The relationship between cell division and elongation during development of the nectar-yielding petal spur in Centranthus ruber (Valerianaceae)

Background and Aims Floral spurs are hollow, tubular outgrowths that typically conceal nectar. By their involvement in specialized pollinator interactions, spurs have ecological and evolutionary significance, often leading to speciation. Despite their importance and diversity in shape and size among angiosperm taxa, detailed investigations of the mechanism of spur development have been conducted only recently.Methods Initiation and growth of the nectar-yielding petal spur of Centranthus ruber ‘Snowcloud’ was investigated throughout seven stages, based on bud size and developmental events. The determination of the frequency of cell division, quantified for the first time in spurs, was conducted by confocal microscopy following 4'',6-diamidino-2-phenylindole (DAPI) staining of mitotic figures. Moreover, using scanning electron microscospy of the outer petal spur surface unobstructed by trichomes, morphometry of epidermal cells was determined throughout development in order to understand the ontogeny of this elongate, hollow tube.Key Results Spur growth from the corolla base initially included diffuse cell divisions identified among epidermal cells as the spur progressed through its early stages. However, cell divisions clearly diminished before a petal spur attained 30 % of its final length of 4·5 mm. Thereafter until anthesis, elongation of individual cells was primarily responsible for the spur’s own extension. Consequently, a prolonged period of anisotropy, wherein epidermal cells elongated almost uniformly in all regions along the petal spur’s longitudinal axis, contributed principally to the spur’s mature length.Conclusions This research demonstrates that anisotropic growth of epidermal cells – in the same orientation as spur elongation – chiefly explains petal spur extension in C. ruber. Representing the inaugural investigation of the cellular basis for spur ontogeny within the Euasterids II clade, this study complements the patterns in Aquilegia species (order Ranunculales, Eudicots) and Linaria vulgaris (order Lamiales, Euasterids I), thereby suggesting the existence of a common underlying mechanism for petal spur ontogeny in disparate dicot lineages.     

Determining the contribution of epidermal cell shape to petal wettability using isogenic Antirrhinum lines

The petal epidermis acts not only as a barrier to the outside world but also as a point of interaction between the flower and potential pollinators. The presence of conical petal epidermal cells has previously been shown to influence the attractiveness of the flower to pollinating insects. Using Antirrhinum isogenic lines differing only in the presence of a single epidermal structure, conical cells, we were able to investigate how the structure of the epidermis influences petal wettability by measuring the surface contact angle of water drops. Conical cells have a significant impact on how water is retained on the flower surface, which may have indirect consequences for pollinator behaviour. We discuss how the petal epidermis is a highly multifunctional one and how a battery of methods, including the use of isogenic lines, is required to untangle the impacts of specific epidermal properties in an ecological context.     

A fine structural analysis of the epidermis and cuticle of the oligochaete aeolosoma Bengalense stephenson

The fine structure of the epidermis and cuticle has been described for the oligochaete Aeolosoma bengalense. The epidermis is a pseudostratified epithelium and consists of the following cell types: ciliated and nonciliated supportive cells, pigment cells and associated satellite cells, mucous cells, basal cells, and ciliated non-supportive columnar cells. Overlying and restricted to the supportive cells is a delicate cuticle composed of: (a) a discontinuous layer of membrane-bounded surface particles; (b) a thin filamentous layer of moderate electron density just under the surface particles; (c) a thicker inner filamentous layer of low electron density. Digestion with pronase effectively removes the cuticle. This, together with the fact that it stains with alcian blue and ruthenium red, indicates that the cuticle contains an acid mucopolysaccharide. Regeneration of the cuticle, following pronase treatment, is marked by the elaboration of numerous microvilli by the supportive cells. Most of the microvilli are transitory and evidence supports a microvillar origin for the cuticular surface particles. The presence of cuticular surface particles may be a characteristic shared in common by all oligochaetes and, perhaps, some polychaetes.     

Why do so many petals have conical epidermal cells?

Background

The conical epidermal cells found on the petals of most Angiosperm species are so widespread that they have been used as markers of petal identity, but their function has only been analysed in recent years. This review brings together diverse data on the role of these cells in pollination biology.

