The fine structure of the phoront of Gymnodinioides pacifica, a ciliated protozoan (Ciliophora, Apostomatida) from euphausiids of the Northeastern Pacific

Phoretic stages of the exuviotrophic apostome Gymnodinioides pacifica were examined using transmission and scanning electron microscopy (TEM and SEM). TEM revealed that the mature cyst wall possesses 2 or 3 layers differing by the presence or absence of the third inner layer. This inner layer may represent a different form of the middle wall material. The inner cyst layer is approximately 0.15 microm thick and has striations with a periodicity of approximately 19 nm. The middle cyst layer has a variable thickness and the outer dense layer is approximately 0.1 microm thick. The 3 layered cyst wall had a thickness of 0.3-0.7 microm and averaged 0.5 microm. Advanced phoront stages were enclosed by fully formed cyst walls or by cyst walls thinned to approximately 0.1 microm, as the phoronts prepared to excyst prior to host ecdysis. Additionally, we report the fine structure of the rosette, trichocysts, nuclei, food plaquettes, oral fiber, and other cytoplasmic inclusions. SEM revealed an outer cyst wall layer connected to the secreted peduncle material, which was observed to extend over a wide (15 microm) area on the host setae. Cysts were usually attached at their posterior ends or, less frequently, along their side.     

Morphological Changes of the Inner Surface of the Blastocoelic Wall before and during Gastrulation in the Newt, Cynops pyrrhogaster

Ultrastructural observations were made of morphological changes in the inner surface of the blastocoelic wall (BW) in newt embryos during the period from morula to gastrula. A scanning electron microscopic investigation distinguished three phases of process formation on the inner surface of the BW. First, small blebs and filopodia were found in the morula. These then changed to large blebs in the blastula and early gastrula. Finally, they acquired lamella-like structures (lamellipodia) in the middle and late gastrula. In addition to the changes in the cell surface, cytoplasmic changes such as of the organization of the cortical microfilamentous layer (CML) were found by transmission electron microscopy (TEM). The morphological changes on the inner surface of the BW and those in the cytoplasmic ultrastructures are considered to be closely related, playing some important roles in the mechanisms of morphogenetic cell movements.     

Ultrastructural Changes During Encystment of Schizopyrenus russelli*

Ultrastructural changes associated with the encystment of Schizopyrenus russelli have been studied by electron microscopy. Before encystment small “black bodies” appear in the cytoplasm and later migrate toward the periphery. The outer cyst wall is secreted at this stage as a thin discontinuous layer which thickens and subsequently becomes continuous. Concomitant with this, the endoplasmic reticulum surrounds the mitochondria. The inner cyst wall later appears as a multilayered structure which presumably is cast off from the plasma membrane. Between the inner and outer layers of the cyst wall, there is a middle, less electron-dense layer wherein extruded cytoplasmic material is found embedded at certain places.     

