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.     

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.     

Six new Eimeria species from vespertilionid bats of North America.

Twenty species of bats (Molossidae, Vespertilionidae) were collected from California, New Mexico, Oregon, South Carolina, Utah, and Baja California Norte (Mexico), and 29 of 404 (7%) animals, including Antrozous pallidus, Eptesicus fuscus, Myotis auriculus, Myotis californicus, Myotis ciliolabrum, Myotis evotis, Myotis lucifugus, Myotis thysanodes, Myotis vivesi, Myotis volans, Myotis yumanensis, and Nycticeius humeralis were infected with Eimeria spp., which represent 6 new species. Sporulated oocysts of a new species from A. pallidus are subspheroidal, 24.8 x 21.6 (22-27 x 19-24) microm with a polar granule and a large globular residuum. The oocyst wall is sculptured, with 2 layers, approximately 1.5 thick. Ovoidal sporocysts are 11.5 x 7.8 (9-13 x 7-10) microm, with Stieda body and residuum of many large granules. Sporulated oocysts of a new species from M. californicus are subspheroidal, 20.7 x 18.2 (19-23 x 16-20) microm, with 1-7 tiny polar granules, but without oocyst residuum. The oocyst wall is rough, with 2 layers, approximately 1.4 thick. Ovoidal sporocysts are 11.2 x 7.3 (10-12 x 7-8) microm, with Stieda body and a globular residuum. Sporulated oocysts of a second new species from M. californicus are subspheroidal, 23.1 x 20.7 (20-26 x 19-23) microm, with residuum and 1 polar granule, but a micropyle is absent. The oocyst wall is rough with 2 layers, approximately 1.5 thick. Ovoidal sporocysts are 12.5 x 7.2 (11-14 x 7-8) microm, with a Stieda body and residuum. Sporulated oocysts of a new species from M. ciliolabrum are subspheroidal, 24.9 x 20.1 (18-27 x 17-23) microm, with 1-2 polar granules, but without micropyle and residuum. The oocyst wall is rough with 2 layers, approximately 1.5 thick. Ellipsoidal sporocysts are 12.5 x 9.0 (8-14 x 7-10) microm, with Stieda and substieda bodies and residuum. Sporulated oocysts of a new species from M. evotis are subspheroidal, 21.3 x 18.6 (20-24 x 15-20) microm, with a prominent polar granule, but without micropyle and residuum. The oocyst wall is smooth with 2 layers, approximately 1.0 thick. Ovoidal sporocysts are 12.2 x 8.0 (11-13 x 7.5-9) microm, with Stieda and substieda bodies and residuum. Sporulated oocysts of the new species from N. humeralis are subspheroidal, 22.4 x 18 (21-24 x 17-20) microm, with 1-3 polar granules, but residuum and micropyle are absent. The oocyst wall is lightly sculptured with 2 layers, approximately 1.4 thick. Ovoidal sporocysts are 10.9 x 7.7 (9-12 x 6-8) microm, with Stieda body and residuum. Sporulated oocysts of E. pilarensis Scott and Duszynski, 1997 and those of at least 12 other morphological forms were seen in the other infected bats; these latter forms were seen in too few numbers to be adequately described as new species.     

First record of parasitism in the mangrove oyster Crassostrea rhizophorae (Bivalvia: Ostreidae) at Jaguaribe River estuary--Ceará, Brazil.

Mangrove oysters Crassostrea rhizophorae were sampled monthly in the estuary of Jaguaribe River, on the east coast of Ceará State, Brazil, between August, 2000 and December, 2001, making up 170 individuals. The water temperature varied from 26 to 30 degrees C and salinity from 21 to 42. The animals' size ranged from 3.4 to 7.2 cm height. Macroscopical and histopathological analyses were carried out in the oysters' tissues. The histological exams showed protozoans and metazoans of genera Nematopsis and Tylocephalum, respectively. Nematopsis prevalence varied from 60 to 100% and it was higher in the gills and mantle. The oocysts presented a mean size of 11.5 microm (+/-1.32) length and 9.1 microm (+/-1.06) width (n = 30), up to 3 oocysts/phagocyte having been observed. Several animals presented focal hemocitical reaction. The percentage of Tylocephalum was 1.7%. In spite of the high infection prevalence by Nematopsis, infected animals did not have their reproductive cycle impaired.     

