Styletogenesis in nemertean worms: The ultrastructure of organelles involved in intracellular calcification

The ultrastructure of cellular organelles involved in stylet formation is examined in six species of nemertean worms by transmission electron microscopy (TEM). Stylets are nail-shaped structures containing calcium phosphate that are assembled intracellularly in large uninucleate cells, called styletocytes. Each stylet develops within a membrane-bound vacuole in the styletocyte cytoplasm. Well developed arrays of Golgi bodies are typically found in the vicinity of developing stylet vacuoles, and fully formed vacuoles are filled with PAS+ material that appears to be derived from the smooth endoplasmic reticulum. At the onset of styletogenesis, a conical sliver of organic material differentiates on the inner surface of the vacuolar membrane. This material displays a species-specific banding pattern in decalcified sections, and apparently acts as a template during calcification of the stylet shaft. After the organic core of the shaft is formed, mitochondria aggregate around the stylet vacuole and presumably help accumulate the calcium used in mineralization of the stylet. A knob-shaped proximal piece is subsequently assembled on the base of the shaft. The proximal piece contains a nonbanded matrix and has electron-dense material at its surface that may help in correctly orienting this region toward the basis during replacement of the central stylet.     

Struktur und Bildung der Stilette bei Haloplanella longatuba und Vejdovskya pellucida (Plathelminthes,Rhabdocoela)

Zusammenfassung Die Penisstilette von Haloplanella longatuba und Vejdovskya pellucida werden intrazellulär und sukzessiv von der Spitze her ausdifferenziert; jedoch bestehen hinsichtlich der Bildungsmodi Unterschiede. Bei H. longatuba wird eine doppelwandige, rohrförmige Hartstruktur in einer ebenfalls rohrförmigen Stilettbildungszelle angelegt. Frei im Cytoplasma werden, an Mikrotubuli angelagert, zahlreiche unregelmäßige Hartelemente gebildet, die später mit der äußeren Stilettwand verschmelzen; diese Wand entsteht durch Anlagerung elektronendichten Materials an die Innenseite der peripheren Zellmembran. Der Aufbau der inneren Schicht um das Lumen der Hartstruktur herum erfolgt an der zentralen Membran der Bildungszelle ohne Anlage von Einzelelementen. Bei V. pellucida kondensiert frei im Cytoplasma der Stilettbildungszelle eine mäßig elektronendichte Substanz in Form unregelmäßig oder faserig geformter Partikel; kurz danach wird an den Innenseiten der Zellmembranen eine dünne homogene Schicht aus dem gleichen Material angelegt. Anschließend werden die Wandungen durch Einlagerung von Hartsubstanz verfestigt. Die Stilettbasis erhält sowohl bei H. longatuba als auch bei V. pellucida eine poröse Struktur und wird in eine intrazelluläre Matrix eingebettet. Beide Stilettbildungsmodi lassen sich trotz ihres sukzessiven Verlaufs von der Holo-Simultanbildung ableiten, die wahrscheinlich zum Merkmalsgrundmuster der Rhabdocoela gehört.

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Structure and formation of the stylets of Haloplanella longatuba and Vejdovskya pellucida (Plathelminthes, Rhabdocoela)

Summary The penial stylets of Haloplanella longatuba and Vejdovskya pellucida are formed intracellularly and in a successive mode starting from the distal end. However, there are obvious differences in the mode of formation. In H. longatuba, a double-walled, tube-shaped hard structure develops in a stylet-forming cell, which is also tubular. In the cytoplasm, numerous irregularly shaped hard elements are formed close to microtubules, and these later fuse with the outer wall layer of the stylet. This wall is built up by the accumulation of electron-dense material at the inner side of the peripheral cell membrane. The inner layer enclosing the lumen of the hard structure is differentiated at the central membrane of the forming cell without individual elements. In V. pellucida, a rather electron-dense substance condenses freely in the cytoplasm of the style-forming cell. Afterwards, a very thin homogeneous layer of the same material develops on the inner sides of the cell membranes. Finally, the walls are strengthened by the deposition of further hard substances. In both H. longatuba and V. pellucida the stylet bases show a porous structure and are embedded in an intracellular matrix. Although both modes of stylet formation are obviously successive, they may be derived from the holo-synchronous formation mode, which is probably part of the ground pattern of the Rhabdocoela.

