Ultrastructure of the cyst and life cycle of Sarcocystis sp. from wild sheep (Ovis musimon)

Sarcocystis sp. (Eimeriina: Sarcocystidae) is described as a heteroxenous coccidian with domestic dogs as an experimental definitive host and wild sheep (Ovis musimon) as natural intermediate hosts. Mature sarcocysts of this Sarcocystis sp. were examined by transmission electron microscopy. Sarcocysts in various muscle tissues were microscopic, had a thin primary cyst wall and septa and measured 81.0 x 30.5 microns. The cysts were located within muscle cells and were limited by a primary cyst wall (PCW). The cyst surface was highly folded forming densely packed projections. Between the PCW projections the surface of the cyst was marked with pit-like invaginations. The ground substance of the cyst formed a layer at the periphery of the cyst, filled the projections and formed septa which divided the cyst into compartments. Sarcocysts contained numerous bradyzoites that were 15.2 x 3 microns and few metrocytes 11.5 x 3.5 microns. Twelve days after ingesting Sarcocystis sp.-infected wild sheep meat, four dogs began passing sporocysts in their feces: two domestic cats did not pass oocysts or sporocysts after ingesting meat from the same animals. Sporocysts measured 14.8 x 9.9 microns.     

Ultrastructural development of the sarcocyst of Sarcocystis rauschorum (Apicomplexa: Sarcocystidae) in the varying lemming Dicrostonyx richardsoni

The development of the sarcocyst of Sarcocystis rauschorum in its intermediate host was studied. Lemmings were orally administered sporocysts of S. rauschorum obtained from snowy owls (Nyctea scandiaca). Beginning at 9 days postinoculation (DPI) and at various intervals to 84 DPI, skeletal muscle tissue taken from the infected lemmings was examined by electron microscopy. At 9 DPI the sarcocysts contained few metrocytes and the cyst wall was flat. The metrocytes underwent endodyogeny, and within a few days the cyst wall of the rapidly growing sarcocyst developed numerous tubulovesicular invaginations into the electron-dense layer, and the wall had a few irregular infoldings. By 21 DPI, banana-shaped bradyzoites appeared, and by 84 DPI the mature cysts were filled with bradyzoites in groups subdivided by septa and by deep infoldings of the cyst wall. The fine structure of the wall remained simple throughout maturation, with no conspicuous invagination or protrusion. The sarcocyst produced in response to S. rauschorum is unlike those from many species of Sarcocystis, which have complex walls that change markedly as the sarcocysts mature; however, its simple appearance is similar to other species that have rodents as intermediate hosts and raptorial birds as definitive hosts.     

Fine structure and development of Sarcocystis aucheniae in llamas

Isolated cysts of Sarcocystis aucheniae of the llama (Lama glama) were fed to one dog and one cat. Only the dog excreted sporocysts, measuring 13.1-15.7 (15.0 +/- 0.54) X 9.0-11.3 (10.4 +/- 0.36) micron after 11 days for 21 days. A second cat, which had ingested meat of a llama containing macrocysts of S. aucheniae as well as sarcosporidial cysts visible only under a microscope also did not excrete sporocysts. The cysts of S. aucheniae are surrounded by a folded primary cyst wall forming cauliflower-like protrusions into the muscle fibre. The protrusions contain numerous microfilaments. In addition, the primary cyst wall forms numerous tiny vesicles. The parasitized muscle fibre is located in a large cavity within the normal muscle tissue. The cyst wall of S. aucheniae is similarly structured to that of S. gigantea of the sheep.     

Electron Microscope Study of Toxoplasma Cysts in Mouse Brain

SYNOPSIS. Toxoplasma aggregates in sub-acutely and chronically infected mouse brain were studied with special regard to interparasitic relationships, encapsulating wall formation and host-parasite interaction. The individual parasites within a cyst are separated from one another by an opaque substance which also appears as a component of the cyst wall. A second constituent in the wall consists of vesicular and membranous structures which presumably are derived from the endoplasmic reticulum of the host cell. In small cysts, the organising are loosely arranged and maintain the typical crescentic shape whereas, in large cysts, they are tightly packed and polygonal in outline. It is concluded from the data obtained that only the term "cyst" correctly designates these parasitic aggregates.     

