Ultrastructural description of an unidentified apicomplexan oocyst containing bacteria-like hyperparasites in the gill of Crassostrea rizophorae

Oocysts of an unidentified coccidian are reported in this study to parasitize the gills of the oyster Crassostrea rizophorae (Mollusca, Bivalvia) collected near the city of Recife (Itamaracá Island, 07 degrees 38' 00" S, 34 degrees 48' 06" W), Brazil. Oocysts appeared as light and dense forms, both containing rod-shaped, bacteria-like hyperparasites (BL). Both light and dense oocysts were spherical, 4.3 to 4.7 pm in diameter, but denser oocysts had irregular contours. Both forms consisted of a thick dense wall (approximately 165 nm thick) consisting of 3 layers. The outermost, a dense and irregular layer about 25 nm thick, possessed numerous bead-like structures and some slender conical projections (up to 1.5 microm long). The inner layer of the wall was formed by a dense and homogenous layer about 125 nm thick. Between these 2 layers, a thin light layer about 12 nm thick was present. Uninucleated sporocysts occupied the internal space of the oocyst and contained some rod-shaped BL and mitochondria surrounded by numerous ribosome-like particles. The dense forms of the oocysts showed the same structures described in the lighter forms and appeared to be the final maturation form of the oocysts. Free sporozoites were occasionally observed among oocysts.     

Fine structure of influenza A virus observed by electron cryo-microscopy.

A rapidly frozen vitrified aqueous suspension of influenza A virus was observed by high resolution electron cryomicroscopy. The influenza particles were grouped into small (diameter < 150 nm) spherical particles with well organized interiors, large spherical ones with less internal organization, and filamentous ones. Envelopes of most of the large virus particles were phospholipid bilayers, and the chromatography fraction containing these large particles was largely devoid of viral activity. The envelopes of most of the filamentous and small spherical virus particles, on the other hand, gave a strange contrast which could be ascribed to a combination of a thin outer lipid monolayer and a 7.2 nm thick protein-containing inner layer. These latter particles represented most of the viral activity in the preparation. Densitometric traces of the near in-focus images confirmed these structural differences. Some viral envelope structures apparently intermediate between these two distinct types of membrane were also detected. A structural model of intact biologically active influenza virus particles was formulated from these results, together with computer simulations.     

香菇病毒的研究 Ⅰ.发生在我国的香菇病毒

我国香菇人工栽培历史悠久;在今天纯种人工栽培同样是食用菌生产的主要方向。经查上海栽培香菇“菌种”内生长不正常的菌丝体制剂中有直径为20nm,长度多为100—200nm的类似棒状病毒颗粒。在接种后14—20天,生长缓慢的菌丝体中有直径为28、36、40nm三种不同大小的类似球状病毒颗粒,未见棒状颗粒。在香菇子实体的制剂中同样见到与菌丝体中一样的球状颗粒以及大小为15—16nm×100—300nm的较细的棒状颗粒。病香菇菌褶超薄切片中显示棒状颗粒结晶及球状颗粒的聚集;在液泡中有直径为15—16nm的棒状颗粒。     

Biophysical studies of vesicular stomatitis virus

McCombs, Robert M. (Baylor University College of Medicine, Houston, Tex.), Matilda Benyesh-Melnick, and Jean P. Brunschwig. Biophysical studies of vesicular stomatitis virus: J. Bacteriol. 91:803-812. 1966.-The infectivity and morphology of vesicular stomatitis virus (VSV) were studied after density gradient centrifugation in cesium chloride (CsCI), potassium tartrate (KT), and sucrose. Centrifugation in CsCl revealed two equally infectious bands corresponding to densities of 1.19 and 1.22 g/ml, and a third (density, 1.26 g/ml) band of low infectivity. Two bands (densities of 1.16 and 1.18 g/ml) were observed in the KT gradient, in which the lighter band contained most of the infectivity. Centrifugation in sucrose resulted in a single broad infectious band (density, 1.16 g/ml). The typical rod-shaped VSV particles were found mainly in the lighter bands obtained in CsCl (1.19 g/ml) and KT (1.16 g/ml) and in the single sucrose gradient band (1.16 g/ml). Bent particles equally as infectious as the rod-shaped particles were a constant finding in the CsCl preparations, and were observed mainly in the second band (density, 1.19 g). Numerous strands 15mmu wide were found in the third CsCl (density, 1.26 g/ml) and the second KT (1.18 g/ml) bands. Similar strands could be liberated from VSV particles after treatment with deoxycholate. Internal transverse striations were found to be a regular feature of VSV particles examined with the pseudoreplication negative-staining technique. For crude virus stocks, the physical particle-to-infectivity ratio ranged from 73 to 194. Several morphological similarities between VSV and myxoviruses were observed, including 10 mmu surface projections, pleomorphic morphological forms, and 15 mmu seemingly nucleoprotein strands.     