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Abkürzungen ae Atriumepithel - ag Atrium genitale - b Basallamina - ba Bakterium - bm Bulbusmuskulatur - bz Stilettbildungszelle - d Ductus ejaculatorius bzw. freies Stilettlumen - f Füllzelle - h Hüllzelle - he Hemidesmosom - mk männlicher Genitalkanal - mm Muskulatur des männlichen Genitalkanals - mv Mikrovilli - n Kern der Stilettbildungszelle - s Stilett - sz1,2,3 Sekretzelle des Typs 1,2,3     

Sensory innervation monitoring movement and position in the mandibular stylets of the aphid, Brevicoryne brassicae

The sensory innervation of the mandibular stylets of the aphid Brevicoryne brassicae (L.) has been examined by electron microscopy. Two groups of sensory neurones are present in the mandible. Each has two neurones, one with a short dendrite extending into the base of the mandible and ending in the base and another with a long microtubular process which extends 500 m? down to the distal tip of the mandible. The two neurones are enclosed by an ensheathing cell comparable to the trichogen cell enveloping the group of neurones innervating pegs and hairs. This ensheathing cell is supported by extensive electron-dense filaments to form a scolopale and is embedded in the mass of stylet-forming cells at the base of the mandible. The inner segments of the dendrites are anchored to the ensheathing cell by desmosome junctions. Desmosome junctions also bind the microtubular outer segments of the short and long dendrite to each other. There is no evidence of a dendritic sheath enclosing the distal portion of the short dendrite which ends while still in the extracellular space within the ensheathing cell. The microtubular process of the long dendrite extends down the lumen of the mandible enclosed by a close-fitting extracellular sheath which penetrates and is attached to the cuticular wall of the mandible tip. Distally this sheath is thickened on one side. Deflection of the mandible would therefore deform the dendritic membrane asymmetrically because the thin walls of the sheath would bend more than the thick walls. This would exert an unequal mechanical strain on the dendritic membrane which could result in depolarization in response to deflection in a particular direction. The arrangement of the dendrites and their sheaths within the mandible is such that deflection to the right would distort one dendrite in the same way as deflection to the left would distort the other.     

Comparative ultrastructure of copulatory organs having a stylet in the Proseriata (Turbellaria)

The ultrastructure of male copulatory organs having a stylet has been studied in some genera of the Proseriata.Within the Monocelididae there was a variety of stylet-like hard structures. The stylet in Monocelis fusca was a differentiation of the basement membrane of the epithelium lining a penis-like muscular papilla. The penis papilla in Ectocotyla consisted of circular muscles surrounded by a thickened basement membrane and an epithelium. Archilopsis sp. and Archilina sp. with a duplex copulatory bulb, had a stylet within a spiny cirrus. The stylet in Archilopsis sp. was a cylindrical muscular protrusion with a thickened basement lamina that lined the cirrus lumen. The stylet structure in Archilina sp. was composed of four long spines which were derivatives of the basement membrane. In Ectocotyla multitesticulata and Dupliminona corsicana, the accessory prostatoid organ was provided with a hook-shaped stylet that was differentiated in the basement membrane and of which the material was continuous with the fibrous matrix between the muscles of the prostatic bulb. The stylet and needles in the Archimonocelis species were intracellular differentiations. The copulatory organ in Carenscoilia biforamen consisted of a tubiform stylet and four needles, all of which were also intracellular specializations.I consider copulatory hard structures in the Turbellaria to be taxonomically significant in terms of structure, differentiation, and location (whether subcellular, in the basement membrane, or intracellular).     