Ultrastructural aspects of hepatic coccidiosis caused by Goussia lusca n. sp. (Apicomplexa: Coccidia) infecting Trisopterus luscus (Gadidae) from the NE Atlantic Ocean

Goussia lusca n. sp. is described from the liver of pouting Trisopterus luscus from the NE Atlantic Ocean in Ibero-Atlantic Portuguese and Spanish waters. Mature oocysts were 31.7 (28.8 to 35.4) microm in diameter. Each oocyst contained 4 ellipsoidal sporocysts arranged in an aleatory position, and measuring approximately 13.7 x 9.2 microm. Each sporocyst contained 2 sporozoites. Ultrastructurally, the sporocyst wall consisted of a dense inner layer 115 nm thick, transversely striated, regularly intercalated by thin grooves with electron-lucent spaces, and separated from the outer layer by a thin, light (electron-lucent) space. The outer layer was multilamellated and consisted of parallel dense bands alternating with light spaces. These lamellae formed filamentous extensions of the wall. The dehiscence suture, a characteristic feature of the genus, was present in the sporocysts. No external clinical signs were observed in the host fish. Parasites observed in the liver tissue were often enveloped in a yellowish-brown matrix, generally known as 'yellow bodies'. Sometimes sporocysts were observed in direct contact with the liver cells. Parasites in degeneration and aggregations of amylopectin granules were frequently observed surrounded by host inflammatory cells. In severe infections, we observed large agglomerations of oocysts encapsulated by layers of concentrically arranged connective tissue forming large granulomas, which caused significant replacement of the host liver parenchyma by the parasite.     

Eimeria from bats of Bolivia: two new species from vespertilionid bats.

Between 1985 and 1987, fecal samples were collected from 71 bats representing 14 species (Desmodontidae, Molossidae, Noctilionidae, Phyllostomidae, Vespertilionidae) from 8 localities in 3 states (Beni, Pando, Santa Cruz) in Bolivia, South America. Of these, 2 black myotid bats (Vespertilionidae), Myotis nigricans, and 1 tent-making bat (Phyllostomidae), Uroderma magnirostrum, had oocysts in their feces that represent undescribed species of Eimeria. The new species from M. nigricans (2/4, 50%) has sporulated oocysts that are subspheroidal, 18.9 x 16.9 (17-23 x 14-20) microm, without a micropyle; oocyst residuum of 6-8 spheroidal globules and 1 highly refractile polar granule are present. The oocyst wall has 2 layers (approximately 1.3 microm thick), with a rough outer layer. Ovoidal sporocysts are 10.1 x 7.4 (7-14 x 5-10) microm, with a Stieda body, substieda body, and a sporocyst residuum. The new eimerian species from U. magnirostrum (1/2, 50%) has sporulated oocysts that are subspheroidal to ellipsoidal, 23.8 x 20.8 (20-26 x 19-24) microm, without micropyle or oocyst residuum, but 1-3 polar granules are present. The oocyst wall has 2 layers (approximately 1.5 microm thick), with a rough, mammilated outer layer. Ovoidal sporocysts are 11.6 x 8.6 (10-12 x 7-10) microm, with a Stieda body, substieda body and a sporocyst residuum.     

Three new species of Eimeria (Apicomplexa: Eimeriidae) from the silvery mole rat Heliophobius argenteocinereus Peters, 1846 (Rodentia: Bathyergidae) from Malawi

