Pneumocystis carinii pneumonia (PCP) occurs at some time in most patients with acquired immunodeficiency syndrome (AIDS). Trimethoprim/sulfamethoxazole or pentamidine isothionate are the traditional modes of therapy for treatment and prophylaxis of PCP. Unfortunately these drugs are associated with a significant incidence of adverse side effects particularly in patients with AIDS. Toxicity and a growing concern that P. carinii strains are becoming resistant to these compounds is providing the impetus for the search for additional drugs to combat P. carinii. Atovaquone, developed as an antimalarial agent, has activity against a wide range of other organisms, including Toxoplasma sp. and P. carinii, with a lower incidence of adverse reactions during clinical trials. Atovaquone inhibits mitochondrial respiration in P. falciparum and P. carinii. In this study transmission electron microscopy (TEM) was used to observe the effects of atovaquone on P. carinii organisms in short term spinner flask culture.

Spinner flask cultures of human embryonic lung cells were inoculated with P. carinii from infected rat lung.

Cisplatin (cis-diamminedichloroplatinum II) is a broad spectrum anti-cancer agent. Its main limitations are its severe toxic side effects. Cisplatin’s major mechanisms of action include inhibition of DNA synthesis by interstrand and intrastrand crosslinking, inhibition of cytokinesis, and macrophage activation. Cisplatin acts through the activation of the immune system by inducing macrophages to form cytoplasmic extensions which seek out and phagocytose tumor cells. More recently, cisplatin has been shown to activate Kupffer cells (macrophages in the liver) by increasing their number and cytoplasmic extensions. Electron microscopic analysis of cisplatin treated rat liver in vivo was performed to examine these extensions in greater detail and relate these to other changes in the liver cells.

Wistar rats (100-130 g) were given intraperitoneal injections of cisplatin (9 mg/kg in 0.9% sodium chloride) over a five day period. Controls were treated with equal amounts of the vehicle of injection. After 9 days the rats were sacrificed. Segments of liver were fixed in 2% glutaraldehyde in 0.1 M cacodylate buffer (pH 7.3).

Cisplatin, a potent broad spectrum anti-cancer agent, has been proven effective in the treatment of different kinds of cancer, such as bladder, lung, ovarian, head, neck, etc.. Drawbacks of this chemotherapeutic drug are its toxic side-effects, which include severe nausea, vomiting, stomach distention and peptic ulcer.

Taxol is another effective chemotherapeutic agent which is usually used in conjection with cisplatin. It has demonstrated impressive activity in breast, ovarian, lung, and head and neck cancers. The toxic side-effects include nausea, vomiting, nephrotoxicity. The mechanism of these gastrointestinal side-effects are still unclear. Because of the role of NOS and gastrin on stomach smooth muscle contractility and gastroprotection, these were studied in the rats after cisplatin and taxol administration.

Wistar rats(100-150 g) were injected with cisplatin(9 mg/kg) and taxol(20 mg/kg) in five divided dosages over a period of 5 days. Rats were killed one day after the last injection. Rat stomach tissues were fixed in Bouin’s solution and processed for light microscopy.

Spiroplasma, a wall-free prokaryote of the class Mollicutes, is host to a small, naked, single-stranded DNA, isometric virus. Spiroplasma virus SpV4 belongs to the Microviridae family, members of which are non-enveloped, have icosahedral capsids, release progeny through a lytic cycle, and contain circular DNA.

Measurements obtained from negatively stained SpV4 particles revealed a nucleocapsid of 27nm in diameter (figure 1). The three-dimensional structure reported here, obtained from unstained particles suspended in a layer of vitreous ice (figure 2), is in agreement with these earlier results, suggesting a 27nm average distance through the nucleocapsid (figure 3). Unreported in earlier studies is the presence of a 6nm, mushroom-shaped protrusion (made up of a stalk, 2.3nm long and 1.3nm wide, and a globular bud of dimensions ≈4.0×4.0×3.7nm) stemming from an ≈1.5nm deep depression at each of the 3-fold icosahedral axes of the virion. A cross section through the longitudinal axis of one protuberance (figure 4) reveals a cylindrical dimple (≈1.0nm in diameter and 2.3nm deep), originating on the axis of the outer surface of the globular bud domain.