Electron microscope observations on intracellular virus-like particles associated with the cells of the Lucké renal adenocarcinoma

The common renal adenocarcinoma of the leopard frog was studied in thin sections with the electron microscope. Approximately a third of the tumors examined were found to contain spheroidal bodies of uniform size and distinctive morphology that are believed to be virus particles. These consist of hollow spheres (90 to 100 mmicro) having a thick capsule and a dense inner body (35 to 40 mmicro) that is eccentrically placed within the central cavity (70 to 80 mmicro). Virus particles of this kind occur principally in the cytoplasm but occasionally they are also found in the nucleus and in the extracellular spaces of the tumor. The intranuclear inclusion bodies that are visible with the light microscope are largely comprised of hollow, spherical vesicles with thin limiting membranes. These are embedded in a finely granular matrix. A few of the thin walled vesicles contain a dense inner body like that of the cytoplasmic virus particles. This suggests that they may be immature virus particles. The inclusion bodies are believed to be formed in the course of virus multiplication but they usually contain very few mature virus particles. Bundles of dense filaments and peculiar vacuolar inclusions also occur in the cytoplasm of the tumor cells. These seem to be related in some way to the presence of virus but their origin and significance remain obscure. These findings are discussed in relation to previous work suggesting that the Lucké adenocarcinoma is caused by an organ-specific filtrable agent. It is concluded that the "virus particles" found in electron micrographs of the tumor cells may be the postulated tumor agent. On the other hand, the possibility remains that the particles described here are not those that are causally related to the tumors.     

Some properties of a new virus of Pieris rapae

A new insect virus of Pieris rapae was purified using a chloroform-butanol treatment followed by two differential and sucrose gradient centrifugations. The sedimentation coefficient of the purified virion was approximately 132 S, and it banded at a density of 1.39 g/cm³ in cesium chloride. The virion has a nonenveloped capsid with icosahedral symmetry. Several virions were shown to have a regular hexagonal contour about 25 nm in diameter and to be composed of many capsomeres. Full and empty viral particles, with 12 capsomeres around the periphery of the capsid, were noted. In some particles a small core has been observed which is spherical, about 15 nm in diameter. Both purified virus and partially purified virus preparations from dead, infected larvae gave only one precipitin band with a reaction of identity when tested against the antiserum to partially purified virus. When crude extracts of uninfected larvae and purified virus were tested against antiserum to partially purified virus, the pure virus produced a precipitin band. The band was formed independently and did not join to the band of the uninfected insect producing a typical reaction of nonidentity.     

