Prevalence and genotype distribution of Enterobius vermicularis among kindergarteners in Shiraz and Khorramabad cities, Iran

Objective: To study the prevalence and genotype of Enterobius (E.) vermicularis from adhesive tape samples in the cities of Shiraz and Khorramabad, Iran. Methods: A total of 1 000 adhesive tape samples from kindergartens in Shiraz (500 samples) and Khorramabad (500 samples) were collected and tested using a microscope to find E. vermicularis egg/s. A questionnaire was filled out for each sample. In order to characterize the genotype of E. vermicularis, the PCR-sequencing method of the mitochondrial cytochrome C oxidase subunit 1 (cox1) gene was used. Genomic DNA was extracted from the positive scotch tape samples of E. vermicularis. The cox1 gene was amplified by the polymerase chain reaction and sequenced. The sequence data were aligned using the BioEdit software and compared with the published sequences in GenBank. Phylogenetic analysis was performed using the maximum likelihood method. Results: The parasitological method showed that 15 out of the 500 samples from Shiraz (3.00%) and 12 out of the 500 samples from Khorramabad (2.40%) were infected with E. vermicularis eggs. BLAST analysis indicated that the sequenced isolates belonged to E. vermicularis genotype B while three different haplotypes were also identified. Conclusions: This is the first study on genotyping E. vermicularis in the cities of Shiraz and Khorramabad. Considering the public health importance of the disease, further studies are necessary to characterize the genotype of E. vermicularis in human populations from other regions of Iran.

Toxocara cati larvae in the eye of a child: a case report

Toxocariasis is a consequence of human infection by Toxocara larvae. There are symptomatic (visceral, ocular) and asymptomatic course of toxocariasis. The ocular form is very rare. We present a 6-year-old patient who developed an ocular form of toxocariasis caused by Toxocara cati. He demonstrated lesions in the peripheral retina of the right eye. White granuloma was present in the superior peripheral retina. A positive immunological assay for toxocariasis essentially completed the outcomes. On the basis of clinical manifestations and conducted examinations, a diagnosis of ocular form of toxocariasis was established. Albendazole and corticosteroids were applied in treatment. Current results clearly highlight the usefulness of excretory-secretory antigens derived from larvae of Toxocara cati for the fine diagnosis ocular larva migrans caused by Toxocara larvae.

Still and moving image evidences for mating of echinococcus granulosus reared in culture media

Echinococcus granulosus cultivation is very important for improvement of different aspect of medical and veterinary researches. Despite many advances in this case, there is a missing link for in vitro life cycle of adult worms and it is fertilization. Regarding the researchers' observations, self-fertilization can be done in worms living in dog intestine, but despite all sorts of experimental techniques, this phenomenon has never been observed in reared worms in culture media. Furthermore cross fertilization has not been observed in vitro and even in parasites with dog intestinal origin; although it theoretically is possible. During a follow-up of cultivated adult worms, evidences of behaviors similar to self-mating [Type 2] and cross-mating were observed in our lab which will be presented here. Protoscoleces were aseptically removed from sheep hydatid cysts, washed twice with PBS and then cultivated in S.10E.H culture medium. The stages of parasite growth were observed using an inverted microscope for two months and all stages and behaviors were microscopically photographed. Different movies have also been made from these behavioral features. After around 55 days post cultivation, some evidences of behaviors similar to self-mating [Type 2] and cross-mating were observed in some of the mature adult worms. However, fertile eggs in these parasites have never been observed. Regarding the above observations, these parasites show tendency to unsuccessful self-mating/fertilization [type 2] which failure could be due to anatomical position and physiological maturation. Also lack of suitable conditions for self-fertilization causes the worms try to do unsuccessful cross- mating/fertilization in culture media

Plagiorchis muris [Tanabe, 1922] in Rattus norvegicus in Iran

Rats are capable to harbor various pathogens, among which certain species of zoonotic parasites are included. A long-term detection of parasite fauna of rats has sporadically been carried out in Iran. Abundance of these vertebrate pests is of great importance as regards public health issue. The present paper is focused on a digenean trematode Plagiorchis muris, obtained during a comprehensive study on rats over the decades in the country. Herein we describe this occurrence in a Rattus norvegicus in northern Tehran, with specific note on its morphological description. P. muris can infect human through consumption of infected marine food items, and has never been observed in Iran

Immune responses to antigens of in vitro reared echinococcus granulosus adult worms in Balb/c mice