On the fine structure of the external glial layer in the isocortex of man

Summary The surface of the external glial layer of the isocortex in the human temporal lobe is generally slightly undulated, with a few protrusions and indentations. The surface is formed by an uninterrupted basement membrane which is continuous over the surface no matter how tortuous it becomes. The overall thickness of the glial layer is generally 15 to 25 m, but diminishes to about 5 m immediately beneath blood vessels. It consists mainly of a variable number of stacked glial cell processes.Two groups of cell bodies are encountered particularly in the middle and lower levels of the glial layer. Most of the cells are specialized fibrous astrocytes. They are characterized by eccentrically placed, rounded nuclei with homogeneously dispersed chromatin, and electron-lucent cytoplasm rich in filaments. Lipofuscin pigment granules occupy large areas of the perikaryon. The astroglial cells give rise to four types of processes: foot-processes, tangential and radial processes, and processes irregular in outline.The foot-processes ascend towards the cortical surface and terminate as flat expansions spreading out immediately beneath the basement membrane. Contiguous terminal expansions are connected by gap junctions. The individual profiles are irregular in form and fit together like in a jig-saw puzzle. The plasmalemma beneath the basement membrane is underlined by a fuzzy material, which is penetrated by glial filaments. In the terminal expansions individual or groups of mitochondria are abundant.The tangential processes are straight and slender and form a lattice within the middle and deep level of the external glial layer. They contain numerous filaments, evenly distributed or fasciculated. The remainder of the lattice is filled up by a considerable number of processes irregular in outline and varying greatly in size. They contain fewer filaments than the tangential processes, coursing in all directions, and glycogen particles. In both types of processes only a few mitochondria are present. These processes are also connected by gap junctions and desmosomes, too.Large cytoplasmic areas of astroglial cells localized in the deepest portion of the glial layer protrude into the neuropil of the molecular layer, giving rise to several radiate processes, which extend deeper into the cortex.The second, heterogeneous group of cell bodies is characterized by elongated nuclei, ovoid or irregular in outline, which are smaller than those of astroglial cells, and contain blocks of condensed chromatin; a thin cytoplasmic rim generating a few appendages surrounds the nucleus. The first sub-type is characterized by a nucleus with large chromatin blocks bordering the inner nuclear membrane and a medium-dense cytoplasmic matrix. The second sub-type displays smaller chromatin condensations at the inner nuclear membrane and many microtubules are scattered throughout an electron-lucent cytoplasm.     

Three-dimensional reconstruction of a pathogenic yeast Exophiala dermatitidis cell by freeze-substitution and serial sectioning electron microscopy

The structure of a budding cell of the pathogenic yeast Exophiala dermatitidis was observed in three dimensions after freeze-substitution, serial ultrathin sectioning and computer reconstruction. The nucleus occupied about 10% of the cell volume. The spindle pole body was composed of two disk elements connected by an intervening midpiece, and occupied about 0.01% of the cell volume. The cell wall consisted of an inner transparent layer, a middle electron-opaque layer, and an outer fibrous layer. The mitochondria occupied about 10% of the cell volume. There were numerous mitochondria in the mother cell and the bud, but no 'giant mitochondrion' was seen. The ratio of mitochondrial volume within the bud to the mitochondrial volume of the cell was close to the ratio of bud:cell cytoplasmic volume. The results emphasize the importance of good cryofixation for 'perfect' preservation of yeast cell structure.     

Atomic structure of the major capsid protein of rotavirus: implications for the architecture of the virion

The structural protein VP6 of rotavirus, an important pathogen responsible for severe gastroenteritis in children, forms the middle layer in the triple-layered viral capsid. Here we present the crystal structure of VP6 determined to 2 A resolution and describe its interactions with other capsid proteins by fitting the atomic model into electron cryomicroscopic reconstructions of viral particles. VP6, which forms a tight trimer, has two distinct domains: a distal beta-barrel domain and a proximal alpha-helical domain, which interact with the outer and inner layer of the virion, respectively. The overall fold is similar to that of protein VP7 from bluetongue virus, with the subunits wrapping about a central 3-fold axis. A distinguishing feature of the VP6 trimer is a central Zn(2+) ion located on the 3-fold molecular axis. The crude atomic model of the middle layer derived from the fit shows that quasi-equivalence is only partially obeyed by VP6 in the T = 13 middle layer and suggests a model for the assembly of the 260 VP6 trimers onto the T = 1 viral inner layer.     

Ultrastructural studies on the sporulation of oocysts of Toxoplasma gondii. II. Formation of the sporocyst and structure of the sporocyst wall

The ultrastructural changes observed during sporocyst formation and the structure of the sporocyst wall was examined in oocysts which had been allowed to sporulate for between 12 and 48 hours at 27 degrees C. As the spherical sporoblast developed into the sporocyst the cytoplasmic mass became ellipsoidal in shape although no change was noted in the organelle compliment, which cosisted of two nuclei plus a number of polysaccharide granules, lipid globules, mitochondria, Golgi bodies, and some rough endoplasmic reticulum. The sporocyst wall consisted of a thin outer layer (15-20 nm) which was formed from two limiting membranes of the sporoblast and an inner layer (40-50 nm) which was comprised of four curved plates. This inner layer was formed under the outer layer and, although no specific cytoplasmic organelle disappeared with its formation, some unit membranes were observed close to the plasmalemma during its formation. Each curved plate has a marginal swelling and an interposing strip of material is present between the margins of adjacent plates. The plates are joined to the interposing strip by a thin band of osmiophilic material. In oblique and tangential sections through the plates two types of cross banding were observed which differed in periodicity.     