Observations on the Fine Structure of the "Cyst Stage" of Besnoitia jellisoni

SYNOPSIS. Light and electron microscope studies of the "cyst" of Besnoitia jellisoni indicate that it consists of an extracellular wall, a large, sometimes multinucleate, host cell, and an intracellular vacuole containing the parasites. The "cyst" wall has fine fibrils and small dense granules embedded in an election-lucid matrix. The wall may be formed from a secretion of the enclosed host cell. The plasma membrane of the host cell is very irregular, being modified into microvillar or pseudopodial extensions. Small vesicles and invaginations of the plasma membrane indicate mioropinocytosis. The one to several large lobular nuclei lie in a thick area of cytoplasm which is filled with rough endoplasmic reticulum and many mitochondria with lamellar cristae. The parasite-containing vacuole is limited by a vacuolar membrane which has many blebs suggesting a transfer of materials into the vacuole.
The "cyst" organisms are crescentic or piriform and are enclosed by a pellicle consisting of outer and inner membranes. Twenty-two subpellicular fibrils extend longitudinally adjacent to the inner membrane from the anterior polar ring to a posterior ring. A micropyle is situated laterally in the pelliole near the level of the nucleus. A conold and several associated paired organelles are present at the anterior end. Microuemes, more abundant in older organisms, are also present in the anterior portion of the parasite. A Golgi apparatus lies adjacent and anterior to the nucleus. One or more mitochondria with saccular cristae, ovoid glycogen bodies, free ribosomes and occasional vacuoles are also present. Organisms within the "cyst" multiply by endodyogeny.     

试验感染水牛体内枯氏住肉孢子虫内生发育的超微结构

通过人工感染试验在电镜下对一种住肉孢子虫在水牛体内的发育进行了研究。结果揭示在血管内皮细胞中的裂殖生殖和在肌细胞内的包囊发育过程与枯氏住肉孢子虫的一致,裂殖体、裂殖子、包囊壁和囊内母细胞、缓殖子的超微结构亦与枯氏住肉孢子虫的相同。首次通过试验感染的发育史研究证明,流行于我国湖南水牛的一种小包囊属于枯氏住肉孢子虫,水牛是黄牛枯氏住肉孢子虫的新宿主。     

Molecular Analysis of the Interactions between Cyst Nematodes and Their Hosts

In order to complete its life cycle, a cyst nematode must stimulate the production of a specialized syncytial feeding site within host root tissues. This process is characterized by major changes in local root morphology, including enlargement of affected nuclei and nucleoli, cell wall degradation, and proliferation of subcellular organelles. At the molecular level very little is known about the processes involved in this host response, but recent evidence suggests that cyst nematodes are able to regulate specific host genes. The host-parasite model system provided by Arabidopsis thaliana and Heterodera schachtii will be fundamental to our future understanding of the formation of syncytia. Molecular biology now offers us the opportunity to study this complex host-parasite interaction in great detail. A better understanding of the host genes regulated by cyst nematodes and the mechanisms by which this regulation is achieved will facilitate the engineering of crop cultivars that possess novel forms of resistance to these adept parasites.     

The fine structure of secretion in Hyalophysa chattoni: formation of the attachment peduncle and the chitinous phoretic cyst wall.

