Mature Cysts Research Articles

Morphology, development and behavior of pneumocystis carinii observed by light-microscopy in nude mice

The present paper describes the morphology, development and behavior of Pneumocystis carinii, especially of the trophozoites in the alveoli of conventionally raised BALB/c nude mice, by using phase-contrast microscopy, paraffin sections and semiultrathin sections embedded in JB-4 plastic. Under phase-contrast microscopy, trophozoites were ameboid in external appearance and 2 to 8 micron in diameter. Usually they have one or more lucid spherical vacuoles and one less lucid nucleus in the cytoplasm. Maturation and independence of intracystic bodies were observed in the developing cysts. The intracystic bodies were polymorphic, i.e. spherical, ameboid or elongated. The paraffin sections using the double staining of P. carinii with Gomori's methenamine silver nitrate (GMS) and Giemsa, showed a small number of mature cysts containing intracystic bodies and a large number of trophozoites within the characteristic honeycombed material in the alveolar spaces. In order to investigate the morphology and parasitizing behavior of the trophozoites and cysts as well as the response of alveolar tissue in more detail, semiultrathin sections of 0.5-2 micron thickness were made from the materials embedded in JB-4 plastic and stained with Giemsa. In lightly infected alveoli, some trophozoites and cysts were found to be closely attached to the Type I alveolar epithelial cells. In heavily infected alveoli, almost all alveoli were filled with trophozoites, cysts, debris of the host cells, and occasionally phagocytic cells (macrophages and neutrophils) containing cysts or trophozoites were found. It was noted that the cysts were very few in number (1%) compared with the number of the trophozoites in nude mice in a 0.4 mm2 area of 2 micron thick lung sections. The host tissue of nude mice in this study was not as strongly affected by the organisms as that in cortisone-treated rats and P. carinii pneumonia patients. In the present study the morphology of P. carinii found in nude mice was not different from that found in rats and in man.

Treatment of hyponatraemic seizures with intravenous 29{middle dot}2% saline

<h3>ABSTRACT</h3> <i>Toxoplasma gondii</i> is a eukaryotic parasite that form latent cyst in the brain of immunocompetent individuals. The latent parasites infection of the immune privileged central nervous system is linked to most complications. With no drug currently available to eliminate the latent cysts in the brain of infected hosts, the consequences of neurons long-term infection are unknown. It has long been known that <i>T. gondii</i> specifically differentiate into a latent form (bradyzoite) in neurons, but how the infected neuron is responding to the infection remain to be elucidated. We have established a new in vitro model resulting in the production of fully mature bradyzoites cysts in brain cells. Using dual, host and parasite, RNA-seq we characterized the dynamics of differentiation of the parasite, revealing the involvement of key pathways in this process. Moreover, we identified how the infected brain cells responded to the parasite infection revealing the drastic changes that take place. We showed that neuronal specific pathways are strongly affected, with synapse signaling being particularly affected, especially glutamatergic synapse. The establishment of this new <i>in vitro</i> model allows to investigate both the dynamics of the parasite differentiation and the specific response of neurons to the long term infection by this parasite.

Growth of Azotobacter vinelandii in dialysed soil medium: Studies upon the life cycle

Azotobacter vinelandii ATCC 12837 cultured in dialysed soil medium with addition of 0.5% glucose showed four distinct morphological cell types: large cells, precyst forms, mature cysts and filterable corpuscles (0.3 micron in diameter). These results indicate that Azotobacter is a bacterium with a complex life cycle under certain culture conditions. Intracellular levels of RNA and poly-beta-hydroxybutyric acid were significantly affected when cells grown in dialysed soil were compared with those obtained after growth on defined medium (N-free). Further studies showed that the chemical composition of filterable corpuscles obtained from dialysed soil medium were different from the composition of normal Azotobacter cells produced in both culture media (dialysed soil and defined media). We suggest that filterable corpuscles represent a stage in the life cycle of Azotobacter in their natural environment.