Fine structure of the cell envelope layers of Flexibacter polymorphus.

Electron microscopy of the filamentous gliding marine bacterium Flexibacter polymorphus demonstrated that the cell envelope consists of an electron-dense intermediate layer located between two unit-type membranes: an outer membrane, presumably of lipopolysaccharide, and an inner cytoplasmic membrane. Separation of living filaments into single cells by lysozyme suggests that a peptidoglycan moiety, possibly corresponding to the intermediate layer, might be situated between the two membranes. Cell division proceeds by invagination of the cytoplasmic membrane and intermediate layer forming a transverse septum. Cells generally fail to separate after the division process, so that a common outer membrane encloses all of the cells in a single filament. There is a continuous layer of macromolecular cup-shaped elements ('goblets') attached to the outermost surface of the lipopolysaccharide membrane. Tangential thin sections, as well as negatively stained preparations of envelope fragments (produced by sonication of autolyzed cells), showed that the goblets are arranged in a close-packed hexagonal array. The presence of electron-dense structures located between the outer and inner membranes, and exhibiting the same periodicity as the goblets, suggests that some part of the goblets penetrates the outer membrane and extends across the periplasmic space to the dense intermediate layer or cytoplasmic membrane. Spontaneous autolysis in aging cultures is accompanied by the formation and release into the culture medium of large numbers of outer membrane vesicles coated with globlets. A tentative reconstruction of the envelope of F. polymorphus, based on the fine-structural data, is presented.     

Light and electron microscopic study of the hindgut of the ant (Formica nigricans, hymenoptera): I. Structure of the ileum

The study of the ileum of the ant Formica nigricans by light and electron microscopy revealed the existence of three differentiated regions: proximal, middle, and distal ileum. The middle region constitutes most of the length of the organ. Its wall is made up by a folded simple epithelium lined by a cuticle, which is surrounded by an inner circular muscle layer and various external longitudinal muscle fibers adjacent to the hemolymph. A subepithelial space is present between the epithelium and the circular muscle layer. Epithelial cells show extensive infoldings of the apical, and to a lesser extent the basolateral plasma membrane. Apical infoldings are characterized by the presence of 10-nm particles (portasomes) covering the cytoplasmic side of the membrane. Mitochondria are abundant throughout the cytoplasm, although they mainly are present underneath the apical infoldings. Lateral borders of epithelial cells display an apical junctional complex, mainly constituted by a long and convoluted pleated septate junction. These features support the view that epithelial cells in the middle ileum are specialized in ion solutes and water transport. The proximal ileum connects with the ampulla into which the Malpighian tubules drain. As opposed to the middle ileum, epithelial cells of the proximal ileum show less developed basolateral infoldings, and the apical plasma membrane is devoid of portasomes and only occasionally invaginates. These features suggest that the proximal ileum plays no relevant role in ion and water transport. The distal ileum penetrates into the rectal sac, forming a valve-like structure; this region presumably controls the amount of urine reaching the rectum.     

