The Fine Structure of Lankesterella garnhami

SYNOPSIS. Thin sections of Lankesterella garnhami were examined by electron microscope and the morphology of trophozoites and sporozoites is described. The envelope of the organism consists of a double membrane, which (in the presumed sporozoite) is broken at one point posteriorly by the microphyle. The anterior end possesses characteristic organelles, which include an open apical ring leading to a collar or conoid and to 30 peripheral fibrils, a paired organelle, and numerous "lankesterellonemes." Typical mitochondria and a Golgi apparatus are found in the cytoplasm and a nucleus of heterogeneous nature.     

The Fine Structure of Trophozoites and Gametocytes in Plasmodium coatneyi*

SYNOPSIS. An electron microscope study of Plasmodium coatneyi in the rhesus monkey supplied information on the fine structure of trophozoites, gametocytes and of the host cell. The trophozoites resemble other mammalian malaria parasites. They do not have typical protozoan mitochondria, but instead a concentric double-membraned organelle, which, it is assumed, performs mitochondrial functions. They feed on the host cell by pinocytosis, engulfing droplets of erythrocytes thru invaginations of the plasma membranes at any region of the cell or thru the cytostome. Digestion of hemoglobin takes place in small vesicles pinched off from the food vacuole proper. Gametocytes can be clearly distinguished into macro- and microgametocytes. Macrogametocytes are covered by 2 plasma membranes, the inner one appearing thicker in some places. The cytoplasm is filled with Palade's particles and has numerous vesicles of endoplasmic reticulum and toxonemes. In microgametocytes most of the inner membrane is thickened, the cytoplasm has few Palade's particles and vesicles of the endoplasmic reticulum and does not have toxonemes. Erythrocytes with trophozoites are irregularly scallop-shaped and have elevated points with knob-like protrusions covered by a double membrane. If these protrusions are sticky they might be in part responsible for clumping and arresting the schizonts and segmenters in the capillaries. The host cell contains numerous Maurer's clefts which in some instances are continuous with the membranes of the parasite suggesting that they might originate from them.     

Fine Structural Observations of the Erythrocytic Stages of Plasmodium chabaudi Landau, 1965*

SYNOPSIS. Electron microscopic examination of Plasmodium chabaudi in mouse erythrocytes revealed many characteristics resembling those observed in other mammalian malarial parasites. A double unit membrane surrounds the trophozoite cytoplasm which contains many ribonucleoprotein particles, a limited amount of endoplasmic reticulum and membraned organelles including sausage-shaped vacuoles and multilaminated membraned bodies. More or less circular double membraned vacuoles, possibly cross sections of the sausage-shaped vacuoles, are common. Typical protozoan mitochondria are lacking. The limiting membrane of the merozoites is triple-layered. Paired organelles and small dense bodies are found in the merozoites along with dense granular masses in the nuclei. Trophozoites have cytostomal structures as well as invaginations of the plasma membrane at sites where no cytostomes are evident. Digestion appears to occur in single membrane-bound vesicles which contain one to several pigment grains. P. chabaudi frequently contains multiple food vacuoles and has polymorphism manifested in part by the presence of cytoplasmic extensions and of nuclei with a variety of shapes. Several apparently free forms are noted, often accompanied by a thin rim of host cytoplasm. “Appliqué” forms are common among the trophozoites as are forms in which 2 or more trophozoites are joined together. Finally, alterations in the host cytoplasm resembling the socalled Maurer's clefts are frequent. Ferritin-containing vacuoles also appear in the host cell.     

Fine structure of trophozoites of the gregarine Leidyana ephestiae (Apicomplexa: Eugregarinida) parasitic in Ephestia kuehniella larvae (Lepidoptera)

The ultrastructure of the eugregarine Leidyana ephestiae, parasitic in the larval gut of the flour moth, Ephestia kuehniella, is described. Guts of experimentally infected larvae of E. kuehniella were fixed and sectioned for electron microscope studies of young and mature trophozoites. Young unsegmented trophozoites were small, oval to ovoid, and possessed a simple, globular epimerite. The plasma membrane covering the epimerite region was continuous with the plasma membrane of the protodeutomerite and was in close contact with that of the host cell. Three cortical membranes covered the protodeutomerite region. Folding of the protodeutomeritic epicyte occurred after about 2 days of development of the gregarine. After 3–4 days the body of the trophozoite became differentiated into three segments. A septum was visible between protomerite and deutomerite, but there was nothing similar to this structure between epimerite and protomerite. Fully developed trophozoites showed a large ovoid epimerite containing many mitochondria and vesicles. The epimerite was situated on a short neck filled with fibrils. The cytoplasm of protomerite and deutomerite was rich in amylopectin granules and electron-dense bodies.     

