Die Feinstruktur des Integumentes und der Muskelansatzstellen vonEchiniscoides sigismundi (Heterotardigrada)

The structure of the integument and the muscle attachments of the marine heterotardigradeE. sigismundi (M. Schultze) was studied by electron microscopy. The cuticle consists of several layers: an outer tripartite (or multilayered) epicuticle, perhaps with an outermost coat; a homogeneous inner epicuticle; a trilaminated layer; an intracuticle; and a fibrous procuticle. These features resemble the cuticle described in Eutardigrada; in contrast, areas on the legs and near the claws, with an outer multilayered epicuticle and a striated layer (inner epicuticle), are — as far as investigated — more similar to the cuticle in Heterotardigrada. The epidermis consists of a single cell layer without glands. The muscle attachments are in line with the general pattern described in the eutardigradeMacrobiotus hufelandi and in Arthropoda.     

Structure of the cuticle of the alpine weta,Hemideina maori (Saussure) (Orthoptera: Stenopelmatidae)

Sclerotized cuticle segments from the thorax, dorsal abdomen, and ventral abdomen of the alpine, weta Hemideina maori (Saussure) (Orthoptera: Stenopelmatidae) were examined by light microscopy and by scanning and transmission electron microscopy. An epicuticle, exocuticle (outer and inner), mesocuticle, endocuticle, and deposition layer are present in transverse sections. The epicuticle is further composed of a cuticulin layer and inner epicuticle, the latter being finely laminated and containing narrow wax canals that terminate below the cuticle surface. Openings to dermal gland ducts are visible on the surface as are large setae and smaller sensory pegs. Frozen fractured cuticle reveals the presence of horizontal ducts or channels that run laterally within the cuticle. The structure of weta cuticle is compared with that of the common house cricket and arthropods in general.     

Structure of the cuticle of the common house cricket with reference to the location of lipids

Cuticle segments from the thorax, abdomen, and jumping legs of the house cricket. Acheta domesticus, were examined using histological techniques for light microscopy, scanning and transmission electron microscopy, and direct examination of frozen-fractured cuticle. The surface of untreated cuticle is covered by a lipid film which obscures fine surface detail. Standard EM preparative procedures, as well as washing the cuticle with ethanol before examination, remove this film exposing previously covered openings to dermal gland ducts and wax canals. An epicuticle, exocuticle, mesocuticle, endocuticle, and a deposition layer were present in all transverse sections of cuticle. Light microscopy showed that the exocuticle and mesocuticle are heavily impregnated with lipids, whereas there is little lipid associated with the endocuticle. Frozen-fractured cuticle clearly shows the ‘plywood’ structure of the meso- and endocuticle, while the exocuticle fractures as if it were a solid sheet. The epicuticle is composed of a dense homogeneous layer, cuticulin, outer epicuticle, and the outer membrane. Superficial wax was detected only in cuticle samples prepared using vinylcyclohexane dioxide as a polar dehydrant. The results were used to construct a comprehensive model of the cuticle of A. domesticus.     

Cuticle structure and systematics of the Macrobiotidae (Tardigrada, Eutardigrada)

The cuticle of tardigrades is characterized by three main layers: epicuticle, intracuticle and procuticle. Pillars are present in the epicuticle of almost all heterotardigrades, but these structures are also known in a few species of eutardigrades. The apparent heterogeneity of the cuticular ultrastructure in several species of the Macrobiotidae (Eutardigrada) prompted us to analyse the structure of the cuticle in this family. Eleven species in several genera were investigated with light and/or electron microscopy. All the species of the genera Murrayon and Dactylobiotus showed pillars in the epicuticle, whereas the examined species of Macrobiotus , Richtersius and Xerobiotus completely lacked pillars. Therefore, in the Macrobiotidae, in contrast to what appears with light microscopy, the cuticle is homogeneous within each genus examined at the electron microscopic level. Considering the absence of pillars in the Macrobiotidae a synapomorphy, we propose the erection of two new subfamilies. Macrobiotinae subfam. n. is characterized by the absence of pillars in the epicuticular layer and includes, in addition to the genera Macrobiotus, Xerobiotus and Richtersius, the genera Pseudohexapodibius, Adorybiotus, and probably also Minibiotus, Calcarobiotus and Pseudodiphascon . Murrayinae subfam. n. is characterized by the presence of pillars in the epicuticular layer and includes the genera Murrayon, Dactylobiotus and, probably, Macroversum.     

