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Transmission electron microscopy of intracellular sporozoites of Eimeria vermiformis (Apicomplexa, Eucoccidiida) in the mouse.

Sporozoites of Eimeria vermiformis from the mouse were first seen in the epithelial cells of villus tips and the crypts of Lieberkühn four hours after inoculation (HAI). They were always within a parasitophorous vacuole. By 12 HAI, most were in crypt epithelial cells between the basement membrane and host cell nucleus. The sporozoites in the villus tips had 26 subpellicular microtubules, two polar rings, two preconoidal rings, two refractile bodies surrounded by amylopectin-like granules, a lamellar Golgi apparatus, numerous micronemes, and rhoptries. The sporozoites in the crypt cells had fewer amylopectin-like granules, micronemes, and rhoptries. A nucleolus was visible, as were pieces broken off from the posterior refractile body. Later, the sporozoites folded over to become U-shaped; the infolded membranes fused; and then the inner membranes disappeared so that spherical meronts were formed. Folding sporozoites were first seen 16 HAI and persisted until 52 HAI.

Animals

Hepatozoon canis: the prevalence of antibodies and gametocytes in dogs in Israel.

A survey for the prevalence of antibodies to Hepatozoon canis and for intraneutrophilic H. canis gametocytes in the peripheral blood neutrophils of dogs in Israel showed that 33.1% were seropositive, while only 1% of the dogs sampled had detectable parasites in their blood smears. Exposure to H. canis is widespread but it appears that most infected dogs undergo a subclinical infection and only a small proportion develop clinical disease.

Animals

The development of Hepatozoon sipedon sp. nov. (Apicomplexa: Adeleina: Hepatozoidae) in its natural host, the Northern water snake (Nerodia sipedon sipedon), in the culicine vectors Culex pipiens and C. territans, and in an intermediate host, the Northern leopard frog (Rana pipiens).

The life cycle of Hepatozoon sipedon sp. nov. was studied in two snake species, the Northern water snake and the Eastern garter snake, in its mosquito hosts Culex pipiens and C. territans, and in the Northern leopard frog. Gametogenesis, fertilization and sporogony occurred within fat body cells in the haemocoel of mosquitoes that had fed on infected water snakes. Mature oocysts averaging 263 microns in diameter and containing more than 500 sporocysts were observed in mosquitoes 28 days post-feeding. Each sporocyst enclosed eight sporozoites. Dizoic cysts were found in the liver of frogs that had been fed infected mosquitoes seven days previously. Two rounds of merogony in various internal organs and intraerythrocytic gamonts were observed in snakes that had been fed frogs which had been orally inoculated with infected mosquitoes. Developmental stages were not seen in snakes that were fed infected mosquitoes directly. A comparison of this life cycle with those described for other Hepatozoon species infecting snakes is presented with reference to the different modes of transmission featured by these parasites.

Animals

Cyclospora cayetanensis: first imported infections in Germany.

Over the last decade increasing numbers of enteritis cases have been attributed to infection with a new coccidian species that was named Cyclospora cayetanensis in 1993. Diarrhea caused by this agent is clinically indistinguishable from cryptosporidiosis, isosporiasis and microsporidiosis, but Cyclospora infections are often very prolonged (up to 15 weeks) and may cause severe weight loss. Diagnosis of infection is important because, in contrast to diarrhea caused by Cryptosporidium and microsporidia, treatment with co-trimoxazole is effective. Here we report the cases of two female patients, aged 70 and 58 years old, respectively, who suffered from severe, prolonged diarrhea after a vacation in Singapore, Java and Bali. Routine microbiological laboratory diagnostics were unable to demonstrate the presence of known enteropathogenic bacteria. Modified Ziehl-Neelsen staining of fecal smears revealed oocysts of Cyclospora cayetanensis, and treatment of the patients with co-trimoxazole was promptly effective. We would like to increase awareness of the possibility of Cyclospora infections in patients with prolonged diarrhea who have travelled to endemic areas.

Aged

The fate of Hepatozoon species naturally infecting Florida black racers and watersnakes in potential mosquito and soft tick vectors, and histological evidence of pathogenicity in unnatural host species.

Haemogregarine parasites, derived from the Florida snakes Coluber constrictor and Nerodia fasciata and ingested by Aedes aegypti, completed sporogony within the hemocoeles of nearly all fed mosquitoes in 14-18 days, and produced oocysts typical of Hepatozoon. However, mortalities and morbidity were high in the Culex which had fed on the Coluber. Oocysts were not found in any Ornithodoros turicata (Argasidae) which fed upon either snake host, but many sections of fed ticks had gametocyte-like cells within the gut lumen. Most lizards, Anolis carolinensis and Anolis sagrei, infected per os with oocysts derived from both snake species developed infections. Infections in the lizards were largely confined to hepatic schizonts with few parasites found in erythrocytes. Unlike naturally infected snake hosts, Hepatozoon schizonts in livers of lizards were often either surrounded by an unidentified dark pigment or heavily infiltrated with mononuclear inflammatory cells.

Animals

Demonstration of common and stage-specific anti-Hepatozoon mocassini antibodies in experimentally infected unnatural lizard hosts.

This study examines the immunological response to Hepatozoon mocassini in lizards. Three lizard species were infected experimentally with H. mocassini. Baseline and post-infection (PI) sera were assayed for anti-H. mocassini meront and gametocyte antibody by immunohistochemistry and IFA. Seroconversion occurred at 38 d PI with endpoint IFA titers of 1:64. Antisera from non-parasitaemic and parisitaemic lizards exhibited similar affinities for merozite and gametocyte antigens. Antibody specific for the membranes of gametocyte-infected erythrocytes was demonstrated exclusively in parasitaemic lizards. The results demonstrate that lizards infected with snake haemogregarines mount an antibody response with common and stage-specific components.