The transfer of lipid in insects from the epidermal cells to the cuticle

The structure of the pore canals and the tubular filaments they contain are described in a series of insects and types of cuticle. In all these cuticles the tubular filaments arise from the plasma membrane of the epidermal cells and they contain argentaffin material, regarded as sclerotin precursors, and lipid-staining material, regarded as wax precursors. These materials are transferred to the inner epicuticle and are exuded over the surface of the outer epicuticle to form the waterproofing layer as described in the preceding paper. They are also transported to those parts of the endocuticle destined to form hard exocuticle. There are no terminations of tubular filaments in the soft cuticle of Manduca larva, in the soft expanding cuticle of Rhodnius, and in the non-sclerotized post-ecdysial endocuticle of Tenebrio. Apis. etc. In the puparium of Calliphora lipid appears to be added by the epidermal cells directly and not by way of tubular filaments. It is confirmed that lipid is a component of sclerotized cuticle.     

The chitin-glucan complex ofSaccharomyces cerevisiae

Differentiation of the cell wall ofSaccharomyces cerevisiae at the site of the future bud was followed. A lentil-like structure originates on the inner side of the cell wall during the first phase. At the same time, an electron-dense layer occurs at the boundary between the inner layer of the cell wall and the lentil-like structure. During the second phase granular material is accumulated at the lower side of the lentil-like structure. During the third phase the lentil-like structure is split apart due to proliferation of the granular material resulting in formation of the base of the encircling region. The marked electron-dense layer observed from the first phase is attached to the surface of the encircling region during differentiation of the latter. During the budding proper the outer layers of the cell wall protrude and the end of the encircling region, together with the adjacent electron-dense layer, acquire their definitive appearance of rings, observed as marked electron-transparent and electron-dense tears on ultrathin sections.     

Studies on the differentiation of the retina receptor cell of toad (Bufo raddei Strauch)

The development of the retinal receptor cell in the young tadpoles (Bufo raddei Strauch), from the stage 20 to the stage 25, was studied by TEM and immunohistochemical method. The morphological differentiation of the photoreceptor cell may be described as follows. The time and the degree of differentiation of the cells in the tadpole retina is asynchronous between central (posterior pole) and peripheral parts of the tadpole retina, namely, they are earlier and higher in the central than in the peripheral. The cells of the outer nuclear layer are undifferentiated at the stage 20. The cells in the posterior part of the retina elongate at the beginning of the stage 21 (Plate I, Fig. 1). This is the first sign of differentiation in the photoreceptor cell. A small hillock-like process forms the inner segment at the scleral pole of the receptor cell. The inner segment is rich in mitochondria, rough-surfaced cytomembrane, free ribosomes, and vesicles. One or two large lipid droplets are also found in the inner segment (Plate I Fig. 2-3). Later on, the connecting structure develops at the tip of the inner segment. The newly formed filaments and the plasma membrane form the outer segment. Its membrane forms some evaginations oriented perpendicularly to the longitudinal axis of the receptor cell. In this way, disks of the outer segment are formed (Plate I Fig. 4-5). The length of the outer segment gradually increases with the number of disks increasing at the base. At the same time, an axon process of receptor cell, extending vitreal, develops synapses with dendrites of the bipolar cell in the outer plexiform layer. At the beginning (the stage 22), the synaptic structure is an immature form that lacks synaptic ribbons and vesicles (Plate II Fig. 8). Later on, ribbons and vesicles are observed in the further developed synaptic structure (Plate II Fig. 9). The toad rhodopsin was prepared by a method of Dewey et al. (1969) and Papermaster & Dreyer (1974) with slight modification. A specific immune serum against the toad rhodopsin was produced in rabbits. Using the indirect Coon's antibody technique, the localization of the rhodopsin antibody and the time when the antibody was seen in the retina of the early developing tadpoles was traced.(ABSTRACT TRUNCATED AT 400 WORDS)     

Feeding Position of Laccifer lucca on Dalbergia balansae and the Influence of Parasitism on Bark Structure