The settling tomite stage of the apostome Hyalophysa chattoni secretes a phoretic cyst wall composed of chitin, mucopolysaccharides, and protein. Within 1 1/2 h after settling, an electron-dense proteinaceous cyst layer (the outer layer) is formed from secretions originating at the base of the kineties and from the thick pellicular layer between the kineties. The inner cyst layer, composed primarily of chitin (acidic and neutral polysaccharides are also present), is secreted across the entire cell surface. Cyst wall formation is completed within 6 h. The fine structure of endocyst secretion resembles stages in the secretion of chitin by fungi, yeasts, and arthropods. A proteinaceous attachment peduncle is secreted to anchor the cell to a shrimp host and is formed by the release of electrondense dense secretory bodies from the cell's ventral surface.     

The Fine Structure of Secretion in Hyalophysa chattoni: Formation of the Attachment Peduncle and the Chitinous Phoretic Cyst Wall

ABSTRACT. The settling tomite stage of the apostome Hyalophysa chattoni secretes a phoretic cyst wall composed of chitin, mucopolysaccharides, and protein. Within 1 1/2 h after settling, an electron-dense proteinaceous cyst layer (the outer layer) is formed from secretions originating at the base of the kineties and from the thick pellicular layer between the kineties. The inner cyst layer, composed primarily of chitin (acidic and neutral polysaccharides are also present), is secreted across the entire cell surface. Cyst wall formation is completed within 6 h. The fine structure of endocyst secretion resembles stages in the secretion of chitin by fungi, yeasts, and arthropods. A proteinaceous attachment peduncle is secreted to anchor the cell to a shrimp host and is formed by the release of electron-dense secretory bodies from the cell's ventral surface.     

The resistance of cysts of Stictodora lari (Trematoda: Heterophyidae) to encapsulation by cells of the fish host

Whereas glass beads are encapsulated by cells within 3 days after implantation into the abdominal cavity of Gambusia affinis, cysts of Stictodora lari in the same site are not encapsulated until 21–23 days after infection. The achievement of encystment in vitro demonstrated that the initial cyst wall is of parasite origin and fluorescent antibody methods showed that it does not mimic fish tissue in composition. Cysts formed in vivo have material, presumably of fish origin, associated with the cyst wall, as do living and fixed in vitro cysts following implantation.It is considered that cysts are not encapsulated for some weeks after infection because they are disguised as host tissue by material of fish origin associated with the cyst wall. An alternative explanation is proposed if the fish material does not have this functional role; the presence of spikes on the initial cyst wall may form an unsatisfactory substrate for the attachment of cells.     

Electron microscopy of stages of Isospora felis of the cat in the mesenteric lymph node of the mouse.

Stages of Isospora felis of the cat in the mesenteric lymph node of the mouse 25 days after oral inoculation with oocysts, have been described at the ultrastructural level. The organisms occurred singly within parasitophorous vacuoles in host cell cytoplasm and were sporozoite-like, having a large crystalloid body up to 5.5 mum in length posterior to the nucleus. The size and appearance of the parasitophorous vacuole varied. Some vacuoles contained numerous, small, electron dense granules about 30 nm in diameter. Because of the aggregation of granules and their arrangement within the parasitophorous vacuole, the impression was sometimes gained by light microscopy that parasites were surrounded by a sheath or cyst wall. However, a cyst wall was not present. In host cells, spherical, membrane-bound bodies with a homogeneous, electron dense core and a maximum diameter of 0.25 mum were filed along the limiting membrane of the parasitophorous vacuole. These extra-intestinal parasites were considered to be waiting stages, with a biological function similar to that of the tissue cyst stage of other general of isosporan coccidia.     

Action lytique des bactéries présentes dans les kystes de Sarcocystis tenella (Sporozoa,Protozoa)

In cysts of Sarcocystis tenella parasitic in the oesophage of sheep, bacteria of the Gram-negative type were found to lyse the limiting membranes of the banana-shaped parasites. In cysts of S. tenella the parasites are enclosed within chamber-like hollows of the ground substance. In old cysts, however, only the peripheral hollows are filled with parasites, whereas those of the midzonal region are empty. There is no explanation for this observation reported by several authors. In the present study we found large numbers of small bacteria (2–2.7 by 0.6–0.8 μm) of the Gram-negative type within the center of the cyst. From this side they were seen to lyse the pellicle of the banana-shaped merozoites. There is no explanation how these bacteria might have penetrated through the muscle tissue into the interior of the cysts, for the parasites at the periphery, the cyst wall and the surrounding host cell were intact. The penetration of the bacteria during preparation can be excluded, too, because the cysts were fixed only seconds after the death of the animals. It might be possible that the bacteria had been present since the beginning of cyst formation.     

