Nectar and pollination drops: how different are they?

Background

Pollination drops and nectars (floral nectars) are secretions related to plant reproduction. The pollination drop is the landing site for the majority of gymnosperm pollen, whereas nectar of angiosperm flowers represents a common nutritional resource for a large variety of pollinators. Extrafloral nectars also are known from all vascular plants, although among the gymnosperms they are restricted to the Gnetales. Extrafloral nectars are not generally involved in reproduction but serve as ‘reward’ for ants defending plants against herbivores (indirect defence).

Scope

Although very different in their task, nectars and pollination drops share some features, e.g. basic chemical composition and eventual consumption by animals. This has led some authors to call these secretions collectively nectar. Modern techniques that permit chemical analysis and protein characterization have very recently added important information about these sugary secretions that appear to be much more than a ‘reward’ for pollinating (floral nectar) and defending animals (extrafloral nectar) or a landing site for pollen (pollination drop).

Conclusions

Nectar and pollination drops contain sugars as the main components, but the total concentration and the relative proportions are different. They also contain amino acids, of which proline is frequently the most abundant. Proteomic studies have revealed the presence of common functional classes of proteins such as invertases and defence-related proteins in nectar (floral and extrafloral) and pollination drops. Invertases allow for dynamic rearrangement of sugar composition following secretion. Defence-related proteins provide protection from invasion by fungi and bacteria. Currently, only few species have been studied in any depth. The chemical composition of the pollination drop must be investigated in a larger number of species if eventual phylogenetic relationships are to be revealed. Much more information can be provided from further proteomic studies of both nectar and pollination drop that will contribute to the study of plant reproduction and evolution.Key words: Nectar, pollination drop, ovular secretion, plant reproduction, proteins, sugars, gymnosperms, angiosperms, plant–animal interaction     

Pollination drop in Juniperus communis: response to deposited material

BACKGROUND AND AIMS: The pollination drop is a liquid secretion produced by the ovule and exposed outside the micropyle. In many gymnosperms, pollen lands on the surface of the pollination drop, rehydrates and enters the ovule as the drop retracts. The objective of this work was to study the formation of the pollination drop in Juniperus communis, its carbohydrate composition and the response to deposition of conspecific pollen, foreign pollen and other particulate material, in an attempt to clarify the mechanism of pollination drop retraction. METHODS: Branches with female cones close to pollination drop secretion were collected. On the first day of pollination drop exposure, an eyelash mounted on a wooden stick with paraffin was used to collect pollen or silica gel particles, which were then deposited by contact with the drop. Volume changes in pollination drops were measured by using a stereomicroscope with a micrometer eyepiece 3 h after deposition. The volume of non-pollinated control drops was also recorded. On the first day of secretion, drops were also collected for sugar analysis by high-performance liquid chromatography. KEY RESULTS: The pollination drop persisted for about 12 d if not pollinated, and formed again after removal for up to four consecutive days. After pollination with viable conspecific pollen, the drop retracted quickly and did not form again. Partial withdrawal occurred after deposition of other biological and non-biological material. Fructose was the dominant sugar; glucose was also present but at a much lower percentage. CONCLUSIONS: Sugar analysis confirmed the general trend of fructose dominance in gymnosperm pollination drops. Complete pollination drop withdrawal appears to be triggered by a biochemical mechanism resulting from interaction between pollen and drop constituents. The results of particle deposition suggest the existence of a non-specific, particle-size-dependent mechanism that induces partial pollination drop withdrawal. These results suggest that the non-specific response may decrease the probability of pollen landing on the drop, reducing pollination efficiency.     