The integument in troglobitic and epigean woodlice (Isopoda: Oniscidea): a comparative ultrastructural study

We compared the ultrastructure and the relative thickness of the integumental cuticle in several species of troglobitic and non-troglobitic woodlice. Measurements of tergal cuticle thickness on histological sections demonstrated that the cuticles in non-troglobites are thicker than those in troglobites of similar body sizes. As revealed by scanning electron microscopy, the endocuticles in troglobites consist of more numerous and thinner lamellae compared to cuticles of similar thickness in non-troglobites. Similar differences in the number and thickness of cuticular lamellae were not found in the exocuticle. As demonstrated by transmission electron microscopy of the epicuticles in troglobitic and non-troglobitic woodlice, the simple inner epicuticle is thinner relative to the total epicuticle thickness in troglobites, but this is not the case for the outer epicuticle. Outer epicuticles consisting of different numbers of sublayers can be found in troglobites as well as in non-troglobites and more complex outer epicuticles are not characteristic of representatives of any of the two ecological groups. Our results indicate that the thickness and structure of the integumental cuticle are important for evolutionary success in the subterranean environment. Nevertheless, the cuticles of troglobites are diverse in their ultrastructural features, likely reflecting different lifestyles of various troglobites.     

Design of the labial cuticle in Cenocorixa bifida Hung. (Hemiptera: Corixidae) with reference to ionic transport

The surface topography and ultrastructure of the labial cuticle of Cenocorixa bifida were examined by scanning and transmission electron microscopy. The dorsal wall of the labium consists of seven sclerotized transverse bars each displaying two rows of semicircular grooves and pores. The cuticle is about 20 microm thick and is composed of epicuticle and lamellate exocuticle and endocuticle, the latter separated from the underlying epidermis by subcuticle containing amorphous material. The epicuticle is subdivided into an electron-dense very thin outer epicuticle and a homogenous thick inner epicuticle, which is penetrated by grooves. The exocuticle is filled with electron-dense blocks of material, which may provide mechanical support to the labial wall. The elongate epidermal cells display extensive infoldings of the apical plasma membrane (facing the cuticle) and contain abundant mitochondria in the cytoplasm. The presence of deep epicuticular grooves and pores in the thin labial cuticle and extensive apical membrane infolding and abundant mitochondria in the epidermal cells suggest that the labium in C. bifida is the site of osmoregulatory ionic uptake.     

An ultrastructural study of the cuticle in the marine annelid Heterodrilus (Tubificidae, Clitellata)

The ultrastructure of the cuticle in four species of the marine Heterodrilus (H. paucifascis, H. pentcheffi, H. flexuosus, H. minisetosus) is investigated with transmission electron microscopy. The noncellular cuticle consists of several parts; closest to the epidermis is a thick zone of collagen fibers embedded in a matrix. The matrix continues outside the fiber zone, forming a layered epicuticle. The external surface of the epicuticle is covered by evenly distributed, membrane-bound bodies, termed epicuticular projections. The epicuticular projections have their longitudinal axis perpendicular to the surface of the cuticle and are attached to the surface by either the surrounding membrane itself or by short pedestals. Microvilli, extensions from the epidermal cells, penetrate and sometimes pass completely through the cuticle. There is interspecific variation in the morphology of the cuticle. The four studied species differ in the arrangement of the collagen fibers, from irregularly distributed fibril bundles to orthogonally arranged fiber layers, as well as in the number and density of layers in the epicuticle. One of the studied species, H. paucifascis, shows intraspecific variation, which is associated with sample locality. The Bahamian specimens of H. paucifascis have four layers in the epicuticle, club-shaped epicuticular projections, and collagen fibers forming a less defined orthogonal grid, while the Belizean specimens have three layers in the epicuticle, epicuticular projections with a bulging part at midlevel, and a distinct orthogonal grid. Based on these findings the variation in the morphology of the cuticle appears to be dependent on both phylogenetic constraints, and functional and environmental factors.     

