The dry and wet stigmas ofPrimula obconica: Ultrastructural and cytochemical dimorphisms

Summary The stigmatic papillae of the distylous speciesPrimula obconica are studied by means of cytochemical, light and electron microscopic techniques. The papillae on thrum stigmas are smaller than those on pin stigmas. At the bud stage, secretory vesicles are not a conspicious part of the cytoplasm, although certain signs of secretory activity are present. The young papillae bear the thin, superficial pellicle typical to dry stigmas. Small vesicles are numerous in mature papillae of both morphs, and seem to originate from the ER. A layer of closely packed, osmiophilic globuli is present in the outermost part of mature walls of pin papillae. At sites with cuticle disruption, the globuli seem to migrate outwards to be incorporated with the copious, blistery exudate. Due to this exudate the pin stigma is characterized as wet. Cytochemical tests suggest that the exudate contains mainly lipids, and different carbohydrates and protein are detected. It reacts positively in tests for peroxidase, acid phosphatase, and non-specific esterases. The thrum stigma remains dry at maturity, with a distinct pellicle also reacting positively to the enzyme tests. Only a few, scattered osmiophilic globuli, sized and situated as those in the pin papillae walls, are found in the thrum walls, and they do not form a proper layer.Thus the generally accepted correlation between dry stigmas and the sporophytic kind of self-incompatibility system is not substantiated withinP. obconica, and the possible influence of the dimorphisms to the pollen/stigma interaction is discussed.     

The control of incompatibility in distylous Pulmonaria affinis Jordan (Boraginaceae)

In P. affinis, pin pollen is shorter on average than thrum pollen. Pins have more coronal hooks on stigmatic papillae than thrums, but gaps between papillae are relatively smaller for thrums. Pin stigmas receive more pollen than thrum stigmas. Thrum stigmas receive more (dissortive) pin pollen, but pin stigmas are assortively pollinated. Pollen only germinates when trapped below papilla coronas. On thrum stigmas, most trapped pollen is pin. Pollen germination is better on thrum stigmas than pin stigmas, and thrum stigmas show a close relationship between numbers of legitimate pollen grains, numbers of germinating grains, and numbers of pollen tubes in the style. There is no inhibition of illegitimate pollen germination. Illegitimate pollen tubes are inhibited in the style. Incompatibility operates by a combination of dissortive pollination, dissortive pollen trapping, and stylar pollen tube inhibition. All heteromorphic features differing between pins and thrums are implicated in the inhibition of within-morph fertilization in thrums.     

Fine Structure and Cytochemistry of the Stigma Surface and Incompatibility in some Distylous Linum Species

The receptive surface of the stigma in distylous Linum grandiflorumand L. pubescens was studied by electron microscopy and cytochemicaltechniques. In both floral morphs a proteinaceous-lipoidal coatingis present on the papilla surface. In thrum stigmas the cuticleis highly irregular and pitted at the papilla tip. The cuticleis dislodged and torn at anthesis and an osmiophilic secretionproduct is released within a pectinaceous matrix. The secretionproduct stains for proteins and lipids and contributes to adhesionof pollen. In the larger pin papillae the cuticle is wavy, continuous,thicker than in thrum papillae and adjoins the cell wall. Inboth species the surface of the two types of pollen grains iscoated with lipids and protein. A similar behaviour of the male gametophyte is observed in incompatiblepollinations of L. pubescens, L. mucronatum and L. grandiflorum.In intramorph thrum pollinations pollen tubes are arrested withinthe stigma. In intramorph pin pollinations the majority of pollengrains fail to adhere to the stigma. Low permeability to waterin pin papillae, as determined by the neutral red test, maybe a factor preventing imbibition of the few adhering grains.Tubes of the few germinated grains are inhibited inside thestigma. On the part of the stigma, the difference in the major siteof inhibition in the two intramorph incompatible combinationsmay be accounted for by the dissimilar properties of the papillae,i.e. the occurrence of wet thrum stigmas and dry pin stigmas.Functionally, the unusual association of sporophytic incompatibilitywith wet thrum stigmas is attributed to retention of the secretorymaterial on individual papillae. Stigmatic papillae, cuticle, pollen coat, distyly, incompatibility, Linum grandiflorum, L. pubescens, L. mucronatum     

Pollen flow inPrimula veris (Primulaceae)

Observations on two natural and one artificial distylous populations ofPrimula veris in Britain revealed very high proportions of intramorph pollen on pin stigmas and moderate proportions of intermorph pollen on thrum stigmas. The average ratio of pin: thrum pollen production per flower was 2.4:1. Large amounts of pollen remained in dehisced anthers, perhaps reflecting low levels of pollinator activity. Population size and density had no clear effect on pollen flow patterns, which are asymmetrical for the two morphs.     

