-based Delft3D-FLOW and SWAN models. Shore-normal shoaling waves over SAG formations were shown to drive two circulation cells: a cell on the lower fore reef with offshore flow over the spurs and onshore flow over the grooves, except near the seabed where velocities were always onshore, and a cell on the upper fore reef with offshore surface velocities and onshore bottom currents, which result in depth-averaged onshore and offshore flow over the spurs and grooves, respectively. The mechanism driving this flow results from the net of the radiation stress gradients and pressure gradient, which is balanced by the Reynolds stress gradients and bottom friction that differ over the spur and over the groove. Waves were the primary driver of variations in modelled flow over SAG, with the flow strength increasing for increasing wave heights and periods. Spur height, SAG wavelength, and the water depth at peak spur height were the dominant influences on the hydrodynamics, with spur heights directly proportional to the strength of SAG circulation cells. SAG formations with shorter SAG wavelengths only presented one circulation cell on the shallower portion of the reef, as opposed to the two circulation cells for longer SAG wavelengths. SAG formations with peak spur heights occurring in shallower water had stronger circulation than those with peak spur heights occurring in deeper water. These hydrodynamic patterns also likely affect coral and reef development through sediment and nutrient fluxes.

     

The role of genes influencing the corolla in pollination of Antirrhinum majus

Studies of pollination ecology have been hindered by an absence of biochemical information about the basis of polymorphism. Using model plants and mutant lines described by molecular genetics may circumvent this difficulty. Mutation of genes controlling petal colour and petal epidermal cell shape in Antirrhinum majus was previously shown to influence fruit set. White flowers set less fruit than magenta flowers and mutants with flat petal epidermal cells set less fruit than flowers with conical cells. Here we analyse the causal pathway underlying this phenomenon through a study of floral characteristics and bee behaviour. Results indicate that bees recognized plants with magenta conical‐celled flowers at a distance and did not approach white flowers or magenta flat‐celled flowers so frequently. Petal cell shape interacted with colour in determining whether an approaching bee landed on a flower within a plot and whether a bee landing on a flower would probe it. The intrafloral temperature of flowers with conical petal cells was shown to increase with solar irradiance, unlike the intrafloral temperature of flowers with flat petal cells. The difference in fruit set may reflect pollinator discrimination between genotypes as a consequence of the effect of intrafloral temperature on nectar quality and quantity.     

Pollinators and nectar robbers cause directional selection for large spur circle in Impatiens oxyanthera (Balsaminaceae)

Nectar spurs have an important role in floral evolution and plant–pollinator coadaptation. The flowers of some species possess spurs curving into a circle. However, it is unclear whether spur circle diameter is under direct selection pressure from different sources, such as pollinators and nectar robbers. In this study, we quantified selection on some floral traits, such as spur circle diameter in Impatiens oxyanthera (Balsaminaceae) using phenotypic selection analysis and compared the relative importance of pollinators and nectar robbers as selective agents using mediation analysis. The study showed that pollinators caused significant selection on corolla length, spur curvature and spur circle diameter while nectar robbers only imposed strong selection on spur circle diameter. Pollinators favored flowers with large corolla, curly spurs and large spur circle while nectar robbers preferred flowers with small spur circle. More pollinator visits resulted in higher female reproductive success, while robbery reduced female fitness. Conflicting selection on spur traits from pollinators and nectar robbers was not found. Mediation analysis showed that selection on floral traits through nectar robbing was stronger than selection through pollination. The results suggested that pollinators and nectar robbers jointly mediated the directional selection for large spur circle, and nectar robbers caused stronger selection than pollinators on floral traits.     

Are spurred cyathia a key innovation? Molecular systematics and trait evolution in the slipper spurges (Pedilanthus clade: Euphorbia, Euphorbiaceae)

The study of traits that play a key role in promoting diversification is central to evolutionary biology. Floral nectar spurs are among the few plant traits that correlate with an enhanced rate of diversification, supporting the claim that they are key innovations. Slight changes in spur morphology could confer some degree of premating isolation, explaining why clades with spurs tend to include more species than their spurless close relatives. We explored whether the cyathial nectar spur of the Pedilanthus clade (Euphorbia) may also function as a key innovation. We estimated the phylogeny of the Pedilanthus clade using one plastid (matK) and three nuclear regions (ITS and two G3pdh loci) and used our results and a Yule model of diversification to test the hypothesis that the cyathial spur correlates with an increased diversification rate. We found a lack of statistical support for the key innovation hypothesis unless specific assumptions regarding the phylogeny apply. However, the young age (hence small size) of the group may limit our ability to detect a significant increase in diversification rate. Additionally, our results confirm previous species designations, indicate higher homoplasy in cyathial than in vegetative features, and suggest a possible Central American origin of the group.     

