Reproduction in Seagrasses: Nature of the Pollen and Receptive Surface of the Stigma in the Hydrocharitaceae

This paper describes the characteristics of the pollen and thereceptive surfaces of the stigmas in the three marine angiospermsincluded in the Hydrocharitaceae. The pollen in Enhalus acoroidesand Thalassia hemprichii is spherical and has an ornamentedexine. An exine layer is not found in Halophila stipulacea wherereniform pollen grains are contained within transparent moniliformtubes. Cytochemical tests show that the pollen wall in the threespecies contains acidic and neutral polysaccharides and acidhydrolase acitivity is detected in the intine of H. stipulaceaand T. hemprichii. In Thalassia, one of the intine enzymes,acid phosphatase, is unambiguously associated with cytoplasmicinclusions. Flowering in Thalassia is coincident with the spring tides andthe pollen is released as a mass suspended in a thecal slimewhich contains approximately 5 per cent by weight carbohydrate,the principal mono-saccharide being mannose. Electrophoreticanalysis of the pollen-free slime shows a single glycoproteincomponent. The stigmas of the three seagrasses are papillate and of the‘dry’ type possessing a continuous protein-aceouspellicle subtended by a cuticle. The stigma pellicle exhibitscytochemically detectable esterase activity and binds the lectinconcanavalin A. Acid phosphatase activity is localized beneaththe cuticle at the tips of the stigma papillae. The discoveries show that the characteristics of the pollenand stigmas in the seagrasses are comparable with those foundin terrestrial flowering plants. The similarity in enzymaticproperites of the pollen wall and stigma pellicle suggests that,intriguingly, a similar mechanism of cuticle erosion might wellfollow compatible pollination both on land and in the sea. Enhalus acoroides, Thalassia hemprichii, Halophila stipulacea, Halophila decipiens, seagrasses pollen wall, stigma surface, hydrolytic enzymes     

异叶苦竹花粉管生长及双受精过程

以异叶苦竹为材料,采用扫描电镜、荧光显微镜技术及传统的石蜡制片技术,解剖观察其花粉管生长途径及双受精过程。结果表明:(1)授粉后,花粉在柱头上吸水膨胀,约30 min即可萌发。(2)授粉1～2 h后花粉管可达到花粉长度的5～10倍,花粉管在柱头分支中进一步伸长,并开始伸入花柱中生长。(3)授粉后5 h,大量花粉管沿引导组织进入花柱基部与子房顶部之间的子房壁,有少量花粉管在子房壁与外珠被之间的缝隙中生长。(4)授粉后8 h,少量花粉管到达珠孔端。(5)授粉后15～18 h,精核与极核融合,形成初生胚乳核;精、卵核融合,形成合子。(6)授粉后20～30 h,仍可在花柱中见到大量呈束状的花粉管。(7)授粉后48 h,子房内的大部分花粉管出现解体,大多数花粉死亡。研究认为,精细胞到达胚珠的时间为8 h。     

Esterases in pollen and stigma of Brassica

The dry type stigma of Brassica is covered with a continuous layer of cuticle. Cutinase and non-specific esterases may be involved in breakdown of this cuticle barrier during pollen-stigma interaction, but only a little is known about their nature and characteristics. We report here the presence of two distinct esterases from stigma and pollen of Brassica. A 33 kD esterase assayed using MU-butyrate substrate shows high activity in stigma papillae. A similar esterase from Tropaeolum pollen has been shown to possess active cutinase activity. The esterase activity in anther tissue is due to a 24 kD enzyme with substrate specificity toward acetate esters. Both enzymes require sulfhydryl groups for their catalytic activity. Immunogold labelling of antibodies raised against these esterases localised the proteins at the subcellular level. Antibodies for MU-butyrate hydrolase gave a positive signal in the cell walls of mature stigma papillae and in the tapetum and microspores during early stages of anther development. In the mature anther, a positive signal in the cytoplasm of pollen grains with some detectable localisation in the exine layer of the pollen wall was obtained. Similar results were obtained with acetate hydrolase antibodies. These esterases are thus spatially and temporally regulated in stigma and anther tissues.Abbreviations MU methyl umbelliferyl - pAbC anti-butyrate hydrolase polyclonal antibodies - pAbE anti-acetate hydrolase polyclonal antibodies     