Identification and mitotic partitioning strategies of vacuoles in the unicellular red alga <Emphasis Type="Italic">Cyanidioschyzon merolae</Emphasis>

Cyanidioschyzon merolae is considered as a suitable model system for studies of organelle differentiation, proliferation and partitioning. Here, we have identified and characterized vacuoles in this organism and examined the partitioning of vacuoles using fluorescence and electron microscopy. Vacuoles were stained with the fluorescent aminopeptidase substrate 7-amino-4-chloromethylcoumarin l-arginine amide, acidotrophic dyes quinacrine and LysoTracker, and 4′,6-diamidino-2-phenyl indole, which, at a high concentration, stains polyphosphate. Vacuoles have been shown to be approximately 500 nm in diameter with a mean of around five per interphase cell. The vacuolar H⁺-ATPase inhibitor concanamycin A blocked the accumulation of quinacrine in the vacuoles, suggesting the presence of the enzyme on these membranes. Electron microscopy revealed that the vacuoles were single membrane-bound organelles with an electron-dense substance, often containing a thick layer surrounding the membrane. Immunoelectron microscopy using an anti-vacuolar-H⁺-pyrophosphatase antibody revealed the presence of the enzyme on these membranes. In interphase cells, vacuoles were distributed in the cytoplasm, while in mitotic cells they were localized adjacent to the mitochondria. Filamentous structures were observed between vacuoles and mitochondria. Vacuoles were distributed almost evenly to daughter cells and redistributed in the cytoplasm after cytokinesis. The change in localization of vacuoles also happened in microtubule-disrupted cells. Since no actin protein or filaments have been detected in C. merolae, this result suggests an intrinsic mechanism for the movement of vacuoles that differs from commonly known mechanisms mediated by microtubules and actin filaments.     

Protein import into the photosynthetic organelles of Paulinella chromatophora and its implications for primary plastid endosymbiosis

The rhizarian amoeba Paulinella chromatophora harbors two photosynthetically active organelles of cyanobacterial origin that have been acquired independently of classic primary plastids. Because their acquisition did take place relatively recently, they are expected to provide new insight into the ancient cyanobacterial primary endosymbiosis. During the process of Paulinella endosymbiont-to-organelle transformation, more than 30 genes have been transferred from the organelle to the host nuclear genome via endosymbiotic gene transfer (EGT). The article discusses step-by-step protein import of EGT-derived proteins into Paulinella photosynthetic organelles with the emphasis on the nature of their targeting signals and the final passage of proteins through the inner organelle membrane. The latter most probably involves a simplified Tic translocon composed of Tic21- and Tic32-like proteins as well as a Hsp70-based motor responsible for pulling of imported proteins into the organelle matrix. Our results indicate that although protein translocation across the inner membrane of Paulinella photosynthetic organelles seems to resemble the one in classic primary plastids, the transport through the outer membrane does not. The differences could result from distinct integration pathways of Paulinella photosynthetic organelles and primary plastids with their respective host cells.     

Freeze-fracture study of the site of attachment of Cryptosporidium muris in gastric glands.

The mode and organization of the attachment site of Cryptosporidium muris to gastric glands of stomach were investigated by the freeze-fracture method. Cryptosporidium muris was enveloped by a double membrane, of host plasma membrane origin, which formed the parasitophorous vacuole. The outer membrane of the double membrane was continuous with host plasma membrane, while the inner membrane was connected with the anterior part of the parasite plasma membrane at the annular ring. The density of intramembranous particles (IMP) was severely altered at the above two junctures. The parasitophorous outer membrane showed low IMP-density when compared to the host plasma membrane, although both membranes were continuous at the dense band. The inner membrane had few IMP, whereas the parasite plasma membrane showed numerous IMP, although both membranes were continuous at the annular ring. The size of dense band and annular ring was similar in diameter. The feeder organelle was clearly visible as membrane folds in freeze-fracture and some of them were connected with small vesicles of cytoplasm, indicating that the feeder organelle may play an important role for incorporation of nutrients from the host cell.     

