The coexistence of binucleate and trinucleate pollen in Mitrephora macclurei Weerasooriya & R. M. K (Annonaceae)

Pollen of angiosperms may be either binucleate or trinucleate at maturity. According to Schürhoff–Brewbaker law, the binucleate state is pleisomorphic occuring in about 70% of all angiosperms, while the trinucleate state is advanced occurring in about 30% of the angiosperms. The derivation of trinucleate pollen from binucleate pollen is irreversible, thus trinucleate or mixed pollen was considered uncommon in early lineage. In the present work, using conventional paraffin section method, the coexistence of both binucleate and trinucleate pollen in the same intact anther were observed in Mitrephora macclurei, a species belonging to the eumagnoliid family Annonaceae. The increasing number of plants with trinucleate or mixed pollen discovered in early lineages, as well as climatic introduced changes in nuclear number challenge analyses of ancestral state and evolution of pollen nuclear number in angiosperms.     

THE DISTRIBUTION AND PHYLOGENETIC SIGNIFICANCE OF BINUCLEATE AND TRINUCLEATE POLLEN GRAINS IN THE ANGIOSPERMS

Studies of the cytology of angiosperm pollen which extend our knowledge of the distribution of binucleate and trinucleate pollen to almost 2,000 species of flowering plants are summarized. Approximately 70% of the species studied release pollen in a binucleate stage, and none of them shed both types of pollen as a constant varietal trait. All phylogenetically primitive taxa are binucleate. The trinucleate trait evidently has originated independently at many times during angiosperm evolution. In no instance must one infer the origin of binucleate taxa from trinucleate ancestors. Most genera are monotypic with respect to pollen cytology, and only 10 genera are known to include both binucleate and trinucleate species. Among the 265 families studied, 179 include only binucleate genera and 54 include only trinucleate genera, while 32 include both types. Nearly all aquatic species with submersed flowers shed pollen in a trinucleate stage. A group of physiological differences which distinguish binucleate and trinucleate pollen is reviewed in relation to the survival and possible selective advantage of trinucleate mutants, and the significance of the binucleate pollen grain in the origin and evolution of flowering plants is discussed.     

SYSTEMATIC SIGNIFICANCE OF POLLEN NUCLEAR NUMBER IN EUPHORBIACEAE,TRIBE EUPHORBIEAE

Pollen nuclear number is determined in 139 species of 5 genera in the Euphorbieae, subtribe Euphorbiinae. The 111 new determinations are tabulated along with previous reports, and the results indicate that the distribution of binucleate (II) and trinucleate (III) pollen is strongly associated with the taxonomic groupings within the Euphorbieae. Although binucleate pollen is probably primitive within the tribe Euphorbieae, as suggested by the nuclear condition in Neoguillauminia, the situation in Euphorbia still requires further elucidation. Within Euphorbia, the morphologically most primitive species studied have III pollen despite the fact that II pollen is presumably the original condition for the subtribe Euphorbiinae. In Euphorbia, II pollen only is reported from nine sections and III pollen only from ten sections, while in four sections (Esula, Goniostema, Aphyllis, and Deuterocalli) both II and III pollen have been found. The New World species of Euphorbia nearly all have III pollen, whereas the vast majority of the African succulents have II pollen. The genera of New World origin, Chamaesyce and Pedilanthus, have III pollen, while the African genera Monadenium and Synadenium have II pollen. Independent derivations of III pollen from II pollen appear to have occurred in sections Goniostema, Aphyllis, and Deuterocalli (all of subg. Euphorbia). There is no evidence that reversals from III to II pollen have occurred.     

