Ultrastructural analysis of Rhynchospora ovules: The first record of Cyperaceae megagametophyte on transmission electron microscope.

Microsporogenesis and microgametogenesis are unusual in sedges (Cyperaceae), the third largest monocotyledonous family, as three microspores are aborted in favor of a single functional microspore. However, studies using light microscopy show that megasporogenesis and megagametogenesis occur normally. Nevertheless, the lack of ultrastructural details limits our knowledge of female gametophyte development in this family. Given the importance of morphological studies of reproductive structures, ovules and megagametophytes of Rhynchospora pubera were analyzed under transmission electron microscopy for the first time. Overall, ovules presented features similar to those described for the family, but ultrastructural details revealed an absence of a clear boundary between the egg cell and the central cell cytoplasm. Most interestingly, antipodal and nucellar cells showed several signs of vacuolar cell death, which suggest that programmed autolysis in sporogenous and gametophytic tissue is common in gametophyte development in the Cyperaceae. This may be related to the reproductive success of this family.

Morpho-palynological study of Cyperaceae from wetlands of Azad Jammu and Kashmir using SEM and LM.

In this study 12 species of Cyperaceae have been studied for quantitative and qualitative observation of pollen grains through Light and scanning electron microscopy. Pollens of 12 species of Cyperaceae from different wetlands of Azad Jammu and Kashmir were collected. Morphological characters of pollen grains were then investigated under the Light and Scanning electron microscope. Two pollen types have been observed apolar and heteropolar. Shape of pollens was prolate (4 spp), sub-spheroidal (7 spp), and oblate (1 spp). Variation observed in exine sculpturing granular (4 spp), reticulate (1 spp), areolate-punctate (3 spp), and psilate (2 spp). Polar to equatorial ratio and fertility percentage of the pollens were also studied. Based on these micromorphlogical characters of pollens taxonomic keys have been made for the accurate identification of the members of Cyperaceae. The characteristics studied in present research work are very much valuable taxonomically and phytochemically for the identification of species of family Cyperaceae. Light microscope (LM) and Scanning electron microscope (SEM) were used for pollen observation, which play vital role in the taxonomical identification of species and provide sufficient information for taxonomist.

Developmental programmed cell death during asymmetric microsporogenesis in holocentric species of Rhynchospora (Cyperaceae).

Members of the Cyperaceae family exhibit an asymmetric microsporogenesis that results in the degeneration of three out of four meiotic products. Efforts have been made previously to describe the resulting structure, named the pseudomonad, but mechanisms concerning the establishment of cell domains, nuclear development, and programmed cell death are largely unknown. Using the Rhynchospora genus as a model, evidence for cell asymmetry, cytoplasmic isolation, and programmed cell death was obtained by a combination of electron microscopic, cytochemical, immunocytochemical, in situ hybridization, and flow cytometric methods. Degenerative cells were identified at the abaxial region, with the cytoskeleton marking their delimitation from the functional domain after meiosis. After attempting to initiate cell division with an unreplicated genome and abnormal spindle assembly, these cells exhibited a gradual process of cytoplasmic contraction associated with hypermethylation of cytosines and differential loss of DNA. These results indicate that the asymmetric tetrad establishes a functional cell, where one nucleus is preferentially selected to survive. Degenerative haploid cells are then eliminated in a multistep process associated with mitotic disorder, non-random elimination of repetitive DNA, vacuolar cell death, and DNA fragmentation.

Evolutionary divergence of ß-expansin structure and function in grasses parallels emergence of distinctive primary cell wall traits.