Ultrastructure and cuticle formation of the carapace in the myodocopan ostracod exemplified by Euphilomedes japonica (Crustacea: Ostracoda)

The ultrastructure and formation of the cuticle of a myodocopan ostracod, Euphilomedes japonica, are investigated utilizing scanning and transmission electron microscopy. The outer lamella cuticle consists of four layers; epicuticle, exocuticle, endocuticle, and membranous layer like in the cuticle of other arthropods. The exocuticle and endocuticle are well-calcified and the organic matrix develops within the both cuticles. The outermost layer of new cuticle (epicuticle) is secreted first and the inner layers (exocuticle, endocuticle and membranous layer) are added proximally in the pre-, and postmoult stages. The calcification takes place in the whole area of carapace at the same time together with the synthesis of organic matrix within the endocuticle. This study demonstrates that the ultrastructure and formation of the cuticle in myodocopans are different from those in podocopans, and that the myodocopan carapaces have achieved a structural diversity for adaptation to different lifestyles.     

Fine structure of the epicuticle of the desert scorpion, Hadrurus arizonensis, with reference to location of lipids.

The surface and transverse sections of the epicuticle of the desert scorpion, Hadrurus arizonensis, were examined by scanning and transmission electron microscopy, respectively. Sclerite cuticle that was untreated prior to normal EM preparative procedures was compared to cuticle subjected to lipid solvents, high temperature, and concentrated alkali. Surface morphology of untreated intersegmental cuticle was also examined. The epicuticle is composed of four sublayers: outer membrane, outer epicuticle, cuticulin, and the dense homogeneous layer. Lipid solvents did not significantly alter the morphology of any of these layers or the contents of the wax canals that penetrate the cuticulin layer even though the solvents effectively remove lipids from the epicuticle for chemical analysis. The surface of the sclerite cuticle contains amorphous particles, crystalline projections, and scattered openings to dermal gland ducts. Perforations that correspond to the opening of wax canals were faintly visible after extraction of surface waxes and clearly visible after KOH treatment. No openings to dermal gland ducts or wax canals were observed in untreated intersegmental cuticle. However, wax canals are likely obscured by surface waxes similar to those present in sclerite cuticle.     

Ultrastructure, development, and homology of insect embryonic cuticles

Ultrastructure and deposition of the cuticles secreted by embryos representing eight insect orders were examined by transmission and scanning electron microscopy. Embryos of the apterygote silverfish Thermobia domestica deposit two embryonic cuticles. Deposition of the first (EC1) is initiated at the beginning of appendage development when the intercalary segment and the neural groove are clearly visible. This cuticle lacks surface microsculpture and consists of an outer epicuticle and an underlying fibrous layer, thought to represent procuticle. At the time of dorsal closure, deposition of a second embryonic cuticle (EC2) begins; this bears sensilla and functions in the first instar larva. In representative embryos of seven pterygote orders (Ephemeroptera, Odonata, Plecoptera, Neuroptera, Coleoptera, Lepidoptera, and Mecoptera), three cuticles were found to be secreted. The first cuticle in pterygotes is homologous to EC1 of T. domestica, but consists solely of outer epicuticle. EC2, the "prolarval cuticle," bears a characteristic surface microsculpture in embryos of some species and egg-teeth and other hatching devices, and consists of outer and inner epicuticles and a more or less reduced procuticle. EC2 is reduced in the embryos of derived endopterygotes, where a procuticle is lacking and the inner epicuticle is reduced. After hatching, when EC2 is shed, the first instar larva is covered by a third embryonic cuticle (EC3), whose deposition was initiated while the insect was still within the egg. Presence of only two embryonic cuticles in cyclorrhaphous flies is due to the total loss of prolarval cuticle. Investigated exopterygote and endopterygote insects excluding flies thus deposit three embryonic cuticles, and their juveniles (exopterygote "nymphs"; endopterygote "larvae") seem to hatch at equivalent stages of development. Differences between the modes of cuticulogenesis in silverfish and pterygote embryos suggest that the apterygote first larval instar was embryonized and became a fully embryonic prolarva in pterygotes.     