POLLINATION DROP IN RELATION TO CONE MORPHOLOGY IN PODOCARPACEAE: A NOVEL REPRODUCTIVE MECHANISM

Observation of ovulate cones at the time of pollination in the southern coniferous family Podocarpaceae demonstrates a distinctive method of pollen capture, involving an extended pollination drop. Ovules in all genera of the family are orthotropous and single within the axil of each fertile bract. In Microstrobus and Phyllocladus ovules are erect (i.e., the micropyle directed away from the cone axis) and are not associated with an ovule-supporting structure (epimatium). Pollen in these two genera must land directly on the pollination drop in the way usual for gymnosperms, as observed in Phyllocladus. In all other genera, the ovule is inverted (i.e., the micropyle is directed toward the cone axis) and supported by a specialized ovule-supporting structure (epimatium). In Saxegothaea there is no pollination drop and gametes are delivered to the ovule by pollen tube growth. Pollination drops were observed in seven of the remaining genera. In these genera the drop extends over the adjacent bract surface or cone axis and can retain pollen that has arrived prior to drop secretion (“pollen scavenging”). The pollen floats upward into the micropylar cavity. The configuration of the cone in other genera in which a pollination drop has not yet been observed directly suggests that pollen scavenging is general within the family and may increase pollination efficiency by extending pollination in space and time. Increased pollination efficiency may relate to the reduction of ovule number in each cone, often to one in many genera, a derived condition. A biological perspective suggests that animal dispersal of large seeds may be the ultimate adaptive driving force that has generated the need for greater pollination efficiency.     

Significance of exine shedding in Cupressaceae-type pollen

In conifers, which have non-saccate Cupressaceae-type pollen, the pollen must land on a pollination drop or be picked up by the pollination drop from the surface of the cone near the ovule before it can be taken into the ovule. After contact with the drop, the pollen intine absorbs moisture from the drop, expands and the exine is shed. In this study the significance of the shedding of the exine is interpreted from experiments in which simulated pollination drops and micropyles were used to determine the movement of pollen and other particles in suspension. The non-expanded pollen, which can be observed upon contact with the pollination drop, sheds the exine, which then functions as a non-elastic particle, while the pollen from which the exine was shed swells and functions as an elastic particle because it is enclosed by the flexible intine. Non-elastic particles are not easily transferred through narrow passages (the micropyle and micropylar canal) and tend to plug these passages. However, elastic particles, such as the swollen pollen, are easily transferred along narrow passages even when non-elastic particles are present. The simulated experiments demonstrate that exine shedding is an important feature in getting pollen through the narrow micropyle and micropylar canal to the nucellus of the ovule.     

Postpollination drop production in hybrid larch is not related to the diurnal pattern of xylem water potential

The hourly production of postpollination drops in the ovules of three hybrid larch trees (Larix x. marschlinsii Coaz) was examined and compared with the measured diurnal pattern of xylem water potential of the same trees under non-stressed conditions during two reproductive seasons. There was no consistent relationship between xylem water potential and ovular drop production in hybrid larch. Individual trees that showed a diurnal drop production in one year did not follow a similar pattern the other year. One tree produced drops in over half of its ovules hourly, regardless of the measured xylem water potential. Variation in number of drops produced per ovule and drop mass was observed between the three trees both years, and between years for two of the trees. This variation was not in response to xylem water potential or environmental conditions. Here we report the first simultaneous measurements of branch water status and the production of ovular secretions for any conifer. We conclude that the postpollination drop production of hybrid larch is not regulated by a tree's overall water status, but is under the control of localized structures such as the cone or the ovule.     

侧柏和北美香柏(柏科)的传粉机制(英)

观察了侧柏（Phaycladusorientalis（L．）Franco）和北美香柏（ThujaoccidentalisL．）散粉后花粉进入珠孔的过程。在散粉期,这两种植物的胚珠均分泌出传粉滴。当花粉落到传粉滴上后,引起传粉滴表面的形状发生改变或减弱胚珠的继续分泌,使得该传粉滴蒸腾加快,导致其比未授粉的传粉滴明显收缩。观察结果表明：不同植物的花粉导致侧柏传粉滴的收缩速率不尽相同。其中,与侧柏亲缘关系较近的植物花粉引起传粉滴的收缩速率和侧柏自身花粉引起的传粉滴收缩速率相似;反之,收缩速率变慢。侧柏传粉滴的收缩可能主要是由于花粉减弱胚珠分泌的结果。     

Proteomic evaluation of gymnosperm pollination drop proteins indicates highly conserved and complex biological functions