Aminergic and Indoleamine Accumulating Neurons in the Retina of the River Lamprey (Lampetra fluviatilis)

Tissues were processed for fluorescence microscopy of biogenic amines according to the method of Falck and Hillarp. Normal animals, and animals injected with α-methylnoradrenaline or 5,6-dihydroxytryptamine were used. Catecholamine containing neurons (junctional cells) occur in the innermost rows of cell bodies of the inner nuclear layer (INL) and close to the vitreous surface. Catecholamine containing fibers occur in three layers: (1) an outer layer around the innermost perikarya of the INL, which is a condition not found in retinas of gnathostome chordates; (2) a middle layer within the outer third of the inner synaptic layer (ISL), separated from the outer layer by ganglion cell axons; (3) a sparse inner layer within the innermost third of the ISL. A few catecholamine containing fibers were seen to extend from the innermost region of the INL to the outer synaptic layer. The position of the junctional cells in the lamprey corresponds to that in gnathostome chordates, but whereas all catecholamine containing fiber layers in gnathostomes are located sclerally to the optic fiber layer and within the ISL, the middle and the inner fiber layers in the lamprey occur vitreally to the optic fiber layer. Indoleamine accumulating neurons occur in the innermost row of perikarya of the INL and close to the vitreous surface. Those of the INL send fine, varicose branches to the ISL forming a network which is somewhat denser at the inner and outer borders of the ISL than in its middle. The indoleamine accumulating terminals do not ramify within the INL in contrast to the catecholamine containing terminals.     

Neuropeptide Y (NPY) immunoreactive neurons in the retina of different species

Neurons displaying Neuropeptide Y (NPY) immunoreactivity were found among amacrine cells in the retina of baboon, pig, cat, pigeon, chicken, frog, trout, carp and goldfish. The immunoreactive cell bodies were located in the middle and the innermost cell rows of the inner nuclear layer with processes forming one, two or three more or less well-defined sublayers in the inner plexiform layer. The location and the density of the sublayers varied with the species investigated. In the frog retina, bipolar-like cell bodies were found in the middle of the inner nuclear layer as well as sparsely occurring ovoid cell bodies in the ganglion cell layer. Like the amacrine cells, these cells emitted processes ramifying in three sublayers in the inner plexiform layer.     

Neuropeptide Y (NPY) immunoreactive neurons in the retina of different species

Summary Neurons displaying Neuropeptide Y (NPY) immunoreactivity were found among amacrine cells in the retina of baboon, pig, cat, pigeon, chicken, frog, trout, carp and goldfish. The immunoreactive cell bodies were located in the middle and the innermost cell rows of the inner nuclear layer with processes forming one, two or three more or less well-defined sublayers in the inner plexiform layer. The location and the density of the sublayers varied with the species investigated. In the frog retina, bipolar-like cell bodies were found in the middle of the inner nuclear layer as well as sparsely occurring ovoid cell bodies in the ganglion cell layer. Like the amacrine cells, these cells emitted processes ramifying in three sublayers in the inner plexiform layer.     

Electron Microscopic Observations on the Structure of the Envelopes of Mature Elementary Bodies and Developmental Reticulate Forms of Chlamydia psittaci

Purified suspensions of Chlamydia psittaci were prepared from L cells. Thin sections of intact elementary bodies and intact developmental reticulate bodies and of their purified envelopes were observed by electron microscopy. In both intact organisms and partially purified envelopes, two membranous structures, each appearing in electron micrographs as two darkly stained layers, were observed. In the elementary body sections, the outer membrane was round, apparently rigid, and was not soluble in 0.5% sodium dodecyl sulfate. The inner layer was irregular in shape and was completely removed by detergent treatment. We interpret these results to indicate that the outer rigid layer of the envelope is the cell wall and the inner layer is the cytoplasmic membrane. When the fragile reticulate body envelopes were similarly studied, the outer cell wall was clearly visible, and some evidence of an inner membrane was seen. After treatment with nucleases and detergent, all evidence of inner or cytoplasmic membrane was removed, but the outer cell wall remained. Thus, it appears that the cell wall of this organism is continuous throughout the growth cycle and that the fragility and lack of rigidity of the reticulate body cell is due to changes in chemical composition or structure of the cell wall.     

Evidence for an S-layer protein pool in the peptidoglycan of Bacillus stearothermophilus.

