Egg-shells in mites

Summary This communication presents results of studies on the formation and structure of the vitelline envelopes in three species of mites: Euryparasitus emarginatus (Gamasida), Erythraeus phalangoides (Actinedida), and Hafenrefferia gilvipes (Oribatida). In E. emarginatus and E. phalangoides, in which the oocytes are not covered with follicular cells, the material of the vitelline envelope appears first in vesicles under the surface of the oocytes prior to secretion by exocytosis. The formed vitelline envelope is built of a homogeneous material which is perforated by numerous channels containing oocyte microvilli. Later, as the microvilli are retracted, the channels disappear. In both of these species the formed vitelline envelope is incomplete and the micropylar orifice occurs as a transitional structure.In H. gilvipes follicular cells encircling the oocyte contain granules filled with material that is subsequently secreted into the perivitelline space forming the vitelline envelope on the oocyte surface. The inner layer of the vitelline envelope is granular, whereas the outer part is more homogeneous. Both lack channels containing microvilli and micropyle.     

Gametangial interaction and oogenesis in the phycomycete Ciliomyces spectabilis (Fungi,Lagenidiales)

Gametangial interaction and oospore formation were studied in Ciliomyces spectabilis, a Lagenidiaceous fungus which is parasitic on ciliate cysts. Electron dense and granular vesicles of the antheridium are engaged in formation of the copulation porus between adjacent thalli. The oosphere is delimited by Golgi-derived cisternae which give rise to the membranes of the oosphere and the periplasm. The contents of the antheridium and the periplasm degenerate. The outer oospore wall is formed by wall vesicles originating from the endoplasmic reticulum. No vesicles are involved in the development of the thick inner oospore wall. Vacuoles with electron dense spherical contents fuse and form the central reserve globule. Lipid bodies aggregate first and disintegrate later into numerous small ones. The number of cytoplasmic organelles decreases. The possibility of wall formation via secretion of soluble wall material is discussed.     

Ultrastructural immunocytochemical localization of vitellogenin in the fat body of the blowfly,Calliphora vicina Rob.-Desv. (erythrocephala Meig.) by use of the unlabeled antibody-enzyme method

Summary The unlabeled antibody-enzyme method was used to demonstrate ultrastructurally the specific localization of vitellogenin in the fat body of Calliphora. In control flies the binding sites to vitellogenin were localized in secretory granules situated in the Golgi complex, and in larger bodies named composite secretory granules. These composite granules appear to be formed when a part of a Golgi complex containing secretory granules and a number of small vesicles become surrounded by a common membrane. Ovariectomized flies, which apparently do not produce secretory granules, exhibited no immunocytochemical staining. Ovariectomized flies in which the administration of ecdysterone induced formation of secretory granules, also revealed specific staining on these granules. This is the first ultrastructural evidence of: (a) the specific localization of vitellogenin in secretory granules of the fat body of an insect; (b) the relationship between the presence of the ovary, and of ecdysterone, and the synthesis of vitellogenin by the fat body.     

Effect of cytochalasin A on actin distribution in the fission yeastSchizosaccharomyces pombe studied by fluorescent and electron microscopy

Summary Actin distribution and ultrastructure of the fission yeastSchizosaccharomyces pombe treated with cytochalasin A (CA) were investigated by fluorescence microscopy using rhodamine-conjugated phalloidin (rh-ph) and freeze substitution electron microscopy. Among the cytochalasins tested, CA was most effective and at 5 g/ ml inhibited the appearance of the actin ring at the cell equator at the stage prior to septum formation and the accumulation of actin dots at the septum-forming site both in wild-type cells and the mutantcdc 11, which is defective in septum formation at restrictive temperature. Freeze substitution electron microscopy of CA-treated cells revealed the displacement and morphological alteration of cytoplasmic vesicles and dictyosomes within 30 min and the appearance of dense bodies in the cytoplasm. A sub-population of cytoplasmic vesicles and dictyosomes were insensitive to CA and maintained their original structure. An electron less dense layer containing filamentous material was noted beneath the plasma membrane and thought to be the area of heavy actin patches stained with rh-ph at the cells ends. These results suggest that CA disrupted an actin network that normally maintains the organization of the secretory pathway involving dictyosomes and vesicles.     