The pollination droplet is a highly conservative pollination mechanism that is observed in all major gymnosperm taxa. Proteomics analysis of the pollination drops was carried out on four gymnosperm species: Juniperus communis (common juniper), Juniperus oxycedrus (prickly juniper), Chamaecyparis lawsoniana (Port Orford cedar), and Welwitschia mirabilis. Pollination drop proteins were purified by SDS-PAGE, and the most abundant proteins were analyzed by mass spectrometry and sequenced. Based on BLAST searching of combined amino acid sequences, the following proteins were identified in the following species: an 83-kDa subtilisin-like proteinase, a 62-kDa glycosyl hydrolase, a 47.5-kDa glucan 1,3-β-glucosidase precursor, a 30-kDa chitinase, and a 25-kDa thaumatin-like protein were identified in J. communis; a 30-kDa chitinase, a 25-kDa thaumatin-like protein, and a 32.5-kDa glucanase-like protein were identified in J. oxycedrus; an 83-kDa subtilisin-like proteinase, a 62-kDa β-d-glucan exohydrolase, a 47.5-kDa glucan 1,3-β-glucosidase, and two 25-kDa thaumatin-like proteins were identified in C. lawsoniana, and a 25-kDa chitinase was identified in W. mirabilis. Based on protein identifications, there is strong evidence that the pollination drop functions in both pathogen defense and pollen development. The discovery of similarities in terms of peptide sequence and protein identifications indicates that ovular secretions are functionally conservative, and that they are essential to reproductive success.     

INFLUENCE OF THE PISTIL ON POLLEN TUBE KINETICS IN PEACH (PRUNUS PERSICA)

Pollen tube growth has been studied in peach and has been related to changes in the pistil structures which the pollen tube has to traverse in its way from the stigma down to the ovule. Growth of the pollen tubes along the pistil is not continuous. While pollen tubes reach the base of the style 7 days after pollination, fertilization does not take place until 12 days later. Pollen tubes stop for 5 days at the top of the obturator and they further stop for 3 days before entering the ovule. The pollen tube growth is heterotrophic; starch, present all along the pistilar tract at anthesis, vanishes as the pollen tubes pass by. Discontinuous pollen tube growth appears to be controlled by the pistil. At anthesis the pistil is not fully matured. Maturation of the pistil implies a number of secretory processes that occur in a basipetal way starting from the stigma down to the style and ending in the ovule. Some of these secretions at the stigma and the style are triggered by pollination; others appear to be a maturative stage of the pistil and are produced in a discrete way. The fact that the pollen tube depends on these secretions together with the fact that these secretions are not continuously produced confer upon the pistil a role of controlling pollen tube kinetics and point out that, for a successful fertilization, male gametophyte development and pistil maturation need to by synchronized.     

云南红豆杉传粉生物学研究

云南红豆杉（Taxus yunnanens／s）是分布于我国西南部及不丹和缅甸北部的红豆杉属植物。用石蜡切片法、电镜扫描和整体透明染色法观察了云南红豆杉成熟小孢子和胚珠的结构,在野外观测了云南红豆杉的花期及传粉滴的活动规律。结果显示：云南红豆杉为单核小孢子,没有气囊,有（无）活力的小孢子水合后都急剧膨胀,胚珠单个生于叶腋。云南红豆杉雄株的散粉期在11月中旬到12月中旬,胚珠下弯期从11月下旬持续到2月中旬。在雄株散粉期内及人工授粉的传粉滴持续存在的时间较短,而且授粉后不再出现,在非散粉期内的传粉滴持续存在的时间较长,而且在消失后又会重新出现。红豆杉属植物可能的传粉机制：（1）传粉滴大而且裸露,持续存在的时间较长,有助于提高传粉效率。（2）小孢子能在常温下较长时间的保持活性,不具备远距离飞行的特征,有利于在一定范围内保持较高的小孢子密度;（3）虽然小孢子散粉期短且易变,但胚珠的成熟期较长,从而保证了花期可遇。传粉期内的降水和低的种群密度可能是影响红豆杉传粉效率的主要因素。     