Vaccinia Virus Intracellular Mature Virions Contain only One Lipid Membrane

Vaccinia virus (VV) morphogenesis commences with the formation of lipid crescents that grow into spherical immature virus (IV) and then infectious intracellular mature virus (IMV) particles. Early studies proposed that the lipid crescents were synthesized de novo and matured into IMV particles that contained a single lipid bilayer (S. Dales and E. H. Mosbach, Virology 35:564–583, 1968), but a more recent study reported that the lipid crescent was derived from membranes of the intermediate compartment (IC) and contained a double lipid bilayer (B. Sodiek et al., J. Cell Biol. 121:521–541, 1993). In the present study, we used high-resolution electron microscopy to reinvestigate the structures of the lipid crescents, IV, and IMV particles in order to determine if they contain one or two membranes. Examination of thin sections of Epon-embedded, VV-infected cells by use of a high-angular-tilt series of single sections, serial-section analysis, and high-resolution digital-image analysis detected only a single, 5-nm-thick lipid bilayer in virus crescents, IV, and IMV particles that is covered by a 8-nm-thick protein coat. In contrast, it was possible to discern tightly apposed cellular membranes, each 5 nm thick, in junctions between cells and in the myelin sheath of Schwann cells around neurons. Serial-section analysis and angular tilt analysis of sections detected no continuity between virus lipid crescents or IV particles and cellular membrane cisternae. Moreover, crescents were found to form at sites remote from IC membranes—namely, within the center of virus factories and within the nucleus—demonstrating that crescent formation can occur independently of IC membranes. These data leave unexplained the mechanism of single-membrane formation, but they have important implications with regard to the mechanism of entry of IMV and extracellular enveloped virus into cells; topologically, a one-to-one membrane fusion suffices for delivery of the IMV core into the cytoplasm. Consistent with this, we have demonstrated previously by confocal microscopy that uncoated virus cores within the cytoplasm lack the IMV surface protein D8L, and we show here that intracellular cores lack the surface protein coat and lipid membrane.     

ELECTRON MICROSCOPE OBSERVATIONS ON INTRACELLULAR VIRUS-LIKE PARTICLES ASSOCIATED WITH THE CELLS OF THE LUCKÉ RENAL ADENOCARCINOMA

The common renal adenocarcinoma of the leopard frog was studied in thin sections with the electron microscope. Approximately a third of the tumors examined were found to contain spheroidal bodies of uniform size and distinctive morphology that are believed to be virus particles. These consist of hollow spheres (90 to 100 mµ) having a thick capsule and a dense inner body (35 to 40 mµ) that is eccentrically placed within the central cavity (70 to 80 mµ). Virus particles of this kind occur principally in the cytoplasm but occasionally they are also found in the nucleus and in the extracellular spaces of the tumor. The intranuclear inclusion bodies that are visible with the light microscope are largely comprised of hollow, spherical vesicles with thin limiting membranes. These are embedded in a finely granular matrix. A few of the thin walled vesicles contain a dense inner body like that of the cytoplasmic virus particles. This suggests that they may be immature virus particles. The inclusion bodies are believed to be formed in the course of virus multiplication but they usually contain very few mature virus particles. Bundles of dense filaments and peculiar vacuolar inclusions also occur in the cytoplasm of the tumor cells. These seem to be related in some way to the presence of virus but their origin and significance remain obscure. These findings are discussed in relation to previous work suggesting that the Lucké adenocarcinoma is caused by an organ-specific filtrable agent. It is concluded that the "virus particles" found in electron micrographs of the tumor cells may be the postulated tumor agent. On the other hand, the possibility remains that the particles described here are not those that are causally related to the tumors.     

OCCURRENCE OF TRI-LAMELLAR BODIES IN ISOLATES OF NOSTOC (CYANOPHYCEAE)1

Tri-lamellar bodies were observed in eight of 29 isolates of Nostoc examined. They appeared identical to the previously described bodies in various species of Anabaena. The bodies consist of three discoid lamellae each ca. 0.3 μm diam and 8 nm thick. The outer lamella (closest to the plasma membrane) is separated from the middle lamella by a 12 nm space whereas the middle and inner lamellae are ca. 8 nm apart. Osmiophilic striations 3 nm wide were generally observed running between the lamellae. Osmiophilic β granules were usually associated with the inner lamella. The bodies were most always located close to the plasma membrane along the longitudinal wall near the junction of the cross and longitudinal walls. In three isolates the bodies located near the cross walls were associated with gas vesicles and possessed a slightly different morphology. These tri-lamellar bodies consisted of three discoid lamellae, each ca. 2 nm thick, ca. 25 nm apart with electron dense material between the inner and middle lamellae. Pores 20 nm diam and ca. 60 nm apart were observed in layer 2 of the cell wall adjacent to the tri-lamellar bodies. These wall pores were also observed in isolates lacking tri-lamellar bodies.     