The localization of nonspecific esterase and cholinesterase activity in germinating pollen and in pollen tube ofVicia faba L. The effect of actinomycin D and cycloheximide

Two types of esterases, nonspecific esterase and cholin esterase have been distinguished in germinating pollen grains and in pollen tubes ofVicia faba using cytochemical methods. The localization of each of them was different. In the hydrated non-germinating pollen grain the nonspecific esterase was present in the cytoplasm and in the intine. The cholinesterase was localized mainly in the sexine and on the outside of the plasma membrane. A particularly large agglomeration of this enzyme was found in the aperture. During germination both types of extracellular esterases were released into the medium. In the pollen tube the activity of the enzymes studied was connected with the wall of its tip. The localization of both types of esterases in the germinating pollen grain and the growing tube were independent of actinomycin D. Cycloheximid prevented the cholinesterase localization at the pollen tube tip.     

Histochemical studies of alpha and beta-glucosidases in the germinating pollen grains ofPortulaca grandiflora

The paper deals with the distribution of alpha and beta-glucosidases in the germinating pollen grains ofPortulaca grandiflora. Both these enzymes are localized in the pollen wall and the cytoplasmic granules. The latter are distributed throughout the pollen cytoplasm, pollen tube and stigma hair. In non-germinating pollen grains, enzymes are concentrated in the pollen wall. Stigma hair sheaths are completely free from enzymes. The functional significance of these enzymes in the hydrolysis of phenolic glycosides and polysaccharides is discussed.     

Localization and release of allergens from tapetum and pollen grains ofBetula pendula

Summary Although intact pollen grains are assumed to be the primary carrier of pollen allergens, specific immunoreactive components have been found in other aerosol fractions, e.g., starch grains and remains of tapetal cells Cryo-scanning-electron-microscopy results demonstrate the presence of a clear network of strands connecting the tapetum with the microspores. The distribution of protein in tapetal orbicules, pollen wall, and pollen cytoplasm was tested by histochemical stains for light microscopy and transmission electron microscopy. The protein is mainly localized at the apertures and starch grains in the cytoplasm of pollen and in the core and on the surface of tapetal orbicules. Monoclonal antibodies Bv-10, BIP3, and BIP4 have been used to locate the cellular sites of pollen and tapetal allergens inBetula pendula (syn.B. verrucosa). The application of rapid-freeze fixation prevented relocation of allergens from their native sites. The allergens are predominantly found in the starch grains and to lesser extent in the exine. We also tested interactions between mature birch pollen and human fluids: saliva, nostrils fluid, and eyes solution. The aim was to mimic more closely the in vivo situation during allergenic response. In all cases we observed several pollen grains that were burst and had released their cytoplasmic contents. In the nose the allergens are released from the pollen within minutes. In rhinitis, nasal pH is increased from the normal pH 6.0 to 8.0. When we used nasal fluid at pH 8.0, the number of ruptured pollen grains increased. The mechanism that might induce formation of small allergen-bearing particles from living plant cells is discussed.     

Evidence of long-distance transport of betula pollen

In April the eastern parts of Fennoscandia are frequently influenced by south to southeastern winds. These air-masses can bring with them large quantities of Betula pollen several weeks before the local flowering season begins. The pollen is transported with air-masses at a high level and then taken down by turbulent transport or washout. The number of pollen grains can be so high that sensitive persons exhibit allergic symptoms. Three localities from Sweden and six from Finland have been selected and numerical air-parcel trajectories have been calculated in order to determine the Betula pollen source. The results indicate that the distant pollen source areas can be identified, and that the travelling times for pollen grains transported in April 1989 were mostly in the range of 9 to 20 hours.     