When Laccifer lacca fed in the bark of Dalbergia balansae, the penetration in the bark by a stylet was mainly intracellular, seldom intercellular. Finally, the stylet arrived at the funtional sieve element, and fed in it. The tip of tt,e stylet was at a distance of 0.48–0.78 mm from the surface of periderm. 70.3% of the stylets fed in the zone of newly-differentiated sieve elements. The fed sieve element had P-protein and callose, and exhibited no serious reaction of injury. The parenchyma cells that were pierced through by the stylet and the neighbouring cells Lad obvious reaction of injury, such as: thickened cytoplasm and plasmolysed; dark stained nuclei; smaller starch grains and intracellular deposition of concentrated golden material. The stylet that pierced through the bark was encircled by a stylet sheath consisted of proteins. The stylet sheath looked like a string of beads as a whole. Branching stylet sheath was observed. Some branches even reached far into the xylem, but the stylet finally reached the sieve element. At the same time, the stylet might penetrate through many sieve elements, finally reach newly-differentiated sieve elements. These results suggest that feeding of Laccifer lacca was a process of initiative choice. Two years after collecting shellac by means of skinning instead of cutting the branch, tb.e stylets and styler sheaths still remained in the bark. Several layers of ceils around them were dead and fully imbued with yellow-brown material. Stylers and styler sheaths in the outer cortex were surrounded by bending phellogen and separated from the living cells, forming many cyst-like structures in the periderm. Such bark should not be further used for feeding.     

Electron microscopic study of glycocalyx formation in different functional states of the frog bladder epithelial cells]

The apical part of the urinary bladder granular cells contains oval granules filled with a homogenous substance; close to the plasma membrane granules with tubular element are disposed. A preliminary 30 min fixation of the bladder epithelium with 0.1--0.5% glutaraldehyde followed by 2.5% glutaraldehyde post-fixation rather increased the number of granules with the orderly distributed inner material. The membrane of these granules includes in to the plasma membrane. The loosening of the inner material takes place and the fibrillar elements similar in structure to the glycocalix fibrilles attach to the external surface of the cell plasma membrane.     

Localization of the ASV src gene product to the plasma membrane of transformed cells by electron microscopic immunocytochemistry.

The cellular location of the src gene product (p60src) of the Schmidt-Ruppin strain of avian sarcoma virus has been determined by electron microscopic immunocytochemistry in Schmidt-Ruppin ASV-transformed NRK cells, and the amount of the protein in different regions of the cell has been quantified. The protein is concentrated on the inner surface of the plasma membrane, particularly under ruffles, and it is highly concentrated on the inner surface of the membrane near junctions connecting adjacent cells. Small amounts of p60src were detected in the cytoplasm and in the perinuclear Golgi region of the cell. No significant localization was detected in control NRK cells or in NRK cells transformed by the Kirsten strain of murine sarcoma virus. The presence of p60src on the inner surface of the plasma membrane indicates that the changes in cell growth, cell shape and cell membrane structure noted in ASV-transformed cells are due to an initial action of p60src at the cell membrane.     