Genetics of floral traits influencing reproductive isolation between Aquilegia formosa and Aquilegia pubescens

Abstract: Reproductive isolation between Aquilegia formosa and Aquilegia pubescens is influenced by differences in their flowers through their effects on pollinator visitation and pollen transfer. Here, we investigate the genetic basis of floral characters differentiating these species. We found that in addition to the effects of flower orientation and the length of nectar spurs previously described, other characters such as flower color or odor affect hawkmoth visitation. Repeatability of measurements in an F2 population ranged from 0.53 to 0.83 among five floral traits, indicating that using the means of multiple measures per plant will substantially increase the power of a quantitative trait locus (QTL) analysis. Integration of floral traits was indicated by significant correlations among traits in an F2 population. In a separate F2 population we found that QTL for different floral traits were often closely associated, indicating that linkage or pleiotropy cause at least some of this integration. In addition, we found QTL for all floral traits examined. Because Aquilegia species are largely interfertile and vary extensively in both floral morphology and ecology, they offer the opportunity for QTL studies of a wide range of characters affecting reproductive isolation.     

Flower reshaping in the transition to hummingbird pollination in Loasaceae subfam. Loasoideae despite absence of corolla tubes or spurs

Many angiosperm lineages present transitions from bee to hummingbird pollination. The flower design in most of these lineages includes either corolla tubes or nectar spurs, structures that commonly experienced an elongation with the acquisition of hummingbird pollination. It is proposed that this increases the fit between the bird head and flower structures, and isolates or partially blocks bees from the interaction. But can this transition occur if the ancestral flower design lacks tubes or spurs? Here we focus on the transition from bee to hummingbird pollination in the Loasaceae subfamily Loasoideae. Loasoideae flowers have radial corollas with separated petals; therefore, they do not display corolla tubes nor nectar spurs. These flowers also present a whorl of nectar scales and staminodes, unique to the subfamily, which is involved in flower–pollinator fit and in nectar harvesting. To explore flower shape adaptation to hummingbird pollination, we tested for correspondence between pollinators and flower shape in Loasoideae. In order to achieve this, we first compared the evolutionary history of flower phenotype and pollination mode, and then used stochastic character mapping and geometric-morphometric variables in a comparison of alternative evolutionary models. The results of our study suggest that the transition from bee to bird pollination was accompanied by changes in the shape of the staminodial complex, along with the evolution of relatively closed corollas. Moreover, while bird pollination seems to be the end point in the evolution of pollination syndromes in many angiosperm lineages, rodent pollinated flowers probably evolved from ancestral bird pollinated flowers in Loasoideae. Our findings suggest that the evolution of bird pollinated flowers from ancestral bee pollinated flowers does not require the presence of corolla tubes or spurs, and can take place as long as the flower design includes structures participating in flower–pollinator fit.     

Floral isolation between Aquilegia formosa and Aquilegia pubescens

The acquisition of floral nectar spurs is correlated with increased species diversity across multiple clades. We tested whether variation in nectar spurs influences reproductive isolation and, thus, can potentially promote species diversity using two species of Aquilegia, Aquilegia formosa and Aquilegia pubescens, which form narrow hybrid zones. Floral visitors strongly discriminated between the two species both in natural populations and at mixed-species arrays of individual flowers. Bees and hummingbirds visited flowers of A. formosa at a much greater rate than flowers of A. pubescens. Hawkmoths, however, nearly exclusively visited flowers of A. pubescens. We found that altering the orientation of A. pubescens flowers from upright to pendent, like the flowers of A. formosa, reduced hawkmoth visitation by an order of magnitude. In contrast, shortening the length of the nectar spurs of A. pubescens flowers to a length similar to A. formosa flowers did not affect hawkmoth visitation. However, pollen removal was significantly reduced in flowers with shortened nectar spurs. These data indicate that floral traits promote floral isolation between these species and that specific floral traits affect floral isolation via ethological isolation while others affect floral isolation via mechanical isolation.