Freeze-Fracture Study of the Site of Attachment of Cryptosporidium muris in Gastric Glands

ABSTRACT The mode and organization of the attachment site of Cryptosporidium muris to gastric glands of stomach were investigated by the freeze-fracture method. Cryptosporidium muris was enveloped by a double membrane, of host plasma membrane origin, which formed the parasitophorous vacuole. The outer membrane of the double membrane was continuous with host plasma membrane, while the inner membrane was connected with the anterior part of the parasite plasma membrane at the annular ring. The density of intramembranous particles (IMP) was severely altered at the above two junctures. The parasitophorous outer membrane showed low IMP-density when compared to the host plasma membrane, although both membranes were continuous at the dense band. The inner membrane had few IMP, whereas the parasite plasma membrane showed numerous IMP, although both membranes were continuous at the annular ring. The size of dense band and annular ring was similar in diameter. The feeder organelle was clearly visible as membrane folds in freeze-fracture and some of them were connected with small vesicles of cytoplasm, indicating that the feeder organelle may play an important role for incorporation of nutrients from the host cell.     

Microscopic Observations on the Filopodia of Entamoeba histolytica*

Living Entamoeba histolytica trophozoites were examined by phase-contrast microscopy. Intact critical point dried trophozoites were examined by transmission electron microscopy at an accelerating voltage of 1000 kV (HVEM) and by scanning electron microscopy (SEM). Half and quarter m? thick sections of epoxy-embedded trophozoites were examined by HVEM. Many of the trophozoites of 2 strains examined had surface filopodia, 1 to over 100 pan in length. The cytoplasm of filopodia was continuous with the cytoplasm and bounded by surface plasmalemma bearing a glycocalyx. Structures called “surface-active lysosomes with trigger,”“dendritic plasmalemmal extensions,” and “extra-amebic vesicles” by previous investigators probably represent portions of filopodia demonstrated in the present study. Filopodia appear to be of frequent normal occurrence in E. histolytica and may function in: (a) endocytosis or pinocytosis; (b) exocytosis; (c) attachment to substratum; (d) penetration of tissue; (e) release of cytotoxic substances; or (f) contact cytolysis of host cells.     

Localization of actin in the electrocyte of Electrophorus electricus L.

Summary The cytoplasm of the electrocyte of Electrophorus electricus possesses a meshwork of 7-nm thick filaments distributed throughout the cell. Observation of stereopairs of transmission electron micrographs shows association of the filaments with the plasma membrane and the membranes of cytoplasmic organelles. Intense fluorescence, indicative of the presence of actin, was observed in the cytoplasm of electrocytes incubated in the presence of NBD-phallacidin or anti-actin antibodies.     

In vitro and in vivo interaction of Entamoeba histolytica Gal/GalNAc lectin with various target cells: an immunocytochemical analysis

The Gal/GalNAc lectin of Entamoeba histolytica trophozoites plays an important role in adhesion. The distribution and final destiny of the lectin during the interaction with host cells are poorly understood. Using monoclonal and polyclonal antibodies against the lectin we studied by immunocytochemistry the in vitro and in vivo interaction of E. histolytica trophozoites with human and hamster hepatocytes. We also analyzed the presence and distribution of the lectin in a mouse model of intestinal amoebiasis. In all cases, trophozoites were highly labeled by anti-lectin antibodies. Cultured human and hamster hepatocytes in contact with, or localized at the vicinity of parasites were also labeled by anti-lectin antibodies. Most of the labeled hepatocytes showed variable degrees of cell damage. Hepatocytes distantly localized from the parasites were also stained with the anti-lectin antibodies. Immunolabeling of tissue sections from different stages of the development of experimental amoebic liver abscess in hamsters showed inflammatory foci containing lectin-labeled trophozoites, hepatocytes, and sinusoidal and inflammatory cells. Lectin-containing hepatocytes had vacuolated cytoplasm with some nuclei with a condensed appearance. Damaged intestinal epithelium also was labeled with anti-lectin antibodies in a mouse model of intestinal amoebiasis. Electron microscopy of axenically cultured trophozoites using gold-labeled monoclonal and polyclonal anti-lectin antibody showed that plasma membrane, vacuole membranes and areas of cell cytosol were labeled. Higher deposits of gold particles in plasma membrane suggestive of cell secretion were observed. Our results demonstrated that Gal/GalNAc lectin was bound and captured by different target cells, and that host cells containing the lectin showed signs of cell damage. The contribution of lectin transfer to host cells in adherence and cell injury remains to be determined.     