Scope

The published effects of conical cells on petal colour, petal reflexing, scent production, petal wettability and pollinator grip on the flower surface are considered. Of these factors, pollinator grip has been shown to be of most significance in the well-studied Antirrhinum majus/bumble-bee system. Published data on the relationship between epidermal cell morphology and floral temperature were limited, so an analysis of the effects of cell shape on floral temperature in Antirrhinum is presented here. Statistically significant warming by conical cells was not detected, although insignificant trends towards faster warming at dawn were found, and it was also found that flat-celled flowers could be warmer on warm days. The warming observed is less significant than that achieved by varying pigment content. However, the possibility that the effect of conical cells on temperature might be biologically significant in certain specific instances such as marginal habitats or weather conditions cannot be ruled out.

Conclusions

Conical epidermal cells can influence a diverse set of petal properties. The fitness benefits they provide to plants are likely to vary with pollinator and habitat, and models are now required to understand how these different factors interact.     

Effect of sowing dates and vernalization on Betavulgaris L. cv. Univers C-leaf structure

This research was conducted to study the effect of three different sowing dates (15th October, 15th November and 15th December) and two vernalization treatments (5 °C and −20 °C) on leaf structure of Betavulgaris L. cv. Univers. The obtained data are summarized as follows:The maximum values of the most studied parameters; lower epidermis + spongy tissue thickness, midrib, mesophyll tissue, vascular bundle, collenchymatous tissue and number of xylem vessels per arm were found as a result of 15th October sowing date treatment compared with the two other sowing dates. Furthermore, effect of the cooling treatments varied according to the recorded character, sowing date and cooling degree. Most of the vernalization treatments at early sowing dates increased the mesophyll tissue, midrib, number of vascular bundles per transverse section, vascular bundle thickness and number of xylem arms per transverse section.The two studied cooling treatments at 15th October sowing date increased both stomatal index and average number of stomata: average number of epidermis cells compared with the control. Furthermore, 15th October under −20 °C treatment led to small epidermal cells and stomata formation, straight epidermal cell walls and closed stomata in comparison to the control.     

Cytochemical studies on the cuticle and epidermis of Tubifex tubifex Müll. with special regard to the localization of polysaccharides, heavy metals and the DAB-reactivity.

The cuticle consits of collagenous fibres and of two types of mucopolysaccharides. The whole cuticle contain PAS-positive polysaccharides, but the acid ones are localized only in the surface zone, both in the epicuticle and in the supracuticular mucoid coat. On the surface-damaged region of the cuticle bacterial infection was observed, those intact mucoid coat may be essential in protection against bacterial infection. Microvilli of the supporting cells play significant role in repairing of cuticular injuries. Acid mucopolysaccharides of the cuticle and epidermis may function as traps for heavy metals, proved by their significant heavy metal content. The cytosol of the epidermal cells possess considerable DAB-reactivity. The enzyme, responsible for the DAB-reaction, may be transported by the microvilli towards the cuticular surface and can play central role in the detoxication of organic foreign compounds.     

ABC-type transporters and cuticle assembly: Linking function to polarity in epidermis cells

The aerial organs of plants are covered with a cuticle, a continuous layer overlaying the outermost cell walls of the epidermis. The cuticle is composed of two major classes of the lipid biopolymers: cutin and waxes, collectively termed cuticular lipids. Biosynthesis and transport of cuticular lipids occur predominantly in the epidermis cells. In the transport pathway, cuticular lipids are exported from their site of biosynthesis in the ER/plastid to the extracellular space through the plasma membrane and cell wall. Growing evidence suggests that ATP-binding cassette (ABC) transporters are implicated in transport of cuticular lipids across the plasma membrane of epidermal cells. The Arabidopsis ABC-type transporter protein CER5 (WBC12) was reported to act as a wax monomers transporter. In recent works, our group and others showed that a CER5-related protein, DESPERADO (DSO/WBC11), is required for cutin and wax monomers transport through the plasma membrane of Arabidopsis epidermis cells. Unlike the cer5 mutant, DSO loss-of-function had a profound effect on plant growth and development, particularly dwarfism, postgenital organ fusions, and altered epidermal cell differentiation. The partially overlapping function of CER5 and DSO and the fact that these proteins are half-size ABC transporters suggest that they might form a hetero-dimeric complex while transporting wax components. An intriguing observation was the polar localization of DSO in the distal part of epidermis cells. This polar expression might be explained by DSO localization within lipid rafts, specific plasma membrane microdomains which are associated with polar protein expression. In this review we suggest possible mechanisms for cuticular lipids transport and a link between DSO function and polar expression. Furthermore, we also discuss the subsequent transport of cuticular constituents through the hydrophobic cell wall and the possible involvement of lipid transfer proteins in this process.Key words: ABC transporter, cuticular lipids, polar expression, plasma membrane, epidermis     