Comparative Study of Thermal Remodeling of Viruses with Icosahedral and Helical Symmetry

Study of the possibilities of virions and viral proteins modifications and structural remodeling is an important problem of the modern molecular virology. A technique of heat treatment of rod-shaped tobacco mosaic virus that allowed producing structurally modified spherical particles consisting of the virus coat protein was previously developed in our laboratory. These particles possessed unique adsorption and immunogenic properties and were successfully used to develop a new candidate vaccine against rubella virus. Later, the possibility of thermal remodeling of the filamentous virions of potato virus X was demonstrated. The present work reports a comparative study of thermal remodeling of viruses with different structure belonging to various taxonomic groups. The generation of structurally modified spherical particles by the heat treatment of rod-shaped virions with helical symmetry (dolichos enation mosaic virus and barley stripe mosaic virus) has been demonstrated. The dependence of the size of spherical particles derived from dolichos enation mosaic virus on the initial virus concentration was revealed. The process of thermal remodeling of the filamentous virions and virus-like particles of alternanthera mosaic virus was studied. Heat treatment of plant viruses with icosahedral symmetry was shown to cause no morphological changes.     

Connective tissue of rat lung. II: Ultrastructural localization of collagen types III, IV, and VI

We localized collagen types III, IV, and VI in normal rat lung by light and electron immunohistochemistry. Type IV collagen was present in every basement membrane examined and was absent from all other structures. Although types III and VI had a similar distribution, being present in the interstitium of major airways, blood vessels, and alveolar septa, as in other organs, they had different morphologies. Type III collagen formed beaded fibers, 15-20 nm in diameter, whereas type VI collagen formed fine filaments, 5-10 nm in diameter. Both collagen types were found exclusively in the interstitium, often associated with thick (30-35 nm) cross-banded type I collagen fibers. Occasionally, type III fibers and type VI filaments could be found bridging from the interstitium to the adventitial aspect of some basement membranes. Furthermore, the association of collagen type VI with types I and III and basement membranes suggests that type VI may contribute to integration of the various components of the pulmonary extracellular matrix into a functional unit.     

Immunoelectrophoretic analysis of major component proteins in cystic fluid of Taenia solium metacestodes.

When cystic fluid of Taenia solium metacestodes (CF) was filtrated through Sephacryl S-300 Superfine, major proteins were in fractions III and IV. Major protein in fraction III was Band C protein of 150 kDa and that in fraction IV was Band N protein (Choi et al., 1990). When CF was electrophoresed in 0.9% agarose gel and reacted with anti-CF rabbit serum (RACF), two main bands, a long outer and a short inner band, were precipitated, together with 8 minor bands. RACF reacted with fraction III forming the long outer band whereas RACF formed the short inner band with fraction IV in immunoelectrophoresis (IEP). The long outer precipitin band of CF fraction III was similar to antigen B in hydatid fluid (HF) of Oriol et al. (1971), while the short inner band of CF fraction IV was similar to HF antigen 5 of Capron et al. (1967). When HF was reacted with RACF, the short inner band was immunoprecipitated without forming the long outer band. Common antigenicity between CF and HF seemed to exist in fraction IV rather than in fraction III of CF. Patient sera of neurocysticercosis reacted more frequently with fraction III than with fraction IV.     

A peculiar mode of formation of the surface lining layer in the lungs of Salamandra salamandra

The pneumocytes of the larva of Salamandra salamandra contain numerous lamellar bodies and their precursors: electron-dense bodies at various stages of development. Both lamellar bodies and electron-dense bodies occur inside the fluid-filled lung. The former are spherical or bell-shaped and possess concentrically arranged smooth membranes, 8 nm thick; the latter have paracrystalline cores composed of alternately oriented clear and dark striations (3.6–3.9 nm and 2.6–3.6 nm, respectively). On all sides such cores separate membranes, which assume a concentric orientation. No tubular myelin was observed in any phase of the transformation of lamellar bodies and electron-dense bodies into the surface lining layer. Fixation of the lungs of adult individuals with tannic acid-containing fixative visualized the surface lining layer, but not tubular myelin.     