Contrasted pollen capture mechanisms in Phyllocladaceae and certain Podocarpaceae (Coniferales)

Comparative study shows that Phyllocladus and representative Podocarpaceae differ in the mechanism by which pollen is introduced into the pollen chamber and onto the apex of the nucellus ("pollen capture"). Both types involve a pollination drop, but only in Podocarpaceae is it consistently inverted and in contact with adjacent surfaces. Phyllocladus has functionally nonsaccate pollen (although a vestigial saccus has been claimed); its pollen is wettable and sinks in water. Podocarpaceae (except Saxegorhaea) have saccate pollen, which is nonwettable and floats on water. In Phyllocladus the pollination drop receives the pollen directly and presence of pollen stimulates complete drop withdrawal, which may be a metabolic process. Once pollinated, an ovule does not resecrete a pollination drop. In Podocarpaceae the drop usually receives the pollen indirectly via pollen scavenging and saccate pollen is preferentially captured. The retraction of the drop appears to be the result of evaporation and is presumably nonmetabolic. Drop secretion can be repeated in the presence of pollen. A major consequence of these contrasted mechanisms is that in Phyllocladus the entire contents of the pollination drop are ingested, whereas in Podocarpaceae only that part of the drop that includes saccate pollen is ingested. Because of differences in repeatability of the secretion process, Podocarpaceae are likely to capture more pollen. In neither mechanism does the process favor 'own" pollen. but in Podocarpaceae all but saccate pollen is excluded. We thus have further evidence for differences in pollen capture mechanisms in conifers with a pollination drop, and differences in the behavior of the pollination drop itself.     

Orientation and withdrawal of pollination drops in Cupressaceae s. l. (Coniferales)

Pollination in the Cupressaceae is studied ex situ, focused on orientation and withdrawal of pollination drops. Orientation of pollination drops is a constant feature in most taxa studied and important for pollen capture. Conspecific pollen causes a withdrawal of pollination drops, varying in time among species from 8 to 24 min, but with little variation within species. Pollination drops of each tested Cupressaceae taxon are also withdrawn when pollinated with foreign, but Cupressaceous pollen. However, they remain unchanged and are not withdrawn immediately when pollinated with pollen of other seed plants. The results clearly indicate that the time for the total withdrawal of pollination drops is strongly influenced by the evolutionary distance of the taxa being involved in the pollination process. Among closely related taxa the withdrawal is much more rapid than in distantly related ones. This points to an effective recognition system regulating the withdrawal of pollination drops, probably controlled by the nucellus. This recognition system can be regarded as an important preadaption for the evolution of a self-incompatibility mechanism. The withdrawal of pollination drops is thus not exclusively a physically induced process as suggested in some earlier studies. Pollination drops of several ovules can fuse to form a large common one, perhaps increasing by this way successful pollen capture.     

The signals to trigger the initiation of ovule enlargement are from the pollen tubes: The direct evidence

In angiosperms, initiation of ovule enlargement represents the start of seed development, the molecular mechanism of which is not yet elucidated.It was previously reported that pollen tube contents,rather than double fertilization, can trigger ovule enlargement. However, it remains unclear whether the signal(s) to trigger the initiation of ovule enlargement are from the sperm cells or from the pollen tubes.Recently, we identified a mutant drop1- drop2-, which produces pollen tubes with no sperm cells. Taking advantage of this special genetic material, we conducted pollination assays, and found that the ovules pollinated with drop1- drop2- pollen could initiate the enlargement and exhibited significant enlarged sizes at 36 h after pollination in comparison with those unpollinated ovules. However, the sizes of the ovules pollinated with drop1- drop2- pollen are significantly smaller than those of the ovules pollinated with wildtype pollen. These results demonstrate that the pollen tube, rather than the sperm cells, release the signal to trigger the initiation of ovule enlargement, and that double fertilization is required for further enlargement of the seeds.     

Comparative Biology of the Pollination Mechanisms in Acmopyle pancheri and Phyllocladus hypophyllus(Podocarpaceae s. l.)