Expansins are wall-loosening proteins that promote the extension of primary cell walls without the hydrolysis of major structural components. Previously, proteins from the EXPA (α-expansin) family were found to loosen eudicot cell walls but to be less effective on grass cell walls, whereas the reverse pattern was found for EXPB (ß-expansin) proteins obtained from grass pollen. To understand the evolutionary and structural bases for the selectivity of EXPB action, we assessed the extension (creep) response of cell walls from diverse monocot families to EXPA and EXPB treatments. Cell walls from Cyperaceae and Juncaceae (families closely related to grasses) displayed a typical grass response ('ß-response'). Walls from more distant monocots, including some species that share with grasses high levels of arabinoxylan, responded preferentially to α-expansins ('α-response'), behaving in this regard like eudicots. An expansin with selective activity for grass cell walls was detected in Cyperaceae pollen, coinciding with the expression of genes from the divergent EXPB-I branch that includes grass pollen ß-expansins. The evolutionary origin of this branch was located within Poales on the basis of phylogenetic analyses and its association with the 'sigma' whole-genome duplication. Accelerated evolution in this branch has remodeled the protein surface in contact with the substrate, potentially for binding highly substituted arabinoxylan. We propose that the evolution of the divergent EXPB-I group made a fundamental change in the target and mechanism of wall loosening in the grass lineage possible, involving a new structural role for xylans and the expansins that target them.

Effects of lead on anatomy, ultrastructure and concentration of nutrients in plants Oxycaryum cubense (Poep. & Kunth) Palla: a species with phytoremediator potential in contaminated watersheds.

Lead (Pb) has been highlighted as a major pollutant of both terrestrial and aquatic ecosystems, causing negative impacts to these environments. The concentration of Pb in plants has increased in recent decades, mainly due to anthropogenic activities. This study has as a hypothesis that the species Oxycaryum cubense (Poep. & Kunth) Palla, abundant in aquatic environments, has the potential to be used a phytoremediator. The plants were grown in a hydroponic system with Pb in increasing concentrations (0, 4, 8, 16 and 32 mg l(-1)) for 15 days. Inductively coupled mass spectrometer (ICP OES) was used to determine the concentration of mineral nutrients and lead. Optical and transmission electron microscopy were used for the analysis of cellular damage induced by lead in roots and leaves. Ultrastructural alterations were observed as disorganization of thylakoids in the chloroplast and disruption of mitochondrial membranes in cells of leaf tissues of plants subjected to increasing Pb concentrations. There was accumulation of Pb, especially in the root system, affecting the absorption and translocation of some mineral nutrients analysed. In roots, there was reduction in the thickness of the epidermis in plants treated with Pb. This species was shown to be tolerant to the Pb concentrations evaluated, compartmentalizing and accumulating Pb mainly in roots. Due to these results, it may be considered a species with phytoremediation capacity for Pb, with potential rizofiltration of this metallic element in contaminated watersheds.

Silica nanoparticles aid in structural leaf coloration in the Malaysian tropical rainforest understorey herb Mapania caudata.

BACKGROUND AND AIMS: Blue-green iridescence in the tropical rainforest understorey sedge Mapania caudata creates structural coloration in its leaves through a novel photonic mechanism. Known structures in plants producing iridescent blues consist of altered cellulose layering within cell walls and in special bodies, and thylakoid membranes in specialized plastids. This study was undertaken in order to determine the origin of leaf iridescence in this plant with particular attention to nano-scale components contributing to this coloration. METHODS: Adaxial walls of leaf epidermal cells were characterized using high-pressure-frozen freeze-substituted specimens, which retain their native dimensions during observations using transmission and scanning microscopy, accompanied by energy-dispersive X-ray spectroscopy to identify the role of biogenic silica in wall-based iridescence. Biogenic silica was experimentally removed using aqueous Na2CO3 and optical properties were compared using spectral reflectance. KEY RESULTS AND CONCLUSIONS: Blue iridescence is produced in the adaxial epidermal cell wall, which contains helicoid lamellae. The blue iridescence from cell surfaces is left-circularly polarized. The position of the silica granules is entrained by the helicoid microfibrillar layers, and granules accumulate at a uniform position within the helicoids, contributing to the structure that produces the blue iridescence, as part of the unit cell responsible for 2 ° Bragg scatter. Removal of silica from the walls eliminated the blue colour. Addition of silica nanoparticles on existing cellulosic lamellae is a novel mechanism for adding structural colour in organisms.

Developmental anatomy of Cyperus laxus (non-Nranz) and Fimbristylis dichotoma (Kranz) (Cyperaceae, Poales) and tissue continuity.