Cuticular encystment of Autographa nigrisigna eggs by epidermal cell migration

It was previously established that Autographa nigrisigna loopers form cuticular cysts at the dorsal site of the 9th (penultimate) abdominal segment after parasitization by the solitary endoparasitoid Campoletis chlorideae and get rid of the parasitoid egg with the old cuticle at ecdysis. The cuticular cyst consists of a space between the old cuticle and new cuticle formed by the epidermis to enclose the parasitoid egg. The fact that A. nigrisigna loopers exclude the oviposited egg from the hemocoel using a cuticular cyst raises the question how the parasitoid egg passes through the epidermis. To exclude the endoparasitoid eggs from the hemocoel, the epidermis is required to move the location of the parasitoid egg. In the current study, we investigated the morphological process of cuticular cyst formation. First, the oviposited egg drifted to the 9th abdominal segment located at the open end of the dorsal vessel as a result of force generated by the hemolymph current from the oviposition site, and formed contacts with the integument containing the fat body (FB). The epidermis, in contact with the egg, then began to move along with the basement membrane formed on the surface of the FB, and settled under the egg, thus altering its location. This inversion was duplicated in vitro using integument from the 9th abdominal segment when parasitoid eggs were inserted between the epidermis and FB. When the integument, without the FB, was incubated on an agar plate, the epidermal cells migrated on the plate. Integument without eggs showed no signs of migration from their original sites. When the actin polymerization inhibitor latrunculin B was added to the cultures, the epidermal cells remained in their original location.     

Development of plant cuticles: fine structure and cutin composition of Clivia miniata Reg. leaves

The fine structure and monomeric composition of the ester-cutin fraction (susceptible to BF₃/CH₃OH transesterification) of the adaxial leaf cuticle of Clivia miniata Reg. were studied in relation to leaf and cuticle development. Clivia leaves grow at their base such that cuticle and tissues increase in age from the base to the tip. The zone of maximum growth (cell expansion) was located between 1 and 4 cm from the base. During cell expansion, the projected surface area of the upper epidermal cells increased by a factor of nine. In the growth region the cuticle consists mainly of a polylamellate cuticle proper of 100–250 nm thickness. After cell expansion has ceased both the outer epidermal wall and the cuticle increase in thickness. Thickening of the cuticle is accomplished by interposition of a cuticular layer between the cuticle proper and the cell wall. The cuticular layer exhibits a reticulate fine structure and contributes most of the total mass of the cuticle at positions above 6 cm from the leaf base. The composition of ester cutin changed with the age of cuticles. In depolymerisates from young cuticles, 26 different monomers could be detected whereas in older ones their number decreased to 13. At all developmental stages, 9,16-/10,16-dihydroxyhexadecanoic acid (positional isomers not separated), 18-hydroxy-9-octadecenoic acid, 9,10,18-trihydroxyoctadecanoic acid and 9,10-epoxy-18-hydroxyoctadecanoic acid were most frequent with the epoxy alkanoic acid clearly predominating (47% at 16 cm). The results are discussed as to (i) the age dependence of cutin composition, (ii) the relationship between fine structure and composition, (iii) the composition of the cuticle proper, the cuticular layer and the non-depolymerizable cutin fraction, and (iv) the polymeric structure of cutin.Abbreviations CL cuticular layer - CP cuticle proper - MX cutin polymer matrix     

Arabidopsis LTPG Is a Glycosylphosphatidylinositol-Anchored Lipid Transfer Protein Required for Export of Lipids to the Plant Surface