In double-stranded RNA (dsRNA) viruses found in animals, bacteria, and yeast, the genome is transcribed within the structurally intact core of the virion with extraordinary efficiency. How is the genome transcribed within the confines of the intact core particles? How are the nascent mRNA transcripts released through the capsid layers? Critical to our understanding of the structural basis of endogenous transcription is the characterization of the internal organization in terms of genome and transcription enzymes in these viruses. Our recent structural studies on rotavirus, a prototypical dsRNA virus, have begun to provide a better understanding of this process.

Rotaviruses are the major pathogens of infantile gastroenteritis. These are complex and relatively large (1000 Å in diameter), nonenveloped icosahedral viruses. The structure of rotavirus is composed of three concentric capsid protein layers enclosing eleven strands of dsRNA (Fig. 1a). Although previous structural studies using electron cryomicroscopy and computer image processing, have described the overall architecture of the rotavirus, the structural details of internal organization has been elusive.

Varicella-zoster virus (VZV) is one of the human herpes viruses. It causes chickenpox in childhood; after chickenpox the virus remains in a latent state in the dorsal root ganglia of the spinal cord. When the virus reactivates in late adulthood, it causes the disease known as shingles or herpes zoster. The VZV genome includes approximately 70 open-reading frames, but only a few of the gene products have been well characterized.

The VZV glycoproteins include gH, gL, gE and gI. The gH:gL complex is an important viral fusogen. Through laser scanning confocal microscopy, we developed an assay to measure VZV-induced fusion and syncytial formation. VZV also encodes a cell surface Fc receptor, glycoprotein gE. VZV gE has previously been shown to display several features common to nonviral cell surface receptors. Attention focused on the ability of VZV gE to undergo receptor-mediated endocytosis. The transient transfection studies demonstrated by confocal microscopy and intemalization assays that VZV gE was endocytosed when expressed in HeLa cells. Endocytosis of gE was shown to be dependent on clathrin-coated vesicle formation within the cells.

Bacteriophage T7 provides a well defined system to study the assembly of a moderately complex virus. The mature virion consists of a 60 nm icosahedral capsid containing the linear 40 kb genome and an internal protein “core”, and a short tail. The capsid originates as a DNA-free prohead, whose assembly requires both the major capsid protein (gpl0, ∼ 420 copies) and the scaffolding protein (gp9, ∼180 copies). Native proheads incorporate an oligomeric ring of the connector/portal protein at one vertex and the inner core, but prohead-like particles assemble when gp10 and gp9 alone are expressed. In maturation -as with other dsDNA phages - the T7 prohead undergoes a major conformational change, in which the shell structure alters dramatically, and the scaffolding protein is expelled. We have used cryo-EM in combination with other techniques to study this transition and to investigate the interactions between scaffold and surface shell in the T7 prohead.

Highly purified preparations of proheads were generated from the co-expression of head and scaffolding proteins from the cloned gene in E. coli. Tail-less mature capsids as well as DNA-filled heads were prepared from infection of a T7 strain that completely lacks the tail genes.

Nuclear magnetic resonance (NMR) spectroscopy was used to determine the three-dimensional structure of an RNA pseudoknot with a sequence corresponding to the 5' end region of the gene 32 messenger RNA of bacteriophage T2. NMR results show that the pseudoknot contains two coaxial A-form helical stems connected by two loops. One of the loops consists of a single nucleotide, which spans the major groove of the seven base pair helical stem 2. The second loop consists of 7 nucleotides, and spans the minor groove of stem 1. A three-dimensional model of the pseudoknot that is consistent with the NMR data will be presented, and features that are likely to be important for stabilizing the pseudoknot structure will be described.

A combination of NMR and phylogenetic methods were used to characterize the structural features of RNA pseudoknots that are associated with frameshift and readthrough sites within the retroviral gag-pro messenger RNA. The majority of the retroviral frameshift and readthrough sites were found to be followed by nucleotide sequences that have the potential to form pseudoknots with structures that are remarkably similar to that of the bacteriophage T2 gene 32 mRNA.

Ruppia maritima L. is a submerged halophyte which is capable of growing in a wide range of salinities. In a previous study, the effects of varying salinity on the ultrastructure of leaf blade epidermal cells of R. maritima were investigated. The ultrastructural morphology of the cells, including organelle number and distribution, differed at different salinities. However, salinity effects on the endoplasmic reticulum (ER) system could not be assessed since this cytoplasmic component was not readily discernible in material processed in the conventional manner for transmission electron microscopy (TEM). In the present study, selective staining of the ER was achieved, and the response of this membrane system to different salinities ascertained.