Isolation and characterization of a polygalacturonase gene highly expressed in Brassica napus pollen

A cDNA clone, Sta 44-4, corresponding to a mRNA highly expressed in Brassica napus cv. Westar stamens, was isolated by differential screening and characterized. Northern blot and in situ analyses demonstrated that Sta 44-4 is synthesized in pollen beginning at the late uninucleate stage and reaches a maximum in trinucleate microspores. Sta 44-4 displayed significant sequence similarity to known pollen polygalacturonase genes. The B. napus pollen polygalacturonase gene was shown to be part of a small gene family and to display some polymorphism among different cultivars.     

Microgametophytic plastid nucleoid content and reproductive and life history traits of tribeTrifolieae (Fabaceae)

Microgametophytic plastid nucleoids were quantified for 18 species representing the four core genera of the tribeTrifolieae (Fabaceae),Medicago, Melilotus, Trigonella, andTrifolium. Generative cells of all taxa contained nucleoids, establishing that biparental plastid inheritance is common in theTrifolieae. Nucleoid number and volumes of pollen grains and generative cell nuclei differed among taxa. Nucleoid number was positively correlated with pollen grain and generative cell nuclear volumes, flower size and style length. These relationships disappeared after adjusting nucleoid number for pollen grain and generative cell nuclear volumes. Adjusted nucleoid numbers provided no evidence to support hypotheses that plastid content is associated with ploidy level, mating system, perenniality or size of the reproductive apparatus.     

Triaperturate pollen in the monocotyledons: configurations and conjectures

Triaperturate pollen are known in at least twenty seven genera of monocotyledons. Differences between aperture type and polarity indicate that the development of three apertures has occurred a number of times. Mode of cytokinesis during microsporogenesis is compared with differences in aperture configuration, to assess the extent to which this appears to influence aperture arrangement. Triapertury in monocot pollen tends to fall into one or another of three situations: 1) it is the normal state, 2) it is fairly common, but pollen with more or less apertures also occur in the taxon or sample, 3) it is a rare, or abnormal state for pollen which usually has less than three apertures. The various forms of triaperturate pollen are described, as well as monosulcate pollen of the orchid genera Cypripedium and Paphiopedilum, often misinterpreted as tri-sulcate, and the unusual extended trichotomosulcate pollen of Agrostocrinum (Hemerocallidaceae). Monosulcy, trichotomosulcy, and zonasulcy, with unusual and rare exceptions of zonasulcy in the eudicots, are aperture states shared exclusively with the basal dicots. Furthermore, to some extent all have links with the triaperturate condition in monocots and basal dicotyledons. This is discussed, as well as the association of tripory with polypory in monocots and basal dicots. The fossil pollen record is considered.This paper is dedicated to Klaus Kubitzki in recognition, not only for his extensive contribution to systematic botany, but also for his firm belief that pollen characteristics contribute to a better understanding of plant systematics and evolution.     

Respiration and Vitality of Binucleate and Trinucleate Pollen

The respiration and vitality of ungerminated bi- and trinucleate pollen were studied in order to determine the influence of relative humidity and temperature on metabolic activity. The gas exchange, germination capacity and staining with tetrazolium bromide were followed under standardized conditions. A constant respiration rate occurred under conditions of high relative humidity (97%). Per mg pollen, the trinucleate grains of Compositae and Gramineae respired 2 to 3 times as intense as 6 species of binucleate grains. Per unit of pollen protein the differences were even larger. In contrast to binucleate pollen, the longevity of trinucleate pollen was very short and the ability to germinate was lost twice as fast as the respiration capacity. This limits the use of tetrazolium bromide as an indicator of viability. At reduced relative humidities respiration was strongly restricted, but the longevity of bi- and trinucleate pollen considerably increased. Pollen of Gramineae, however, was very sensitive to changes in relative humidity; short exposure to low relative humidity decreased both the vitality and the capacity to respire.     