Nonoccluded baculovirus- and filamentous virus-like particles in the spotted cucumber beetle,diabrotica undecimpunctata (coleoptera: chrysomelid)

Nonoccluded baculovirus-and filamentous virus-like particles were found in nuclei of hemocytes or midgut cells of field-collected spotted cucumber beetles. Each type of particle was associated with a different type of virogenic stroma containing various viral components similar to those referred to as capsid, nucleocapsid, viroplasm, and viral envelope in other known baculovirus infections. Nucleocapsids of the virus which occured only in hemocytes were rod-shaped particles approximately 230 nm long and 52 nm wide and were enveloped singly by a trilaminar unit membrane. Enveloped and partly enveloped particles appeared to be released from the nucleus to the cytoplasm by budding through the nuclear envelope acquiring additional membranes. The nucleocapsids of the virus which occurred only in nuclei of midgut cells were filamentous particles with an average diameter of 25 nm and variable length up to 2 μm. Some extremely long particles were bent almost 360° near the middle, resulting in a hairpin-like configuration. The particles were always enveloped singly. No particles budding through the nuclear envelope were observed.     

Ultrastructural Studies of Barley Mesophyll Cella lnfected with Barley Yellow Mosaic Viruses

lexuous filamentous, rod-shaped particles, and laminated, pinwheel inclusions were observed in the mesophyll cells of the barley plants naturally infected with barley yellow mosaic viruses. These virus particles had a length of 480–920 nm and a width of 10–20 nm. In addition, bundles of filamentous structures which consisted of many particles with more 2000 nm in length were found in the leaves of the infected barley plants. The ultrastructural alterations of the infected mesophyll cells were rather conspicuous. The cytoplasmic matrix was lost seriously, and the chloroplast membrane system was destroyed. The cristae and matrix of the mitochondrium were decreased and some of them became vacuoles. The endoplasmic reticulum (ER) expanded teristic membranous network structures occurred in the cytoplasm of infected cells. The virus particles were often associated at one end with ER and with the membranes of network structures. The nucleus, membrane and wall of ceils also had somewhat variation.     

Phage-like infectious virus of the green alga Chlorococcum

A pure culture of the green eukaryotic alga Chlorococcum sp. and a virus infectious for it were isolated. Infection of the culture leads to the destruction of most of the cells, although complete destruction of the culture is not usually observed. The virus particles are similar in morphology to bacteriophage and consist of a hexagonal head 220 nm long and 180 nm wide and a tail which is inside the head in a majority of the particles, but which is on the outside in some. The virus is not adsorbed on the cell membranes, but evidently penetrates into the zoospores through the flagellar channels, where particles of it are regularly found. Destruction of the nucleus and membrane structures of the cytoplasm occurs during growth of the virus, but the chloroplasts and mitochondria are retained. The membranes of infected cells are partially destroyed and the virus particles are released into the medium. The virus is designated as PLVCH (L)--page-like virus of Chlorococcum.     

Physical, chemical and morphologic dimensions of human hepatitis A virus strain CR326 (38578).

CR326 human hepatitis A virus purified by isopycnic banding from infected marmoset sera was shown to consist of 27 nm spherical particles on electron microscopic examination. The particles were identified as hepatitis A virus by tests for infectivity and by specific neutralization of infectivity with convalescent human hepatitis A serum. Also, indentical 27 nm viruses in liver extracts gave specific reactions with hepatitis A antisera when tested by immune electron microscopy. The bouyant density of the virus in CsCl was 1.34 and it was heat (60 degrees), ether and acid stable but was destroyed by heat (100 degrees), formalin (1:4000) and ultraviolet irradiation. Electron microscopic studies of sections of infected marmoset liver showed intracytoplasmic virus particles, usually in vesicles. Presumptive findings for RNA, together with the intracytoplasmic location of the virus, indicated the virus to be of RNA-type. The attributes of the virus indicate it is closely related to the enterovirus family and not to hepatitis B virus.     

Paracrystalline inclusions in various isolates of the blue-green bacteria Nostoc and Anabaena.