Changes in Respiratory Activity during Encystment of Azotobacter vinelandii

Vegetative cells of Azotobacter vinelandii were induced to encyst in the presence of β-hydroxybutyrate. Although modal cell volume fluctuated (2·7 to 4 μm3) markedly during exponential growth, the final modal volume attained by mature cysts (1·13 to 1·3 μm3) was relatively constant. Respiratory activity of encysting cultures fell rapidly 3 h after initiation of encystment. This was accompanied by decreased inhibition of respiration by KCN. NADH oxidase activity of membrane preparations fell from approximately 450 to 50 nmol O2 consumed min–1 (mg protein)–1 during differentiation and was also less sensitive to KCN. Difference spectra of cytochromes in membrane preparations revealed a number of changes. Amounts of cytochromes b, c and a 1 all fell during encystment; cytochrome o levels increased whilst cytochrome oxidase d was gradually lost from the membrane during the first 22 h of morphogenesis.

Changes in cell surface proteins and glycoproteins during the encystation of Entamoeba invadens

Changes in cell surface components of axenically grown trophozoites of Entamoeba invadens which occur during encystation were followed. Protein patterns of trophozoites, immature and mature cyst forms, were analyzed by sodium dodecyl sulphate gel electrophoresis. Total protein profiles of trophozoites and cyst forms stained by Coomassie blue gave similar patterns. In contrast, a number of different bands were observed in gels stained with the carbohydrate-specific Schiff's reagent as well as when nitrocellulose blottings were treated with 125I-radiolabelled wheat germ or soybean agglutinins. The most notable differences were bands at 250 and 95–105 kDa present in the cyst forms and absent in the trophozoites, and two bands at 70 and 75 kDa present in the latter and missing in the cysts. Labelling of trophozoites and cyst cell surfaces by iodination with lactoperoxidase revealed a number of protein bands which were exposed on the trophozoite surface and missing in the cysts. Moreover, gel electrophoresis patterns of non-reduced or reduced samples also differed considerably, indicating that a number of proteins are linked by disulphide bonds. This study shows that specific glycoproteins are produced during cyst formation.

Scanning Electron Microscopy of Developmental Stages of Malpighamoeba mellificae Prell in the Honey Bee

ABSTRACTPrimary trophozoites of Malpighamoeba mellificae Prell in the process of penetrating the cell membrane were found among the brush border of epithelial cells of the midgut of Apis mellifera L. They were long and slender with an average diameter of 3.68 μm. Their surfaces had some wrinkles and the cytoplasm contained some vesicles. Secondary trophozoites were found in the lumen of Malpighian tubules. Their size was variable and their shapes were highly irregular; some had pseudopodia. The smooth surface of trophozoites possessed numerous small protrusions and pits. Mature cysts were small and oval‐shaped; they measured from 2.65 times 3.62‐3.4 times 4.6 μm. Their surface was usually smooth but some had wrinkles.

Ultrastructure of cyst formation inBlastocrithidia familiaris inLygaeus pandurus (Hemiptera: Lygaeidae)

The ultrastructural study of encystment ofBlastocrithidia familiaris in the hostLygaeus pandurus is described. Cysts were found attached to flagella of epimastigotes and free in the gut lumen. Most mature cysts were observed in the ileum and rectum, but small numbers were also observed in the ventriculus. Encystment inB. familiaris is characterised by condensation of the nuclear chromatin, kinetoplast and cytoplasm. Morphological changes seen in the plasmalemma include the formation of a dense surface coat around the outer membrane and a dense layer around the inner membrane on the side facing the cytoplasm. The subpellicular microtubules are not seen in mature cysts. A comparison is made with recent ultrastructural studies of encystment ofBlastocrithidia triatomae in vitro and excystment ofB. triatomae in artificially infected hosts. This report is the first known ultrastructural description of encystment in a phytophagous hemipteran and in a naturally infected host.

