THE STIGMATIC PAPILLAE OF AMYEMA (LORANTHACEAE): DEVELOPMENTAL RESPONSES TO PROTANDRY AND SURFACE ADAPTATIONS FOR BIRD POLLINATION

The flowers of Amyema miquelii and A. miraculosum are protandrous and pollinated by birds. Their dry-type stigmatic surface is composed of unicellular papillae. At the male phase, these papillae are constricted with rugulose surfaces. During the transition to the female (pollen receptive) phase these cells expand, almost doubling in width while their surface becomes much smoother. Beneath the thin proteinaceous pellicle, the papillar wall consists of an extraordinarily thick bi-layered cuticle overlying the primary wall. The two layers of the cuticle are stained by lipid dyes, but are distinguished by their different responses to other cytochemical tests. The reaction product for the enzyme esterase is present within crenulations on the papillar surface in small amounts, and in dense deposits in the cuticular clefts at the base between papillae. Not surprisingly, pollen tubes are unable to penetrate the thick papillar cap and enter the style through these clefts. The unusual thickness of the cuticle is interpreted as an adaptive response to pollination by perching birds (passerines) probing for nectar.     

DEVELOPMENT OF THE STIGMATIC SURFACE OF PRUNUS AVIUM L., SWEET CHERRY

The stigmatic papillae of sweet cherry were examined to determine developmental characteristics of the wet-stigma surface. Early stages of secretion are detectable 1 wk prior to anthesis by using a 1% crystal violet solution. The number of stainable cells and the amount of interstitial staining subsequently increase, although secretions are not visible on unstained specimens until anthesis. Auto-fluorescence above 500 nm (excited by 335–480 nm) becomes microscopically detectable at floral maturity and grows more intense after anther dehiscence. Light microscopy of plastic sections shows that papillae degenerate in peripheral regions of unpollinated mature stigmas, and that this is even more pronounced in pollinated ones. The distal portions of the papillae are covered with a homogeneous cuticular cap, which when viewed with electron microscopy encloses a subcutaneous secretion prior to cuticle exfoliation. Other exudates observed with electron microscopy prior to anthesis are interstitial electron-translucent globules and surrounding matrix, and assorted vesicles, lipid globules, and starch grains which are present at floral maturity. Flowers observed under field conditions in the terminal secretion stage accumulate trichomatous structures. Our observations indicate that the stigma of Prunus avium L. is characterized by several phases of secretion which appear to be facilitated by mechanical abrasion. A model for the primary pollen-receptive area is proposed and suggestions are made concerning the origin of the secretions.     

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     

Comparative Proteomics Studies of Insect Cuticle by Tandem Mass Spectrometry: Application of a Novel Proteomics Approach to the Pea Aphid Cuticular Proteins

The cuticle is a biological composite material consisting principally of N‐acetylglucosamine polymer embedded in cuticular proteins (CPs). CPs have been studied and characterized by mass spectrometry in several cuticular structures and in many arthropods. Such analyses were carried out by protein extraction using SDS followed by electrophoresis, allowing detection and identification of numerous CPs. To build a repertoire of cuticular structures from Bombyx mori, Apis mellifera and Anopheles gambiae the use of SDS and electrophoresis was avoided. Using the combination of hexafluoroisopropanol and of a surfactant compatible with MS, a high number of CPs was identified in An. gambiae wings, legs and antennae, and in the thoracic integument cuticle of Ap. mellifera pupae. The exoskeleton analysis of B. mori larvae allowed to identify 85 CPs from a single larva. Finally, the novel proteomics approach was tested on cuticles left behind after the molt from the fourth instar of Acyrthosiphon pisum. Analysis of these cast cuticles allowed to identify 100 Ac. pisum CPs as authentic cuticle constituents. These correspond to 68% of the total putative CPs previously annotated for this pea aphid. While this paper analyzes only the recovered cuticular proteins, peptides from many other proteins were also detected.     