ULTRASTRUCTURE OF THE RED TIDE DINOFLAGELLATE GYMNODINIUM BREVE. I. GENERAL DESCRIPTION1,2,3

Gymnodimium breve Davis, an unarmored marine dinoflagellate has a cell covering (theca) composed of four membranes. The inner two membranes represent a vesicular layer and in tangential section, the theca appears composed of polygonal areas. Unusual threat ridges are located in the cingular region between the epi- and hypocone. This osmotically sensitive species is extremely vesiculate with dispersed areas of cytoplasm containing typical eukaryotic organelles as well as other organelles found only in dinoflagellates. The non-vesiculated cytoplasm is continuous in serial sections. The chloroplasts can contain either quasi-radial or parallel lamellae typically consisting of three thylakoids each. The pyrenoid is multiple-stalked and lacks a starch cap. The dinophycean pusule is simple and similar to those found in several unarmored marine species. The nucleus is typically dinophycean but the chromosomes appear to lack nonfibrillar material.     

The Fine Structure of Rhynchocystis pilosa (Sporozoa, Eugregarinida)

SYNOPSIS. The trophozoite of Rhynchocystis pilosa obtained from the seminal vesicles of the earthworm Lumbricus terrestris was studied by light and electron microscopy. The trophozoite's cortical organization is particularly interesting because of its unusual evaginations and associated fibrillar structures. The pellicle is formed by 2 concentric membranes elevated into 60–70 alternating primary and secondary ridges extending posteriad. Numerous long ‘hairs’ or cytopilia originate along the primary ridges and each contains a system of fibrils originating from an underlying longitudinal myoneme. Longitudinal rows of pores lie between adjacent pollicular ridges. Three systems of fibrils lie in the cortex of the trophozoite. A longitudinal myoneme consisting of 12–18 fibrils lies below each primary pellicular ridge. Circular myonemes lie below the pellicle in a parallel array along the length of the organism. Each myoneme consists of 4–8 fibrils structurally similar to those of the longitudinal myonemes. Pairs of fine filaments also lie in the inner pellicular membrane along the apex of each ridge. The trophozoite's anterior end is modified as an attachment organelle consisting of 30–35 delicate pellicular folds which originate at the base of an anterior papilla. The folds extend approximately 15 μ posteriad where they become continuous with the primary pellicular ridges. The nucleus lies in the cytoplasm near the posterior level of the attachment organelle and is surrounded by a double membrane perforated by numerous pores. The cytoplasm contains numerous small vesicles which may be found in dense aggregations. These aggregations often occur in proximity to Golgi complexes and certain membrane-bound bodies. Mitochondria are abundant in the cytoplasm as are large, ovoid paraglycogen bodies. Occasionally layers of granular membranes are arranged parallel to the surface of the paraglycogen bodies but also occur thruout the cytoplasm.     

The plastid in Plasmodium falciparum asexual blood stages: a three-dimensional ultrastructural analysis

The plastid in Plasmodium falciparum asexual stages is a tubular structure measuring about 0.5 micron x 0.15 micron in the merozoite, and 1.6 x 0.35 microns in trophozoites. Each parasite contains a single plastid until this organelle replicates in late schizonts. The plastid always adheres to the (single) mitochondrion, along its whole length in merozoites and early rings, but only at one end in later stages. Regions of the plastid are also closely related to the pigment vacuole, nuclear membrane and endoplasmic reticulum. In merozoites the plastid is anchored to a band of 2-3 subpellicular microtubules. Reconstructions show the plastid wall is characteristically three membranes thick, with regions of additional, complex membranes. These include inner and outer membrane complexes. The inner complex in the interior lumen is probably a rolled invagination of the plastid's inner membrane. The outer complex lies between the outer and middle wall membranes. The interior matrix contains ribosome-like granules and a network of fine branched filaments. Merozoites of P. berghei and P. knowlesi possess plastids similar in structure to those of P. falciparum. A model is proposed for the transfer of membrane lipid from the plastid to other organelles in the parasite.     