Production of fertile tobacco pollen from microspores in suspension culture and its storage for in situ pollination

Summary A simple procedure is described for the in vitro production of tobacco (Nicotiana tabacum L.) pollen from microspores isolated just before entering mitosis. During a 3-day culture period in a liquid medium containing pyrimidine nucleosides these microspores develop into young pollen grains to the stage of starch deposition. Pollen maturation and transition to dormancy is achieved during a further 2- to 3-day culture period in the same medium stepwise supplemented by a concentrated solution of sucrose and l-proline. Upon transfer of the pollen to a simple germination medium containing sucrose and boric acid, up to 40% of the grains were observed to produce relatively long tubes. The in vitro-matured pollen grains can be stored at-20° C either suspended in 1.17 M sucrose and 100 mM l-proline or separated from the medium on filter paper discs. The stored pollen germinated both in vitro and on the stigma, the pollen tubes grew through the style into the ovary and pollination produced up to 300 viable seeds per pod. The procedure is of interest for pollen developmental studies and various fields of pollen manipulation, such as in vitro pollen selection.     

Diurnal variation of biological and non-biological particles in the atmosphere of Córdoba, Spain

In this work, the daily distribution of biological and non-biological particles in the atmosphere of the city of Córdoba, Spain, is analysed in order to know at what time of day the concentrations of solid suspended particles in the air is maximum. This knowledge can be useful for all those people who suffer of respiratory diseases and can help them to plan their outdoors activities. The concentrations of non-biological material have been estimated by using spectrophotometrical techniques. With this method, the amount of non-biological material is expressed in percentage of light absorbance in values oscillating from 0.00: absence of particles to 1.00: extreme presence of particles. As the absorption of light depends on the colour of the particle, it is also possible to distinguish dark particles (soot, diesel exhaust particles, sand) from hyaline or light coloured ones (pollen grains and several fungal spores types). The results have shown that the maximum peaks of material are achieved early in the mornings and late in the evenings. Aerobiological methodology has been used when dealing with material of biological origin. In this latter case, pollen grains and fungal spores have been differentiated from the rest of solid material. Due to their importance as agents causing pollen allergy in the area and the high percentage that they represent with respect to the total pollen spectrum, Olea europaea and grass pollen types have been differentiated from the rest of pollen types. The results show that the peaks of non-biological particles in the air throughout the day are related to the activities carried out by men in the city: commercial and working hours or social activities in the different seasons of the year. As regards the biological material, the pattern of fungal spores is clearly influenced by the climatic characteristics of the area, presenting the maximum concentrations in the late afternoon. The pollen grains follow a pattern coinciding with the diurnal maximum temperatures and sunshine hours. The values of the curve oscillate from 200 grains/m3 of air in night hours to maximum over 1000 grains/m3 at midday. The diurnal variation of Olea europaea is quite similar to this general pattern, presenting the maximum concentrations of pollen grains at central hours of day. The grasses show some differences due to the high number of species included in this family.     

The role of allergenic proteins Pla a 1 and Pla a 2 in the germination of Platanus acerifolia pollen grains

Platanus acerifolia (Aiton) Willdenow is a plane tree, widely grown as an ornamental tree in many cities of the United States and Western Europe, which has become an important source of airborne allergens in our cities. The aim of the present study is to immunolocalize the major allergens in the pollen grain and to examine their potential function in the fertilization process. Observations were made in mature and hydrated, activated pollen of P. acerifolia for 5, 15, 30 min and 2 h in the germination medium. Specimens were fixed using freezing protocols for transmission electron microscopy (TEM). For immunogold labelling, cryosections and resin-embedded ultrathin sections were incubated using rabbit antisera against the purified pollen allergens Pla a 1 and Pla a 2. Elution of P. acerifolia allergens took place after 5 min of pollen incubation in buffered medium. Intense labelling of Pla a 1 and Pla a 2 was detected after pollen exudates were released. In pollen grains, Pla a 1 was predominantly localized in concentric cisternae of the endoplasmic reticulum (ER), situated between the vegetative nucleus and the generative cell, and was released from pollen grains 5 min after hydration; cytoplasmic localization decreased 15 min after hydration. In pollen grains, glycoprotein Pla a 2 was abundant in association with Golgi cisternae and vesicles situated in the apertural periphery of the mature pollen grains. Pla a 2 proteins were also detected in ER and in the generative cell wall. Immunolabelling of Pla a 2 decreased 5 min after pollen hydration but was again intense after 15–30 min in germination medium, presumably as a consequence of renewed expression and glycosylation of this protein. Pla a 1 belongs to a new class of allergens related to proteinaceous invertase and pectin methyl esterase inhibitors (PII, PMEI) which could be involved in membrane protection and pectin de-esterification control during pollen hydration. Pla a 2 has an exopolygalacturonase (PG) enzymatic activity consistent with pollen-stigma adhesion mechanisms or compatibility systems. Moreover, the expression of Pla a 2 found 15–30 min after hydration might contribute to pollen-tube growth and the modification of transmitting tissue cell walls. The abundant production and elution of Pla a 1 and Pla a 2 proteins may alter the environment in which pollen tube elongation occurs, thus promoting a potential crosstalk between the pollen and the gynoecium.     