The natural delimitation ofCentaurea (Asteraceae: Cardueae): ITS sequence analysis of theCentaurea jacea group

Taxonomic complexity has hindered partitioning the genusCentaurea into natural subdivisions, even though it has long been recognized as an unwieldy, artificial assemblage. Most of the remaining difficulties center in theCentaurea jacea group, whose taxa share a common advanced type of pollen. Because it comprises half the species of the genusCentaurea, as well as five other disputed genera previously segregated fromCentaurea (Chartolepis, Cheirolepis, Cnicus, Grossheimia andTomanthea), theCentaurea jacea group is a significant taxonomic challenge. Newer molecular approaches are useful for resolving complex relationships because they provide more precise inferences of evolutionary relationships than traditional morphological characters. Sequences of the Internal Transcribed Spacers (ITS) of nuclear ribosomal DNA were analyzed for a comprehensive sample of this group. Results indicated that theCentaurea jacea group is monophyletic and includes the segregated genera, but not two other genera (Oligochaeta andZoegea), whose inclusion in theCentaurea jacea group was doubtful. In addition to pollen morphology, the ITS phylogeny is also supported by karyological evidence and by good correlation with biogeographic distribution of the species. The monophyly of theCentaurea jacea group suggests that a natural delimitation ofCentaurea that minimizes nomenclatural changes is possible, but only if a new type of the genus is designated.     

RAPID SCREENING METHOD TO DETECT POTENTIAL BIPARENTAL INHERITANCE OF PLASTID DNA AND RESULTS FOR OVER 200 ANGIOSPERM SPECIES

We have developed a diagnostic method to screen rapidly for plant species potentially capable of biparental inheritance of plastid DNA using the DNA fluorochrome 4′,6-diamidino-2-phenylindole (DAPI) in conjunction with epifluorescence microscopy. Pollen shed from 235 plant species (including about 50 of agronomic importance) representing 80 families were screened. Putative plastid DNA was detected in the generative and/or sperm cells of pollen from 26 genera (43 species) representing 15 families. Plastid DNA was not detected in the generative or sperm cells of pollen from 192 plant species, thereby strongly suggesting that these species have only maternal inheritance. Our cytological diagnosis corroborated the known genetic evidence in 42 plant species and conflicted with the genetic reports in five species, which are discussed. The data suggest that biparental inheritance of plastids is rare; overall, it may occur in about 14% of flowering plant genera, examples of which are scattered among 19% of the families examined. This methodology also readily reveals whether pollen is bi- or trinucleate.     

Protein synthesis of binucleate and trinucleate pollen and its relationship to tube emergence and growth

Under humid conditions, both bi- and trinucleate pollen species incorporate, on the average, very low amounts of leucine, e.g., 0.4 pmol min^-1mg pollen^-1. During germination in vitro, however, the two types of pollens greatly differ in their capacity for protein synthesis.Binucleate pollen species such as Typha, which are characterized by slow respiration in humid air and prolonged lag periods during germination in vitro, contain large amounts of monoribosomes at dehiscence. Polyribosomes are formed soon after the pollen is wetted in the germination medium, and a considerable incorporation of leucine is initiated after 10–15 min. More rapidly respiring, binucleate pollen showing a short lag period, such as Tradescantia, may already contain many polysomes at dehiscence and incorporate leucine within 2 min of germination. However, rapidly respiring, trinucleate Compositae pollen contains very limited amounts of ribosomal material and never attains any substantial level of incorporation. Cycloheximide completely inhibits both protein synthesis and tube emergence and growth in the slowly respiring, binucleate pollen species. The more rapidly respiring types are less dependent on protein synthesis, while germination of the phylogenetically advanced, trinucleate Compositae pollen proceeds completely independently. It is concluded that the level of phylogenetic advancement of the male gametophyte is characterized by its overall state of metabolic development at dehiscence rather than by the number of its generative cells.Abbreviations BSA bovine serum albumin - CHI cycloheximide - EGTA ethyleneglycol-bis(-aminoethyl ether) N,N-tetraacetic acid - RH relative humidity - TCA trichloroacetic acid     