Freeze-etching observations of trophozoites of pathogenic Entamoeba histolytica.

Pathogenic Entamoeba histolytica trophozoites were studied by the freeze-etching (FE) technique of electron microscopy. Surface replicas of intact cell membranes were highly convoluted with numerous invaginations, evaginations, and undulations. Sperical depressions and elevations varying from 0.5 mu to 1.0 mu in diameter were commonly present on the external cell membrane and appeared to represent an extracellular secretory mechanism of trophozoites. Cleaved surfaces of amebae exhibited a granular and lumpy cytoplasm in which there were many vesicles and vacuoles that ranged in diameter from 0.2 mu to 9.0 mu. Some vacuoles contained tightly enveloped bacteria, while others contained bacteria and host cytocomponents. Occasional vesicles and vacuoles appeared to be fused to each other. Replicas of FE nucleus were enclosed by double nuclear membranes which were fenestrated by numerous sperical pores measuring approximately 640 A in diameter and spaced at intervals of 650 A. Counts of nuclear pores were possible and indicated 35 pores per square micron on the nuclear envelope. Golgi apparatus, mitochondria and well formed endoplasmic reticulum were absent in FE replicas. This was in agreement with electron microscope observations on thin sections previously reported by other investigators.     

A GREEN DINOFLAGELLATE WITH CHLOROPHYLLS a and b: MORPHOLOGY,FINE STRUCTURE OF THE CHLOROPLAST AND CHLOROPHYLL COMPOSITION1

A green-colored marine unicell has been grown in unialgal culture and its morphology, chloroplast fine structure, and chlorophyll composition investigated. The organism is typical of dinoflagellates in its shape, flagellation, nucleus, mitochondria, and trichocysts. It is similar to Gymnodinium but possesses fine body scales. Chloroplasts and two kinds of vesicles bounded by double membranes, but no organelles obviously identifiable as nuclei or mitochondria, are associated in ribosome-dense cytoplasm separated by a double membrane from the dinophycean cytoplasm. The chloroplasts are unlike any previously reported for dinoflagellates. Each is enclosed by an envelope consisting of a double membrane. Chloroplast lamellae consist of three appressed thylakoids. Interlamellar pyrenoids are present. Pigment analysis reveals chlorophylls a and b but not chlorophyll c. It seems likely that the organism is an undescribed dinoflagellate containing an endosymbiont with chlorophylls a and b and that the reduction of the endosymbiont nucleus and mitochondria has permitted a more initmate symbiosis.     

THE FEEDING MECHANISM OF AVIAN MALARIAL PARASITES

Electron microscope studies of the erythrocytic forms, including gametocytes and asexual schizonts, of the protozoa Plasmodium fallax, P. lophurae, and P. cathemerium, have revealed a "cytostome," a specialized organelle of the pellicular membrane which is active in the ingestion of host cell cytoplasm. In material fixed in glutaraldehyde and postfixed in OsO₄, the cytostome appears in face view as a pore limited by two dense circular membranes and having an inside diameter of approximately 190 mµ. In cross-section, the cytostome is a cavity bounded on each side by two dense segments corresponding to the two dense circles observed in face view; its base consists of a single unit membrane. In the process of feeding, the cytostome cavity enlarges by expansion of its membrane, permitting a large quantity of red cell cytoplasm to come into contact with the cytostome wall. Subsequent digestion of erythrocyte cytoplasm occurs exclusively in food vacuoles which emanate from the cytostome invagination. As digestion progresses, the food vacuoles initially stain more densely and there is a marked build-up of hemozoin granules. In the final stage of digestion, a single membrane surrounds a cluster of residual pigment particles and very little of the original host cell cytoplasm remains. The cytostome in exoerythrocytic stages of P. fallax has been observed only in merozoites and does not seem to play the same role in the feeding mechanism.     