The fine structure of Gnathostomulid reproductive organs

Summary The male copulatory organs of five species of Gnathostomulida Scleroperalia have been studied by TEM techniques. These observations provide a more solid basis for classification in the light microscope: inLabidognathia longicollis (fam. Mesognathariidae) the stylet is composed of eight, and inSemaeognathia sterreri, Gnathostomula jenneri, Gnathostomula mediterranea andGnathostomula microstyla (Gnathostomulidae) of ten stylet rods. Each rod consists of a microtubule-filled inner rod, and of an outer rod, filled with crystallized inclusions. The inner rods are continuous with eight — or ten — rod formation cells which are located in the proximal stylet sack. Bipartition of rods occurs by a longitudinal invagination of the basement lamina, underlying the rod cells and the gland cells and continuous with that of the body wall epithelium. InLabidognathia, the outer rods are interlocked, in Gnathostomulidae, the stylet rods are surrounded by an extracellular (cuticular) tube-like stylet sheath of variable fine structure, which is believed to provide extra rigidity. In the species investigated, one single stylet gland, consisting of a monolayered epithelium showing different gland cell types, surrounds the stylet. In the apical gland cell portions, medially and distally membrane-bound secretory granules lie adjacent to the stylet sheath. In Gnathostomulidae, two anterior gland cells are seen in connection with the formation of the stylet sheath. In the muscular sheath the cross-striated fibers, basically derived from the longitudinal body wall musculature, show a tendency towards helical and circumferential arrangement. Musculature is especially prominent in the proximal stylet sack, which is rather a propulsive element than a sperm-storing vesicle, and lacks glands. InGnathostomula species, atrial cells underlie the distal tip of the stylet. The entrance into the male opening is lined with ciliary receptor cells and specialized gland cells.Stylet evolution in Scleroperalia is characterized by progressive differentiation of the muscular sheath, in particular of the proximal stylet sack, and of the stylet — the occurrence of a stylet sheath is seen in connection with increasing diversity of stylet shape.Abbreviations ac atrial cell(s) - ag anterior gland cell(s) - b bursa - bl basal lamina - c rod-crystal in outer rod - cj cuticle of jaw - d desmosome - di dictyosome - e body wall epithelium - ej pharyngeal epithelium - g stylet gland (cell) - gm median gland cell - i gut (cell) - ir inner rod - jc junctional complex - m muscular layer - mo male opening - mv microvillar protrusions - nu nucleus - o ovary - or outer rod - po proximal opening of the proximal stylet sack - ps proximal stylet sack - r stylet rod - rc rod cell - sg secretory granule - sj septate junction - sp sperm - ss stylet sheath - st stylet - te testes - v ventral - z centriole     

Ultrastructure of root cortical cells parasitized by the ring nematodeCriconemella xenoplax

Summary Individuals of the plant-parasitic nematodeCriconemella xenoplax, monoxenically cultured on root expiants of clover, carnation, and tomato, fed continuously for up to 8 days from single cells in the outer root cortex. Individual cortical cells parasitized by nematodes were modified into discrete food cells in all hosts examined. The nematode's stylet penetrated between epidermal cells and frequently through a subepidermal cortical cell. Electron-transparent callose-like material continuous with the cell wall enveloped the portion of the stylet that traversed subepidermal cortical cells. Food cells were typically located in the first or second cell layers of the cortex. The stylet penetrated 5–6 m through the wall of the food cell without penetrating the plasma membrane. Electron-transparent callose-like deposits formed between the invaginated plasma membrane and stylet, except at its aperture. The plasma membrane of the food cell was appressed tightly to the wall of the stylet aperture creating a 130–160 nm hole in the membrane. This opening provided continuity between the lumen of the stylet and the food cell cytosol for ingestion of nutrients by the nematode. Ribosomes were dissociated from the cisternae of the endoplasmic reticulum in food cells and accumulated with other cell organelles in a zone of modified cytoplasm around the stylet. A fibrillar material appeared to form a barrier in the cytosol around the stylet aperture that limited movement of cell organelles toward the aperture. Electron-dense secretory components were secreted into the food cell by the nematode. Clusters of putative nematode secretory components consisting of 20–40 nm diameter, electron-dense particles were dispersed in the densely particulate zone of cytoplasm around the stylet tip. The cytosol immediately around the stylet aperture in the center of the modified cytoplasm was finely granular.Plasmodesmata connecting the cytoplasm of the food cell with the cytoplasm of neighboring cells were greatly modified in a way that could facilitate solute transport into the food cell. The plasma membrane-lined canals of the modified plasmodesmata appeared to be increased in diameter and lacked desmotubules. Additionally, they frequently were lengthened by electron-transparent callose-like deposits projecting from the wall into the cytoplasm of the food cell. An electron-dense cap that formed an apparent tight seal with the plasma membrane developed over the entrance of each modified plasmodesma in the neighboring cells. These caps excluded all cell organelles from the cytosol contained within them. The nucleus of the food cell was usually enlarged and atypically shaped with dense peripheral clumps of condensed chromatin. Our results show thatC. xenoplax induces elaborate cellular modifications in host tissue to support sustained ingestion of nutrients from a single food cell.     