Factors affecting in vitro maturation of isolated maize microspores

Summary An in vitro method to simulate pollen development was developed in maize (Zea mays L.). Microspores at the late uninucleate to early binucleate stage were isolated and cultured under various conditions. Cell viability, starch content and the formation of the three nuclei as found in normal mature pollen were monitored during the course of the culture. Media composition was modified in order to promote starch accumulation and frequency of mitosis, while maintaining the viability of the microspores. Under the best conditions, up to 12% of the microspores matured in vitro into trinucleate, starch-filled viable pollen grains which were unable to germinate or produce seeds. At different stages during in vitro maturation, proteins patterns were analyzed and compared with their in vivo equivalent and the patterns were only partially similar.     

Vergleichende Karyologie der Gräser-SubtribenAristaveninae undAirinae (Poaceae — Aveneae)

The chromosome numbers of nearly all species of the grass subtribesAristaveninae andAirinae from Europe and northern Africa are presented. Among theAristaveninae the genusAristavena has 2n = 14 chromosomes, whereasDeschampsia forms a polyploid series with the basic number x = 13. In the subtribeAirinae the basic number x = 7 predominates.Avenella includes a polyploid series up to dekaploidy, whilst the lowest diploid value so far known in grasses — caused by descending dysploidy — exists in the annual generaAiropsis andPeriballia with 2n = 8.From both subtribes 12 different karyotypes are described and depicted as idiograms. The basic karyotypes ofCorynephorus, Periballia andVahlodea differ from each other by different chromosome length. SAT-chromosomes in theAirinae vary somewhat. Some marker chromosomes eludicate phylogenetic relationships. Amphiplasty appears in various genera and was studied particularly in the amphidiploidAira caryophyllea. Karyological and genomatic trends are considered in relation to evolutionary strategies of annuals and perennials.The nuclear DNA content of some species has been determined cytophotometrically. In subtribeAirinae a positive correlation exists between chromosome volume, pollen diameter, and DNA content. A comparison of the duration of microsporogenesis and microgametogenesis in annual and perennial species with their nuclear DNA content has shown that a primary nucleotypic influence is not recognizable.

     

Floral morphology and taxonomic relations among the genera of Amaranthaceae in the New World and the Hawaiian Islands

Twenty-three genera of Amaranthaceae occur in the New World. Two endemic genera occur in the Hawaiian Islands. Among the genera of the subfamily Amaranthoideae, Celosia, Cyathula and Achyranthes have their main distributions in the Old World; the two last-named genera are represented in the Americas only by widespread weeds. All the New World genera of the subfamily Gomphrenoideae are mainly or entirely restricted to this region. Characters of androecium and gynoecium are fundamental in the recognition of genera within the family. Androecia of different genera may be structurally and phylogenetically more similar than would appear from a cursory examination. It is suggested that the type of staminal tubes found in Pseudogomphrena and Froelichia can be derived from that in Alternanthera and Froelichiella by reduction of filament length and a fusion of pseudostaminodia with the filaments. The staminal tube in Gomphrena could result from a further decrease in distance between pseudostaminodia of the Pseudogomphrena type, and a deeper forking of the pseudostaminodia; each so-called apical filament lobe in Gomphrena would then be homologous with half a pseudostaminodium in Pseudogomphrena. Much of the variation in the androecia of these and other genera, as well as within genera such as Pfaffia, can be explained as the combined results of coalescence and splitting-up tendencies. Splitting up of staminal tubes may not necessarily take place along the borders of phylogenetically original filaments and pseudostaminodia. The Amaranthus-type of pollen is found in the majority of genera of the subfamily Amaranthoideae, but also in the Chenopodiaceae. A group of genera within the subfamily Gomphrenoideae also has pollen very similar to, or identical with, this type. Most genera of the subfamily Gomphrenoideae have pollen of the Gomphrena-type. Pseudoplantago has unilocular (at anthesis) anthers, a characteristic of the subfamily Gomphrenoideae, but floral structure as well as pollen morphology connect the genus to a group of genera within the Amaranthoideae, subtribe Achyranthinae. The combination of subcuboidal shape and opercula with radially arranged hooked protuberances, makes the pollen of Pseudoplantago unique among the angiosperms studied so far. Floral morphology and palynological characteristics indicate a close relationship between Pfaffia and Alternanthera. Both genera, as currently accepted, are relatively homogeneous from pollen morphological points of view. There are no correlations between pollen morphology and the variation in the androecium in Pfaffia, nor would pollen structure support recognition of Hebanthe as a distinct genus. Woehleria and Irenella may be derived from, or be of the same origin as, Dicraurus and Iresine. All four genera are placed in the subfamily Gomphrenoideae because of the bisporangiate anthers, but their pollen structure is very close to, or identical with, that of the Amaranthus-type. Pseudogomphrena combines characteristics of Gomphrena and Pfaffia.     