Distyly and pollen dimorphism in<Emphasis Type="Italic"> Damnacanthus</Emphasis> (Rubiaceae)

We document distyly in Damnacanthus (Rubiaceae) and investigate pollen morphology in the genus. Within nine taxa in Japan (four species, four varieties and one form), five show distyly. Non-distylous taxa of Damnacanthus always have a long style and short stamens. We examined the pollen morphology of all taxa of Damnacanthus in Japan. The pollen grains of Damnacanthus are 3- to 5-orthocolporate or sometimes loxocolporate. Polymorphic pollen is sometimes observed even within a single flower. Pollen dimorphism is associated with distyly; pollen grains from short-styled (thrum) flowers are significantly larger than those from long-styled (pin) flowers, and the muri are smooth in pin flowers whereas the pollen grains of thrum flowers have minute granules on the top and/or on the sides. In non-distylous taxa, in which the style is longer than stamens, the muri are always provided with minute granules. Interestingly, the pollen grains with granules in the non-distylous taxa contrast with the pollen of the distylous taxa, because granulate pollen grains are found only in thrum flowers. Although Damnacanthus is a stenopalynous genus, the number of colpi is useful for delimiting taxa. Based on palynological characters, we conclude that Damnacanthus should be revised. Electronic Publication     

Ambrosia pollen in the air of Lublin, Poland

Studies on Ambrosia pollen concentrations were carried out in Lublin in the period 1995–2004. The effects of a number of meteorological factors were analysed. In the first period of the study, the gravimetric method was used (1995–1999), while in the second period, the volumetric method was applied. The results show an increasing trend in the amount of airborne pollen. The Ambrosia pollen season in Lublin lasts from August to October. Over a period of 5 years, the highest number of pollen grains was recorded in September (53%), followed by August (44%) and October (3%). There were wide variations in annual totals. The annual total pollen counts was 167–1180 grains, with the peak value in 2002. Maximum daily pollen concentrations (56–312 pollen grains m⁻³) were recorded in the first half of August and in the first half of September. On the days when high Ambrosia pollen concentrations occurred, the temperature was above 21°C and the winds were mainly from the southeast, south and east. Maximum intradiurnal concentrations of pollen grains occurred in the afternoon hours. These results indicate, to some degree, that Ambrosia pollen is transported for long distances before descent.     

Are the birch trees in Southern England a source of <Emphasis Type="Italic">Betula</Emphasis> pollen for North London?

Birch pollen is highly allergenic. Knowledge of daily variations, atmospheric transport and source areas of birch pollen is important for exposure studies and for warnings to the public, especially for large cities such as London. Our results show that broad-leaved forests with high birch tree densities are located to the south and west of London. Bi-hourly Betula pollen concentrations for all the days included in the study, and for all available days with high birch pollen counts (daily average birch pollen counts >80 grains/m³), show that, on average, there is a peak between 1400 hours and 1600 hours. Back-trajectory analysis showed that, on days with high birch pollen counts (n = 60), 80% of air masses arriving at the time of peak diurnal birch pollen count approached North London from the south in a 180 degree arc from due east to due west. Detailed investigations of three Betula pollen episodes, with distinctly different diurnal patterns compared to the mean daily cycle, were used to illustrate how night-time maxima (2200–0400 hours) in Betula pollen counts could be the result of transport from distant sources or long transport times caused by slow moving air masses. We conclude that the Betula pollen recorded in North London could originate from sources found to the west and south of the city and not just trees within London itself. Possible sources outside the city include Continental Europe and the Betula trees within the broad-leaved forests of Southern England.     