The fine structure of distal receptors on the labium of the aphid,Brevicoryne brassicae L. (Homoptera)

Summary Short peg receptors located at the distal tip of the aphid labium have the structure of mechanoreceptors. Each peg is innervated by a single sensory nerve which is anchored eccentrically to a basal cuticular tube and terminates in electron-dense material in the base of the peg. The arrangement and eccentric insertion of the eight pegs in the labial wall on one side of the stylet groove, with the eccentric insertions of their innervating neurones, provide a mirror image of the receptors on the opposite side. On the basis of a comparison of the structure of these receptors with that of tactile receptors for which electrophysiological data on sensitivity are available, it is possible to predict that the receptors detect both surface contact (pressure) and surface profile; and that the bilateral symmetry in the receptor arrangement facilitates the detection of vein contours which are preferred settling sites on the leaf. The structure of the dendritic terminal and its insertion is that of a well reinforced cytoskeleton designed to transmit tension to the cell membrane, in agreement with the concept that transduction is a membrane related phenomenon. The distal microtubules, fifty per-cent of which originate as well as terminate in the tubular body, are packed in electron-dense material which binds to the cell membrane. The membrane in turn is attached to cuticular components of the receptor. Abrupt changes in dimension of the dendritic outer segment may be designed to modulate the conduction of a membrane potential. On the other hand, lack of continuity in the microtubules makes these organelles poor candidates for the transduction of excitation from a distal site of stimulation to a proximal region.Supported by operating grants Nos. A 6063 and A 9856 from NRCC     

Release of chromaffin granular content from staphylococcal enterotoxin B (SEB)-treated and-untreated PC12 cells

Summary The release of chromaffin granular content from staphylococcal enterotoxin B (SEB)-treated and-untreated PC12 cells was studied by electron microscopy. The treatment of the cells with SEB at the concentration of 20 μg/ml caused marked increase of the chromaffin granules that either bound to the plasma membrane by the characteristic rods, measuring 15 to 20 nm in length and showing a tubular structure, or budded off at the free cell surface, surrounded by a layer of rod-containing cytoplasm and enclosed by the plasma membrane. The binding between the granular and plasma membranes by the rods did not lead to membrane fusion and exocytosis of the granular content. Many of the bound granules showed vesiculation with loss of the electron-dense core material; at the same time, some of the binding rods contained intraluminal electron-dense material similar to the granular core material. These findings suggested that the electron-dense material (i.e., norepinephrine) of the bound granules was released extracellularly through channels within the rods. Although the granules were bound to the plasma membrane with equal frequency at the free and contiguous cell surfaces, the granular budding occurred only at the free cell surface, indicating that it occurred incidentally to some granules bound at the free cell surfaces. On the basis of the morphological observations, it is postulated that the electron-dense material of the bound granule is selectively released extracellularly through the rods, leaving the vesiculated granules behind in the cytoplasm. The same mode of release of the granular content was observed, though less frequently, in the untreated control cells. No morphological evidence that indicated that the granular content was released extracellularly by exocytosis was found in the treated and control cells. The present observations indicated that the SEB treatment of PC12 cells stimulated the binding of chromaffin granules to the plasma membrane by the rods and the budding of the bound granules at the free cell surface.     

Ultrastructure of the copulatory organ of Haplopharynx quadristimulus and its phylogenetic significance (Plathelminthes,Haplopharyngida)