Plant epidermal cells dedicate more than half of their lipid metabolism to the synthesis of cuticular lipids, which seal and protect the plant shoot. The cuticle is made up of a cutin polymer and waxes, diverse hydrophobic compounds including very-long-chain fatty acids and their derivatives. How such hydrophobic compounds are exported to the cuticle, especially through the hydrophilic plant cell wall, is not known. By performing a reverse genetic screen, we have identified LTPG, a glycosylphosphatidylinositol-anchored lipid transfer protein that is highly expressed in the epidermis during cuticle biosynthesis in Arabidopsis thaliana inflorescence stems. Mutant plant lines with decreased LTPG expression had reduced wax load on the stem surface, showing that LTPG is involved either directly or indirectly in cuticular lipid deposition. In vitro 2-p-toluidinonaphthalene-6-sulfonate assays showed that recombinant LTPG has the capacity to bind to this lipid probe. LTPG was primarily localized to the plasma membrane on all faces of stem epidermal cells in the growing regions of inflorescence stems where wax is actively secreted. These data suggest that LTPG may function as a component of the cuticular lipid export machinery.     

Repellent,larvicidal and adulticidal activities of essential oil from Dai medicinal plant Zingiber cassumunar against Aedes albopictus

Zingiber cassumunar is an important plant used in traditional medicine and as a natural mosquito repellent. However, the compounds responsible for the repellent activity of the plant are still unknown. The aim of the study is to identify the components of Z. cassumunar essential oil that show repellent activity against Aedes albopictus. We also evaluated the larvicidal and adulticidal activities of Z. cassumunar essential oil against Ae. albopictus. In-cage mosquito repellent experiments showed that Z. cassumunar essential oil possessed moderate repellent activity with a minimum effective dose (MED) of 0.16 ± 0.01 mg/cm², compared to reference standard N,N-diethyl-3-methylbenzamide (DEET, 0.03 ± 0.01 mg/cm²). Bioassay-guided fractionation identified the major active compound of Z. cassumunar essential oil as (−)-terpinen-4-ol (1) (MED: 0.19 ± 0 mg/cm²). We also found that Z. cassumunar essential oil showed moderate larvicidal activity against first instar larvae of Ae. albopictus with a LC₅₀ (50% lethal concentration) of 44.9 μg/L after 24 h. Fumigation bioassays showed that Z. cassumunar essential oil exhibits moderate adulticidal activity against Ae. albopictus with a LC₅₀ of 5.44%, while (−)-terpinen-4-ol showed significant adulticidal activity with a LC₅₀ of 2.10% after 24 h. This study verifies that the Z. cassumunar essential oil has mosquito repellent activity, and that (−)-terpinen-4-ol is mainly responsible for this activity. Furthermore, this study provides scientific support for the folk usage of Z. cassumunar essential oil as mosquito repellent and indicates that Z. cassumunar essential oil and (−)-terpinen-4-ol can be used as plant-derived repellents and insecticides for mosquito control.     

Description and SEM Observations of a New Species of Cyst Nematode Heterodera goldeni (Nematoda: Heteroderidae) Attacking Panicum coloratum in Egypt

A cyst nematode, Heterodera goldeni n. sp., is photographed and described from Qasabagrass roots (Panicum coloratum L.) in Alexandria, Egypt. It is characterized in having second-stage juveniles with body length of 546 µm (450-612), stylet length of 22.6 µm (22-23.5) with anchor-shaped knobs, lateral field with 3 lines, tail 60-75 µm, hyaline tail terminus 38.4 µm (33-43); cysts are lemon-shaped, dark to light brown with an extensive sub-crystalline layer covering the entire cyst, cuticular midbody pattern zig-zag, cysts ambifenestrate, well-developed underbridge with finger-like projections, bullae present, vulva slit measuring 44-48 µm long. Males are absent, and females have heavy punctations on the cuticle. Its relationship to H. graminophila described from Florida and Louisiana and H. leuceilyma described from Florida are discussed. The present known distribution is restricted to Alexandria, Egypt. Its economic importance in rangeland grasses and cultivated crops such as rice is not known.     

