Endogenous Developmental Cycle of the Human Coccidian Cyclospora cayetanensis.

Cyclospora cayetanensis is a coccidian parasite of humans of known and growing importance. However, we are surprisingly naïve as to our understanding of how to diagnose it and how it develops inside the human body. Here we provide details of the developmental stages of C. cayetanensis in the gallbladder of a 33-yr-old male with human immunodeficiency virus. The gallbladder was removed surgically in 2001 because of severe abdominal pain. For the present study, the archived paraffin block of gallbladder was processed for light microscopy and transmission electron microscopy (TEM). Histological sections were examined after staining with hematoxylin and eosin (HE) or using the periodic acid Schiff (PAS) reaction. Immature and mature asexual stages, gamonts, and oocysts were seen in epithelial cells, both in the superficial epithelium and in glands. The merozoites were present singly, in pairs, and 3 or more in a single parasitophorous vacuole in the host cytoplasm. Up to 6 nuclei were seen in immature schizonts without evidence of merozoite formation. Mature schizonts were 7.6 × 5.1 µm and contained up to 10, 3-4 µm long merozoites. Merozoites were 0.6 to 2.0 µm wide, and their shape varied from pear-shaped to slender. Merozoites were generally PAS-positive; however, some were intensely positive, some had only minute granules, while others were PAS-negative. The microgamonts (male) were 6.6 × 5.2 µm and contained fewer than 20 microgametes around a residual body. The microgametes were up to 2 µm long and were flagellated. Macrogamonts (female) contained distinctive eosinophilic wall-forming bodies that varied in size and were less than 1 µm in HE-stained sections. Macrogamonts were 5.8-6.5 × 5.3-6.5 µm. Oocysts in sections were unsporulated and had a diameter of 5.7-7.5 µm. The TEM examination confirmed the histologic findings. The DNA extracted from paraffin sections was confirmed as C. cayetanensis with real-time PCR. The detailed description of the life cycle stages of C. cayetanensis reported here in an immunosuppressed patient could facilitate histopathologic diagnosis of this parasite. We have shown that the parasite's development more closely resembles that of Cystoisospora than Eimeria and that the parasite has multiple nuclei per immature meront indicating schizogony, and we have undermined evidence for a Type II meront.

Silver nanoparticles as a therapeutic agent in experimental cyclosporiasis.

Cyclosporiasis is an emerging worldwide infection caused by an obligate intracellular protozoan parasite, Cyclospora cayetanensis. In immunocompetent patients, it is mainly manifested by self-limited diarrhea, which is persistent and may be fatal in immunocompromised patients. The standard treatment for cyclosporiasis is a combination of two antibiotics, trimethoprim and sulfamethoxazole. Gastrointestinal, haematologic and renal side effects were reported with this combination. Moreover, sulfa allergy, foetal anomalies and recurrence were recorded with no alternative drug treatment option. In this study, silver nanoparticles were chemically synthesized to be evaluated for the first time for their anti-cyclospora effects in both immunocompetent and immunosuppressed experimental mice in comparison to the standard treatment. The effect of silver nanoparticles was assessed through studying stool oocyst load, oocyst viability, ultrastructural changes in oocysts, and estimation of serum gamma interferon. Toxic effect of the therapeutic agents was evaluated by measuring liver enzymes, urea and creatinine in mouse sera. Results showed that silver nanoparticles had promising anti-cyclospora potentials. The animals that received these nanoparticles showed a statistically significant decrease in the oocyst burden and number of viable oocysts in stool and a statistically significant increase in serum gamma interferon in comparison to the corresponding group receiving the standard treatment and to the infected non-treated control group. Scanning electron microscopic examination revealed mutilated oocysts with irregularities, poring and perforations. Biochemical results showed no evidence of toxicity of silver nanoparticles, as the sera of the mice showed a statistically non-significant decrease in liver enzymes in immunocompetent subgroups, and a statistically significant decrease in immunosuppressed subgroups. Furthermore, a statistically non-significant decrease in urea and creatinine was recorded in all subgroups. Thus, silver nanoparticles proved their effectiveness against Cyclospora infection, and this will draw the attention to its use as an alternative to the standard therapy.

Molecular identification of Cycospora spp. using multiplex PCR from diarrheic children compared to others conventional methods.

Cyclospora organisms named C. cercopitheci, C. colobi & C. papionis were identified in stool samples from several primates. They were morphologically indistinguishable from C. cayetanensis but genetically different. In the present work, Cyclospora infection was diagnosed among 140 diarrheic children with three conventional diagnostic methods and was confirmed using nested PCR. The possibility of infection with not only C. cayetanensis but the three Cyclospora species of primates was identified by multiplex PCR among all cyclosporiasis patients diagnosed by different methods. The results showed that Cyclospora was detected in 25 (17.8%), 31 (22.2%), 32 (22.9%) & 35 (25%) patients using modified Kinyoun stain, auto-fluorescent characteristics, sporulation process of the oocysts and nested PCR respectively. The sensitivity, specificity, accuracy, positive predictive value, negative predictive value and Kappa test were calculated in relation to nested PCR. The single nucleotide polymorphisms (SNPs) in the 18S rRNA gene of C. cayetanensis were identified in 35 cyclosporiasis patients and only one patient had the possibility of human coinfection with primates Cyclospora species (C. cercopitheci, C. colobi & C. papionis) and C. cayetanensis by appearance of a 361-bp. Scanning electron microscopy proved no morphological differences could be detected among Cyclospora oocysts isolated from this patient.

[A human case of cyclosporiasis after traveling in the subtropics]. / Yurtdisi seyahat hikayesi olan bir cyclosporiasis olgusu.

In this study, Cyclospora oocysts were detected in a 64-year old man who complained of persistent diarrhea, abdominal pain, nausea and vomiting after visiting the Greek Islands in a sailing boat. Round oocysts about 8-9 microm in size with wrinkled walls that varied in color from red to pale pink after staining were found in Kinyoun's modified acid fast stained stool samples. Public health offices and laboratories, general practitioners, and medical microbiology labs should be informed that more attention should be paid to cyclosporiasis causing diarrheal illness and which requires specific screening methods with experienced microscopists in laboratories.

Experimental studies on cyclosporiosis.

Swiss albino mice get Cyclospora infection after orally inoculated with sporulated oocysts. Two weeks post inoculation, most of them passed numerous acid fast immature oocysts in their stools. One week later, light microscopic examination of their intestinal H & E stained sections revealed parasitic stages in a supranuclear location within enterocytes. They were most prominent in the mucosal villi. Lamina propria was expanded by an inflammatory infiltrate. The combined parasitological and histopathological present studies helped in distinguishing Cyclospora which is often confused with Cryptosporidia in stool samples and with Isospora in intestinal sections. Electron microscopy demonstrated both sexual and asexual developmental stages of Cyclospora in ultrathin infected sections. Therefore, Cyclospora species require only a single host to complete its entire life cycle