UDP-N-acetylglucosamine 4'-epimerase from the intestinal protozoan Giardia intestinalis lacks UDP-glucose 4'-epimerase activity

The protozoan parasite Giardia intestinalis has a simple life cycle consisting of an intestinal trophozoite stage and an environmentally resistant cyst stage. The cyst is formed when a trophozoite encases itself within an external filamentous covering, the cyst wall, which is crucial to the cyst's survival outside of the host. The filaments in the cyst wall consist mainly of a beta (1-3) polymer of N-acetylgalactosamine. Its precursor, UDP-N-acetylgalactosamine, is synthesized from fructose 6-phosphate by a pathway of five inducible enzymes. The fifth, UDP-N-acetylglucosamine 4'-epimerase, epimerizes UDP-N-acetylglucosamine to UDP-N-acetylgalactosamine reversibly. The epimerase of G. intestinalis lacks UDP-glucose/UDP-galactose 4'-epimerase activity and shows characteristic amino acyl residues to allow binding of only the larger UDP-N-acetylhexosamines. While the Giardia epimerase catalyzes the reversible epimerization of UDP-N-acetylglucosamine to UDP-N-acetylgalactosamine, the reverse reaction apparently is favored. The enzyme has a higher Vmax and a smaller Km in this direction. Therefore, an excess of UDP-N-acetylglucosamine is required to drive the reaction towards the synthesis of UDP-N-acetylgalactosamine, when it is needed for cyst wall formation. This forms the ultimate regulatory step in cyst wall biosynthesis.     

A Light Microscopic Comparison of The Cysts of Four Species of Sarcocystis Infecting The Domestic Reindeer (Rangifer Tarandus) in Northern Norway

Fresh preparations of micro-isolated sarcocysts from skeletal and cardiac muscle of 12 reindeer were examined by light microscopy. On the basis of cyst structure and cyst wall structure 4 Sarcocystis spp. could be differentiated. New names have been proposed for 2 previously unnamed Sarcocystis spp. of reindeer, and S. grueneri has been redefined. S. rangiferi n. sp. had macroscopic cysts in skeletal muscle measuring 2106×403 µm. The cyst wall protrusions were finger-like and measured 13.2×6.7 µm. The cysts were surrounded by a layer of fibrillar material. S. tarandi n. sp. had micro- to macroscopic cysts primarily in skeletal muscle, but a few cysts were found in the heart of one animal. In skeletal muscle the cysts measured 999×75µm; in the heart the cysts were shorter and wider. The cyst wall protrusions were fingerlike and measured 9.2×2.2 µm. S. grueneri had micro- to macroscopic cysts in cardiac muscle measuring 581×137 µm. The cyst wall was thin and relatively smooth with no visible protrusions. Sarcocystis sp. had micro- to macroscopic, slender cysts in skeletal muscle measuring 916×64 µm. The cyst wall had tightly packed, short, knob-like protrusions. The cysts of this species were previously classified as cysts of S. grueneri.     