Aedes

Characterization of the clinical and anatomical pathological changes associated with Hepatozoon mocassini infections in unnatural reptilian hosts.

Laboratory-reared Aedes aegypti mosquitoes were employed in the successful transmission of Hepatozoon mocassini from a cotton-mouth moccasin (Agkistrodon piscivorus leucostoma) to 3 lizard species (Sceloporus undulatus, Eumeces obsoletus and Sceloporus poinsetti). Marked to severe lethargy and anorexia developed in the S. undulatus, E. obsoletus and S. poinsetti at 15, 38, and 96 days postinfection (PI), respectively. All 3 lizards developed a leukocytosis and had increased plasma aspartate aminotransferase activity (AST) by 14 days PI. Multifocal random hepatocellular necrosis and intrahepatic aggregates of heterophils centered on mature H. mocassini meronts were demonstrated in all 3 lizards. The pulmonary interstitium was multifocally thickened by aggregates of heterophils centered on meronts. No comparable clinical or anatomical pathological changes were demonstrated in naturally infected snakes. The results of this study suggest that H. mocassini is capable of inducing necrotizing inflammatory by lesions in unnatural reptilian hosts.

Aedes

Concurrent Hepatozoon canis and Toxoplasma gondii infections in a dog.

A female 1-year-old dog died suddenly and was submitted for necropsy. Numerous grey-tan-colored nodules were seen in the lungs, brain and lymph nodes. Microscopically, the predominant lesion was necrosis associated with numerous Toxoplasma gondii tachyzoites. The parasites reacted positively with anti-T. gondii serum in an immunohistochemical test. Schizonts of Hepatozoon canis were seen in sections of lymph nodes and the spleen.

Animals

Ultrastructure of Hepatozoon canis in the dog.

The ultrastructure of several stages of Hepatozoon canis found in dogs with clinically diagnosed infections was determined using transmission electron microscopy. Stages were found in skeletal muscle tissue that corresponded to the 'onion skin' cyst stage, as described at the light microscopic level, and were composed of an electron-transparent material that appeared to radiate from a central core. Larger cysts, walled off by fibroblasts, contained a transformed host cell located centrally within them. The parasitic stage within these transformed cells contained numerous organelles including mitochondria and Golgi apparatus, and was singularly nucleated. In cardiac muscle, a meront was observed which produced merozoites by ectomerogony. Adjacent to this meront was a granuloma containing merozoites within mononuclear phagocytes which may either serve as a reservoir of parasites for reinfection of the host or differentiate into the circulating gamont stage of the parasite. Gamonts were found within parasitophorous vacuoles inside circulating neutrophils. They had a condensed cytoplasmic appearance and were extremely electron dense with respect to other observed parasite stages. In vitro cultivation of parasitized neutrophils resulted in the appearance of a stage of the parasite with altered ultrastructure compared with gamonts found in circulating neutrophils. This stage was judged to be a possible gamete stage of the parasite.

Animals

Diarrhea associated with Cyclospora Sp. in Bangladesh.

A spherical acid-fast organism measuring approximately 10 microns in diameter (Cyclospora sp.) has recently been implicated in diarrheal diseases in many parts of the world. We detected this organism in the stools of six Bangladeshi patients with diarrhea. Four patients had chronic diarrhea and two had acute diarrhea at the time of presentation. This is the first report of infection with this organism in the indigenous population from this region.

Adult

Ultrastructural studies on oocysts, sporulation and sporozoites of Schellackia cf. agamae from the intestine of the starred lizard Agama stellio.

Young intracellular oocysts of Schellackia cf. agamae in the gut epithelium of agama stellio were bound by several fine membranes. Later-stage oocysts and sporoblasts in the lamina propria were intercellular and were bound by a thin but firm tri-layered wall. Oocysts had a large central refractile body which, during sporulation, sent extensions into the developing sporozoites. Sporozoites escaped into the gut tissue, leaving a large oocyst residuum with the remains of a refractile body. These sporozoites invaded a variety of connective tissue cells, endothelial cells and circulatory leucocytes in the lamina propria. Sporozoites caused lysis of the host cell cytoplasm at their perimeter and multiple sporozoite infections led to complete degradation of the host cell.

Animals

Identification of intracellular stages of Cyclospora species by light microscopy of thick sections using hematoxylin.

Cyclospora sp is a recently identified coccidia responsible for enteric infection in humans. Most reports have failed to detect this parasite in intestinal biopsies by light microscopy, although the different stages have been ultrastructurally described in jejunum enterocytes. Very recently, some investigators have reported the detection by light microscopy of parasitophorous vacuoles in intestinal biopsies; however, only transmission electron microscopy (TEM) could clearly identify the parasitic stages. To improve the histological diagnosis without using TEM, we have tested different staining methods in biopsies obtained from a patient infected by the human immunodeficiency virus who was shedding Cyclospora oocysts. Hematoxylin stain alone for 15 minutes on 5 micrometer-thick sections of duodenal biopsies was found to be the most efficient method for observing different stages of the parasite. In particular, the banana-shaped merozoites were visualized and appeared very similar to the human coccidia Isopora belli. This simple technique may be useful in diagnosing Cyclospora infection.

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