An unusual homostyle inHedyotis caerulea (Rubiaceae)

Hedyotis caerulea, a spring flowering herb widely distributed in eastern North America, has distylous flowers that differ by a number of morphological and physiological traits. The presence of a strong incompatibility system is indicated by the fact that intramorph crosses or self-pollinations produced little or no seed and intermorph crosses produced copious seed. An unusual homostyle was located that had most floral characters intermediate between pin and thrum flowers, although its pollen size was that of pin. The homostyle was only moderately self-compatible, and its pollen did not behave as pin pollen on thrum stigmas. It appears that, despite their intermediate position and morphology, the homostyle stigmas were fundamentally pin. Although the exceptional rarity of the homostyle suggests that it is an inadaptive failure, workers should watch for its further occurrence since additional study of it may provide insights into the genetic and physiological basis of heterostyly in theRubiaceae.     

Heterostyly and Pollen-tube Interactions in Luculia gratissima (Rubiaceae)

Observations on the floral biology of Luculia gratissima (Rubiaceae)showed that this species is distylous with complementary positioningof anthers and stigmas in the two floral forms. Unusual featuresof distyly in this species include the larger size of the corolla,the stigma surface and the stigmatic papillae in the thrum flowerscompared to the pin ones. Stigmatic surfaces have similar secretionsbut they appear more copious in thrum than pin. The floral dimorphismwas accompanied by a very effective self-incompatibility systemand no seed was set on selfing. Seed number per capsule on crossingwas significantly greater in thrum flowers compared to pin.Incompatible pollen tubes were inhibited within 24 h at thebase of the stigma/top of the style in both morphs. Amputationof this region of the gynoecium removed the self-incompatibilityreaction in thrum but not pin flowers. Pollination with a mixtureof compatible and incompatible pollen and sequential pollinationwith self followed by cross pollen showed that there were interactionsbetween the two types of pollen tube. The presence of compatibletubes was found to cause the excessive swelling of the pollen-tubetip of the incompatible ones. The incompatible tubes did notappear to have any effect on the growth of compatible ones. Luculia gratissima, distyly, floral biology, self-incompatibility, pollen-tube interactions     

Self-compatibility in distylousHedyotis salzmannii (Rubiaceae)

Hedyotis salzmannii (Rubiaceae) is distylous. Experimental pollination revealed that both morphs are self-compatible. However, the thrum morph proved to be more self-compatible. Hand cross-pollination between different plants of the same floral form produced less seed-sets per pollination than hand cross-pollination between pin and thrum or thrum and pin pollination. Two species ofHymenoptera, oneApidae and oneHalictidae, and two species ofDiptera (Syrphidae) were observed as pollinators.     

HETEROSTYLY,POPULATION COMPOSITION,AND POLLEN FLOW IN HEDYOTIS CAERULEA

Hedyotis caerulea is a perennial, spring-flowering herb native to eastern North America with distylous flowers that differ in a number of morphological and physiological traits. However, the pin and thrum morphs produce the same numbers of buds, flowers, and fruits per plant, although it is possible that differences in these may occur in some populations at certain times of the flowering period. The two morphs are self-incompatible and cross-compatible. Most populations contained an excess of pins over thrums (anisoplethy); less commonly pins and thrums were equally represented (isoplethy). Populations change from anisoplethy to isoplethy and in the reverse direction. The spatial distribution of pin and thrum flowers in populations was random in some populations but non-random in others. There is some indication that the two morphs in some populations have somewhat different flowering periods. Pollinators seem to be chiefly bombyliid flies and perhaps thrips, but insects were rarely observed visiting flowers. In some populations, the two morphs produced equal numbers of pollen grains per flower; in others they did not. The average pollen viability varied, but on the average there was a moderate level of pollen sterility. High numbers of pollen grains remained in dehisced anthers, probably as a result of low pollinator activity. However, between 5% and 9% of the pollen produced participated in pollination. Stigmas of most pin flowers received more pin pollen grains than thrum pollen grains; on stigmas of thrum flowers pin pollen grains outnumbered thrum pollen grains. Thus, compatible pollen flow from pins to thrums was greater than in the reverse direction. Anisoplethic and isoplethic populations had the same pollen flow patterns. A plant-by-plant examination of stigmas indicated that many stigmas bore few or no pollen grains. Seed production of the two morphs was equal. Despite the inequities in pollen flow patterns, the widespread and occasionally weedy nature of H. caerulea suggests that its breeding system must be viewed as a successful one.     