EVOLUTIONARY AND ECOLOGICAL SIGNIFICANCE OF STARCH STORAGE IN POLLEN OF THE ARACEAE

Starch content was qualitatively assessed for pollen of 79 of the 111 currently recognized genera of the family Araceae—one of three monocot families known to exhibit both starchy and starchless pollen. Although 73% of the genera investigated had exclusively starchy pollen, character correlation suggests that starchless pollen is the primitive type for the family Araceae, as well as for monocots in general. Pollen starch content is a highly conservative character at the generic level in Araceae; only a single genus (Schismatoglottis) clearly exhibits both character states. The distribution of starchy pollen among aroid genera is consistent with what have here been termed Bakers' Starch Laws. Aroid pollen below a certain critical diameter—17-25 μm—is almost invariably starchless. Larger pollen is nearly always starchy, except where insect pollinators may use pollen nutritionally. There is strong evidence that the trend from starchless to starchy pollen in Araceae is reversible, according to the constraints imposed by the aforementioned factors.     

Pollen-ovule patterns in tribeTrifolieae (Leguminosae)

Relative pollen and ovule production in the genera of the legume tribeTrifolieae is explored particularly as to howMedicago with its explosive pollination mechanism compares with its allies.Medicago produces much larger, although much fewer pollen per ovule than the other five genera in the tribe; this is interpreted as a consequence of its highly specialized, irreversible pollination mechanism, which allows only one effective exchange of pollen with pollinators.Melilotus andTrifolium produce a comparatively large quantity of pollen; this can be related to their floral characteristics requiring abundant pollen to achieve fertilization. InMedicago andTrigonella, annuals convert a higher proportion of pollen grains and ovules into seeds than perennials.     

The pollen morphology ofOdontites (Scrophulariaceae) and its taxonomic significance

The pollen morphology of 29 species ofOdontites and related genera was examined by light and scanning electron microscopy. Three major pollen types differing fundamentally in exine sculpturing were found. In the parasitic tribePedicularieae retipilate sculpturing is the most widespread type representing a plesiomorphic character state from which the two other types are derived. Of these, reticulate sculpturing is confined toOdontites, whereas a complex retirugulate pattern was found only in the monotypic Near East genusBornmuellerantha. Within the retipilate and the reticulate major types eight minor types were distinguished based on the differential correlation of exine surface morphology, size, shape and amb form. The pollen data are generally well correlated with macromorphological features and furnish important taxonomic characters at the genus—as well as at the specieslevel.     