Tormogen cell and receptor-lymph space in insect olfactory sensilla

Summary (1) The basiconic sensilla on the antennae of Calliphora resemble other insect epidermal sensilla; one or several bipolar sense cells are surrounded by three non-neural cells. (2) The apical cell membrane of the tormogen cell(one of the three accessory cells) forms microvilli coated internally with particles. (3) In the (extracellular) outer receptor-lymph space hyaluronic acid can be demonstrated histochemically. (4) Demonstration of non-specific alkaline phosphatase, Mg²⁺-activated ATPase, and the presence of mitochondria in the apical part of the tormogen cell suggest active transport processes through these cells into the outer receptor-lymph space.Supported by the Deutsche Forschungsgemeinschaft     

A critical evaluation of the relationship between the presynaptic network,synaptic vesicles and dense projections in central synapses

Summary Synapses of the oculomotor nucleus of Echidna have been examined ultrastructurally with the aim of integrating data obtained from osmicated and nonosmicated PTA stained material. Particular emphasis has been laid on the relationship between the synaptic vesicles of the osmicated material and the presynaptic network and vesicular grid of the PTA material. This relationship has been explored qualitatively by examining osmicated material of varying qualities of fixation. Such material contains dense projections in addition to synaptic vesicles, and various vesicular network appearances. A variety of measurement techniques have shown that the PTA network is characterised by reticular strands, spaces, and regular hexagonal units smaller than vesicles, these observations prompting the formulation of a vesicle-network coincidence model of the presynaptic terminal. This model has been tested by tracing the profiles of vesicles within the PTA network and comparing their size and shape frequency distributions with those of osmicated synaptic vesicles. The distributions have been found to be essentially similar, suggesting that vesicles can be located within the network, and that the hexagonal network units are formed only in the presence of an underlying vesicular matrix.Additionally, the following points have emerged: 1) the dense projections in the two types of material appear to be equivalent; 2) a loose correlation exists between dense projections and vesicles in osmicated terminals, increase in the area of the dense projections being associated with a decrease in the area of the vesicles; 3) network and dense projection units are similar. In view of the similarity between network and dense projection units, the demonstrated vesicular basis of the network raises the question of whether dense projections are entirely independent structures, or whether they depend in part for their existence on the nearby presence of synaptic vesicles.This work was supported by the Arnold Yeldham and Mary Raine Research Foundation of the University of Western Australia and by the Australian Research Grants Committee and the Nuffield Foundation     

Changes in the activity of the Golgi apparatus during the cell cycle in antheridial filaments ofChara vulgaris L.

Summary The number of dictyosomes found in one central cell section in antheridial filaments ofChara vulgaris increases proportionally to the cell length during interphase. The activity of Golgi apparatus was expressed by a number of Golgi vesicles surrounding a single dictyosome. These vesicles are most numerous during mitosis and cytokinesis,i.e., prior to and during cell plate formation. In the middle and late S phase the number of Golgi vesicles decreases by about 25%; subsequently, during the early and middle G₂, it increases again. At the end of the G₂ phase, Golgi vesicles are the scarcest.The increase in the number of Golgi vesicles during the G₂ phase coincides with the period of intense cellular elongation, and, thus, it is probably related to the enhanced synthesis of cell wall components.Coated vesicles are most numerous in prophase, metaphase, and early telophase, and during interphase in both late S and G₂ phase. It was found that the number of coated vesicles is proportional to the degree of condensation of nuclear chromatin.This work was supported by the Polish Academy of Sciences within the project 09.7.3.1.4.     