Morphology of Sarcocystis gigantea in experimentally-infected sheep

The development of the parasite and lesions was studied in 32 sheep killed 10 days to 47 months after inoculation with Sarcocystis gigantea sporocysts from cats. At 21–42 days post-inoculation (d.p.i.), there was a mild encephalitis, but organisms were not seen in the brain. Immature sarcocysts were detected from 40–84 d.p.i. The cyst wall was not measurable by light microscopy at 40 d.p.i., but was 1.5 –2 μm thick at 84 d.p.i. At 119 d.p.i. both immature cysts containing only metrocytes, and mature cysts containing both metrocytes and merozoites, were present. These mature cysts did not have a secondary cyst wall. A mature cyst, 350 μm in length, was found in a sheep killed at 8 1 2 months p.i. At 10 m.p.i. cysts were up to 0.5 mm long and a secondary cyst wall was present. At 47 m.p.i. cysts were 2–5 × 4.5–7.5 mm, and were found only in the muscles of tongue, oesophagus, pharynx and flank.

The Ultrastructure of Meiosis in Woronina pythii (Plasmodiophoromycetes)

During cleavage of cystogenous plasmodia, nuclei of Woronina pythii underwent one meiosis I-like division. The division was centric, closed with polar fenestrations, and characterized by the formation of synaptonemal complexes. Pachytene nuclei exhibited nuclear projections which were heretofore undocumented structural modifications of the nuclear membrane. Hypothetical functions of these structures are proposed. During diakinesis, fragmentation of the nuclear envelope occurred at the poles through the apparent growth of microtubules. Intranuclear membranous vesicles appeared to bud from the inner membrane of the original nuclear envelope during metaphase and anaphase, and coalesced on the surfaces of the separating chromatin masses, thereby forming new daughter nuclear envelopes within the intact original nuclear envelope. Similar descriptions of the spindle apparatus and events relating to nuclear envelope breakdown and reformation have been reported for the mitotic divisions of W. pythii. Nuclei of some mature cysts contained intranuclear cisternae which were continuous with the inner membrane of the nuclear envelope, and these cisternae frequently partitioned the nuclei into two or three separate compartments. The formation of intranuclear cisternae was presumed to result in the eventual fission of nuclei, which may represent functionally the second meiotic division.

Myositis and death in bobwhites, Colinus virginianus (L.), due to hemorrhagic cysts of a haemosporozoan of undetermined taxonomic status.

An epizootic of myositis and death in pen-reared bobwhites occurred at a hunting club in California. The myositis was caused by myriads of elongate protozoan cysts. The cysts were in various stages of development and when mature, contained spherical zoites 1 micron in diameter. Sinuous compartments were present in all cysts. The walls of the compartments were composed of a material similar to the cyst walls. Mature cysts were filled with blood. The histologic and ultrastructural morphology of the parasite revealed the parasite to be a haemosporozoan. The parasite has numerous similarities to both Akiba caulleryi (Mathis and Leger, 1909) and organisms that cause aberrant leucocytozoonosis in other species of birds. Further studies are needed to determine if the parasite is a part of the normal parasite fauna of quail or if it represents a parasite in an aberrant host.

Cytochemical and Electrophoretic Studies on the Cyst Wall of a Ciliate, Histriculus muscorum Kahl

ABSTRACTMThe localization of proteins and polysaccharides in the cyst wall of a ciliate, Histriculus muscorum, was examined by light and electron microscope cytochemistry and by using an enzyme digestion test on thin sections. The endocyst and ectocyst were digested by treatment with pepsin or protease VI. The endocyst was intensely PAS‐positive and alcian blue‐positive. The ectocyst was also PAS‐ and alcian blue‐positive, but the reaction was weaker than that of the endocyst. At the electron microscope level, an intensely positive reaction to methenamine silver was observed in the endocyst and a weak reaction in the ectocyst of the mature cyst. In the ectocyst of the encysting organism, however, the reaction to methenamine silver was more intense than that of the mature cyst. These results demonstrate the possible presence of glycoproteins in the ectocyst and endocyst. The mesocyst was negative to all cytochemical and enzyme digestion tests examined. The cyst wall, isolated by sonication, was analyzed by SDS‐polyacrylamide gel electrophoresis. Two bands, 190 and 140 kilodaltons, were specific for the cyst wall. The 190 kilodalton band was the only PAS‐positive band and its localization in the cyst was was discussed.