Three species of Eimeria Schneider are described from feces of the African bathyergid rodent, Heliophobius argenteocinereus, from Malawi. Oocysts of Eimeria heliophobii n. sp. are broadly ellipsoidal; 27.9 (22-31) x 22.3 (18-24.5) microm with a brownish, heavily pitted oocyst wall, and vacuolar oocyst residuum. Sporocysts are oval, 12.8 (12-14) x 8.4 (8-9) microm with Stieda and substieda bodies. Eimeria nafuko n. sp. has subspherical oocysts; 15.5 (15-16) x 12.8 (12-13) microm with a smooth, colorless oocyst wall. Sporocysts are oval, 9.2 (9-10) x 5.3 (5-6) microm, with a small Stieda body; the substieda body is not visible. Oocysts of Eimeria yamikamiae n. sp. are broadly ellipsoidal to subspherical; 20.8 (19-22) x 17.5 (15.5-19) microm, with slightly yellowish, very faintly pitted oocyst wall. The majority of oocysts contained a single spherical vesicular oocyst residuum and numerous very small granules. Sporocysts are oval, 10.7 (10-11) x 6.8. (6-7) microm, with a dome -like Stieda body and a subspherical to lentil-like substieda body. Typically, infected rodents shed oocysts of more than 1 species of Eimeria.     

Investigation of Nematopsis spp. oocysts in 7 species of bivalves from Chonburi province, Gulf of Thailand

This is the first detailed report of Nematopsis spp. in Thai bivalves. A monthly survey was conducted on 7 species of commercial bivalves from Chonburi province, on the eastern seaboard of Thailand, from November 2000 to November 2001 to investigate the prevalence of the apicomplexan parasite Nematopsis Schneider, 1892. Nematopsis spp. sporozoites were found in the cultivated bivalves Arcuatula arcuatula, Anadara granosa and Perna viridis as well as the locally harvested Paphia undulata. They were not found in Donax faba, Meretrix meretrix or Saccostrea cucullata. Using light microscopy, we were able to identiby 4 oocyst morphotypes of the gregarine Nematopsis spp. Prevalence of Nematopsis spp. during the 13 mo sampling period was highest in A. arcuatula (91.8%; n = 110) and lowest in A. granosa (59.2%; n = 130). The morphology of the oocysts differed between hosts, with an average (x +/- SD) length/width of 16.28 +/- 0.64/12.01 +/- 0.35 microm (n = 50) for A. arcuatula, 16.90 +/- 0.71/12.69 +/- 0.33 microm (n = 50) for A. granosa, 17.61 +/- 0.69/12.72 +/- 0.36 microm (n = 50) for P. viridis, and 11.21 +/- 0.62/8.55 +/- 0.52 microm (n = 50) for P. undulata. Identification of oocysts of these apicomplexan gregarines to species was not attempted. The prevalence of infection in relation to habitat and time of sampling is discussed.     

Observations on the life history and descriptions of coccidia (Apicomplexa) from the western chorus frog,Pseudacris triseriata triseriata,from eastern Nebraska

Two hundred and twenty-four anurans of 6 species (47 adults and 16 tadpoles of Rana blairi, 35 R. catesbeiana, 31 Hyla chrysoscelis, 30 adults and 46 tadpoles of Pseudacris triseriata triseriata, 11 Bufo woodhousii, and 8 Acris crepitans) from Pawnee Lake, Lancaster County, Nebraska, were surveyed for coccidian parasites during March 2001 to May 2002. Of these, 23 of 30 (77%) adults and 4 of 46 (9%) tadpoles of P. t. triseriata shed oocysts of Isospora cogginsi n. sp. Oocysts of I. cogginsi were ovoid, 19.3 x 15.1 (18-23 x 11-20) microm, with a thin, smooth, colorless, single-layered wall, with no micropyle or oocyst residuum. Sporocysts were ovoid, 13.3 x 9.9 (11-15 x 9-13) microm, with a thin, colorless, smooth wall, and Stieda body absent. Sporocyst residuum was present, 5.5 x 5.3 (4-7 x 4-7) microm, consisting of numerous granules. Histological examination of frogs and tadpoles infected with the new species revealed endogenous stages including mature meronts, developing microgamonts, mature microgametes, mature macrogamonts, and young unsporulated oocysts located in the cytoplasm of the epithelial cells of the small intestine. Concurrently, 2 adult P. t. triseriata shed oocysts of Eimeria streckeri. Oocysts of E. streckeri were spherical, 15.7 x 15.4 (14-17 x 14-19) microm, with a thin, smooth, single-layered, colorless wall with an oocyst residuum composed of numerous granules surrounding a large vacuolated area, with a previously undescribed globularlike body present within the vacuole, and no micropyle. Sporocysts were ovoid, 9.1 x 6.1 (7-10 x 5-7) microm, with a thin, colorless, smooth wall with a Stieda body and sporocyst residuum. Our results are the first to document infection of adult and tadpole stages of frogs of the same species with the same species of coccidian, indicating that adult frogs may contaminate breeding ponds with oocysts during their breeding season and infect tadpoles directly by the ingestion of sporulated oocysts.     