Intact cells of Bacillus stearothermophilus PV72 revealed, after conventional thin-sectioning procedures, the typical cell wall profile of S-layer-carrying gram-positive eubacteria consisting of a ca. 10-nm-thick peptidoglycan-containing layer and a ca. 10-nm-thick S layer. Cell wall preparations obtained by breaking the cells and removing the cytoplasmic membrane by treatment with Triton X-100 revealed a triple-layer structure, with an additional S layer on the inner surface of the peptidoglycan. This profile is characteristic for cell wall preparations of many S-layer-carrying gram-positive eubacteria. Among several variants of strain PV72 obtained upon single colony isolation, we investigated the variant PV72 86-I, which does not exhibit an inner S layer on isolated cell walls but instead possesses a profile identical to that observed for intact cells. In the course of a controlled mild autolysis of isolated cell walls, S-layer subunits were released from the peptidoglycan of the variant and assembled into an additional S layer on the inner surface of the walls, leading to a three-layer cell wall profile as observed for cell wall preparations of the parent strain. In comparison to conventionally processed bacteria, freeze-substituted cells of strain PV72 and the variant strain revealed in thin sections a ca. 18-nm-wide electron-dense peptidoglycan-containing layer closely associated with the S layer. The demonstration of a pool of S-layer subunits in such a thin peptidoglycan layer in an amount at least sufficient for generating one coherent lattice on the cell surface indicated that the subunits must have occupied much of the free space in the wall fabric of both the parent strain and the variant. It can even be speculated that the rate of synthesis and translation of the S-layer protein is influenced by the packing density of the S-layer subunits in the periplasm of the cell wall delineated by the outer S layer and the cytoplasmic membrane. Our data indicate that the matrix of the rigid wall layer inhibits the assembly of the S-layer subunits which are in transit to the outside.     

The cell envelope of Mycobacterium smegmatis: cytochemistry and architectural implications

In sections stained for localizing both carbohydrates (Thiery's method) and lipids (the O.T.O. method), the cell envelope of Mycobacterium smegmatis appeared to consists of an asymmetric cytoplasmic membrane surrounded by a three-layered cell wall. The outer layer of the cytoplasmic membrane was found to contain more glycoconjugate molecules (probably phosphatidyl inositol mannosides) than the inner one. The cell wall consists of the peptidoglycan (the innermost layer) surrounded by a layer containing both arabinogalactan and mycolates (the electron-dense layer), whereas the outermost layer was unstainable. There is clearly a difficulty in reconciling such a cell wall organization with the models so far proposed.     

The structure of the cuticle and sheath of the infective juvenile of Trichostrongylus colubriformis

The infective third-stage juvenile of Trichostrongylus colubriformis is surrounded by its own cuticle as well as the incompletely moulted cuticle of the second-stage juvenile, which is referred to as the sheath. The sheath comprises an outer epicuticle, an amorphous cortical zone, a fibrous basal zone and an inner electron-dense layer. The basal zone of the sheath consists of three layers of fibres; the fibres are parallel within each layer, but the fibre direction of the middle layer is at an angle to that of the inner and outer layers. The cuticle comprises a complex outer epicuticle, an amorphous cortical zone and a striated basal zone. The lateral alae of the cuticle and the sheath are aligned and overlie the lateral hypodermal cords. The lateral alae of the sheath consist of two wing-like expansions of the cortical zone with associated specializations of the inner electron-dense layer which form a groove. The cuticular lateral alae consist of two tube-like expansions of the cortical zone. The lateral alar complex of the cuticle and the sheath may maximise locomotory efficiency and prevent rotation of the juvenile within the sheath.     

Histochemical and ultrastructural studies on the metacercarial cyst of Echinostoma revolutum (Trematoda).

Histochemical and ultrastructural studies were made on the metacercarial cyst of Echinostoma revolutum obtained from the kidney of experimentally infected Physa and Lymnaea snails. Ultrastructural studies revealed three cyst walls, an outer, middle and inner. The outer wall was more electron-dense than the middle, and contained coarser granules than those found in the middle layer. The inner wall was lamellated and contained membranous whorls. Collagenous fibers presumably of host origin surrounded the outer cyst wall. The outer and middle cyst walls stained identically with all histochemical procedures used. These walls contained acid mucopolysaccharides and glycoprotein, whereas the inner cyst wall contained glycoprotein. All cyst walls stained positively with a variety of protein stains.     