POLLEN FLOW AND FECUNDITY IN POPULATIONS OF LITHOSPERMUM CAROLINIENSE

Stigmatic pollen loads of pin and thrum forms of distylous Lithospermum caroliniense (Walt.) MacMill. generally have very large proportions of incompatible pollen grains. Assortative pollination probably results from large components of self-pollination rather than geitonogamous pollinations, because pollen loads of large and small individuals are the same, although large individuals would be expected to experience more geitonogamous pollinator flights than small individuals. The distylous reproductive system appears relatively insensitive to changes in pollinator activity and preferential pollen flow between pins and thrums cannot be demonstrated. In contrast to the relatively uniform proportions of pin and thrum pollen on stigmas, absolute pollen loads vary substantially in different years and localities. Extremely low fecundity, which appears to be characteristic of this species, may result in part from lack of pollinator activity. However, even under optimal conditions for pollination, fecundity is less than 20%. Although L. caroliniense appears to have an inefficient pollination system, intrinsic low fecundity may represent a more critical feature of the life history.     

Pollen-pistil interaction in Linum grandiflorum

A detailed investigation of the receptive surface of the stigma of a dimorphic taxon, Linum grandiflorum, was carried out using scanning electron microscopy, cytochemistry and polyacrylamide gel electrophoresis. The stigma surfaces of the pin and thrum morphs showed distinct differences. The stigma of the pin morph was of the dry type and the papillae were covered with a uniform cuticle-pellicle layer. The stigma of the thrum morph, on the other hand, resembled the wet type; the cuticle-pellicle layer was disrupted at places and a secretion product was released onto the surface of the stigma. Coomassie blue staining material was present on the surface of only the thrum stigma. Although esterases and acid phosphatases were present on the stigma of both the morphs, their activity was invariably higher on the thrum stigma. Polyacrylamide gel electrophoresis of stigma leachates also showed distinct differences in the protein profiles of the two morphs.     

FEATURES OF POLLEN FLOW IN DIMORPHIC SPECIES OF LYTHRUM SECTION EUHYSSOPIFOLIA

The North American Lythrum lineare L., L. alatum Pursh, L. curtissii Fernald, and L. californicum T. & G. have distylous flowers that differ in a number of traits. In the two species examined, pins have a stronger incompatibility system than thrums. Pins outnumber thrums in natural populations, but the ratio of the two may change radically during the course of a year, probably as a result of sexual reproduction. Exotic honeybees visiting L. californicum exhibited preferential collection of pin pollen over thrum pollen. The relative proportion of pin and thrum pollen grains transferred to stigmas is not dependent upon the proportion in which these pollen grains are produced in a population. Analyses of stigmatic pollen loads of L. lineare and L. californicum indicate very high levels of intramorph pollen flow. Although distyly is likely derived from tristyly in Lythrum, pollen flow patterns in trimorphic species seem to operate more “efficiently” than in dimorphic ones. It is probable that distyly has evolved from tristyly several times in the Lythraceae, but comparisons between pollen flow patterns in distylous species and those in a tristylous species provide no clear insights into the selective pressures that have led to this fundamental change in floral morphology and reproductive system.     

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.     

Differential localization of enzymes in the stigmatic exudates ofPrimula obconica

Summary The stigmatic dimorphisms in the distylous speciesPrimula obconica, which has been dealt with in an earlier paper, has been described further. An ultrastructural localization of various enzymes in the exudates is hereby revealed, and evidence is given for intermorph differences. Esterases are confined to the lipid phase of the exudates, including the lipid globuli in the papillae walls of both morphs, but they are not found in the pellicle of the dry thrum stigma. However, this pellicle exhibits acid phosphatase activity, as does the lumen of the blisters in the viscous exudate on pin stigmas. The blisters are presumed to hold the watery phase of the pin secretion. From the present findings and from previous results with LM cytochemical methods it is suggested that the sites of peroxidase activity resemble those of acid phophatases.     