Eimeria pavoaegyptica sp. nov. (Apicomplexa: Eimeriidae) in faeces of Indian peacocks, Pavo cristatus Linnaeus, 1758 (Galliformes: Phasianidae) from Egypt

Coprological examination of 15 Indian peacocks, Pavo cristatus, revealed the presence of a coccidium species of the genus Eimeria, which apparently represents a previously undescribed species. Sporulation is exogenous and fully developed oocysts of Eimeria pavoaegyptica sp. nov. are ellipsoidal, with a dimension of 15 (13-16) × 12 (10-12.9) microm and with a shape index of 1.25 (1-1.3). The sporulated oocysts have no micropyle but enclose one large rectangular-shaped polar granule and an oocyst residuum. The oocysts have a distinct two-layered wall, which is ~approximately1.7 microm thick. The outer layer has a smooth texture; it fills ~? of the total thickness and appears bicolored. The sporocysts are boat-shaped, of about 10 (9-11) × 4 (4-4.7) microm; their average shape-index is 2.5 microm with a small pointed Stieda body and a smooth, thin single-layered wall. No substieda body is detected. The sporocysts contain numerous, nearly uniform granular residua. The sporozoites are banana-shaped, 6 × 3 microm and each has two different-sized refractile bodies.     

Life Cycle of Goussia pannonica (Molnar, 1989) (Apicomplexa, Eimeriorina), an Extracytoplasmic Coccidium from the White Bream Blicca bjoerkna

ABSTRACT Life cycle stages of Goussia pannonica from naturally-infected white bream Blicca bjoerkna were studied by light and electron microscopy. Fourteen of the sixteen fish examined were infected, with developmental stages found in all parts of the intestine. Merogonial, gamogonial, and sporogonial stages were localized intracellularly and extracytoplasmically in the microvillous region of enterocytes. They were separated from the gut lumen by closely apposed enterocyte and parasitophorous vacuole membranes. There were two types of extracytoplasmic attachment: 1) monopodial, with a single zone of attachment, and 2) spider-like, with several isolated zones of attachment to the host cell. First-generation merozoites were formed by ectomerogony. Second- or third-generation merozoites were formed by endodyogeny and endopolygeny. Thirty to 50 biflagellated microgametes developed at the periphery of a microgamont. Macrogamonts contained lipid inclusions, amylopectin and dense granules; however, granules comparable to wall-forming bodies type I and II were absent. At the beginning of sporogony, the sporont cytoplasm detached from two layers which subsequently became constituents of the oocyst wall. After the rupture of enterocyte and parasitophorous vacuole membranes, the sporont was released into the water where exogenous sporulation was completed within 48 h. The thin sporocyst wall contained a small longitudinal suture. Sporocyst and oocyts walls were of similar structure.     

A new myxozoan parasite from the Amazonian fish Metynnis argenteus (Teleostei, Characidae): light and electron microscope observations

Myxobolus metynnis n. sp. (Phylum Myxozoa) is described in the connective subcutaneous tissues of the orbicular region of the fish, Metynnis argenteus (Characidae), collected in the lower Amazon River, near the city of Peixe Boi, Pará State, Brazil. Polysporic, histozoic plasmodia were delimited by a double membrane with numerous microvilli on the peripheral cytoplasm. Several life-cycle stages, including mature spores, were observed. An envelope formed by numerous fine and anastomosed microfibrils was observed at the spore surface. The spore body presented an ellipsoidal shape and was about 13.1 microm long, 7.8 microm wide, and 3.9 microm thick. Elongated-pyriform polar capsules were of equal size, measuring 5.2 microm in length, 3.2 microm in width, and possessing a polar filament with 8-9 turns around the longitudinal axis. The binucleated sporoplasm contained a vacuole and numerous sporoplasmosomes. These were circular in cross-section, showing an adherent eccentric, dense structure, with a half-crescent section. Based on the morphological differences and host specificity, we propose that the parasite is a new species named Myxobolus metynnis n. sp.     