The pollination mechanisms of Acmopyle pancheri(Brongn. &Gris) Pilg. andPhyllocladus hypophyllus Hook.f. were investigatedby conventional microscopical techniques and by nuclear magneticresonance (NMR) imaging. Dissimilarities include the orientationof the ovule and type of pollen;Phyllocladus has erect ovulesand wettable pollen with vestigial sacci, whereas Acmopyle hasmore-or-less erect ovules and non-wettable, functionally saccatepollen. Similarities include the mode of formation of the pollinationdrop and its response upon pollination. In both genera, pollinationtriggers pollination drop retraction and drop secretion ceases.Neither NMR imaging nor conventional histology of Phyllocladusovules revealed any specific tissue beneath the ovule whichcould be responsible for pollination drop retraction. It ismore likely, therefore, that the drop is channelled into thevascular supply or the apoplast. These findings invalidate thetaxonomic value of the pollination mechanism as a suite of characterstraditionally used to separate Phyllocladaceae from Podocarpaceae.Copyright 2000 Annals of Botany Company Acmopyle pancheri, gymnosperms, NMR imaging, nuclear magnetic resonance imaging, Phyllocladaceae,Phyllocladus hypophyllus , Podocarpaceae, pollination drop, pollination mechanism     

Ovulate cone,pollination drop,and pollen capture in Sequoiadendron (Taxodiaceae)

In Sequoiadendron ovules are borne inside the ovulate cone, and pollination drops secreted from these ovules collect pollen. We examined: (1) the relation between ovular position and pollen capture; (2) pollen behavior when in contact with a pollination drop; and (3) ultrastructure of ovules during pollination drop secretion. During wet periods a water sheet forms on the surface of the cone due to bract shape and wettability. Pollination drops persist inside the wetted cone, and pollen capture resumes immediately after drying. Pollen landing on a pollination drop is taken inside the drop and carried into the micropyle when the drop contracts. Several notable ultrastructural features appear in the nucellus, integument, chalaza, and bract lamina during pollination-drop secretion. The abaxial surface of the lamina is covered by a membrane that may contribute to the wettable nature of the surface.     

Post-pollination prefertilization drops affect germination rates of heterospecific pollen in larch and Douglas-fir

Pollen of larch (Larix?×?marschlinsii) and Douglas-fir (Pseudotsuga menziesii) was used in homospecific and heterospecific crosses. Germination of heterospecific pollen in ovulo was reduced in post-pollination prefertilization drops. This provides evidence of selection against foreign pollen by open-pollinated exposed ovules in these two sister taxa, which share the same type of pollination mechanism. Of the other prezygotic stages in pollen-ovule interactions, uptake of pollen by stigmatic hairs did not show any selection. Pollen tube penetration of the nucellus was similar for hetero- and homospecific pollen tubes, but heterospecific tubes only delivered gametes in one cross. To test for differences in the post-pollination prefertilization drops of each species, drops were gathered and analysed. Glucose and fructose were present in similar amounts in Douglas-fir and larch, while sucrose was found in larch only. Other carbohydrates such as xylose and melezitose were species-specific. In P. menziesii, sucrose is absent due to its conversion to glucose and fructose by apoplastic invertases. In contrast, Larix?×?marschlinsii drops have sucrose because they lack apoplastic invertases. The presence of invertase activity shows that the composition of gymnosperm post-pollination prefertilization drops is not static but dynamic. Drops of these two species also differed in their calcium concentrations.     

Mating systems in two species ofPotentilla from Alaska

Aspects of reproductive ecology have been studied in two species ofPotentilla (Rosaceae) from interior Alaska. Data were collected to determine mating system traits such as seed: ovule and pollen: ovule ratios, and experiments were carried out in the field in order to study the effect of caging, emasculation, and manual self-pollination on seed set and seed quality. Mating system strategies differ between the two species in the populations examined. High pollen: ovule ratio, high seed: ovule ratio, good autodeposition ability, and unpredictable pollination in concert with self-incompatibility indicate thatP. hookeriana in the population studied has adopted a more or less pseudogamous mating system. In contrast, the data suggest thatP. uniflora in the studied population is approaching a sexual, facultatively outbreeding species. Because both species exhibit ploidy variation, a trait common among agamospermous plants, it is concluded that both species are likely to be facultatively agamospermous but that sexual reproduction is much more important in the population ofP. uniflora than it is inP. hookeriana.     