Ultrastructure of the epidermis of adult and embryonic Paravortex species (Turbellaria,Eulecithophora)

The epidermis and associated structures of adult and embryonic Paravortex cardii and Paravortex karlingi, internal parasites of Cerastoderma edule, have been examined using scanning and transmission electron microscopy. The cellular epidermis of adult Paravortex bears cilia and microvilli which differ in number and distribution between P. karlingi and P. cardii. Cellular organelles include mitochondria, lipid bodies, Golgi bodies, and ultrarhabdites. Epidermal nuclei are located in the proximal portion of the cells. The development of the tegument of embryo Paravortex has been described and a possible origin for the embryo capsule is suggested. These findings are discussed in relation to the phylogenetic status of the Turbellaria in relation to other Platyhelminthes and in the functional adaptation of the epidermis for a parasitic mode of life.Abbreviations bb- basal bodies - bl- basal lamella - c- cilia - cp- capsule - dc- dark cells - e- embryos - ep- epidermis - g- Golgi bodies - int- interdigitation (of cells) - l- lipid - lf- lamellar fold - mc- migrating cell - mf- membranous folds - mt- mitochondria - mv- microvilli - n- nucleus - nb- neoblasts - p- projections of epidermis - par- parenchyma of mother - pr- primary rootlet - rc- rhabditogen cells - sr- secondary rootlet - ur- ultrarhabdites - vt- vitelline material     

Ultrastructure of the tegumental microvilli (microtriches) of Hymenolepis diminuta

Summary The ultrastructure of microtriches of the rat tapeworm, Hymenolepis diminuta, was examined with a number of electron-microscopic techniques. Fixatives containing different buffers, non-ionic detergents, chelators, tannic acid and various concentrations of aldehydes were tested for ability to stabilize cytoskeletal components while extracting background material. These methods revealed features unique to these specialized microvilli, and permitted construction of a detailed model of microthrix architecture. The microtriches of H. diminuta are comprised of a microfilament-containing base, a dense cap and a complex junctional region between the base and cap. The microfilaments of the base are contiguous distally with a tubular structure (the junctional tubule) within the junctional region; proximally, the microfilaments end abruptly: a terminal web appears to be absent. A beveled bilayered cylinder of dense material (the core tunic) encircles the microfilamentous core. The core tunics and junctional tubules of the microtriches are specifically and uniformly aligned along the strobila. Microtriches therefore can be distinguished from other microvilli (e.g., those of enterocyte brush borders) by their complex ultrastructure and precise orientation upon the cytoplasmic surface.     

Fine structure of probable mechano- and chemoreceptors in the caudal epidermis of the lugworm Arenicola marina (Annelida,Polychaeta)

Summary The caudal part of the lugworm Arenicola marina shows numerous epidermal papillae formed by a thick glandular epidermis in which ciliated sensory buds have been found. These buds comprise supporting cells and two types of receptors, R1 and R2, which are primary sensory cells whose axons are connected to the basiepidermal nerve plexus. The receptors possess several typical cilia projecting into the surrounding seawater, stout intracuticular microvilli filled with filaments, and they contain dense vesicles. The R1 cells, more numerous, show features of chemosensory cells. The rarer R2 cells have large striated rootlets surrounded by a dense sheath of fibrillar material and are probably mechanoreceptors. The physiological functions of these receptors are discussed.     

F0 cysteine,bCys21, in the Escherichia coli ATP synthase is involved in regulation of potassium uptake and molecular hydrogen production in anaerobic conditions

The single cysteine in the b subunit of the membranous F₀ sector and the 19 cysteines in extramembranous F₁ sector of the Escherichia coli ATP synthase were replaced by alanine. When cells were grown under anaerobic conditions on glucose, the k _cat for ATP hydrolysis of membrane vesicles containing the bCys21Ala mutant enzyme, but not enzymes with other cysteine replacements, was lower, while ATP-driven H⁺ pumping was unchanged. However, the ATP-dependent increase in the number of accessible thiol groups in membrane vesicles was negated. Furthermore, K⁺ uptake and molecular hydrogen production by whole cells and protoplasts was greatly decreased. These results indicate a role for the F₀ subunit bCys21 in the functionality of F₀F₁ and coupling to other membranous activities under fermentative conditions.     