The Cyperaceae species are present in different ecosystems and constitute the herbaceous extract. Of the approximately 5,500 species of the family; a third has Kranz anatomy, representing an important characteristic of the taxonomy and phylogeny of the group. In Cyperus laxus L. (non-Kranz) and Fimbristylis dichotoma Vahl (Kranz), development begins with germination that is marked by the emergence of the coleoptiles, followed by the primary root, which is ephemeral. The rhizome originates from the mesocotyl and it promotes the vascular connection between the roots, leaves and scapes. The continuity of the tissues is evidenced by the presence of an endodermis and pericycle in all vegetative organs. Leaves and scapes differ between the two species by the arrangement of mesophyll cells, which is regular in Cyperus laxus (non-Kranz) and arranged radially in Fimbristylis dichotoma (Kranz). Also differ in the number of bundles sheaths: two in Cyperus laxus (non-Kranz) and three in Fimbristylis dichotoma (Kranz). The outer bundle sheath in both species constitutes the endodermis, and the inner sheath in Cyperus laxus and the middle and inner sheaths in Fimbristylis dichotoma constitute the pericycle.

Respiratory flexibility and efficiency are affected by simulated global change in Arctic plants.

Laboratory studies indicate that, in response to environmental conditions, plants modulate respiratory electron partitioning between the 'energy-wasteful' alternative pathway (AP) and the 'energy-conserving' cytochrome pathway (CP). Field data, however, are scarce. Here we investigate how 20-yr field manipulations simulating global change affected electron partitioning in Alaskan Arctic tundra species. We sampled leaves from three dominant tundra species - Betula nana, Eriophorum vaginatum and Rubus chamaemorus - that had been strongly affected by manipulations of soil nutrients, light availability, and warming. We measured foliar dark respiration, in-vivo electron partitioning and alternative oxidase/cytochrome c oxidase concentrations in addition to leaf traits and mitochondrial ultrastructure. Changes in leaf traits and ultrastructure were similar across species. Respiration at 20°C (R(20)) was reduced 15% in all three species grown at elevated temperature, suggesting thermal acclimation of respiration. In Betula, the species with the largest growth response to added nutrients, CP activity increased from 9.4 ± 0.8 to 16.6 ± 1.6 nmol O(2) g(-1) DM s(-1) whereas AP activity was unchanged. The ability of Betula to selectively increase CP activity in response to the environment may contribute to its overall ecological success by increasing respiratory energy efficiency, and thus retaining more carbon for growth.

Asymmetric cytokinesis guide the development of pseudomonads in Rhynchospora pubera (Cyperaceae).

The late stages of microsporogenesis in the family Cyperaceae are marked by the formation of an asymmetrical tetrad, degeneration of three of the four nuclei resulting from meiosis and the formation of pseudomonads. In order to understand the cytological changes involved in the development of pseudomonads, a combination of 11 different techniques (conventional staining, cytochemistry procedures, immunofluorescence, FISH and transmission electron microscopy: TEM) were used to study the later stages of microsporogenesis in Rhynchospora pubera. The results demonstrated the occurrence of two cytoplasmic domains in the pseudomonads, one functional and the other degenerative, which are physically and asymmetrically separated by cell plate with an endomembrane system rich in polysaccharides. Other changes associated with endomembrane behaviour were observed, such as a large number of lipid droplets, vacuoles containing electron-dense material and concentric layers of endoplasmic reticulum. Concomitant with the isolation of degenerative nuclei, the tapetal cells also showed evidence of degeneration, indicating that both tissues under programmed cell death (PCD), as indicated by immunofluorescence and TEM procedures. The results are significant because they associate cellular polarisation and asymmetry with different cytoplasmic domains, and hence open new possibilities for studying cellular compartmentalisation and PCD.

Leaf- and cell-level carbon cycling responses to a nitrogen and phosphorus gradient in two Arctic tundra species.