Latisemenia longshania,gen. et sp. nov., a new Late Devonian seed plant from China

The earliest known ovules in the Late Devonian (Famennian) are borne terminally on fertile branches and are typically enclosed in a cupule. Among these ovules are some that have terete integumentary lobes with little or no fusion. Here, we report a new taxon, Latisemenia longshania, from the Famennian of South China, which bears cupulate ovules that are terminal as well as opposite on the fertile axis. Each ovule has four broad integumentary lobes, which are extensively fused to each other and also to the nucellus. The cupule is uniovulate, and the five flattened cupule segments of each terminal ovule are elongate cuneate and shorter than the ovule. Associated but not attached pinnules are laminate and Sphenopteris-like, with an entire or lobate margin. Latisemenia is the earliest known plant with ovules borne on the side of the fertile axis and may foreshadow the diverse ovule arrangements found among younger seed plant lineages that emerge in the Carboniferous. Following the telome theory, Latisemenia demonstrates derived features in both ovules and cupules, and the shape and fusion of integumentary lobes suggest effective pollination and protection to the nucellus. Along with other recent discoveries from China, Latisemenia extends the palaeogeographic range of the earliest seed plants.     

Effective Pollination Period and Nature of Pollen-collecting Apparatus in the Gymnosperm, Larix leptolepis

In controlled environment, the effective pollination periodin Larix leptolepis is characterized by its brevity, lastingfor less than 1 d. It is associated with the opening of thepapillae of the pollen collecting apparatus which possess surfaceesterase activity, a condition that has its parallel in thesurface pellicle of angiosperm stigmas. Larix leptolepis, ovule receptivity, morphometric analysis, pollination, stigma surface, esterase activity     

Integration and macroevolutionary patterns in the pollination biology of conifers

Integration influences patterns of trait evolution, but the relationship between these patterns and the degree of trait integration is not well understood. To explore this further, we study a specialized pollination mechanism in conifers whose traits are linked through function but not development. This mechanism depends on interactions among three characters: pollen that is buoyant, ovules that face downward at pollination, and the production of a liquid droplet that buoyant grains float through to enter the ovule. We use a well‐sampled phylogeny of conifers to test correlated evolution among these characters and specific sequences of character change. Using likelihood models of character evolution, we find that pollen morphology and ovule characters evolve in a concerted manner, where the flotation mechanism breaks down irreversibly following changes in orientation or drop production. The breakdown of this functional constraint, which may be facilitated by the lack of developmental integration among the constituent traits, is associated with increased trait variation and more diverse pollination strategies. Although this functional “release” increases diversity in some ways, the irreversible way in which the flotation mechanism is lost may eventually result in its complete disappearance from seed plant reproductive biology.     

Pollination biology of Indotristicha ramosissima (Podostemaceae: Tristichoideae)

Floral morphology, phenology and mode of pollination have been studied in Indotristicha ramosissima (Wt.) van Royen. Although the plants are submerged, self-pollination (autogamy) occurs above water. This is aided by considerable elongation of the pedicel (20 mm) prior to pollination. The filaments of the stamens also elongate rapidly (∼6.5 mm/h) before and after pollination. The flowers are typically trimerous. Each anther contains 1743 ± 187 pollen grains. These are spherical, multiporate, 3-celled and ∼97% fertile at the time of shedding. Germination of pollen on the stigma and the growth of pollen tubes have been traced in both naturally and manually (self- as well as cross-) pollinated pistils. The pollen:ovule ratio is ∼72:1 and the ovule: seed ratio is ∼2:1. Mature fruits are 8 or 9 ribbed and open by 2 or 3 longitudinal slits that release ∼32 seeds.