Sensorische Eingänge und synaptische Verbindungen im Zentralnervensystem von Insekten

Zusammenfassung Bei Calliphora vicina und Periplaneta americana wurde mit Hilfe experimentell erzeugter anterograder Degeneration der Verlauf von Axonen antennaler Rezeptorzellen im Oberschlundganglion verfolgt. Nach Abtrennen einer Antennengeißel (die beiden proximalen Segmente bleiben intakt) findet man degenerierte Nervenfortsätze in den Glomeruli des ipsilateralen Deutocerebrum. Bei Calliphora laufen viele dieser Axone über eine Bahn dorsal des Oesophagus in die Glomeruli des contralateralen Deutocerebrum. Ein großer Ast des Antennennerven Calliphora zieht vorbei an der Region der Glomeruli in posteriore Regionen des Oberschlundganglions. Nach Abtrennen der ganzen Antenne einschließlich der proximalen Segmente findet sich in diesem Trakt eine große Zahl degenerierter Axone. Die Synapsen in der Region der Glomeruli ähneln anderen Synapsen, wie sie im Zentralnervensystem von Insekten bisher beschrieben wurden.

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Sensory inputs and synaptic connections in the insect CNSExperimental degeneration in the antennal afferent pathway in the supraoesophageal ganglia of flies and cockroaches

Summary Pathways of axons from antennal receptor organs into the brain have been traced by means of anterograde experimental degeneration in Calliphora vicina and Periplaneta americana. After removal of one antennal flagellum (with the two proximal segments left intact) degenerating nervous processes were found in the glomeruli of the ipsilateral deutocerebrum. In Calliphora a great number of these axons run via a pathway dorsal to the oesophagus into the glomeruli of the contralateral deutocerebrum. A big branch of the antennal nerve of Calliphora runs toward a posterior region of the brain bypassing the glomerular region. After removal of the whole antenna including the proximal segments degenerations can be seen also in this branch on the ipsilateral side. The synapses in the glomerular region of the deutocerebrum resemble other central synapses in insects described so far (electron dense cleft, often ribbon-like dense structures within the presynaptic element near or at the synaptic membrane, clear vesicles etc.).

Mit Unterstützung des Schweizerischen Nationalfonds für die wissenschaftliche Forschung Nr. 3807.     

Ultrastructural changes in suspension culture cells of Panicum maximum during cryopreservation

A Panicum maximum cell suspension was used to study ultrastructural changes during cryopreservation. Pregrowing the cells in mannitol caused reduction in the vacuolar volume by redistribution of the large central vacuole into a number of smaller vesicles. Invaginations were formed in the plasma membrane of the cells, to accommodate the reduced cell volume. Swelling of organelles occurred during different stages of cryopreservation. The cisternae of the endoplasmic reticulum dilated and formed vesicles. Although some damage was apparent, organelles were still recognizable in cells frozen slowly and freeze-fixed at –10°C. The cells were able to repair such damage within two days in culture, and regained their normal appearance. Cells frozen slowly without any cryoprotection, and cells frozen rapidly by direct immersion into liquid nitrogen after cryoprotection, were lethally damaged by destruction of membranous structures. Osmiophilic granules were found along the plasma membrane of lethally damaged cells, indicating that their formation is a consequence of freeze damage, rather than a mechanism to prevent injury.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - DMSO dimethyl sulfoxide     

In vivo shedding of apical plasma membrane in the thyroid follicle cells of the mouse