THE TRANSMITTING TRACT IN GLADIOLUS. I. THE STIGMA AND THE POLLEN-STIGMA INTERACTION

The stigma papillae in Gladiolus are of the “dry” type and are highly vacuolated cells with an organelle-rich peripheral cytoplasm. The cell wall of each papilla is overlain by a distinctive cuticle possessing an irregularly scalloped inner margin. Between the cell wall and cuticle is a layer of amorphous sub-cuticular material. Lipids are detected on the papilla surface. A pollen grain will hydrate and germinate only on a papilla and not on any other (non-papillate) portion of the stigma. The pollen tube penetrates the papilla cuticle, which is forced away from the papilla cell wall by sub-cuticular pollen tube growth. As the cuticle lifts away, the sub-cuticular material disperses. At the base of the papilla, the pollen tube grows onto the adaxial non-papillate surface of the stigma lobe. At this site, the cuticle has been lifted away from the underlying cells by release of a mucilaginous substance from the latter, and the pollen tube grows within this substance beneath the detached cuticle. The cytological features of Gladiolus papillae are compared with other stigma papillae described in the literature. Also, a review of the literature, as well as some of the findings of the present study, suggest that certain prevalent interpretations of dry stigma structure and function may be open to question.     

The Occurrence of a Secondary Cuticle in Libertia elegans (Iridaceae)

A study of the formation of cuticle in Libertia elegans hasshown that the cuticle develops centripetally and is producedcontinuously at its juncture with the underlying pectic layer.Peroxidase activity may be demonstrated in this region duringdevelopment. Growth occurs in two discrete phases; initiallywhilst the underlying epidermal cells are expanding and laterwhen such expansion is complete. The phases are morphologicallydistinct and it is proposed that the terms primary and secondarybe applied to them. Cuticular channels with a pit-like appearancein T.S. are described here for the first time as is the fissuringand active replacement of inelastic secondary cuticular materialto accommodate costal expansion and the growth of papillae.     

Disentangling the assembly mechanisms of ant cuticular bacterial communities of two Amazonian ant species sharing a common arboreal nest

Bacteria living on the cuticle of ants are generally studied for their protective role against pathogens, especially in the clade of fungus‐growing ants. However, little is known regarding the diversity of cuticular bacteria in other ant host species, as well as the mechanisms leading to the composition of these communities. Here, we used 16S rRNA gene amplicon sequencing to study the influence of host species, species interactions and the pool of bacteria from the environment on the assembly of cuticular bacterial communities on two phylogenetically distant Amazonian ant species that frequently nest together inside the roots system of epiphytic plants, Camponotus femoratus and Crematogaster levior. Our results show that (a) the vast majority of the bacterial community on the cuticle is shared with the nest, suggesting that most bacteria on the cuticle are acquired through environmental acquisition, (b) 5.2% and 2.0% of operational taxonomic units (OTUs) are respectively specific to Ca. femoratus and Cr. levior, probably representing their respective core cuticular bacterial community, and (c) 3.6% of OTUs are shared between the two ant species. Additionally, mass spectrometry metabolomics analysis of metabolites on the cuticle of ants, which excludes the detection of cuticular hydrocarbons produced by the host, were conducted to evaluate correlations among bacterial OTUs and m/z ion mass. Although some positive and negative correlations are found, the cuticular chemical composition was weakly species‐specific, suggesting that cuticular bacterial communities are prominently environmentally acquired. Overall, our results suggest the environment is the dominant source of bacteria found on the cuticle of ants.     

The setae of parasitic Liphyra brassolis butterfly larvae form a flexible armour for resisting attack by their ant hosts (Lycaenidae: Lepidoptera)

Caterpillars of the lycaenid butterfly, Liphyra brassolis, live inside the nests of arboreal weaver ants, Oecophylla smaragdina, and eat their brood. Observations of mature larvae suggest that they are impervious to relentless ant molestation, yet they lack sclerotized cuticular plates. We document a novel form of integumental defence that imparts protection from ant attack whilst maintaining the flexibility necessary to walk with a hydraulic skeleton. Analysis of the trunk integument and cuticular structures of early and late instars of L. brassolis using light microscopy, scanning electron microscopy, and histology revealed three new setae types (disc, clavate, and lanceolate), as well as three new cuticular structures (pored sockets, cuticular pores, and cuticular domes). The unique cuticle is covered with lanceolate setae, which act as endocuticular struts, and overlapping scale‐like sockets, which form a hard, flexible integument. The imperfect armour of the early‐instar larvae suggests that abundant, putatively secretory pores are likely to be homologous to pore cupola organs (PCOs) found in other lycaenid larvae and thus may exude semiochemicals to allay ant aggression. The importance of these pores presumably wanes as structural (setal) cuticular defenses are reinforced in later instars, when adult ants have been observed attacking caterpillars to no avail. The caterpillar's antennae are unusual and seem to be involved in manipulating ant larvae into the caterpillar's mouth. Behavioural observations indicate that the dexterity of these structures is associated with eating ants (myrmecophagy).     