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

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

Identification and characterization of a novel secretory granule calcium-binding protein from the early branching eukaryote Giardia lamblia

Giardia lamblia is a flagellate protozoan that infects humans and other mammals and the most frequently isolated intestinal parasite worldwide. Giardia trophozoites undergo essential biological changes to survive outside the intestine of their host by differentiating into infective cysts. Cyst formation, or encystation, is considered one of the most primitive adaptive responses developed by eukaryotes early in evolution and crucial for the transmission of the parasite among susceptible hosts. During this process, proteins that will assemble into the extracellular cyst wall (CWP1 and CWP2) are transported to the cell surface within encystation-specific secretory vesicles (ESVs) by a developmentally regulated secretory pathway. Cyst wall proteins (CWPs) are maintained as a dense material inside the ESVs, but after exocytosis, they form the fibrillar matrix of the cyst wall. Little is known about the molecular mechanisms involved in granule biogenesis and discharge in Giardia, as well as the assembly of the extracellular wall. In this work, we provide evidences that a novel 54-kDa protein that exclusively localizes to the ESVs is induced during encystation similar to CWPs, proteolytically processed during granule maturation, and able to bind calcium in vitro. The gene encoding this molecule predicts a novel protein (called gGSP for G. lamblia Granule-specific Protein) without homology to any other protein reported in public databases. Nevertheless, it possesses characteristics of calcium-sequestering molecules of higher eukaryotes. Inhibition of gGSP expression abolishes cyst wall formation, suggesting that this secretory granule protein regulates Ca(2+)-dependent degranulation of ESVs during cyst wall formation.     

Intracellular Parasitism and the Problem of Sarcocystosis

The obligatory heterogenous tissue cyst-forming coccidia of the genus Sarcystosis are regarded as an excellent example of the specific coexistence of two organisms, i.e., the host and parasite. These parasitic protozoans are known as causative agents of the chronic, often life-threatening disease, sarcocystosis, which still cannot be effectively controlled. In Sarcocystis, the entire phase of asexual multiplication was transferred to the intermediate host. Of special interest is the parasite's ability to persist in this host at the stage of tissue cyst or sarcocyst. This is a giant meront, in which unidirectional development proceeds starting from a little differentiated metrocyte, through intermediate cells, and towards highly differentiated cyst merozoites (gamonts) unable to further divide. The life span of the sarcocyst depends, to a great extent, on self-regulation within the cyst itself and on relations between the cyst and its immediate environment. A totally new field of research into Sarcocystis was initiated by the discovery that the intracellular parasite damages both cyst harboring and intact muscle cells, apart from the adjacent connective and nervous tissue. The previously unknown cytopathological effects of sarcocysts have been described and characterized. The changes observed within and outside the sarcocysts have been analyzed in terms of general biological processes: proliferation, differentiation, and programmed cell death.     

Mining the Giardia lamblia genome for new cyst wall proteins

The Giardia lamblia cyst wall (CW), which is required for survival outside the host and infection, is a primitive extracellular matrix. Because of the importance of the CW, we queried the Giardia Genome Project Database with the coding sequences of the only two known CW proteins, which are cysteine-rich and contain leucine-rich repeats (LRRs). We identified five new LRR-containing proteins, of which only one (CWP3) is up-regulated during encystation and incorporated into the cyst wall. Sequence comparison with CWP1 and -2 revealed conservation within the LRRs and the 44-amino-acid N-flanking region, although CWP3 is more divergent. Interestingly, all 14 cysteine residues of CWP3 are positionally conserved with CWP1 and -2. During encystation, C-terminal epitope-tagged CWP3 was transported to the wall of water-resistant cysts via the novel regulated secretory pathway in encystation-secretory vesicles (ESVs). Deletion analysis revealed that the four LRRs are each essential to target CWP3 to the ESVs and cyst wall. In a deletion of the most C-terminal region, fewer ESVs were stained in encysting cells, and there was no staining in cysts. In contrast, deletion of the 44 amino acids between the signal sequence and the LRRs or the region just C-terminal to the LRRs only decreased the number of cells with CWP3 targeting to ESVs and cyst wall by approximately 50%. Our studies indicate that virtually every portion of the CWP3 protein is needed for efficient targeting to the regulated secretory pathway and incorporation into the cyst wall. Further, these data demonstrate the power of genomics in combination with rigorous functional analyses to verify annotation.