Summary Clusters of luminal dense bodies, limited by a triple-layered membrane, were found in all follicle lumina in thyroid glands of mice. After thyroxine treatment the number of luminal dense bodies increased, especially in the periphery of the lumen, where the intraluminal bodies often displayed a striking resemblance to microvilli. In hyperplastic goiters, obtained by feeding mice with propylthiouracil, luminal dense bodies were replaced by intraluminal vesicles. During goiter involution the vesicles were gradually replaced by luminal dense bodies; the presence of intermediate forms suggests that vesicles and dense bodies are basically the same formations. Luminal dense bodies were observed in colloid droplets indicating their removal by endocytosis. As demonstrated by electron-microscopic cytochemistry, luminal dense bodies contain a membranebound peroxidase, and electron-microscopic autoradiography after administration of ¹²⁵I indicate that they possess an iodinating capacity.Our observations on mouse thyroid glands suggest that the luminal dense bodies, which appear as vesicles in hyperplastic glands, are formed by shedding of the apical plasma membrane of the follicle cell. The shedding process might be of importance for the turnover of plasma-membrane material.This study was supported by Grant No. 12X-537 from the Swedish Medical Research Council.     

The formation of intercellular bridge coating in the hairy whirligig beetle,Orectochilus villosus

Summary The ultrastructure of the intercellular bridges in the ovaries of Orectochilus villosus has been studied. In young egg chambers the inner filamentous coating of the bridge is thin. Characteristic vesicles are observed in close contact with this coating. The inner coating of the vitellogenic chambers is fully developed although vesicles are only sporadically associated with it. These observations suggest that the vesicles are involved in the formation of the filamentous coating.This work was supported by Government Problem Grant MR II, 1.6.4.     

ULTRASTRUCTURE OF TETRASPOROGENESIS IN THE PARASITIC RED ALGA LEVRINGIELLA GARDNERI (SETCHELL) KYLIN12

Ultrastructural studies on tetraspore formation in Levringiella gardneri revealed that 3 stages may be recognized during their formation. The youngest stage consists of a uninucleate tetraspore mother cell with synaptonemal complexes present during early prophase of meiosis I. Mitochondria are aggregated around the nucleus, dictyosome activity is low, and chloroplasts occur in the peripheral cytoplasm. A 4-nucleate tetraspore mother cell is formed prior to tetrahedral cell cleavage, and an increase in the number of chloroplasts and mitochondria occurs. Small straight-profiled dictyosomes secrete vesicles into larger fibrous vesicles or contribute material to the developing tetraspore wall. During the second stage of tetraspore formation, striated vesicles form within endoplasmic reticulum, semicircular profiled dictyosomes secrete vesicles for fibrous vesicles or wall material, and starch formation increases. The final stage is characterized by the disappearance of striated vesicles, presence of straight, large dictyosomes which secrete cored vesicles, and an abundance of starch grains. Cleavage is usually complete at this stage and the tetraspore wall consists of a narrow outer layer of fibrillar material and an inner, electron transparent layer. These spores are surrounded by a tetrasporangial wall which was the original wall surrounding the tetraspore mother cell.     

Etudes sur l'épicuticule des insectes

Résumé L'épicuticule de l'adulte de Tenebrio molitor est composée de deux couches distinctes dénommées épicuticule externe et épicuticule interne. L'épicuticule externe est la première couche cuticulaire sécrétée sous forme de petites plaques s'agrandissant par leurs bords pour recouvrir toute la surface cellulaire. Au moment de sa sécrétion, cette couche est formée de quatre lames denses A, B, C1 et C2. La lame B, très fine, disparaît par la suite et les lames C1 et C2 deviennent très nettes. L'épicuticule externe de l'adulte est donc formée de trois lames denses séparées par deux lames claires.L'épicuticule interne est formée de lames superposées denses et claires de structure complexe, qui sont masquées pendant la sécrétion des premières couches de cuticule lamellée (procuticule). Cette structure correspond à un arrangement moléculaire hautement organisé.La forme de la surface cuticulaire des sternites est déterminée par la forme de la surface cellulaire avant le dépôt de l'épicuticule.

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The development of the epicuticle in the adult Tenebrio molitor L.