Heterostyly inPrimula. 1. Fine-structural and cytochemical features of the stigma and style inPrimula vulgaris huds

Summary The stigmas of the heterostylous genusPrimula are of the dry type without a free-flowing surface secretion. The papillae of the stigma surface cells of the two morphs, in pin (stigma exserted) and thrum (stamens exserted), bear a thin proteinaceous surface pellicle, overlying a discontinuous cuticle. The vacuoles of the papillate cells contain tannins, and tannin cells extend in files through the stigma heads and form a loose sheath surrounding the pollen-tube transmitting tract in the styles. The cells of the transmitting tissue in the stigma heads have a normal complement of organelles, and abundant ribosomal endoplasmic reticulum. The intercellular spaces contain an internal secretion which reacts cytochemically for both carbohydrate and protein. The transmitting tract in the styles forms a central core surrounded by several vascular strands. The cells are elongated, and the intercellular spaces here also have a carbohydrate-protein content. In a compatible pollination, thrum pollen tubes enter the stigma by penetrating the cuticle at the tip or on the flank of the pin papilla. Pin tubes on the thrum stigma enter between adjacent papillae, penetrating the thin cuticle at the base. The tubes grow through the transmitting tracts in the intercellular material.     

INCOMPATIBILITY AND THE POLLEN ECONOMY OF JEPSONIA PARRYI

Jepsonia parryi (Saxifragaceae) has heterostylous flowers and is strongly self-incompatible. Pin flowers have long styles, large stigmas, short stamens, and numerous, small pollen grains with finely sculptured walls. Thrum flowers have short styles, small stigmas, long stamens, and fewer, larger pollen grains with coarsely sculptured walls. Pin plants and thrum plants occur in a 1:1 ratio in field populations. Although the insect pollinators of J. parryi transfer ample compatible pollen to pin and thrum stigmas to account for full seed production, much of the pollen deposited on stigmas is incompatible. Analysis of the pollen deposits on stigmas collected from field populations indicates that compatible “legitimate” pollination of pin and thrum flowers is essentially random and is not obviously aided by floral dimorphism. It is suggested that although heterostyly had a positive adaptive value in the past evolutionary history of Jepsonia it is no longer adaptive under the present pollination regime, although it is maintained because of its strong genetic fixity.     

绣球茜的二型花柱及其传粉生物学初步研究

首次对绣球茜(Dunnia sinensis Tutch)的花药和柱头高度、花粉和柱头形态,以及人工辅助授粉的结实情况等进行了研究.结果表明:绣球茜在同一居群中同时具有长花柱(Pin 型)和短花柱(Thrum)两种花型,长花柱植株的柱头和雄蕊高分别为9.61±0.36 mm和6.79±0.38 mm,短花柱则为6.11±0.56 mm和8.96±0.59 mm,表现出互补式雌雄异位的花部特征.通过人工辅助授粉,两种花型植株的自交和同型植株间异交都不结实,仅在型间异交方能正常结实,说明绣球茜中存在极强的自交不亲和和型内不亲和现象.在花粉与柱头形态方面:长柱型花的单花花粉量(38555.6±6258.4)显著地小于短柱型花(52145.45±8924.5)(t=4.009,P＜0.01),长柱花和短柱型花的胚珠数分别为38.73±1.85和39.1±2.25(t=-4.13,P＞0.01);长柱型花花粉粒的大小(极径21.27±0.70 μm)显著地小于短柱花(21.92±0.79μm)类型(t=3.890,P＜0.01),而柱头乳突大小长柱类型要大于短柱类型,表现出明显的二型性.研究结果表明绣球茜是一种典型的二型花柱植物.此外,对其传粉昆虫及昆虫访花行为的观察表明,蝶类(青风蝶属Graphium)和蜂类(熊蜂Bombus sp.和木蜂Xylocopa magnifica Smith)为绣球茜的主要访花昆虫和传粉昆虫,它们能够在两种不同花型植株之间有效地传粉.     

Comparative reproductive biology of Primula merrilliana Schltr. and P. cicutariifolia Pax

The reproductive biology of Primula merrilliana Schltr. and P. cicutariifolia Pax that are sister taxa and the only two members of Primula sect. Ranunculoides were studied. The results showed that P. merrilliana is typically a distylous species not only in morphology but also in function; it has an incompatibility system that restricts the legitimate pollination to that between pin and thrum flower morphs, but self-fertilization may occur under certain conditions. P. cicutariifolia is self-compatible. The pin morph of P. merrilliana has longer stigma papillae and smaller pollen than the thrum morph. The results stated above support the view that P. filchnerae should be moved from sect. Pinnatae to sect. Auganthus.     