Two new species of coccidia (apicomplexa: Eimeriidae) from the bearded false chameleon Chamaeleolis barbatus (Sauria: polychridae) from cinco pesos, Pinar Del Río, Cuba.

Parasitological examination of bearded false chameleons Chamaeleolis barbatus freshly imported from Cuba revealed the presence of 2 species of coccidia that are described as new. Oocysts of Isospora chamaeleolidis n. sp. are spherical to slightly subspherical, 16.1 (13-21) x 15.6 (13-19) microm, with a brownish and bilayered wall approximately 1.0-1.5 microm thick; outer layer markedly pitted. 0.75-1.0 microm thick. One, rarely 2, globular polar granules, 1.5 in diameter are present in the sporulated oocysts. Sporocysts are ellipsoidal, 10.8 (10-13) x 7.8 (7-9) microm, with a smooth, colorless, and unilayered sporocyst wall. Stieda body and substieda bodies are present. A sporocyst residuum is present, consisting of small granules of irregular size scattered among the sporozoites. Oocysts of Eimeria chamaeleolidisbarbati n. sp. are broadly oval, 19.0 (17-21) x 15.7 (15-17) microm, with a bilayered, colorless oocyst wall approximately 0.75 thick; outer layer of oocyst wall is smooth, 0.5 microm thick. One or 2, rarely 4, globular, irregular polar granules, approximately 1.5 microm in diameter, are present in sporulated oocysts. Sporocysts are broadly oval, 7.4 (7-8.5) x 6.1 (5.5-7) microm, with a smooth, colorless, and unilayered sporocyst wall, composed of 2 valves joined by suture; Stieda body and substieda bodies are absent.     

Penetration of the mosquito (Aedes aegypti) midgut wall by the ookinetes of Plasmodium gallinaceum.

We observed Plasmodium gallinaceum ookinetes in both intracellular and intercellular positions in the midgut epithelium of the mosquito Aedes aegypti. After epithelial cell invasion intracellular ookinetes lacked a parasitophorous vacuolar membrane and were surrounded solely by their own pellicle. Thus, the ookinete in the midgut epithelium of the mosquito differs from erythrocytic and hepatic stages in that the parasite in the vertebrate host is surrounded by a vacuole. The midgut epithelial cytoplasm around the apical end of invading ookinetes was replaced by fine granular material deprived of normal organelles. Membranous structure was observed within the fine granular area. Most ookinetes were seen intracellularly on the luminal side and intercellularly on the haemocoel side of the midgut epithelial cells. These observations suggest that the ookinete first enters into the midgut epithelial cell, then exists to the space between the epithelial cells and moves to the basal lamina where the ookinete develops to the oocyst.     

Ultrastructure of tachyzoites, bradyzoites and tissue cysts of Neospora caninum

Dividing tachyzoites of Neospora caninum were 4 x 3 microns and had ultrastructural characteristics typical for the cyst-forming coccidia. Unusual ultrastructural characteristics of fully-formed tachyzoites included no micropores, 8-12 anterior and 4-6 posterior rhoptries, and a few posterior micronemes. Most tachyzoites were located free in the host cell cytoplasm; only a few occurred within a parasitophorous vacuole. Parasite multiplication appeared to be rapid because most organisms were in various stages of endodyogeny. Neural tissue cysts of N. caninum were 24.3 x 19.2 microns and contained 50-200 bradyzoites (7.3 x 1.5 microns), which lacked micropores. The cyst wall was 0.74-1.12 microns thick and consisted of the primary cyst wall (the parasitophorous vacuole membrane) and a thick granular layer with electron-dense vesicles.     