Fine structure of the cuticle of the desert scorpion, Hadrurus arizonensis.

The structure of the sclerite and intersegmental cuticle of the opithosoma of the desert scorpion, Hadrurus arizonensis, has been examined by transmission electron microscopy. The sclerite cuticle contains a four-layered epicuticle, a hyaline exocuticle, an inner exocuticle and an endocuticle. The outer part of the hyaline exocuticle and the whole of the inner exocuticle are constructed of helicoidally arranged planes of microfibrils. Within the endocuticle, the overall architecture is not helicoidal as previously assumed, but consists of bundles of microfibrils oriented horizontally and vertically. Microbibrils of the inner exocuticle and the endocutile are seen as simple unstained rods, but those of the hyaline exocuticle are electron dense rods with an unstained central core. The intersegmental cuticle contains a four-layered epicuticle and a procuticle. In detail, its fine structure differs in most respects from that of the sclerite cuticle. Electron microscopy reveals that hyaline exocuticle, previously assumed to be continuous from sclerite to intersegmental membrane, is absent in the latter.     

Comparative study of the cuticle in some aquatic oligochaetes (Annelida: Clitellata)

An examination of the cuticle of six aquatic oligochaete species using transmission electron microscopy revealed a larger morphological variation than previously known. Three freshwater species, Aulodrilus pluriseta, Spirosperma ferox (both Tubificidae), and Pristina breviseta (Naididae), and three marine species, Clitellio arenarius, Heterochaeta costata (both Tubificidae), and Paranais litoralis (Naididae), were investigated. The arrangement of the collagen fibers in the cuticle differs among the studied species. Only S. ferox shows an "orthogonal grid," i.e., layers of parallel fibers perpendicular to each other, as earlier described for lumbricids and enchytraeids. Clitellio arenarius and H. costata have fibers arranged in layers, while A. pluriseta and P. litoralis have irregularly distributed fibers. Pristina breviseta lacks cuticular fibers. The matrix surrounding the collagen fibers (when present) continues outside the fiber layer, making up a thin epicuticle, which has a unique banding in each of the studied species. The external surface of the epicuticle is covered with epicuticular projections. Their number, shape, and attachment to the epicuticle vary among the studied species. Furthermore, a distinctive internal substructure of the projections was observed in H. costata, A. pluriseta, S. ferox, and P. breviseta. Microvilli, extensions from the epidermal cells, penetrate the cuticle and terminate at its outer surface. In three species microvilli were observed to pinch off the epicuticular projections. The size, number, and shape of the latter vary; no typical microvilli were observed in S. ferox.     

Ultrastructure of cuticular exudations in parasitic juvenileHeterodera schachtii,as related to cuticle structure

Summary The development ofHeterodera schachtii inside roots of a cruciferous host plant grown under monoxenic conditions in an agar medium was observed with video-enhanced contrast light microscopy. One to 6 days after inoculation, roots were excised and processed for electron microscopic observations. Exudates were present on the cuticle surfaces of J 2 and early J 3 juveniles located at feeding sites. Fibrillar exudations were correlated with similar fibrillar patterns in the epicuticle, exocuticle, intermediate zone, and the striated endocuticle. Secretion vesicles assembled at many Golgi sites in the hypodermis, appeared to coalesce and form large electron translucent vesicles in the cytoplasm. We propose that secretion vesicles migrate toward the cuticle, contact the plasmalemma and transfer their contents by exocytosis or a similar mechanism to a secretion accumulation site. These contents are associated with cuticle structure and emerge as surface exudations.     