The timing and effectiveness of sequential pollinations in Hibiscus moscheutos

In many species with animal-pollinated flowers, pollen arrives on the stigma in pulses and late-arriving pollen may be precluded from fertilizing ovules. When seed set per fruit is not pollen limited, the fate of pollen from later cohorts is affected by the time between pollinator visits, variation in pollen tube growth rates, and the amounts of pollen deposited relative to the availability of stigmatic area and unfertilized ovules. In a natural population of Hibiscus moscheutos, we found that consecutive pollinator visits to individual flowers occurred within 15 min of each other in more than half of our observations. We then conducted hand-pollination experiments using equal doses of early and late pollen bearing unique allozyme markers for paternity analysis (each dose was more than sufficient to result in full seed set). When pollen was applied 15 or 30 min after an earlier pollen load, the proportions of seeds sired by late-arriving pollen were reduced by 13–30% and 21–57%, respecitively. A few pollen-tubes from the late pollen load sired seeds even when a large dose of pollen was applied 1–2 h earlier, suggesting that the performance of pollen grains within each cohort was highly variable. The advantage of arriving early was greater when the first pollen load was applied at 0930 hours as compared with 1130 hours. We found no effect of previous pollination on the growth rates of late-arriving pollen-tubes. Taken together, these results demonstrate that pollen from later visits can compete with pollen from earlier visits, but the success of late-arriving pollen typically declines to very low levels after an interval of about 2 h. Given that surplus pollen often arrives on stigmas by mid-morning, we conclude that male reproductive success is more likely to be affected by the timing of pollen dispersal and pollen-tube competitive ability than by the total amount of pollen that is exported from flowers.     

Società Botanica Italiana Congresso Sociale 1982 Parto—12-14 Ottobre Relazioni

Abstract

« Harmomegathy »: a still open problem not yet well-known. - Wodehouse used the term « Harmomegathy » to indicate the form and volume change in pollen grains, due to water loss or assumption. Roundish and tricolporate grains in the anther become oval and decrease in volume during the dehydration preceeding anthesis. Viceversa during rehydration on the stigma they undergo the opposite process.

Pollen water content at an thesis varies from species to species and it usually ranges between 10 and 20%; nevertheless in some families as Zingiberaceae and Araceae it is higher. Anther and pollen dehydrations are affected by environmental factors, such as humidity of air and temperature, before the anthesis and when pollen is suspended in air. Besides, to avoid the further loss of water during pollen flow, an oily layer derived from tapetum degeneration is deposited outside the pollen grains.

During dehydration and rehydration pollen grains undergo a stress due to shape and volume changes expecially in the colpus and/or pore areas. Pollen walls are structured in order to avoid their own damage, which could be followed by protoplast death and/or irregularities in grain shape. Intine, formed by pectocellulosic filaments, has elastic properties, as sporopollenin, the component of exine.

Stigma can be dry or wet, namely covered or not by stigmatic exudate giving place to different hydration patterns. The structure of intine and exine has to match that of the stigma, in fact pollen will find the best place and way to proper hydration only on stigmas of its own species. Grains structure also must be sterically conform to stigma: this is well-shown in species as Linum, Lytrum and Primula which have an heteromorphic autoincompatibility.

The right keeping of shape and structure of grains, despite the hydration and dehydration, is therefore essential to the physical-morphological recognition between pollen and stigma, which preceedes the biochemical recognition.     

Evaluation of Viability,Shedding Pattern,and Longevity of Pollen from Genetically Modified (GM) Herbicide-tolerant and Wild-type Zoysiagrass (<Emphasis Type="Italic">Zoysia japonica</Emphasis> Steud.)

Knowledge about pollen viability is important when evaluating the risk of genetically modified (GM) plants. Here, staining via iodine potassium iodide (IKI) or triphenyl tetrazolium chloride (TTC) could not distinguish between live and dead pollen from Zoysia japonica. Therefore, to obtain a reliable assessment of such viability and longevity, we developed an optimum germination medium containing 20% sucrose and 50 ppm H₃BO₃. Pollen grains transferred to the germination medium at about 1000 hours had a germination rate of >90%. Pollen was most predominantly shed at approximately 1000 hours, with viability declining to nearly 0% at 1200 hours. All germinability was lost within 150 min when stored at 25°C. No significant difference was found between GM and non-GM plants in their pollen viability or longevity.     