Leaf material from plants growing at two salinities (5‰and 20‰) was examined. The zinc iodide-osmium tetroxide (ZIO) impregnation technique was used to stain the ER, and its morphology and distribution determined in leaf blade epidermal cells exposed to the different salinities. Leaf samples were fixed in glutaraldehyde, post fixed in an aqueous mixture of ZIO, dehydrated, and embedded in resin. Controls were processed in exactly the same way, but without ZIO post fixation. Sections with dark gold to purple interference colours were cut and examined unstained with a Phillips 301 TEM.

Soil removal and flooding are serious problems for many urban trees. The main purpose of this paper is to outline some changes which occur in leaves of cherrybark oak (Quercus falcata var. pagodaefolia) and water oak (Quercus nigra) during soil removal and flooding treatments. There has not been any prior documentation of leaf tissue of trees under these conditions. Two-year-old saplings of cherrybark oaks and water oaks were randomly planted in twenty-four experimental plots. Trees were randomly subjected to a combination of soil removal and flooding treatments for one year. Leaf tissues were collected from trees in controlled and treated plots. Tissues were fixed overnight at 4° C in 4% glutaraldehyde buffered with 0.07M cacodylate at pH of 7.2 and post-fixed for one hour in 1% osmium tetroxide in the same buffer. After washing thoroughly in the buffer, tissues were dehydrated in a graded ethanol series and embedded in Eponate 12 resin. Thin sections of 75nm thick were cut with a MT-6000 ultratome and placed on 300 mesh copper grids. Sections were then positive stained with uranyl acetate followed by lead citrate. All examinations of the cell structure were made using a Zeiss EM 10C transmission electron microscope.

Three near isolines of soybean [Glycine max (L.) Merr.], glabrous, normal, and dense, were examined by light, scanning electron, and transmission electron microscopy, as well as by energy dispersive x-ray spectroscopy. This was done in order to determine the morphology and possible functions, in addition to conveying insect resistance, of their non-glandular trichomes.

The uppermost fully expanded trifoliate leaf from greenhouse grown juvenile plants was chosen for examination. Fixed tissue was examined by LM, TEM and SEM as previously described. Some tissue was prepared for energy dispersive spectrometry by fixing in glutaraldehyde but omitting osmium. The tissue was then dehydrated, critical point dried, and coated with carbon. Si x-ray maps were made on the adaxial and abaxial leaf surfaces of all three biotypes.

LMs of Spurr embedded toluidine blue stained semithin sections of normal (Fig. 1) and glabrous (Fig. 2) trichomes appear similar in structure.

P-proteins are group of proteins that express specifically in the sieve elements or parenchymatous cell of phloem tissues. They are filamentous, spherical, tubular, crystalline and amorphous in shape and their ontogenesis varied from species to species. Some of them are plant lectins and their coding genes have been sequenced. In the previous study, a 24 kDa phloem protein was obtained from phloem exudate of Luffa cylindrica fruits and it was immunofluoresciently located in the bicollateral phloem tissues of vascular bundles. In the present study, the intracellular localization of 24 kDa p-protein was examined with colloidal-gold immunolabelling.

Fruits of L. cylindrica were harvested for study. The outer parts of fruits containing exocarp and mesocarp were firstly cut into small cubes in dimensions of 2 x 2 x 6 mm, prefixed in 0.1 M phosphate buffer containing 4 % aldehyde mixture of paraformaldehyde and glutaraldehyde (3:1) for 4 h and washed with distilled water. The washed cubes were then trimmed into 1 mm cubes and postfixed in OSO4 for 2 h, dehydrated in a graded ethanol series and embedded in Spurr's resin.

Various species of endophytic fungi produce a variety of potentially useful new Pharmaceuticals in host tissue of different plants. As an example, the anti-cancer drug, taxol is produced by several species of fungi isolated from various host species.

A Phoma was isolated from the inner bark of Cephalotaxus fortunei growing near Hangzhou, China. After 7-10 days of incubation at 23C cultures were hyphal tipped and transferred to PDA and gamma-irradiated carnation leaves. Cultures which produced bioactive substances were studied further. This Phoma sp. is particularly active against Pythium ultimum. Another endophytic Phoma sp. isolated from Taxus wallachiana also makes antimetabolite compounds. Presumably these endophytic Phoma spp. exist in a symbiotic relationship with various tree species contributing substances that inhibit or destroy invading pathogenic fungi and bacteria.