PREMISE OF THE STUDY: Consequences of global climate change are detectable in the historically nitrogen- and phosphorus-limited Arctic tundra landscape and have implications for the terrestrial carbon cycle. Warmer temperatures and elevated soil nutrient availability associated with increased microbial activity may influence rates of photosynthesis and respiration. • METHODS: This study examined leaf-level gas exchange, cellular ultrastructure, and related leaf traits in two dominant tundra species, Betula nana, a woody shrub, and Eriophorum vaginatum, a tussock sedge, under a 3-yr-old treatment gradient of nitrogen (N) and phosphorus (P) fertilization in the North Slope of Alaska. • KEY RESULTS: Respiration increased with N and P addition-the highest rates corresponding to the highest concentrations of leaf N in both species. The inhibition of respiration by light ("Kok effect") significantly reduced respiration rates in both species (P < 0.001), ranged from 12-63% (mean 34%), and generally decreased with fertilization for both species. However, in both species, observed rates of photosynthesis did not increase, and photosynthetic nitrogen use efficiency generally decreased under increasing fertilization. Chloroplast and mitochondrial size and density were highly sensitive to N and P fertilization (P < 0.001), though species interactions indicated divergent cellular organizational strategies. • CONCLUSIONS: Results from this study demonstrate a species-specific decoupling of respiration and photosynthesis under N and P fertilization, implying an alteration of the carbon balance of the tundra ecosystem under future conditions.

Multiple origins of congested inflorescences in Cyperus s.s. (Cyperaceae): developmental and structural evidence.

PREMISE OF THE STUDY: The understanding of homoplasic structures becomes more relevant when they are complex and define large angiosperm taxa. Inflorescence architecture usually fulfills both features, as happens with Cyperus, a genus with two taxonomical subdivisions characterized either by alternative expressions of Kranz anatomy (C(3) or C(4)) or inflorescence shape (condensed or lax). Those subdivisions are not completely congruent because at least one of these presumed characters has evolved several times. We focused a SEM study on the inflorescence development in species with condensed inflorescences and different photosynthetic anatomy to test the possibility that condensed inflorescences of subgen. Anosporum (C(3) anatomy) have evolved independently from those of subgen. Cyperus (C(4) anatomy). METHODS: Freshly collected inflorescences of C. entrerianus, C. eragrostis, C. oxylepis, and C. incomtus were studied using stereoscopic and scanning electron microscopy. KEY RESULTS: Condensed inflorescences of Cyperus species with C(3) and C(4) anatomy had differences in structure and development: (1) mature structure, (2) position of second-order branching initiation in the first developmental stage of the inflorescence, (3) main axis development and elongation, and branching development, (4) types of ramifications, (5) phyllotaxis and symmetry. CONCLUSIONS: Results support multiple origins of condensed inflorescences in Cyperus, based especially on differences in timing during development and elongation of the main axis and branches, branching pattern and phyllotaxis. Structure and development may be the key to using inflorescence morphology as an external feature to distinguish large natural groups within Cyperus based on vegetative anatomy.

Selective microspore abortion correlated with aneuploidy: an indication of meiotic drive.

Selective megaspore abortion (monomegaspory) probably arose once in seed plants and occurs routinely in more than 70% of angiosperm species, representing one of the key characters of a heterosporous life history. In contrast, selective microspore abortion leading to pollen dispersal as pseudomonads (here termed monomicrospory) apparently arose at least twice independently within angiosperms, though it occurs in a limited number of taxa. Remarkably, similar examples of monomicrospory occur in members of two distantly related angiosperm families: the sedge family (Cyperaceae) and the epacrid subfamily (Styphelioideae) of the eudicot family Ericaceae. In sedges, monomicrospory is derived directly from normal tetrads, whereas epacrid pseudomonads apparently evolved via an intermediate stage, in which variable sterility occurs in a single tetrad. Our comparison of these two examples of selective microspore abortion highlights a correlation with aneuploidy, indicating that non-random chromosome segregation caused by monomicrospory could drive chromosomal mutations to rapid fixation through meiotic drive.

Long-term effects of elevated UV-B radiation on photosynthesis and ultrastructure of Eriophorum russeolum and Warnstorfia exannulata.