Summary The epicuticle of adult Tenebrio consists of two distinct layers named outer and inner epicuticle.The outer epicuticle is the first cuticular layer to be deposited in form of small patches on top of the microvilli. These initial patches are composed of four dense laminae (A, B, C1 and C2) separated by three light spaces. The outer epicuticle grows by densification of diffuse material at the edges of the patches until the entire area is covered.The thickness of outer epicuticle remains constant (175 Å) during the development of the pharate adult, lamina B however rapidly disappears. Thus, the adult outer epicuticle is fivelayered (three dense laminae: A, C1 and C2).After being deposited, the inner epicuticle shows a complex laminar structure interpreted to represent a highly organized molecular system. The laminae are masked during the formation of the first procuticle lamellae.During the deposition of the epicuticle, lamina A is covered by a component of the moulting fluid, forming an irregular dense layer which disappears after the resorption of this fluid. Perhaps this layer protects the new epicuticle from lytic enzymes of the moulting fluid.In adult animals, there is an additional superficial layer, the signification of which is not clear. The possibility of remains of cement or wax is discussed.The development of the surface patterns of the sternal and pleural cuticle is determined before the epicuticle formation by the shape of the epidermal surface. The rate of outer epicuticle deposition appears to depend on the size of the microvilli: epicuticle deposition seems to proceed faster over high microvilli.

Nous tenons à remercier notre directeur de recherche, le Professeur Noirot pour ses encouragements et ses conseils et Madame Curie pour son aide technique efficace.     

Characterization of fumonisin toxicity in orally and intravenously dosed swine

Fumonisin B₁ (FB₁), a recently identified mycotoxin produced by Fusarium moniliforme in corn, has been shown to cause death in swine due to pulmonary edema, an apparently species specific effect, and to interfere with sphingolipid metabolism in vitro. Here we characterize the toxicity of fumonisins, using female cross-bred swine weighing 6 to 13 kg, and present a hypothesis regarding the mechanism of fumonisin-induced pulmonary edema in swine. FB₁ was given daily intravenously (IV) to pig 1 for 9 days for a total of 72 mg (7.9 mg/kg) and to pig 2 for 4 days for a total of 67 mg (4.6 mg/kg). Pig 3 (control) was given saline IV for 9 days. Corn screenings naturally contaminated with FB₁ (166 ppm) and FB₂ (48 ppm) were fed to pigs 4, 5, and 6, and ground corn was fed to pigs 7 and 8 (controls). Pigs 4 and 7 were killed on day 5; pig 5 was found dead on day 6; and pigs 6 and 8 were killed on day 15. Pigs 4 and 5 had ingested 187 and 176 mg total fumonisins, respectively, while pig 6 had ingested 645 mg. Feed consumption had decreased in pigs fed corn screenings, with an additional sharp decrease prior to onset of clinical signs. Increases in serum liver enzymes, total bilirubin, and cholesterol were present, but electrocardiograms, heart rate, and body temperature were unaffected. Pigs dosed IV with FB₁, developed mild intermittent respiratory abnormalities, while those fed screenings developed respiratory distress within 5 days. Mild interstitial pulmonary edema was observed in pig 1. Severe interstitial pulmonary edema, pleural effusion, and increased lung wet/dry weight ratio were observed in pigs 4 and 5. All pigs given fumonisin (either IV or orally) had hepatic changes characterized by hepatocyte disorganization and necrosis; pancreatic acinar cell degeneration was also observed. Ultrastructural changes in orally dosed swine included loss of sinusoidal hepatocyte microvilli; membranous material in hepatic sinusoids; and multilamellar bodies in hepatocytes, Kupffer cells, pancreatic acinar cells and pulmonary macrophages. Pulmonary intravascular macrophages (PIMs) contained large amounts of membranous material. Thus, the target organs of fumonisin in the pig are the lung, liver, and pancreas. At lower doses, slowly progressive hepatic disease is the most prominent feature, while at higher doses, acute pulmonary edema is superimposed on hepatic injury and may cause death. We hypothesize that altered sphingolipid metabolism causes hepatocellular damage resulting in release of membranous material into the circulation. This material is phagocytosed by the PIMs thus triggering the release of mediators which ultimately results in pulmonary edema.Presented in part at the 1991 Annual Meeting of the Society of Toxicology. The Toxicologist 11: 143 (A499).