Ascosphaera osmophila Sp. Nov., An Australian Spore Cyst Fungus

Ascosphaera osmophila sp. nov. is described. Septa occur often close together and remain intact when the mycelium disintegrates. A fairly good production of mature spore cysts occurs only on media containing 10% sugar or more. A. osmophila lives in association with the mason bee, Chalicodoma mystaceana, and with leafcutter bees, Megachile spp., in Queensland.

Formation of cystlike structures by iron-limited Azotobacter vinelandii strain UW during prolonged storage

Slant or liquid cultures of nonencapsulated Azotobacter vinelandii strain UW survived at least 2 years at 20 °C, whereas parallel cultures died after 6 months storage at 4 °C. Survival was decreased at 20 °C when the growth medium was acidic and it was the least at 4 °C when the growth medium was iron limited. Although strain UW was capable of forming cyclopropyl fatty acids and alkyl resorcinols, these indicators of encystment were neither abundant nor present, respectively, in 1.7-year-old cells. Only lauric acid was more abundant in lipid extracts from old cells than from vegetative cells or mature cysts. The old cells were very fragile and rapidly lost viability upon dilution or dehydration. Electron microscopy revealed the production of fragile cystlike structures in iron-limited medium after 1.7 years at 20 °C. These structures had a distinct exine layer composed of membranelike plates, a poorly defined intine, and an extremely dense central body lacking storage granules or internal membranes. Iron-sufficient cultures produced similar but free central bodies, although cystlike structures were rarely seen.

Development of Sarcocystis fayeri in the Equine

Eight ponies and a horse were inoculated orally with sporocysts of Sarcocystis fayeri from dogs. They were examined for clinical signs of infection and killed 10, 15, 20, 25, 30, 50 (horse), 77, 101, and 156 days after inoculation (DAI). Elevated temperature was observed in three ponies 20 and 26 DAI and anemia was observed in three ponies and the horse 15 to 69 DAI. Schizonts were found in or near cells lining capillaries or arteries of the heart, brain, and kidney 10, 20, and 25 DAI. Immature cysts containing only metrocytes were first found in muscles 50 DAI. Mature intramuscular cysts containing metrocytes and zoites or zoites alone were found 77, 101, and 156 DAI and produced patent infections in dogs fed infected meat.

Morphological evidence of a permeability barrier in urodele testis

The testis of Triturus cristatus is multilobular and each lobe is composed of individual transient cysts in which Sertoli cells develop parallelly to the maturation of germ cells. The junctions between Sertoli cells in immature and mature cysts were studied with lanthanum used as a tracer and surface replicas. Immature cysts appeared as an “open” compartment, as they were penetrated by the tracer, and both sections and replicas showed that Sertoli junctions were differentiating. Mature cysts appeared as a “tight” compartment, as the tracer never reached the lumen and was stopped within inter-Sertoli specializations. Replicas evidenced ridges and grooves, coincident across face transition and continuous over a long distance around the surface of Sertoli membranes. These findings reinforce the idea that a barrier exists, regardless of the general organization of the gonad, and its establishment is related to the differentiation of the haploid male cells.

The fine structure of Acanthamoeba astronyxis, with special emphasis on encystment.

The fine structure of the trophozoite, encysting cells, and the cyst of Acanthamoeba astronyxis has been examined. In the trophic form a microtubule organizing center was associated with a well developed Golgi complex. During encystment the organelles of the amoeba changed considerably. The profiles of rough endoplasmic reticulum elongated and were often arranged in circles of multilayered concentric systems, enclosing mitochondria, the nucleus, or other inclusions. The mitochondria showed a tendency toward elongation and constriction. One or two nucleolus-like bodies appeared in the nucleus. Lipid droplets increased considerably in amount and were distributed individually or as aggregates. The mature cyst was star-shaped and surrounded by an almost circular exocyst and an endocyst that was closely apposed to the cell membrane. Both walls differed in their thickness and granulation. The exocyst was continuous over the entire cyst, while the endocyst was interrupted by gaps, ostioles, in the region of the rays. Within the ostioles was a bell-shaped structure, the operculum. The latter was composed of a granular material comparable in electron density to that of the endocyst.