Summary The male reproductive system in Haplopharynx quadristimulus consists of paired testes, sperm ducts, seminal vesicles, seminal ducts, a copulatory organ containing prostatic vesicle and stylet apparatus, and the male canal. By electron microscopy all components appeared to be regional specializations of a canal extending from the testes to the body wall and lined by a multiciliated epithelium. The epithelium of the stylet apparatus contained six different cell types. One cell type (matrix syncytium) formed the stylet and the other five were located distal to the stylet/prostatic-vesicle junction along the male system epithelium. Each cell type was attached to the supporting intercellular matrix at a different level along the stylet apparatus. All cell types extended to the distal end of the stylet apparatus regardless of where they originated along its length. The cells in the apparatus lacked cilia, but one of the cell types contained rootlets. Modified rootlets or rootlet derivatives were possibly present in another cell type in the form of rootlet-like ribbons. The findings support the monophyly of the Macrostomida Haplopharyngida (by common occurrence of a matrix syncytium) and at the same time suggest their separation as two distinct taxa (by differences in the structure of the prostatic vesicle and other parts of the stylet apparatus).Abbreviations a accessory spine - c circular muscle - ce centriole - ci cilium - di dictyosome - e epithelial cell - ed ejaculatory duct - ep epidermal cell - f rootlet-like ribbon - g prostatic gland cell neck - g1 type I gland cell granules - g2 type II gland cell granules - g3 type III gland cell granules - h hemidesmosome - i intercellular matrix - im internal muscle - j septate junction - l stylet apparatus lumen - le spine lateral extension - lm longitudinal muscle - m matrix syncytium - mc male-canal epithelial cell - me male canal - mp male pore - mt microtubules - mv microvilli - n nucleus - nc nerve cell body - np nerve process - om oblique muscle - p prostatic vesicle epithelial cell - pv prostatic vesicle - r rootlet - s stylet - sa stylet apparatus - sc sensory receptor - sd sperm duct - se seminal duct - sl stylet lumen - sp spot desmosome - sr sperm - sv seminal vesicle - t terminal web - te testis - u ultrarhabdite - z zonula adhaerens - 2 cell type 2 - 3 cell type 3 - 4 cell type 4 - 6 cell type 6     

Characteristics of the cytodifferentiation of omega-particle symbiotic bacteria from the micronucleus of Paramecium caudata clone M1-48]

The structure of the omega-particle-bacteria, growing in the micronucleus of Paramecium caudatum (Ciliata, Protozoa), was studied by electrom microscopy in the course of their life cycle. The cytoplasm of the spindle-shaped vegetative cells contains a large number of dense particles and transparent regions comprising the fibrillar material. Such cells, via several intermediate stages, are transformed into elongated twisted cells that are regarded as spores. The spore consists of two parts: homogeneous, and that containing the membrane system and rounded light bodies. The membranes are often double and connected with the fibrils. The cell wall is constructed, during all stages, of the outer membrane layer and the inner electron-dense layer.     

Structure of the shell and tertiary membranes of eggs of softshell turtles (Trionyx spiniferus)

Eggs of the turtle Trionyx spiniferus are rigid, calcareous spheres averaging 2.5 cm in diameter. The eggshell is morphologically very similar to avian eggshells. The outer crystalline layer is composed of roughly columnar aggregates, or shell units, of calcium carbonate in the aragonite form. Each shell unit tapers to a somewhat conical tip at its base. Interior to the crystalline layer are two tertiary egg membranes: the outer shell membrane and the inner shell membrane. The outer shell membrane is firmly attached to the inner surface of the shell, and the two membranes are in contact except at the air cell, where the inner shell membrane separates from the outer shell membrane. Both membranes are multi-layered, with the inner shell membrane exhibiting a more fibrous structure than the outer shell membrane. Numerous pores are found in the eggshell, and these generally occur at the intersection of four or more shell units.     