Vavraia lutzomyiae n. sp. (Phylum Microspora) infecting the sandfly Lutzomyia longipalpis (Psychodidae, Phlebotominae), a vector of human visceral leishmaniasis

Vavraia lutzomyiae (Microsporida; Pleistophoridae) is a new species parasitic in the tropical phlebotomine sandfly, Lutzomyia longipalpis (Diptera, Psychodidae, Phlebotominae), a major vector of Leishmania chagasi in Latin America where human visceral leishmaniasis is endemic. Infected larvae and pupae were parasitized in the abdomen, and some adults were parasitized in Malpighian tubules and midgut. The sporogonial plasmodium divided by multiple divisions into up to 64 uninucleate sporoblasts. These stages were surrounded outside the plasmalemma by a thick, amorphous dense coat and transformed into a merontogenetic sporophorous vesicle within which the sporonts developed into sporoblasts. The mature microsporidian spores were broadly ellipsoidal and measured 6.1+/-0.43 x 3.1+/-0.15 microm. The spore wall consisted of a transparent endospore (approximately 100 nm) and a thin electron dense exospore (approximately 30 nm) with the outer limit slightly undulated. Spores contained a polar filament arranged peripherally in a single layer of eight to nine wide anterior coils (approximately 125 nm diameter), and three to four narrow posterior coils (approximately 70 nm diameter). Transverse sections revealed a concentric layer organization with the internal layer surrounded by numerous (up to 25) longitudinal microfibrils. The angle of tilt of the polar filament was about 65-68 degrees.     

The fine structure of the developing oocyst of Pterospora floridiensis (Apicomplexa,Urosporidae)

Developing oocysts of the gregarine Pterospora floridiensis Landers 2001 were examined by transmission electron microscopy. Each oocyst had an outer capsule and an inner capsule that contained 8 sporozoites. In early stages of development the inner capsular wall was separated from the developing sporozoites and residual mass, and was not appressed to the sporozoites. Early stage sporozoites were connected to a residual mass and were filled with endoplasmic reticulum, golgi and numerous developing secretory vesicles. In late stages of oocyst and sporozoite development, the inner capsular wall was closely appressed to the sporozoite surface. The inner capsular wall was ～60-100 nm thick and the outer capsular wall was ～160-320 nm thick. There were no extensions on the outer wall for which the genus was named. Late stage sporozoites had no residual mass connection, were more electron dense, and contained three distinct types of dense secretory structures: 1) small oval/spherical dense vesicles, 2) large (350-400 nm) vesicles near the anterior end, and 3) elongated dense tubular bodies that converged at the apex. Few ultrastructural reports exist of developing gregarine oocysts and sporozoites, and as more studies are completed these morphological characteristics may be important in interpreting molecular phylogenetic analyses.     

Immunodetection of the microvillous cytoskeleton molecules villin and ezrin in the parasitophorous vacuole wall of Cryptosporidium parvum (Protozoa: Apicomplexa)

Microvilli - actin - villin - ezrin - Cryptosporidium parvum The sporozoites and merozoites of the Apicomplexan protozoan Cryptosporidium parvum (C. parvum) invade the apical side of enterocytes and induce the formation of a parasitophorous vacuole which stays in the brush border area and disturbs the distribution of microvilli. The vacuole is separated from the apical cytoplasm of the cell by an electron-dense layer of undetermined composition. In order to characterize the enterocyte cytoskeleton changes that occur during C. parvum invasion and development, we used both confocal immunofluorescence and immunoelectron microscopy to examine at the C.parvum-enterocyte interface the distribution of three components of the microvillous skeleton, actin, villin and ezrin. In infected cells, rhodamine-phalloidin and anti-villin and anti-ezrin antibodies recognized ring-like structures surrounding the developing parasites. By immunoelectron microscopy, both villin and ezrin were detected in the parasitophorous vacuole wall surrounding the luminal and lateral sides of the intracellular parasite. In contrast, anti-beta and anti-gamma actin antibodies showed no significant labelling of the vacuolar wall. These observations indicate that the parasitophorous vacuole wall contains at least two microvillus-derived components, villin and ezrin, as well as a low amount of F-actin. These data suggest that C.parvum infection induces a rearrangement of cytoskeleton molecules at the apical pole of the host cell that are used to build the parasitophorous vacuole.     