The integumental ultrastructure of Lamippe rubra Bruzelius and Enalcyonium rubicundum Olsson (Copepoda,Poecilostomatoida)

The integument of Lamippe rubra Bruzelius and of Enalcyonium rubicundum Olsson has been studied with the electron microscope.Most of the cuticle covering the body of Lamippe is represented by the epicuticle, which shows an average thickness of about 2.0 µm, but in sclerified zones it consists of a thin epicuticle (0.2 µm) and a stratified laminated procuticle (0.5–1.5 µm) without bow-shaped structure. A complex system of epithelial microvilli or a well-developed system of membranes running parallel to the cuticle is also present.The cuticle of Enalcyonium consists of a thin procuticle (0.4–0.5 µm) covered with a uniform fibrillar coat (0.5 µm), whereas in sclerotized areas it is composed of a stratified procuticle (0.7–3.5 µm) with bow-shaped structures.In both species, cuticular hairs and gland vents occur at the dorsal and ventral surfaces. Some of the hairs are considered to be sensory in nature.The cuticular ultrastructure of L. rubra and of E. rubicundum is compared with that of some other copepods.     

一号冰川地区四种藓类植物的解剖学研究

运用石蜡切片技术和扫描电镜方法,对一号冰川地区生长的4种藓类植物茎、叶的内部结构及叶表皮角质层褶皱、疣和中肋等进行观察,结果表明：红扭口藓(Barbula asperifolia Mitt.)茎横切面呈多棱形;叶背、腹面角质层厚,粗疣不分叉,但顶端朝细胞凹陷处倾斜;叶背面细胞壁凹陷深,象张开的气孔,粗疣也藏在其中,中肋突出明显。丛叶扭藓(Tortella humilis (Hedw.) Jenn.)茎横切面呈椭圆形;叶背、腹面中上部均密被鹿角状粗疣,这些分叉的粗疣,从凹陷的细胞壁处成束突起,顶端向下弯曲成钩状,在叶的下部疣状突起则逐渐减少至无,中肋较宽。异叶提灯藓(Mnium heterophyllum (Hook.)Schwagr.)茎横切面呈五棱形;叶细胞一层,呈不规则多边形,细胞壁凹陷使叶片呈网状;中肋红色。大灰藓(Hypnum plumaeforme Wils.)茎的横切面呈圆形;叶背、腹面均密被疣状突起,且差异不大,粗疣顶部均倒向孔口呈遮盖状;中肋短而弱。     

The ultrastructure of the cuticle of Nematoda

 The ultrastructure of the body cuticle in species of six of seven representative genera of Stilbonematinae (Eubostrichus, Catanema, Laxus, Robbea, Leptonemella, and Stilbonema) was investigated using SEM and TEM techniques. Additionally, one species of Spirinia (Spiriniinae) and one of Desmodora (Desmodorinae) were studied for outgroup comparison. The body cuticle of all investigated stilbonematids shows a consistent pattern composed of specific elements in a characteristic arrangement to each other. This pattern does not occur in Stilbonematinae alone, but also in Desmodorinae and Spiriniinae. Furthermore, a comparison within the Desmodorida reveals that this cuticular pattern apparently is present in the cuticle of representatives of Monoposthiidae, Epsilonematidae, and Draconematidae. The present results lead to the following conclusions: (1) the cuticle of Stilbonematinae contains no autapomorphic characters for this taxon, (2) there is a common cuticular pattern within the Desmodorida, and (3) this pattern is an autapomorphic character for the order Desmodorida. Accepted: 4 February 1996     

The structure of the postantennal organ in Onychiurus sp. (Insecta: Collembola) and its connection to the central nervous system

Summary The postantennal organ in Onychiurus (group armatus) is a sensory organ comprising one sensory cell, several enveloping cells and cuticular structures.The perikaryon of the sensory cell is located in the central nervous system and distally gives off a dendrite in which one inner and two outer segments are distinguishable. Two ciliary structures connect the outer dendritic segments with the inner segment. The outer segments divide repeatedly, basal to the cuticular structures, into small branches which end distally beneath the cuticular wall. The wall of the cuticular structures is very thin and is pierced by numerous funnel-shaped pores. The pores are filled with electron-dense material which forms a continuous sheath underneath the cuticle. This material encases the small dendritic branches and the processes of the enveloping cells which occupy the lumen of the cuticular structures. There are three types of enveloping cells: one inner, several outer and one basal. Their processes differ in the manner in which they envelop the various regions of the dendrite.At the beginning of moulting outer dendritic branches are not found within the cuticular structures of the organ. They may be assumed to retract inwardly. However, in the later stages, when the cuticle is fully formed, the outer dendritic segments appear to divide. It is assumed that the small dendritic branches reach their targets before ecdysis. The electrondense material which clogs the intermoult cuticular pores is absent until the final stages of the moulting cycle.Supported by a grant from the Deutscher Akademischer Austauschdienst.     