The fungus produces globose dark brown, thin-walled pycnidia which are semi-immersed in carnation leaf pieces (Fig. 1). Cream-colored conidia (Fig. 2), which average 4.5-5 × 1.75-3 μm are produced in pycnidia with a single ostiole normally obscured by a gelatinous mass (Fig. 3), which is characteristically not evident on old pycnidia (Fig. 4). Conidia are formed enteroblastically from discrete phialidic conidiogonous cells attached directly to the pycnidial wall. Sclerotia (ca. 1.5-2 mm) are also produced on PDA.

Most tissues of a plant are derived from meristems that reside at the tips of the shoot and root. To identify genes that are involved in the organization and in the coordination of cell division and differentation in the meristem, mutations that affect meristem development have been sought. Several mutations that affect meristem development have been characterized in Arabidopsis and maize; however, the angiosperm meristem is small, inconspicuous and difficult to characterize in a non-destructive way. Gametophytes of the fern Ceratopteris richardii have a meristem that is simple, small and conspicuous and is an ideal system for studying meristem development. In this study, the meristems of wild-type and various mutant gametophytes of Ceratopteris are described using cold stage SEM of hydrated gametophytes.

The haploid gametophytes of the fern Ceratopteris are either male or hermaphroditic and are easily distinguished by size and shape as well as by type of sex organ produced. Hermaphrodites form a distinct meristem; newly divided cells of this meristem have only four possible cells fates: they may differentiate as antheridia, the sperm-forming organs; differentiate as archegonia, the egg-forming organs; enlarge to contribute to the growing two-dimensional sheet of cells; or remain within to maintain the meristem (Fig. 1; an=antheridia; ar=archegonia). Male gametophytes lack a meristem and are thus much smaller than hermphrodites. Almost all cells of the male differentiate as antheridia, the sperm-forming organs (Fig. 2). Because the gametophytes of Ceratopteris are small (<3 mm), conspicious, free-living, and haploid, it is very simple to select mutations that affect the sex of the gametophyte as well the meristem.

Bryozoans are among the most common multicellular animals that attach to submerged surfaces in fresh waters. Zooids of the colony capture and digest suspended food particles; they also produce distinctive asexual buds called statoblasts. Measuring 0.35 to nearly 1 mm in diameter, statoblasts have a chitinous exterior and are the only truly hard parts of the bryozoan colony. Like the mastax of rotifers or mouthparts of midge larvae, statoblasts show a variety of taxonomically significant features. Scanning electron microscopy reveals clear patterns in statoblast surface topography that suggest many more species than the 60 or so normally recognized.

Statoblast surface features were first used to distinguish species of the genus Fredericella. The European F. sultana, with a smooth statoblast surface, had been assumed to occur worldwide. SEM micrographs showed the statoblast surface of New World Fredericella to be finely pitted, identical to that of F. indica in India (Figure 1). Another Fredericella species, not yet named, is distinguished only by its highly rugose statoblast surface (Figure 2). Such features are retained through generations of laboratory rearing under various controlled conditions.

Relatively benign or mildly pathogenic strains of bovine Theileria, an intraerythrocytic protozoan parasite, have been reported in many parts of the world including T. mutans in Africa, T. buffeli in Australia, T. orientalis in Britain, and a Theileria sp. from the Southwestern United States (US). Although the Theileria found in the US has not been specifically speciated, it has been referred to as T. mutans-like and as T. orientalis (USA). Although cattle in the US infected with these Theileria spp. usually have a circulating parasitemia of less than 1% and are asymptomatic, recently an animal suffering from severe anemia was admitted to the Large Animal Clinic at Texas A&M University with a circulating parasitemia of over 50%.

Parasites in Giemsa stained blood films were pleomorphic in shape, with some parasites apparently connected to the erythrocyte membrane (Fig. 1) by structures which appeared similar to the bar structure found within eland (Taurotragus oryx) erythrocytes parasitized by T. taurotragi. Examination by TEM revealed that in addition to the main body of the parasite, parasitized erythrocytes contained parasites which possessed cytoplasmic extensions and an interaction between the parasite and the erythrocyte membrane which involved invaginations of the erythrocyte membrane which appeared to be surrounded by parasite cytoplasm (Fig. 2).