Effect of Ethylenediaminetetraacetate upon the Surface of Escherichia coli

The effect of ethylenediaminetetraacetate (EDTA) on the envelope of two strains of Escherichia coli (B and Cla) was studied with freeze-fracturing methods. Untreated cells showed the outer membrane's outer surface with a fine texture of randomly spaced depressions of about 4.5-nm diameter; small areas with symmetrical arrangements of structural surface elements were also observed. The outer membrane's fracture plane revealed a random distribution of particles on its "concave" plane, only occasionally interrupted by particle-free areas. The "convex" aspect of the outer membrane's fracture plane showed only a few scattered particles. The cleavage plane of the inner membrane was often interrupted by many localized elevated plateaus, at which the cleaving process had, for short distances, switched to the outer membrane. The effects of EDTA treatment were mainly seen in the structure of the freeze-etched outer membrane: (i) the pits as well as the symmetrical surface elements of the outer membrane's outer surface had disappeared; (ii) a number of plateaus (about 20 to 50/cell) were seen at which a cleavage plane within the inner membrane had switched to the hydrophobic portion of the outer membrane (outer membrane's fracture plane). These plateaus were also visible in untreated cells; however, EDTA treatment apparently caused an increased exposure of plateaus. Surface areas, exposed by freeze-etching, revealed the underlying plateaus as elevations in the surface contour of the cell, suggesting a slower etching rate in the zones of the plateaus relative to the rest of the outer membrane. Well-defined, particle-free patches in the outer membrane's fracture plane, concave, were more frequent and larger in size after EDTA treatment than in the controls. In the presence of glycerol, the cells often cleaved in the outer membrane's fracture plane, but isolated plateaus were rarely observed. After metabolic poisoning of cells for 15 to 25 min at 37 degrees C, the plateaus had widened. These data suggest that the material of the plateaus has a slow rate of lateral diffusion. Placement of EDTA-treated cells in fresh medium at 37 degrees C caused, after 3 to 5 min, the reoccurrence of the pitted surface structure. We propose that the plateaus represent localized zones, at which newly synthesized lipopolysaccharide has been inserted.     

Encapsulation of proteins into human erythrocytes: a kinetic investigation

Moderate osmotic shocks of human erythrocytes by hypotonic dialysis (0.06 mosmol/kg) induce cell swelling and formation of pores, without causing apparent lysis. Using 125I-labeled macromolecules of different molecular weight and net charge, we followed the kinetics and efficiency of their encapsulation into erythrocytes. After a 20-30 min period of cell dialysis, macromolecules of up to 50 kDa begin diffusing into the swollen cells by a process which can be described by a first-order two-compartment kinetics. Adsorption to the external cell surface was insignificant, while adsorption to the inner membrane surface was substantial (15-20%) only for positively charged proteins, at physiological pH. After resealing, pores of a 12-14 kDa cut-off might remain open allowing some release of entrapped material (20-30%), depending on the final cytocrit, while the remaining might be associated with inner membrane or cytosolic components. Although the method of hypotonic dialysis is known to affect minimally the biophysical and immunological properties of red blood cell membranes, the interaction of encapsulated material with cell constituents would need to be further assessed when considering red cells as macromolecular carriers.     

Ultrastructure of infection-thread development during the infection of soybean by Rhizobium japonicum

The location and topography of infection sites in soybean (Glycine max (L.) Merr.) root hairs spot-inoculated with Rhizobium japonicum have been studied at the ultrastructural level. Infections commonly developed at sites created when the induced deformation of an emerging root hair caused a portion of the root-hair cell wall to press against an adjacent epidermal cell, entrapping rhizobia within the pocket between the two host cells. Infections were initiated by bacteria which became embedded in the mucigel in the enclosed groove. Infection-thread formation in soybean appears to involve degradation of mucigel material and localized disruption of the outer layer of the folded hair cell wall by one or more entrapped rhizobia. Rhizobia at the site of penetration are separated from the host cytoplasm by the host plasmalemma and by a layer of wall material that appears similar or identical to the normal inner layer of the hair cell wall. Proliferation of the bacteria results in an irregular, wall-bound sac near the site of penetration. Tubular infection threads, bounded by wall material of the same appearance as that surrounding the sac, emerge from the sac to carry rhizobia roughly single-file into the hair cell. Growing regions of the infection sac or thread are surrounded by host cytoplasm with high concentrations of organelles associated with synthesis and deposition of membrane and cell-wall material. The threads follow a highly irregular path toward the base of the hair cell. Threads commonly run along the base of the hair cell for some distance, and may branch and penetrate into subjacent cortical cells at several points in a manner analagous to the initial penetration of the root hair.