The structure and distribution of cyanoglucoside-storing cuticular cavities in Pryeria sinica Moore (Lepidoptera,Zygaenidae)*

The dorsal and lateral integument of last instar larvae of Pryeria sinica Moore, 1877 contains two different types of cuticular chambers which are used for the storage of a cyanogenic defensive secretion. Both types of cavities are found to be homologous with those found in the larva of Zygaena trifolii (Esper, 1783), another zygaenid moth with a completely different aposematic pattern. In contrast to the condition in Zygaena trifolíi, the type I cavities in Pryeria sinica bear two different types of opening structures, the smaller one of these being homologous with similar structures found in Zygaena. The aposematic pattern of the Pryeria larva is not contiguous with the cuticular cavities, as it is in Zygaena. The development of the opening structures in the four larval instars of Pryeria is described. It is suggested that the two types of opening structures of cuticular cavities, which form an apomorphic character of the Zygaeninae, have developed independently, the type I mechanisms representing the phylogenetically older system.     

Sensory Spots of Echinoderes capitatus (Zelinka, 1928) (Kinorhyncha,Cyclorhagida)

The sensory spots of Echinoderes capitatus from the Gulf of Trieste were examined by transmission and scanning electron microscopy. Their arrangement is bilaterally symmetrical and is species-specific. At the cuticle surface the sensory spot appears as a rounded to ovoid area of small cuticular papillae in which two pores open. The sensory organ consists of two different sensory cells, the monociliary receptor and the collar receptor, and one sheath cell. The course of the axons and their connections to the nervous system are described. A survey of collar receptors among invertebrates is given. A comparison of the sensory spots within Kinorhyncha and a comparison with the flosculi of Priapulida and the N-flosculi of Loricifera is made. A possible homology of these three structures is discussed.     

Ultrastructural observations of Beauveria bassiana infection in Xylotrechus rusticus larvae

In the present study we investigated the infection process of Beauveria bassiana on Xylotrechus rusticus larvae using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The SEM results showed that B. bassiana spores germinated on the surface of the larval body and invaded the larva as an appressorium. The hyphae then germinated from the spores and spread throughout the larval body. After the death of the larva, conidiophores formed at one end of the hypha on the surface of the larval body and prepared for a new round of infection. The TEM results showed severe damage to the larval cuticle after hyphae infection. The structure of the cuticle became thinner and eventually flocculent; muscle tissues were dissociated and eventually stuck to the hyphae, and the corpus adiposum was loose and deformed, and eventually degraded.     

Clearing pigmented insect cuticle to investigate small insects' organs in situ using confocal laser-scanning microscopy (CLSM)

Various microscopic techniques allow investigating structures from submicron to millimeter range, however, this is only possible if the structures of interest are not covered by pigmented cuticle. Here, we present a protocol that combines clearing of pigmented cuticle while preserving both, hard and soft tissues. The resulting transparent cuticle allows confocal laser-scanning microscopy (CLSM), which yields high-resolution images of e.g. the brain, glands, muscles and fine cuticular structures. Using a fluorescent dye, even single labeled neurons can be visualized and resolved up to an imaging depth of 150 μm through the cleared cuticle. Hydrogen-peroxide, which was used to clear the cuticle, does not preclude immunocytochemical techniques, shown by successful labeling of serotonin-immunoreactive neurons (5HT-ir) in the ants' brain. The ‘transparent insect protocol’ presented here is especially suited for small arthropods where dissection of organs is very demanding and difficult to achieve. Furthermore, the insect organs are preserved in situ thus allowing a more precise three-dimensional reconstruction of the structures of interest compared to, e.g., dissected or sectioned tissue.     

