Ultrastructural aspects of Cystoisospora belli (syn. Isospora belli) in continuous cell lines.

Cystoisospora belli is an opportunistic protozoan that causes human cystoisosporiasis, an infection characterized by diarrhea, steatorrhea, abdominal pain, fever, and weight loss. The lack of animal models susceptible to C. belli, and the difficulty in obtaining clinical samples with fair amounts of oocysts have limited the research pertaining to the basic biology of this parasite. This study aimed to describe the ultrastructure of endogenous stages of C. belli in Monkey Rhesus Kidney Cells (MK2) and Human Ileocecal Adenocarcinoma cells (HCT-8). Zoites of C. belli exhibited typical morphological features of coccidia, which included a trilaminar pellicle, an apical complex formed by a conoid, polar rings, rhoptries, and micronemes, in addition to dense granules and the endoplasmic reticulum. No crystalloid body was observed but various lipid and amylopectin granules were usually present in the cytoplasm of zoites. We observed a tendency of the endoplasmic reticulum of the host cell to be located near the parasitophorous vacuole membrane. Merozoites were formed by endodyogeny and during replication, the apical complex of the mother cell remained intact. The formation of gametes or oocysts was not observed. The ultrastructural findings of C. belli are further evidence of its proximity to Sarcocystidae family members and corroborate their reclassification as Cystoisospora spp.

Lack of infectivity of a Brazilian Anaplasma marginale isolate for Boophilus microplus ticks.

Previous studies have shown that one Brazilian Anaplasma marginale isolate presents an inclusion appendage (tail), while other isolates do not present such inclusion. Studies on tick transmission have been carried out with tailless isolates but little is known about transmission of tailed isolates by Boophilus microplus. Two splenectomized calves were experimentally inoculated with the tailed A. marginale isolate. During ascending rickettsemia, B. microplus larvae, free from hemoparasites, were fed on the calves and the resulting nymphs, adult males and engorged females were examined by optic and electronic microscopy. No A. marginale colonies were observed in the gut cells of engorged females and the larvae originated from them did not transmit A. marginale to susceptible calves. In addition, no colonies of A. marginale were seen in the gut cells or in salivary glands of adult males and nymphs. These results suggest that B. microplus is not the biological vector for this tailed isolate.

Molecular fluorescent approach to assessing intraerythrocytic hemoprotozoan Babesia canis infection in dogs.

The development of recent flow cytometry-based protocols for the diagnosis of canine babesiosis, Babesia gibsoni in particular, has encouraged us to investigate its applicability to detect B. canis-infected erythrocytes as well as optimize the hydroethidine-flow cytometry methodology (HE-FC), using peripheral blood samples from naturally and experimentally infected dogs. Our data demonstrated that HE at 25 microg/ml provided the most outstanding fluorescence profile, able to discriminate between infected and uninfected dogs with no alterations in cell properties such as forward scatter and unspecific fluorescence. The results were expressed as the percentage of positive fluorescent erythrocytes (PPFE) for each individual sample, with 1.53% of PPFE as the cut-off determined between infected and uninfected animals. B. canis-infected erythrocytes during both acute and chronic experimental infection were identified through HE-FC, validating its use for diagnosis purposes in endemic areas for canine babesiosis. In a clinical trial, 22.8% out of 162 dogs showed to be positive to Babesia infection through this approach. Such prevalence was similar to that estimated for altered hematological profiles (HT) < or = 30% (29%), but highly distinct from the prevalence provided by direct blood smear (BS) examination (1.8%) or immunofluorescent assay (IFA) (60.5%). Furthermore, our findings indicate that positive PPFE data was associated with HT < or = 30%, emphasizing that, in clinical practice, the haematocrit should be used as a screening test followed by HE-FC, suitable to confirm hypotheses of canine babesiosis.