The depletion of stratospheric ozone above the Arctic regions may increase the amount of UV-B radiation to which the northern ecosystems are exposed. In this paper, we examine the hypothesis that supplemental UV-B radiation may affect the growth rate and photosynthesis of boreal peatland plants and could thereby affect the carbon uptake of these ecosystems. In this study, we report the effects of 3-year exposure to elevated UV-B radiation (46% above ambient) on the photosynthetic performance and ultrastructure of a boreal sedge Eriophorum russeolum and a moss Warnstorfia exannulata. The experiment was conducted on a natural fen ecosystem at Sodankylä in northern Finland. The effects of UV-B radiation on the light response of E. russeolum CO(2) assimilation and the maximal photochemical efficiency of photosystem II in a dark-adapted state (F(v)/F(m)) were measured in the field. In addition, the effect of supplemental UV-B radiation on organelles of photosynthetic cells was studied by electron microscopy. The UV-B treatment had no effect on the CO(2) assimilation rate of either species, nor did it affect the structure of the cell organelles. On chlorophyll fluorescence, the UV-B exposure had only a temporary effect during the third exposure year. Our results suggested that in a natural ecosystem, even long-term exposure to reasonably elevated UV-B radiation levels does not affect the photosynthesis of peatland plants.

Spikelet structure and development in Cyperoideae (Cyperaceae): a monopodial general model based on ontogenetic evidence.

BACKGROUND AND AIMS: In Cyperoideae, one of the two subfamilies in Cyperaceae, unresolved homology questions about spikelets remained. This was particularly the case in taxa with distichously organized spikelets and in Cariceae, a tribe with complex compound inflorescences comprising male (co)florescences and deciduous female single-flowered lateral spikelets. Using ontogenetic techniques, a wide range of taxa were investigated, including some controversial ones, in order to find morphological arguments to understand the nature of the spikelet in Cyperoideae. This paper presents a review of both new ontogenetic data and current knowledge, discussing a cyperoid, general, monopodial spikelet model. METHODS: Scanning electron microscopy and light microscopy were used to examine spikelets of 106 species from 33 cyperoid genera. RESULTS: Ontogenetic data presented allow a consistent cyperoid spikelet model to be defined. Scanning and light microscopic images in controversial taxa such as Schoenus nigricans, Cariceae and Cypereae are interpreted accordingly. CONCLUSIONS: Spikelets in all species studied consist of an indeterminate rachilla, and one to many spirally to distichously arranged glumes, each subtending a flower or empty. Lateral spikelets are subtended by a bract and have a spikelet prophyll. In distichously organized spikelets, combined concaulescence of the flowers and epicaulescence (a newly defined metatopic displacement) of the glumes has caused interpretational controversy in the past. In Cariceae, the male (co)florescences are terminal spikelets. Female single-flowered spikelets are positioned proximally on the rachis. To explain both this and the secondary spikelets in some Cypereae, the existence of an ontogenetic switch determining the development of a primordium into flower, or lateral axis is postulated.

Early meiosis in Rhynchospora pubera L. (Cyperaceae) is marked by uncommon ultrastructural features.

The family Cyperaceae has an unusual microsporogenesis in which tetrad formation does not occur. In addition, other cytological features are important, such as the occurrence of holokinetic chromosomes and post-reductional meiosis. We have examined the ultrastructural features of the pollen mother cell (PMC) of Rhynchospora pubera. Anthers of several sizes were analyzed using light and transmission electron microscopy. The PMC before meiosis presented a central nucleus and a regular profile of the nuclear envelope. During prophase I, the nucleus was in the abaxial region of the cell. This cellular polarization was accompanied by other marked ultrastructural features in the nuclear envelope. Morphological changes involved dilations of perinuclear cisterns and polarization of the nuclear pore complexes. The results show that polarization occurs in the initial phases of microsporogenesis in R. pubera, unlike other plant species.

The floral scales in Hellmuthia (Cyperaceae, Cyperoideae) and Paramapania (Cyperaceae, Mapanioideae): an ontogenetic study.