Fine structure of the cuticle of the black widow spider with reference to surface lipids

The surface and transverse sections of the cephalothorax, abdomen, and walking leg cuticle of the black widow spider, Latrodectus hesperus, were examined by scanning and transmission electron microscopy. Cuticle that was untreated prior to normal EM preparative procedures was compared with cuticle subjected to lipid solvents and/or concentrated alkali. The surface of untreated dorsal cephalothorax cuticle contained droplets and a lipid film that obscured fine surface detail. Immersing the cuticle in chloroform: methanol removed the droplets and lipid film, exposing previously covered openings to dermal gland ducts. An epicuticle, exocuticle, and endocuticle were present in all transverse sections of cuticle as was a complex system of pore and wax canals that connected the epidermis with the cuticle surface. The epicuticle of the walking leg was composed of three sublayers: outer membrane, outer epicuticle, and the dense homogeneous layer. A cuticulin layer was not observed. Lipid solvents did not significantly alter the morphology of any of these layers or the contents of the wax/pore canals.     

Early pattern of calcification in the dorsal carapace of the blue crab, Callinectes sapidus

The pattern of calcium carbonate deposition was observed in the dorsal carapace of premolt (D2-D3) and early postmolt (0-48 h) blue crabs, Callinectes sapidus, using scanning (SEM) and transmission (TEM) electron microscopy. Samples of dorsal carapace for SEM were quick-frozen in liquid nitrogen, subsequently lyophilized, and viewed using secondary and backscattered electrons as well as X-ray maps of calcium. Pieces of lyophilized cuticle were also embedded in epoxy resin and subsequently sectioned and viewed with TEM and SEM. Fresh pieces of dorsal carapace for TEM were also fixed in 2.5% glutaraldehyde in phosphate buffer followed by postfixation in 1% OsO4 in cacodylate buffer. Calcium concentrations were determined using atomic absorption spectrophotometry and quantitative X-ray microanalysis. Calcium accumulation began in the cuticle at 3 h postmolt at the epicuticle/exocuticle boundary and at the distal and proximal margins of the interprismatic septa (IPS). The bidirectional calcification of the IPS continued until the two fronts met at 5-8 h postmolt. The roughly hexagonal walls of the IPS formed a honeycomb-like structure that resulted in a rigid cuticle. The walls of the canal containing sensory neurons also calcified at 3 h, thereby imparting rigidity to the structure and additional strength to the cuticle. Examination of thin sections of lyophilized cuticle and fixed cuticle revealed that the first mineral deposited is more soluble than calcite and is probably amorphous calcium carbonate. The amorphous calcium carbonate is transformed to calcite along a front that follows the original deposition and is probably controlled by a specialized matrix within the IPS. Since amorphous calcium carbonate is isotropic, it would also make the mineral in the exocuticle stronger by an equal distribution of mechanical stress.     

Cuticular cycle and molting hormone levels during the metamorphosis of Tenebrio molitor (Insecta Coleoptera)

The cuticular cycle of Tenebrio molitor (apolysis, synthesis of outer and inner epicuticle, fibrous cuticle deposition) was studied during the last larval and pupal stages by electron microscopy. Concurrently, molting hormone (MH) titers in the hemolymph were determined by a radioimmunoassay method. It appears, both in larvae and in pupae, that the MH peak coincides with the initiation of pre-ecdysial cuticle deposition (i.e., outer epicuticle synthesis). Thus MH is involved in the induction of cuticular synthesis; however, its role in inducing larval-pupal apolysis is questionable. We note that this peculiar apolysis occurs long before MH release.     