The formation of the epicuticle and associated structures inOniscus asellus (Crustacea,Isopoda)

Summary The integument of the woodlouse,Oniscus asellus, consists of a two-layered epicuticle, a largely lamellate procuticle — itself divided into two regions (pre-and postecdysial cuticles), and the epidermis. At the initiation of new cuticle production the epidermal cells become vacuolated and retract away from the cuticle. Apolysis occurs immediately after the cessation of postecdysial cuticle production. The formation of the epicuticle is unique among the arthropods since material aggregates along the distal epidermal membrane. By indenting, doubling back on itself, and incorporating septa, the epicuticle forms surface structures such as plaques and tricorns.The innervation, and so the receptive function of the tricorns is confirmed, but since there is no connection between the old and new receptors during premoult, sensory information from these exoreceptors must be severely curtailed. This may explain the biphasic moult in all isopods since it ensures that only half the body experiences this sensory deprivation at any one time. In terrestrial species there is the additional advantage of restricting the area of permeable new cuticle. The frequency of moulting may be due to the need to renew disrupted receptor surfaces.Tricorns do not appear to be the mechanoreceptors involved in the marked thigmotactic response of woodlice since they do not have the typical internal structure of such receptors; rather, the dendrite —which extends into the lumen of the tricorn —is protected from deformation by the previously unreported combination of a dendritic sheath and a cuticular tube. The modality of tricorns is possibly one of hygro-perception. One of the behavioural responses of woodlice to desiccation is aggregation. The numerical distribution of tricorns over the body surface is admirably suited to assist in the formation and maintenance of such aggregates during desiccation and to their observed dispersal when the relative humidity rises.     

Ultrastructure of an integumental organ with probable sensory function in Paragordius varius (nematomorpha)

The cuticle of late parasitic stages of Paragordius varius (Leidy, 1851) is composed of a layer with large fibres and a second layer (often named the areolar layer) distal from it. In this paper, organs are described that start at the basal side of the epidermis, pass the epidermis and the fibrous layer of the cuticle and merge with large, cushion‐like structures in the distal layer of the cuticle. The epidermal part of the organs is composed of darkly stained cells, which are probably in contact with the basi‐epidermal nervous system. Up to four processes of this cell traverse the cuticle. These processes might include cilia, because they contain microtubule‐like structures. The probable connection to nerve cells and the connection to the cushion‐like structures in the outer cuticular layer make it likely that the organs described here are sensory in function.     

Pigment distribution, light reflection and cell structure in petals

Petal structure and the distribution of pigments in petals were studied in relation to the functional anatomy of petals and the ways in which petals absorb and reflect light. We examined 201 species from 60 angiosperm families. Anthocyanins, betalains and ultraviolet-absorbing flavonoids were normally confined to the epidermal cells, occurring in solution in the vacuole; carotenoids were found in the epidermis and in smaller quantities in the mesophyll, normally in chromoplasts. In a few species, mainly blue-flowered members of the Boraginaceae and Liliaceae-Scilleae, anthocyanins were confined to the mesophyll.
Six basic kinds of petal epidermis anatomy were found, sometimes in combination; papillate (112 species) and multiple-papillate (13 species), in which the conical-papillate form of the cells traps incident light and scatters emergent light, with surface striations aiding these functions in many cases; reversed-papillate (4 species), multiple reversed-papillate (29 species), lenticular (32 species) and flat (11 species), all with surface striations in some cases. Light is usually reflected from petals mainly by an aerenchymatous unpigmented reflective mesophyll; in certain species this is replaced by a reflective layer of starch grains in the upper mesophyll.     

Ultrastructural investigation of the vesicular glands in Scutigera coleoptrata (Chilopoda,Notostigmophora)

In the notostigmophoran centipedes, two pairs of vesicular glands have evolved. These paired glands are situated in the first and second trunk segment and open via cuticular ducts in the upper part of the particular pleura. The vesicular glands of Scutigera coleoptrata were investigated using light and, for the first time, electron microscopical methods. The glands consist of wide sac‐like cavities that often appear vesicular. The epithelia of both glands are identically structured and consist of numerous glandular units. Each of these units consists of four different cells: a single secretory cell, a small intermediary cell, and one proximal and one distal canal cell. The intermediary cell forms a conducting canal and connects the secretory cell with the canal cells. Proximally, the intermediary cell bears microvilli, whereas the distal part is covered with a distinct cuticle. The cuticle is a continuation of the cuticle of the canal cells. This investigation shows that the ultrastructure of glandular units of the vesicular glands is comparable to that of the glandular units of other epidermal glands in Chilopoda and Diplopoda, although the glands look completely different in the light microscope. Thus, it is likely that the vesicular glands and epidermal glands share the same ground pattern. With regard to specific differences in the cuticular lining of the intermediary cells, a common origin of epidermal glands in Myriapoda and Hexapoda is not supported. J. Morphol. 2009. © 2008 Wiley‐Liss, Inc.     