Evolution of porencephalic cysts from intraparenchymal hemorrhage in neonates: sonographic evidence.

Ninety-four low-birth-weight neonates were screened consecutively with real-time sonography for evidence of cerebroventricular hemorrhage. Among them, 13 were found to have intraparenchymal hemorrhage beyond the area of the germinal matrix. Weekly follow-up sonograms revealed progression to porencephaly in every surviving infant. Four different sonographic patterns were observed during the evolution from the original hemorrhage to the porencephalic cyst. The size of the mature porencephalic cyst correlated directly with the size of the intraparenchymal hemorrhage. After ventriculoperitoneal shunting, the porencephalic cysts became smaller or remain unchanged. One small cyst was no longer identified after successful shunting, but reappeared with shunt failure.

Changes in Two Acid Hydrolase Levels During Cyst Differentiation of a Ciliate, Histriculus muscorum1

ABSTRACTCellular levels of protein and two acid hydrolases, acid phosphatase (EC 3.1.3.2) and acid proteinase, were followed during cyst differentiation, arbitrarily divided into five stages, in the ciliate Histriculus muscorum Kahl. Extracellular enzyme activities were also measured. Protein content decreased gradually during cyst differentiation. In mature cysts the protein content was ca. 60% that of stationary phase organisms. The activities of both acid hydrolases remained unchanged during stage 1 and then decreased gradually; acid proteinase decreased more rapidly. Both enzymes remained slightly active in the mature cysts. The acid proteinase activity of stage 1 was reduced by cycloheximide treatment at time zero, whereas the enzyme was no longer sensitive to the inhibitor when treated at 1.5 h (late stage 1) after the first wash with encysting medium. Acid phosphatase activity was insensitive to the inhibitor. Extracellular release of acid phosphatase increased linearly at least until stage 5, although the extracellular release of acid proteinase was not detected. Cycloheximide blocked the extracellular release of acid phosphatase after stage 1. These results suggest that de novo synthesis of acid proteinase occurs during stage 1 and that lysosomes may play an important role during early stages of cyst differentiation.

Growth phase dependent alterations in the surface coat of Acanthamoeba castellanii

Application of ruthenium red, cationized ferritin and concanavalin A to exponentially growing trophozoites reveals on their plasma membrane negatively charged surface coat bearing sugar residues. In the coat of trophozoites from advanced stationary growth phase no sugar residues can be visualized. In mature cysts the external layer of their wall is negatively charged, however, on their protoplast surface no terminals reacting with the 2 polycations, or with concanavalin A can be revealed, even though the penetration of the reagents has been ensured by enzymatic impairing of the cyst wall. The results are confronted with the known facts concerning alterations of physiological properties of plasma membrane occurring during the life cycle of Acanthamoeba.

Further Light Microscopic Studies on Morphology and Development of Pneumocystis carinii

In order to add more advance in light microscopic investigation of P. carinii, phase contrast microscopy partly followed by wet giemsa stain and semiultrathin section of the lungs embedded in JB-4 plastic were studied. In phase contrast microscopy, small and large sized trophozoites of P. carinii were clearly recognized. Although movement of trophozoite was not found, rhythmic movement of intracystic bodies with filopodia was often seen in mature cyst. Those living organisms were then directly stained with Giemsa by infiltrating under the coverglass. Thus the organism could be investigated both in unstained and stained conditions. It is noticed with interest that 8 intracystic bodies seem to fill up the cavity of cyst when cell division is completed, then they liberate and become independent into spherical bodies, followed by banana-shaped or amoeboid forms with motility. An emphasis was done that semiultrathin section made from JB-4 plastic embedded lungs was quite useful for investigation of P. carinii infection. Several sizes of mononuclear thin-walked trophozoites, mature and immature cysts, and empty cysts were more clearly distinguished than any other light microscopical method ever reported.