Ultrastructural organization of hypodermis and formation of cuticle in pharate larva of Leptotrombidium orientale (Acariformes: Trombiculidae)

The ultrastructural organization of hypodermis and the process of cuticle deposition is described for the pharate larvae of a trombiculid mite, Leptotrombidium orientale, being under the egg-shell and prelarval covering. The thin single-layered hypodermis consists of flattened epithelial cells containing oval or stretched nuclei and smooth basal plasma membrane. The apical membrane forms short scarce microvilli participating in the cuticle deposition. First of all, upper layers of the epicuticle, such as cuticulin lamella, wax and cement layers, are formed above the microvilli with plasma membrane plaques. Cuticulin layer is seen smooth at the early steps of this process. Very soon, however, epicuticle starts to be curved and forms particular high and tightly packed ridges, whereas the surface of hypodermal cells remains flat. Then a thick layer of the protein epicuticle is deposited due to secretory activity of hypodermal cells. Nearly simultaneously the thick lamellar procuticle starts to form through the deposition of their microfibrils at the tips of microvilli of the apical plasma membrane. Procuticle, as such, remains flat, is situated beneath the epicuticular ridges and contains curved pore canals. Cup-like pores in the epicuticle provide augmentation of the protein epicuticle mass due to secretion of particular substances by cells and to their transportation through the pore canals towards these epicuticular pores. The very beginning of the larval cuticle formation apparently indicates the starting point of the larval stage in ontogenesis, even though it remains for some time enveloped by the prelarval covering or sometimes by the egg-shell. When all the processes of formation are over, hungry larvae with a fully formed cuticle are actively hatched from two splitted halves of prelarval covering.     

Cuticle secretion during larval growth in Drosophila melanogaster

The moulting cycle and growth of the larval integument of Drosophila melanogaster has been studied by light and electron microscopy. Growth during the first, second and third larval instars is accompanied by 3.0-, 3.4- and 3.7-fold increases in surface area, respectively. Growth in surface area occurs continuously during the larval stages, with no detectable relationship to the moulting cycle. Measurements of the thickness of the cuticular layers show that the endocuticle grows in thickness by apposition and in surface area by stretching. The pre-apolytic epicuticle remains at fairly constant thickness during the increase in surface area, indicating that it grows by intussusception of new components. Post-apolytic epicuticle becomes thinner and increases in surface area by stretching. The epicuticle and pre-ecdysial endocuticle are traversed by filaments, but these do not penetrate the endocuticle secreted after ecdysis. We suggest that the filaments transport breakdown products from the old cuticle inward to the epidermis for reutilization. The growth and deposition of cuticle in two larval growth mutants, lethal (2) giant larvae and Chubby Tubby, involves mechanisms similar to those found in wild-type larvae, but in Chubby Tubby the endocuticle contains inclusions which are ultrastructurally similar to dense epicuticle.     

The extracellular matrix of the cuticle of Gordius panigettensis (Gordioiidae,Nematomorpha): observations by TEM,SEM and AFM

The cuticle of Gordius panigettensis (Sciacchitano, 1955) was studied by scanning electron microscopy (SEM), transmission electron microscopy (TEM) and atomic force microscopy (AFM). The cuticle is composed of 30-50 compact layers. The number of the layers is higher in the central part of the animal's body and decreases at the extremities. Each layer is composed of parallel tightly packed fibres approximately 640 nm in diameter and of indefinite length. The fibres run strictly parallel within each layer, while in adjoining layers they run at a variable angle from 45 degrees in the central body to 90 degrees in the extremities. Each fibre shows a barely detectable filamentous inner structure and is enveloped in a thin highly regular net formed by hexagonal meshes. Our results suggested that these fibres should be proteinaceous although non-collagenous. Thinner radial fibres run among the large fibres and across all the layers and span the whole thickness of the cuticle from the epithelial layer located deep underneath the large fibres up to the epicuticle on the external surface of the animal.