Functional micromorphology of petals of <Emphasis Type="Italic">Chaenomeles japonica</Emphasis> exposed to humid and cold season

In this study, micro- and nano-traits of petal epidermises of flowers of Chaenomeles japonica extended under environmental conditions, during the humid and cold period of the year, are presented. The outer (abaxial) and the inner (adaxial) epidermises of petals of C. japonica consist of convex and papillae cells, respectively, that are covered by epicuticular wrinkled relief further ornamented by submicron motifs, forming interfaces between floral tissues and environment. Structural epidermal features of the petal relief at the nanoscale level reveal different functionality on the two sides of the corolla. The cuticular folds of convex epidermal cells display declining water retention on the outer petal surface and the exposed side of the corolla to the environmental conditions. The cuticular folds of papillae epidermal cells increase in size the inner petal surface, in comparison with the outer surface; such traits facilitate light absorption and enhanced the contact area among folds and curvatures at the inner side of the corolla. It appears that nanometric surface structures of petals may be important adaptive features of C. japonica flowers, contributing to their performance in the field.     

Evaluation of Dermal Substitute in a Novel Co-Transplantation Model with Autologous Epidermal Sheet

The development of more and more new dermal substitutes requires a reliable and effective animal model to evaluate their safety and efficacy. In this study we constructed a novel animal model using co-transplantation of autologous epidermal sheets with dermal substitutes to repair full-thickness skin defects. Autologous epidermal sheets were obtained by digesting the basement membrane (BM) and dermal components from rat split-thickness skins in Dispase II solution (1.2 u/ml) at 4°C for 8, 10 and 12 h. H&E, immunohistochemical and live/dead staining showed that the epidermal sheet preserved an intact epidermis without any BM or dermal components, and a high percentage of viable cells (92.10±4.19%) and P63 positive cells (67.43±4.21%) under an optimized condition. Porcine acellular dermal matrixes were co-transplanted with the autologous epidermal sheets to repair full-thickness skin defects in Sprague-Dawley rats. The epidermal sheets survived and completely re-covered the wounds within 3 weeks. Histological staining showed that the newly formed stratified epidermis attached directly onto the dermal matrix. Inflammatory cell infiltration and vascularization of the dermal matrix were not significantly different from those in the subcutaneous implantation model. Collagen IV and laminin distributed continuously at the epidermis and dermal matrix junction 4 weeks after transplantation. Transmission electron microscopy further confirmed the presence of continuous lamina densa and hemidesmosome structures. This novel animal model can be used not only to observe the biocompatibility of dermal substitutes, but also to evaluate their effects on new epidermis and BM formation. Therefore, it is a simple and reliable model for evaluating the safety and efficacy of dermal substitutes.     

Cuticulogenesis correlated with ultrastructural changes in oenocytes and epidermal cells in the late cockroach embryo

During late embryogenesis in a cockroach, the epidermal cells secrete two cuticles: the embryonic cuticle and the pharate first larval cuticle. Late embryogenesis begins with the deposition of the cuticulin layer of the embryonic cuticle. The embryonic cuticle is an atypical one. It remains relatively thin and a well lamellated endocuticle is usually lacking. After general apolysis of the embryonic cuticle the epidermis secretes the epicuticle of the first larval cuticle and, subsequently, a typical lamellate procuticle. During the penultimate phase of late embryogenesis (i.e. before general apolysis) the epidermis becomes larvally committed. Some epidermal cells start to differentiate into specialized structures of the dermal glands, whereas the differentiated oenocytes appear to have acquired some stability. Nevertheless, shortly before general apolysis some oenocytes display signs of an increased alteration of the SER. When general apolysis occurs, the oenocytes contain a well-developed SER. The whole of the oenocyte population is programmed to regress after epicuticle deposition of the first larval cuticle. The correlation of oenocyte regression with available data on cuticulogenesis, ecdysteroid titres and cuticular lipid synthesis is discussed.