Asymmetry in pollen flow promotes gender specialization in morphs of the distylous neotropical herb <Emphasis Type="Italic">Arcytophyllum lavarum</Emphasis> (Rubiaceae)

Distyly is a floral polymorphism, characterized by a reciprocal positioning between stigmas and anthers in different flowers, where two floral morphs, long-styled (pin) and short-styled (thrum) occur within the population. Distyly is suggested as one of the routes leading to the evolution of separate sexes in plants. In this evolutionary pathway, pollinators may disrupt the complementarity of pollen transfer between morphs. Consequently, the floral morphs gradually specialize as either male or female. A key process required for gender specialization in distylous plants is a deviation of the realized functional gender (i.e. the proportion of genes transmitted to the next generation via pollen donation and seed production) from the potential functional gender (i.e. the expected contribution of male and female function to reproductive success from the number of ovules or pollen grains produced by each morph). I selected the distylous herb Arcytophyllum lavarum (Rubiaceae) to determine if asymmetry in pollen flow promotes differences in seed production, pollen donation and a discrepancy between the potential and the realized functional genders in pin and thrum floral morphs. Pollen flow in A. lavarum is asymmetric and the pin morph is more efficient at performing cross-pollination than the thrum morph. Conversely, the thrum morph produced two times more seeds than the pin morph. Male and female contributions to the potential functional gender were equivalent in both morphs. However, the pin morph transmitted more genes through pollen donation and the thrum morph more through seed production than expected from their potential functional genders. These results support the hypothesis that if pollinators consistently promote asymmetric pollen flow between morphs over generations, it is possible that gender specialization may evolve to the extreme of dioecism from an original distylous condition.     

濒危植物安徽羽叶报春两种花型的繁育特性及其适应进化

二型花柱的维持机制和自然选择压力多年来一直是生态学和进化学研究领域的热点之一。通过实验室栽培和野外观察统计相结合的方法,对安徽羽叶报春两种花型(长柱花和短柱花)的形态特征、花粉活力、柱头可授性、花粉/胚珠比、自然授粉及结籽能力、自交亲和性等繁育特性进行了比较研究。结果表明:长、短柱花的花冠直径和裂片宽无明显差异,而花冠筒、雌蕊和雄蕊高、花粉数目及大小、P/O比均有显著差异。在自然条件下,长柱花所接受的总花粉数要明显高于短柱花的总花粉数,但所接受的异型花花粉数和平均每果结籽数两者无显著差异。长柱花和短柱花的花粉和柱头活力相似,均能在较长时间内维持较高活力,仅在开花末期显著下降。两种花型的花在自花授粉、同型异花授粉、异型花授粉条件下均能结籽,但异型花授粉的结籽数均明显高于自花授粉和同型异花授粉结籽数。在长柱花各种授粉方式中,花粉萌发率无明显差异,但异型花花粉管的生长速度明显比同型异花花粉和自花花粉的快,而在短柱花柱头上表现为异型花授粉的萌发率最高,但只要萌发后在花柱中的生长速度无显明差异。此外,综合上述结果,对二种花型花部综合征的维持机制及自然选择压力进行了探讨。     

Cryptic self‐incompatibility and distyly in Hedyotis acutangula Champ. (Rubiaceae)

Distyly, floral polymorphism frequently associated with reciprocal herkogamy, self‐ and intramorph incompatibility and secondary dimorphism, constitutes an important sexual system in the Rubiaceae. Here we report an unusual kind of distyly associated with self‐ and/or intramorph compatibility in a perennial herb, Hedyotis acutangula. Floral morphology, ancillary dimorphisms and compatibility of the two morphs were studied. H. acutangula did not exhibit precise reciprocal herkogamy, but this did not affect the equality of floral morphs in the population, as usually found in distylous plants. Both pin and thrum pollen retained relatively high viability for 8 h. The pollen to ovule ratio was 72.5 in pin flowers and 54.4 in thrum flowers. Pistils of pin flowers remained receptive for longer than those of thrum flowers. No apparent difference in the germination rate of pin and thrum pollen grains was observed when cultured in vitro, although growth of thrum pollen tubes was much faster than that of pin pollen tubes. Artificial pollination revealed that pollen tube growth in legitimate intermorph crosses was faster than in either intramorph crosses or self‐pollination, suggesting the occurrence of cryptic self‐incompatibility in this species. Cryptic self‐incompatibility functioned differently in the two morphs, with pollen tube growth rates after legitimate and illegitimate pollination much more highly differentiated in pin flowers than in thrum flowers. No fruit was produced in emasculated netted flowers, suggesting the absence of apomixis. Our results indicate that H. acutangula is distylous, with a cryptic self‐incompatibility breeding system.