The large eggs of Haematopinus phacochoeri are found attached to the long dorsal mane bristles of the diurnal warthog. The eggs are generally exposed to great environmental stresses such as excessive sunlight and temperature as well as the occasional plunge under water or mud during grooming. This investigation was therefore undertaken to determine how the eggs of H. phococoeri are adapted to tolerate the various environmental changes it is constantly exposed to. To our knowledge no other SEM study on the eggs of this species has been performed.

Specimens still attached to the bristles were fixed in 70% ethanol, dehydrated and ultrasonically cleaned. Some eggs were longitudinally sectioned with a razor blade to expose the chorionic layers. The eggs were critical point dried in liquid CO2, mounted and viewed by means of a Leica Stereoscan 420 scanning electron microscope.

The general structure of the shell consists of two layers of chorion, i.e. the endo-and exochorion. These two layers are attached at the pre-formed line of weakness at the operculum and again near the hydropyle. Elsewhere these two layers are completely separated by the respiratory layer.

The tick Dermacentor variabilis returns much of the water and ions from the bloodmeal back into the host as saliva produced in paired salivary glands. These glands are made up of acini and a system of ducts (Fig. 1). Bloodsucking alternates with salivation, but saliva is constantly produced by the acini and expelled into the ducts where it accumulates during bloodsucking. Ducts consist of a layer of epithelial cells, a cuticular coil, and an epicuticular lining of the lumen (Figs. 2, 3). Actin-like filaments run from the coil into the epithelial cells as shown by TEM and CLSM (Figs. 3, 4). These filaments are absent in cytochalasin D treated ducts.

Nitric oxide synthase (=NADPH diaphorase) is present in duct epithelial cells as shown by histochemistry (Fig. 5). We measured nitric oxide (NO) indirectly by the conversion of L-arginine to L-citrulline. NO production and production of saliva is dopamine dependent. NO activates guanylyl cyclase as measured by an RIA for cGMP. The lumen width in dopamine stimulated isolated ducts increased by 19.7% as measured optically. Ducts pre-treated with 10 μM L-NAME, an NOS specific inhibitor, showed no significant increase in width. The addition of 10 μM L-arginine to these ducts partially restored the increase in lumen diameter.

Microwaves of 2450 MHz have been used as a heating procedure in food technology, tissue processing for histological studies (1), in medicine (2) and as a sterilization agent for medical devices (3). Experiments were carried out exposing yeast cells of Candida albicans suspended in growth media to different times of radiation at 750 watts in a commercial microwave oven. Batches of 5 tubes containing the cells suspension were radiated from 15 seconds to 3 minutes and colony counts were done after exposure. Viability ceased totally after one minute radiation. Samples were obtained from unexposed controls and tubes radiated for 15, 30, 45 and 60 seconds, centrifuged and the pellets fixed in 1.5% potasium permanganate. After embbeding in Epon, thin sections were obtained and stained with uranyl acetate. Micrographs were taken with a Zeiss EM-109 electron microscope. Cells showed alteration of internal membranes at early times, and later changes indicative of cytoplasmic coagulation and organelle disorganization were seen. At 15 seconds mitochondrial matrix showed alterations followed by damage of its membranes.

Distribution of glucose and mannose moieties of Acanthamoeba sp. (W4) Philippine isolate was detected using fluorescem isothiocyanate (FITC)- labeled Concannavalin A (Con A) lectin. Green fluorescent patches around the plasma membrane of agglutinated trophozoites (Fig.1) were observed. Isolated Acanthamoeba cyst exhibited strong fluorescence on the cyst wall Brighter fluorescence was detected on the site of adherence between the Acanthamoeba (W4) cysts and trophozoites (Fig. 1,3). These lectin receptors were concentrated at the uroidal region of the trophozoite. The fluorescence, however, was absent in the newly forming hyaline cap (Fig.4). Upon addition of α -methyl-mannoside (0.5 M), Con A binding to sugar moieties in cyst and trophozoites was blocked and no fluorescence was observed.

The binding specificity of Con A-FITC and Acanthamoeba cell surface mannose moieties demonstrate topographical distribution of lectin receptor sites. Ultrastructurally, ferritin-labeled Con A at cell adhesion sites showed clustering of lectin receptors. Occurrence of fluorescence in Naegleria sp. using Con A-FITC has been shown to concentrate at the uroidal region but no fluorescence was seen at the anterior of newly formed pseudopodia.