Substrate-dependent modulation of ATPase activity by Na+ and K+ in roots of Plantago species

The equal rates of water vapour absorption by both bi- and trinucleate pollen indicate that their widely-differing rates of respiration have an intrinsic, biochemical basis. This was investigated with various metabolic inhibitors that were previously introduced into dry pollen via anhydrous acetone. The uncoupler, carbonyl cyanide m-chlorophenyl hydrazone, inhibited the O₂ uptake of rapidly respiring pollen and stimulated that of slowly respiring types to similar absolute values, that probably reflect the rates of substrate transport across the mitochondrial membranes. The extent of inhibition of the O₂ uptake by oligomycin, dicyclohexyl carbodiimide, antimycin A, and salicyl hydroxamic acid, alone and in combinations, indicates that hardly any oxidative phosphorylation and anabolic activities occur in slowly respiring, binucleate pollen species, having low-developed mitochondria and high energy charge values. The presence of the alternative pathway was insignificant. In other binucleate pollen species, characterized by recognizable mitochondria and low energy charge values, a limited ATP synthesis was established. The low energy charge values point to imbalance between phosphorylative and anabolic activities. In rapidly respiring, trinucleate pollen, containing well-developed mitochondria, a significant activity of the alternative oxidase was found. The energy charge values were high notwithstanding the large demand for ATP, mounting to 1.7 μmol h^?1 (mg pollen)^?1. In some pollen species, oligomycin highly stimulated the flow of electrons through the cytochrome pathway, which made an estimation of the ATP synthesis impossible.     

Synchronized dichogamy and dioecy in neotropicalLauraceae

In hermaphrodite neotropicalLauraceae a highly evolved dichogamous system is present which represents a kind of temporal dioecy. This system involves the existence of two flower morphs which are characterized by reciprocal phases of receptivity of the stigmas and pollen release. In some genera (Persea, Cinnamomum), nectar is produced as a reward for the flower visitors, while in other genera (Aniba, Clinostemon, Licaria), nectar is absent and pollen seems to be the only reward. This implies that in this case the flowers in the female stage must be deceptive flowers. In dioecious species of the generaOcotea andNectandra, both the male and female flowers attract the visitors with nectar. The pollen-ovule ratio of theLauraceae is comparatively low. — The type of reproductive system that characterizes theLauraceae, comprising functional dioecy, small, inconspicuously coloured flowers, pollination by small bees, and large, one-seeded fruits dispersed by birds, is quite prominent among trees of various families in the tropical lowland forest. The relationship between the different modes of flowering within theLauraceae and the causes for the correlation of their reproductive traits are discussed.     

Ontogeny,function and evolution of extreme floral construction inMonimiaceae

In at least 4 genera of theMonimiaceae (Tambourissa, Wilkiea, Kibara, Hennecartia) extremely specialized flowers with a hyperstigma occur, i.e. a secretory zone in the narrow entrance of the floral cup. The mucilaginous secretion of the hyperstigma and of the carpels produces a transmitting medium for pollen tubes continuous from the mouth of the floral cup to the ovules. As to their floral morphology, the two extreme types,Hortonia andTambourissa, are connected gradually by various other genera. Possible evolutionary trends and systematic problems are outlined.     

Notes on the floral morphology in Vivianiaceae (Geraniales)

The functional floral morphology of the three genera of Vivianiaceae (= Ledocarpaceae, Geraniales), Rhynchotheca, Viviania and Balbisia, is compared. Likely pollination mechanisms are inferred from morphology and field observations. The flowers of Viviania are nectariferous and apparently zoophilous with nectar as the (primary) pollinator reward. Balbisia has pollen flowers without nectaries, its showy corolla indicates that it is also zoophilous with pollen as sole pollinator reward; bees were observed as flower visitors. One taxon (B. gracilis) may be anemophilous. Rhynchotheca has flowers without petals, with large, pendulous anthers and lacks nectaries. It shows synchronous mass flowering in its natural populations and is evidently anemophilous. A comparison with other Geraniales shows that nectar flowers with small anthers are likely the ancestral condition in Vivianiaceae. This suggests that the pollen flowers with larger anthers of Balbisia and Rhynchotheca may represent an apomorphic condition. The documentation of pollen flowers and anemophily in Vivianiaceae expands the range of known floral and pollination syndromes in Geraniales.