Ultrastructure de la Grégarine Callynthrochlamys phronimae Frenzel; Étude Comparée de son Noyau avec Celui de Thalicola salpae (Frenzel) (Eugregarina)

SYNOPSIS. The structure of the gregarine Callynthrochlamys phronimae has been studied with the electron microscope. The cuticular complex is not different from those previously described in other species of gregarines. It is composed of 2 layers of different thickness delimited by 3 unit membranes and constitutes series of oblique folds at the surface of the deutomerite. Longitudinal rods of dense material surrounded by a slight pellicle are seen under the cuticle. Pinocytic vacuoles are present under the surface of the gregarine. Cytoplasmic organelles include mitochondria, Golgi complexes, endoplasmic reticulum, vacuoles and dense bodies from different sizes. There is a connection between the different features of the cytoplasm in the protomerite and deutomerite and the corresponding cuticular organization.
A characteristic feature of the species is the peculiar differentiation of the nuclear membrane. The nucleus is surrounded by a typical double membrane of which the inner one has a dense fibrillar layer apposed to it. In mature trophozoites, tubular expansions without inner layer arise from the double membrane; the same type of nuclear membrane occurs in another species, Thalicola salpae.     

Shape,size, and distribution of cell structures by 3-D graphics reconstruction and stereology

This report discusses fundamental limitations in attempting to derive cell size, shape, or distribution from the two-dimensional images provided by conventional electron microscopy. Morphometric or stereologic measurement of random thin sections is a convenient way to obtain some information of this type. However, it cannot provide complete, objective information about real size, shape, or connectivity of cells containing irregular or unevenly distributed structures or nonuniform populations of cells. Anisotropic structures require analysis of a complete set of serial sections. The analysis may utilize either stereo, mono, or tilted optical slices, and subsequent integration of this information into a single 3-D computer data set. In this study, we analyze stereo pairs of high-voltage electron micrographs of serial thick sections (0.5 μm) and critical-point-dried whole-cell mounts of rat brain astroglial cell cultures. The Z-axis resolution is increased by digitizing contours at discrete levels within each stereo view. This is accomplished with a new type of stereoscopic contouring device. We calculated area and volume changes accompanying hypo-osmolar swelling and spontaneous reversal of the swelling. (Regulatory Volume Decrease-RVD). An understanding of the mechanism of swelling of astroglial cells is important for improving the treatment of brain injury. The total cell-volume results are comparable with results previously obtained using nonmetabolized, radioactively tagged compounds that diffuse into various cell compartments. Our serial-section and whole-cell data also provide new information about the relative swelling of nucleus, cytoplasm, and individual organelles such as mitochondria. The basic biological problem being approached is whether homeo-stasis of cell function is accompanied by surface area and volume regulation of enzyme-rich membranes and organelles. Conversely, it is proposed to explore the possibility that abnormal organelle areas and volumes are indicators of perturbations of cell division, metabolism, or gene expression.     

Development of Plasmodium chabaudi in Mouse Red Blood Cells: Structural Properties of the Host and Parasite Membranes1

The structure of both the host and parasite membranes during stages in the asexual development of Plasmodium chabaudi in mouse red blood cells is examined by transmission electron microscopy of thin sections and freeze-fracture preparations. The erythrocyte's plasma membrane, the membrane of the parasitophorous vacuole, and the plasma membrane of the parasite exhibit different structural properties in terms of membrane width and the frequency and diameter of the typical intramembrane-particles (IMP) populating the membrane's fracture faces. The difference between the parasitophorous vacuolar membrane and host cell's plasma membrane is remarkable because the vacuolar membrane is formed from an invagination of the erythrocyte's plasma membrane. The vacuolar membrane has significantly reduced frequencies and diameters of IMP's on both faces and reveals a marked temperature response manifesting itself as large IMP-devoid domains emerging on both faces on chilling to 4°C. In contrast, cooling induces only some very small IMP-devoid patches on both faces of the host plasma membrane. Neither of these membranes changes significantly as parasite development progresses. In contrast, the parasite's plasma membrane shows local alterations during its development, forming compaction domains with the nuclear envelope in ca. 30% of the ring-stages and trophozoites. These compaction domains disappear in late uninuclear trophozoites and schizonts. Furthermore, the plasma membrane of the host cell, the vacuolar membrane, and the parasite's plasma membrane do not respond to externally applied Ca²⁺, and their temperature-response remains unaltered during the infection cycle. Thus, modification of these three membranes as a consequence of invasion and development of the parasites, as recently found in the primate malaria caused by P. knowlesi, can be detected neither directly nor indirectly via temperature- and/or Ca²⁺-response in the rodent malaria caused by P. chabaudi.