Three new species of Isospora schneider, 1881 (Apicomplexa: Eimeriidae) from the double-collared seed eater, Sporophila caerulescens (Passeriformes: Emberizidae), from Eastern Brazil

Three isosporan species are described from the double-collared seedeater, Sporophila caerulescens from Eastern Brazil. Isospora sporophilae n. sp. oocysts spherical to subspherical; oocyst wall bi-layered, smooth, inner layer colorless to pale yellowish, 21.6 x 20.9 (19.20-23.20 x 18.40-22.60) microm, shape-index 1.03 +/- 0.02 (1-1.10), with no micropyle or oocyst residuum. Polar bodies splinter-like or comma-like. Sporocysts ovoidal, 15.2 x 10.6 (17.40-12.80 x 12.60-8.40) microm, shape-index 1.43 +/- 0.14 (1.17-1.81), with knob-like Stieda body and residuum. Large crystalloid body in the center of the sporocyst. Isospora flausinoi n. sp. oocysts spherical to subspherical, oocyst wall bi-layered, smooth, colorless, 17.30 x 16.53 (14-20 x 13.60-20) microm, shape-index 1.05 +/- 0.04 (1-1.21). Micropyle and oocyst residuum absent; presence of a large polar body. Sporocystpiriform, 14.88 x 10.70 (11.80-18 x 8-12.40) microm, shape-index 1.40 +/- 0.18 (1.07-1.77), with smooth, thin, single-layered wall. Sporocyst with rounded Stieda body with no substieda body, and residuum composed of granular material. Isospora teixeirafilhoi n. sp. oocysts spherical to subspherical, oocyst wall bi-layered, smooth, colorless, 17.41 x 16.81 (15.60 - 19.40 x 14.20-18.80) microm. Shape-index 1.04 +/- 0.08 (1-1.12). Micropyle and oocyst residuum absent; presence of a small double-lobuled polar body. Sporocyst ovoid, 11.74 x 8.12 (9-14.20 x 6.20-9.40) microm. Shape-index 1.46 +/- 0.23 (1.06-1.88). Sporocyst with knob-like Stieda body, no sub-Stieda body and residuum composed of granular material.     

Three new species of Isospora Schneider, 1881 (Apicomplexa: Eimeriidae) from the lesser seed-finch, Oryzoborus angolensis (Passeriformes: Emberizidae) from Brazil

Three new coccidian (Apicomplexa: Eimeriidae) species are reported from the lesser seed-finch, Oryzoborus angolensis from Brazil. Sporulated oocysts of Isospora curio n. sp. are spherical to subspherical; 24.6 x 23.6 (22-26 x 22-25) microm, shape-index (SI, length/width) of 1.04 (1.00-1.15). Oocyst wall is bilayerd, approximately 1.5 microm thick, smooth and colourless. Micropyle and oocyst residuum are absent. The sporocysts are ovoid, 13.2 x 10.9 (15-17 x 10-13) microm, SI = 1.56 (1.42-1.71), with a small Stieda body and residuum composed of numerous granules scattered among the sporozoites. Sporozoites are elongated and posses a smooth surface and two distinct refractile bodies. Oocysts of Isospora braziliensis n. sp. are spherical to subspherical, 17.8 x 16.9 (16-19 x 16-18) microm, with a shape-index of 1.06 (1.00-1.12) and a smooth, single-layered wall approximately 1 microm thick. A micropyle, oocyst residuum and polar granules are absent. Sporocysts are ellipsoid and slightly asymmetric, 13.2 x 10.8 (12-14 x 9-12) microm, SI = 1.48 (1.34-1.61). Each sporocyst contains a barely visible Stieda body and a residuum composed numerous of granules. Sporozoites are elongated and each of them contains two distinct refractile bodies. Oocysts of Isospora paranaensis n. sp. are subspherical to broadly ellipsoid 24.3 x 19.8 (22-26 x 18-22) microm, SI = 1.22 (1.15-1.38) with smooth single-layered wall approximately 1.5 microm thick. A micropyle and oocyst residuum are absent, but one distinct ellipsoid polar granule (2.5-3.5 x 1.5-2.5 microm) is present. Sporocyst are ovoid, 15.7 x 10.1 (14-18 x 8-12) microm, SI = 1.46 (1.31-1.72), with distinct Stieda and sub-Stieda bodies. Each sporocyst contains a spherical sporocyst residuum, 4 microm in diameter All described isosporan species represent a possible cause of acute coccidiosis for O. angolensis in captivity.