Ultrastructure of the Membrane System in Lactobacillus plantarum

Electron microscopic study of Lactobacillus plantarum revealed mesosomes in different stages of maturation and structural relation with other cell organelles. Small, immature mesosomes were bounded by a prominent electron-dense layer with another extremely faint layer on the outside. This corresponds to the appearance of the cytoplasmic membrane. Large mature mesosomes were surrounded by a triple-layered unit membrane having electron-opaque layers of approximately equal density, suggesting that the composition of the boundary membrane alters during development of this structure. Three-dimensional observations derived from serial sections indicated that mesosomes always maintain a connection between the cytoplasmic membrane and the comparable layers of their boundary. The cytoplasmic membrane also consisted of a triple-layered unit membrane, the innermost layer of which was less electron-opaque and was usually hidden by the relatively dense background of the cytoplasm. The innermost layer of the cytoplasmic membrane was most clearly seen in plasmolyzed cells. Only mature mesosomes made distinct contacts with, or were partially immersed in, the nucleoplasm. The boundary of such mesosomes frequently seemed to be discontinuous, suggesting that the mesosome interior was in direct contact with the nucleoplasm. Mesosomes involved in cross-wall formation at a division plane increased in size and passed through a sequence of positions which led ultimately to an association with the nucleoplasms of the daughter cells. The inner surface of the cell wall was lined by a thin, electron-dense layer whose composition and function are unknown. Under the cultural conditions used, this organism regularly contained a polyphosphate granule.     

Histological study of ibotenic acid-induced modifications of rat retina and their attenuation by diazepam

Summary After 2 h intraocular injections of 19 and 190 nmoles ibotenic acid in the rat retina produced an intensive vacuolization of the inner plexiform layer and cellular alterations, in the inner nuclear layer and ganglion cell layer. These alterations consisted of either cytoplasmic swelling accompanied by clumping of the nuclear chromatin or darkening of the cytoplasm along with nuclear condensation. A week later the retinas were thinner than the controls due to the disappearance of the affected cells. Pre-treatment with diazepam prevents the morphological alterations induced by 19 nmoles ibotenic acid; mainly the swelling, which was completely prevented, while the darkening was reduced drastically, although some vacuolization of the inner plexiform layer is still present.     

Freeze-Etch Study of Pseudomonas aeruginosa: Localization Within the Cell Wall of an Ethylenediaminetetraacetate-Extractable Component

A freeze-etch study of normal cells of Pseudomonas aeruginosa and of cells after incubation with ethylenediaminetetraacetate (EDTA) and tris(hydroxymethyl)aminomethane (Tris) was performed. When cells were freeze-etched without a cryoprotective agent, a smooth outer cell wall layer, which showed a regular array of subunits, and the presence of flagella and pili were observed. These features were not observed in cells freeze-etched after cryoprotection with glycerol. Four fracture surfaces, which resulted from splitting down the center of the outer wall membrane and of the inner cytoplasmic membrane, were revealed in freeze-etched glycerol-protected cells. The murein layer was seen in profile between the outer cell wall membrane and the cytoplasmic membrane. Spherical units and small rods composed of the spherical units were observed in the inner layer of the outer cell wall membrane. These spherical units appeared to be attached to, or embedded in, the inner face of the outer layer of the outer cell wall membrane. These spherical units were removed from cells on exposure to EDTA-Tris, resulting in cells that were osmotically fragile. The spherical units were detected via electron microscopy of negatively stained preparations in the supernatant fluid of cellular suspensions treated with EDTA-Tris. Upon addition of Mg(2+), the spherical units were reaggregated into the inner layer of the outer cell wall membrane and the cells were restored to osmotic stability. The spherical units were shown to consist primarily of protein. These data are thought to represent the first ultrastructural demonstration of reaggregation of cell wall components within a living cell system.