A number of different crystalline inclusions were observed in various isolates of Anabaena and Nostoc. Membrane-limited crystalline bodies were observed in 7 of 20 isolates of Anabaena and 19 of 29 isolates of Nostoc. These are spherical, single membrane-limited bodies from 0.6 to 0.1 micron in diameter. In most of the isolates they contained needle-like crystals 20 A in thickness and up to 80 nm in length. In 9 of the isolates the inclusions contained granular and fibrillar material. The number of bodies per cell varied in the different isolates from only a few, observed in many sections, up to 5 in a single section of A. subtropica (B1618). Crystalloids were observed in the cytoplasm of Anabaena sp. (1551), N. calcicola (B382), Nostoc sp. (588), and N. punctiforme (1629). In Anabaena sp. (1551) the roughly cuboidal inclusions 0.6 micron in diameter were composed of 100 A thick osmiophilic striations spaced to produce a 150 A periodicity. In Nostoc sp. (588) the elongate, 0.1 micron by 2.5 micron, crystalloids were composed of 100 A thick osmiophilic striations spaced to produce a 200 A periodicity. N. punctiforme (1629) and N. calciola (B382) contained intrathylakoidal crystalloids which consisted of short curved segments with 100 A thick osmiophilic striations producing a 200 A periodicity. Granular areas were observed in 2 isolates of Anabaena and 5 of Nostoc. These bodies found in various locations in the cells, were interpreted to be elongate structures 0.2 micron thick, 1.2 micron long and about 5 micron in depth. These inclusions were composed of 15 nm diameter granules which in some section planes appeared in rows spaced 20 nm apart. Spherical bodies up to 0.7 micron in diameter and of medium electron density were observed in 4 isolates of Anabaena and 2 of Nostoc. Convoluted inclusions were found in N. calcicola (B382) and Anabaena sp. (1551). These roughly spherical bodies up to 0.8 micron in diameter contain lighter swirled areas.     

Ultrastructural analysis of the freeze-etched spore envelope of the microsporidian,Nosema apis zander

The outer limiting layer of the spore coat ofNosema apis is relatively smooth. The inner limiting layer shows two fractured faces, the concave face carrying many stud-like projections, 120 nm long and 50 nm high, while the convex face carries numerous depressions which are complementary to the projections. In addition, the convex face bears 7 nm particles. In between the outer and inner limiting layers lies the thick homogeneous portion of spore coat which is comprised of numerous microfibres, each 9 nm in diameter. These microfibres resemble those in the freeze-etched host endocuticle. Next to the inner limiting layer of the spore coat are double spore membranes. The convex faces of these spore membranes have a dense population of particles, each 7 nm in diameter.     

Lateral diffusion of redox components in the mitochondrial inner membrane is unaffected by inner membrane folding and matrix density

We report the first lateral diffusion measurements of redox components in normal-sized, matrix-containing, intact mitoplasts (inner membrane-matrix particles). The diffusion measurements were obtained by submicron beam fluorescence recovery after photobleaching measurements of individual, intact, rat liver mitoplasts bathed in different osmolarity media to control the matrix density and the extent of inner membrane folding. The data reveal that neither the extent of mitochondrial matrix density nor the complexity of the inner membrane folding have a significant effect on the mobility of inner membrane redox components. Diffusion coefficients for Complex I (NADH:ubiquinone oxidoreductase), Complex III (ubiquinol: cytochrome c oxidoreductase), Complex IV (cytochrome oxidase), ubiquinone, and phospholipid were found to be effectively invariant with the matrix density and/or membrane folding and essentially the same as values we reported previously for spherical, fused, ultralarge, matrix-free, inner membranes. Diffusion of proton-transporting Complex V (ATP synthase) appeared to be 2-3-fold slower at the greatest matrix density and degree of membrane folding. Consistent with a diffusion-coupled mechanism of electron transport, comparison of electron transport frequencies (productive collisions) with the theoretical, diffusion-controlled, collision frequencies (maximum collisions possible) revealed that there were consistently more calculated than productive collisions for all redox partners. Theoretical analyses of parameters for submicron fluorescence recovery after photobleaching measurements in intact mitoplasts support the finding of highly mobile redox components diffusing at the same rates as determined in conventional fluorescence recovery after photobleaching measurements in fused, ultralarge inner membranes. These findings support the Random Collision Model of Mitochondrial Electron Transport at the level of the intact mitoplast and suggest a similar conclusion for the intact mitochondrion.