Cuticular anatomy of Sphenobaiera huangii (Ginkgoales) from the Lower Jurassic of Hubei, China

Sphenobaiera huangii (Sze) Hsü is typical Early Mesozoic fossil foliage of Ginkgoales in China. It has been recorded from the Upper Triassic to the Lower Jurassic. The cuticular anatomy is investigated based on material from the type locality, Lower Jurassic Hsiangchi Formation, Zigui County, Hubei Province. The specimens are similar to S. huangii, but contain new information about leaf morphology and cuticular anatomy. Lower and upper cuticle is investigated using light and electron microscopy (LM, SEM, and TEM). Many features are described for the first time, including general structures of lower and upper cuticle, stomata, papillae, and cuticular ultrastructure. At the ultrastructural level, two layers have been distinguished in both lower and upper cuticle, including a homogeneous outer layer with granules and a heterogeneous inner layer with fibrils. Based on a literature comparison between S. huangii and other relevant species of Sphenobaiera, S. huangii may represent the best-known taxon in the genus Sphenobaiera in both leaf morphology and cuticular structures. This study provides the first detailed ultrastructural data on the leaf cuticle of Sphenobaiera, one of the oldest foliage taxa of Ginkgoales, and offers further evidence for potential discussion on the taxonomic relationships of S. huangii with other ginkgoalean taxa.     

Ultrastructure of the nymphal integument in the camel tick Hyalomma (Hyalomma) dromedarii (Ixodoidea: Ixodidae)

The ultrastructure of the nymphal integument in the ixodid tick Hyalomma (Hyalomma) dromedarii is compared for stages of development during and after feeding, and up to the first step of molting, apolysis. The integument comprises a cuticular layer and underlying epidermal cells. The body cuticle, which consists of both sclerotized and non-sclerotized parts, is divided into an outer, thin epicuticle, and an inner, thick, fibrillar procuticle. Pore canals in the procuticle are continuous with wax canals which traverse the epicuticle. As feeding progresses, the parallel, extensible epicuticular folds disappear due to the gut filling with ingested blood. The procuticular zone, however, becomes subdivided into an exocuticle, similar to the previously seen procuticle, and a lamellate endocuticle. Pore canals lose their parallel pattern and appear to have become deformed by stretching of the cuticle. The flat epidermal cells grow asynchronously during feeding; their cytoplasm becomes packed with well-developed rough endoplasmic reticulum (RER), while the cell apices project long microvilli extending deep into the procuticle. The RER undergoes ultrastructural changes indicating synthetic activity. Dense material released through the microvilli may serve to lyse the endocuticle and thus cause separation of the cuticle from the epidermis during apolysis. The lysed area, the exuvial cavity, is filled with lysed components which are probably withdrawn by endocytosis into the apical coated vesicles which appear in epidermal cells. Two types of integumental glands, which may participate in wax production, are observed in this study. The ultrastructure of their previously undescribed cuticular ducts is described, in addition to other hypodermal structures including epidermis-muscle attachments and sensory receptors.     

La formation des canaux cuticulaires chez l'adulte de Tenebrio molitor L. étude ultrastructurale et remarques histochimiques

The sclerotized cuticle of adult Tenebrio shows (1) an exocuticle composed of rotating lamellate layers and of columns of cuticular material, the fibres of which run perpendicularly through the lamellae, (2) an endocuticle composed of layers with preferred orientation. In the exocuticle, the pore canals are numerous and run along the columns; they do not rotate with the lamellate layers. They show several filaments some of which leave the canals and form a dense intracuticular network. In the last layers of exocuticle, the pericolumnar canals fuse and form large endocuticular canals which rotate in phase with the cuticular fibres. The formation of columns and canals is in relation with cellular expansions which penetrate into the cuticle during cuticle deposition. Exocuticular columns seem characteristic of highly sclerotized cuticles and the intracuticular filaments may have a role in the transport of sclerotisation precursors.     

STRUCTURE OF THE PEAR LEAF CUTICLE WITH SPECIAL REFERENCE TO CUTICULAR PENETRATION

Study of the pear leaf cuticle (Pyrus communis L. ‘Bartlett‘), in both intact and enzymatically isolated forms, has revealed that the cuticular membrane is separated from the underlying epidermal cell wall by a layer of pectic substances which extend into but not through the membrane. A layer of embedded birefringent waxes occurs towards the outer surface of the cuticular membrane. Platelet-like epicuticular waxes are deposited on the outer surface. The upper cuticular membrane is astomatous. The lower epidermis is stomatous, and the outer cuticular membrane is continuous with that lining the substomatal cavity. The lower cuticular membrane is also generally thicker than the upper, and both the upper and lower cuticular membranes are thicker over veinal than over mesophyll tissue. The birefringence frequently is discontinuous over anticlinal walls and over veinal tissue. The lower cuticle appears to contain fewer embedded waxes (as indexed by birefringence) than the upper. Enzymatic isolation of the cuticular membrane from the underlying tissues does not appear to cause any discernible change in structure as viewed with a light microscope. These findings are discussed in light of current knowledge concerning penetration of foliar applied substances into the leaf.