BACKGROUND AND AIMS: In 1976 the monotypic genus Hellmuthia was placed in the Hypolytreae s.l., but was subsequently ascribed to the Mapanioideae, tribe Chrysitricheae, mainly because of the presence in Hellmuthia of two lateral, mapanioid-like floral scales with ciliated keels, the anatomy of the nutlet, the embryo and the inflorescence. Recently, based on cladistic analyses and supported by pollen ontogenetic evidence, Hellmuthia was transferred to a Cyperaceae, tribe Cypereae, clade mainly consisting of Ficinia and Isolepis. In this study, the floral ontogeny in Hellmuthia was investigated and compared with the floral ontogeny in Paramapania, with special attention for the floral scales. METHODS: Freshly collected inflorescences of Hellmuthia membranacea and Paramapania parvibractea were investigated using scanning electron and light microscopy. KEY RESULTS: In the conical 'spikelet' in Hellmuthia, proximal bracts occur, each axillating an axis with empty glume-like structures, or a reduced spikelet. Hence, it is a reduced partial inflorescence. In Hellmuthia, the stamen primordia originate before the primordia of the perianth-gynoecium appear. Moreover, a third adaxially positioned 'floral scale' was observed for the first time. The position and relative time of appearance of the floral scales in Hellmuthia are typical for perianth parts in Cyperoideae. The basal position of Hellmuthia within a clade of species with usually perianthless flowers, allows the presence of rudiments of a perianth in Hellmuthia to be interpreted as a primitive state. Development of the lateral 'scales' in Paramapania follows a different pattern. Therefore, it was decided that the lateral 'scales' in Paramapania are different from the lateral perianth parts in Hellmuthia. The pollen grains in Hellmuthia are cyperoid, with one polar and five lateral apertures, of which the membrane is covered with sexinous bodies. The pollen surface is granulate and perforate with microspines. CONCLUSIONS: The floral ontogeny in Hellmuthia occurs according to the general cyperoid pattern. The lateral scales in Hellmuthia are perianth parts, and they are not homologous to the lateral 'scales' in Paramapania.

Ozone effects on the ultrastructure of peatland plants: Sphagnum mosses, Vaccinium oxycoccus, Andromeda polifolia and Eriophorum vaginatum.

BACKGROUND AND AIMS: Ozone effects on peatland vegetation are poorly understood. Since stress responses are often first visible in cell ultrastructure, electron microscopy was used to assess the sensitivity of common peatland plants to elevated ozone concentrations. METHODS: Three moss species (Sphagnum angustifolium, S. magellanicum and S. papillosum), a graminoid (Eriophorum vaginatum) and two dwarf shrubs (Vaccinium oxycoccus and Andromeda polifolia), all growing within an intact canopy on peat monoliths, were exposed to a concentration of 0, 50, 100 or 150 ppb ozone in two separate growth chamber experiments simulating either summer or autumn conditions in central Finland. After a 4- or 5-week-long exposure, samples were photographed in a transmission electron microscope and analysed quantitatively using image processing software. KEY RESULTS: In the chlorophyllose cells of the Sphagnum moss leaves from the capitulum, ozone exposure led to a decrease in chloroplast area and in granum stack thickness and various changes in plastoglobuli and cell wall thickness, depending on the species and the experiment. In E. vaginatum, ozone exposure significantly reduced chloroplast cross-sectional areas and the amount of starch, whereas there were no clear changes in the plastoglobuli. In the dwarf shrubs, ozone induced thickening of the cell wall and an increase in the size of plastoglobuli under summer conditions. In contrast, under autumn conditions the cell wall thickness remained unchanged but ozone exposure led to a transient increase in the chloroplast and starch areas, and in the number and size of plastoglobuli. CONCLUSIONS: Ozone responses in the Sphagnum mosses were comparable to typical ozone stress symptoms of higher plants, and indicated sensitivity especially in S. angustifolium. The responses in the dwarf shrubs suggest stimulation of photosynthesis by low ozone concentrations and ozone sensitivity only under cool autumn conditions.