Histochemical studies on the distribution of non-specific esterase in the germinating pollen grains ofPortulaca grandiflora

The study deals with the distribution of non-specific esterase in germinating pollen grains ofPortulaca grandiflora. Intense activity of the enzyme is seen in small granules distributed homogeneously in pollen grains stigma hairs and throughout the length of pollen tubes. Further the walls of pollen grains also demonstrate intense activity. The functional significance of the enzyme in these locales has been discussed.     

Accelerated Degradation of Dipentyl Phthalate by Fusarium oxysporum f. sp. pisi Cutinase and Toxicity Evaluation of Its Degradation Products Using Bioluminescent Bacteria

The efficiency of two lipolytic enzymes (fungal cutinase and yeast esterase) in the degradation of dipentyl phthalate (DPeP) was investigated. The DPeP degradation rate of fungal cutinase was surprisingly high, i.e., almost 60% of the initial DPeP (500 mg/L) was decomposed within 2.5 hours, and nearly 40% of the degraded DPeP disappeared within the initial 15 minutes. With the yeast esterase, despite the same concentration, >87% of the DPeP remained even after 3 days of treatment. The final chemical composition after 3 days was significantly dependent on the enzyme used. During degradation with cutinase, most DPeP was converted into 1,3-isobenzofurandione (IBF) by diester hydrolysis. However, in the degradation by esterase, pentyl methyl phthalate, in addition to IBF, was produced in abundance. Toxicity monitoring using various recombinant bioluminescent bacteria showed that the degradation products from yeast esterase contained a toxic hazard, causing oxidative stress and damage to protein synthesis. Ji-Young Ahn, Yang-Hoon Kim are contributed equally to this work     

Pollen size in Carex: The effect of different chemical treatments and mounting media

Pollen size and pollen aperture size for ten species of the genus Carex L., native to Estonia, have been measured using light microscopy. The species selected represent different sections of the genus, a range of habitats and different chromosome numbers. The effects of two basic chemical treatments, two mounting media and the effect of chemically induced dehydration with tertiary butyl alcohol (TBA) on the size of pollen grains were then recorded.

In general pollen size and pollen aperture size of the species examined is highly variable at both intraspecific and interspecific levels. Carex hirta has notably larger pollen grains than any of the other species investigated and, although correlations between size and chromosome number in the species examined are limited, it also has the highest chromosome number. Statistically significant size differences resulted from variations in chemical treatment, mounting media and tertiary butyl alcohol (TBA) induced dehydration. Acetolysed pollen grains are larger than potassium hydroxide (KOH) treated pollen grains. Pollen grains dehydrated after chemical treatment with TBA are larger than pollen grains not dehydrated. Pollen grains mounted in silicon oil are smaller than grains mounted in glycerine. But considering the great size variation of Carex pollen grains, the size changes caused by preparation procedures fall within the size variation range of the species examined.

All the samples contained a high number of deformed pollen grains and pollen grains with hardly distinguishable or no lateral apertures.     

COSTS OF SELF-POLLINATION IN A SELF-INCOMPATIBLE PLANT,POLEMONIUM VISCOSUM

Plants of Polemonium viscosum are strongly self-incompatible, yet have floral features that enforce spatial and temporal separation of pollen and stigma presentation. In this study, we address the widely held view that such barriers to selfing are maintained because they reduce interference by self pollen in performance of outcross grains. In the first year of the study we found that 24 hr precedence of self pollen on the stigma (24S), reduced outcross pollen germination by 32%, and seed set by 40% in flowers of P. viscosum. In the second year significant negative effects of self pollen on compatible pollen germination were seen when self pollen was applied four hours before (4S) or immediately prior (S×) to outcross pollen. Surprisingly, however, in the second year of study, seed set was not reduced with as much as 50% inhibition of compatible pollen germination. Results suggest that by interfering with normal functioning of outcross grains, incompatible self pollen can increase pollen limitation in P. viscosum. Reduction in seed production as a result of this phenomenon should select for displacement of pollen presentation from stigma receptivity.