Cystic echinococcosis [CE], also known as echinococcosis/hydatidosis, is one of the most important parasitic diseases in the world. It enhances both humoral and cellular [Thl and Th2] responses in infected host. Different antigens of the worm may favor the Thl or Th2 immune responses in CE patients. To evaluate the humoral and cellular immune responses of Balb/c mice against the crude and excretory/secretory [E/S] antigens of in vitro reared Echinococcus granulosus adult worms. A total of 20 Balb/c mice divided into 5 groups of 4 mice each. Three groups of mice [n=4] were immunized with crude, E/S and an immunodominant antigen of in vitro reared Echinococcus granulosus adult worms on day 1 and 28. The fourth and the fifth groups were negative control groups and received PBS plus adjuvant, or nothing, respectively. Two weeks after the second injection, the mice were killed and their blood was collected for determining antibody responses, and their spleens were employed for proliferation assay. Total IgG were measured by indirect ELISA. Spleen cells of immunized mice were cultivated and exposed to different antigens of adult worms including E/S and crude antigens. Level of IFN-y, IL-12, IL-4 and IL-10 were measured in the recovered cell culture supernatants by capture ELISA. Total IgG assay showed the highest level of antibody produced in mice immunized with crude antigens. Proliferation assay showed a statistically significant production of cytokines in the mice immunized with crude antigens [p<0.05]. The highest levels of IFN-gamma, IL12 and IL-4 were produced in mice immunized with crude antigen of the in vitro reared Echinococcus granulosus adult worms followed by E/S antigens. Immunodomonant antigen induced the lowest levels of cytokines [IL-12, IFN-gamma, IL-4 and IL-10] in immunized mice. A significant levels of Thl related cytokines [IFN-gamma and IL-12] were produced in Balb/c mice immunized with crude antigen of the in vitro reared Echinococcus granulosus adult worms

Experimental Toxocara cati Infection in Gerbils and Rats

Mongolian gerbils and Wistar rats were inoculated orally with 240 and 2,500 Toxocara cati embryonated eggs, respectively, to evaluate the larval recovery in different tissues and organs, such as the liver, lungs, heart, kidney, and skeletal muscles after 5, 30, 49, 70, and 92 days post-infection (PI). Larval recovery rates were 1.7-30.0% in Mongolian gerbils on days 5-92 PI and 0.2-3.8% in rats on the same days. These results indicate that Mongolian gerbils and Wistar rats are suitable experimental paratenic hosts for the study of neurological toxocariasis as well as visceral toxocariasis.

Serum Antigen and Antibody Detection in Echinococcosis: Application in Serodiagnosis of Human Hydatidosis

Diagnosis of hydatidosis is based on immunodiagnostic methods along with radiological and ultrasound examinations. The objectives of the present study were to develop a specific and simple antigen-based ELISA method for diagnosis of hydatidosis and compare it with antibody detection method. The subjects in this study included 89 patients in the following groups: surgically confirmed hydatidosis patients (35 cases), control with other parasitic diseases (29 cases), and healthy controls (25 cases). Hyperimmune serum was raised against hydatid cyst fluid in rabbits. Anti-hydatid cyst IgG was purified by affinity chromatography using protein A column and labeled with horseradish peroxidase. Collected sera were assessed for hydatid cyst antigens and antibody by ELISA. Circulating hydatid antigen was found in 9 out of 35 patients with surgically confirmed hydatidosis. A sensitivity of 25.7% and a specificity of 98.0% were calculated for the antigen detection assay. Antibody detection by indirect ELISA, using antigen B, showed that 94.2% of patients (33 cases) have anti-hydatid cyst antibodies in their serum while cross reaction was noted in a few of non-hydatidosis patients. A sensitivity of 94.2% and specificity of 81.6% were found for the antibody detection assay. Findings of this study indicated that antibody detection assay is a sensitive approach for diagnosis of hydatid cyst while antigen detection assay might be a useful approach for assessment of the efficacy of treatment especially after removal of the cyst.

Giardia spp. and Sarcocystis spp. status in pet dogs of Shiraz, Southern part of Iran

A study was carried out in order to find out the status of Giardia spp. and Sarcocystis spp. in pet dogs and stray cats of Shiraz, Fars Province of Iran. Faecal samples of 147 pet dogs and 112 stray cats of different age groups, breeds, and sexes were tested. The stools were examined with the following techniques: direct faecal smears using normal saline, zinc sulfate flotation and formalin-ether concentration technique. Out of a total of 147 pet dogs examined, only one case (0.68%) of Giardia spp. was observed. A total of 3 (2.04%) pet dogs were found positive for Sarcocystis spp. Specimens from stray cats were also examined, however no Giardia spp. trophozoite or cyst was observed in these specimens.

Prevalence of Toxocara cati and other intestinal helminths in stray cats in Shiraz, Iran

Toxocariasis is a parasitic zoonosis with worldwide distribution that affects both cats and dogs. Necropsy of 114 stray cats from Shiraz revealed that 106 (92.9%) stray cats were infected at least with one of the intestinal helminth species. The overall infection rates in stray cats infected with cestoda and nematoda were 105(99.1%) and 101(95.3%) respectively. The detected cestodes were Joyeuxiella pasqualei (34.3%), Dipylidium caninum (49.5%), Taenia taeniaeformis (12.3%), Spirometra sp. (3.8%) and the detected nematodes were Physaloptera sp. (44.6%), Toxocara cati (42.6%) and Toxascaris leonina (12.9%). The study revealed that T. cati was one of the most frequently detected intestinal helminths, which is an important source of zoonotic helminths.

In vitro protoscolicidal effects of hypertonic glucose on protoscolices of hydatid cyst

To evaluate the protoscolicidal effects of various concentrations of hypertonic glucose, live protoscolices of sheep were exposed to 10%, 15%, 25% and 50% glucose solutions. Cetrimide (0.5%), silver nitrate (0.5%) and hypertonic saline (20%) were used as positive controls, while physiological saline was used as a negative control. After 1, 2 and 5 min, the protoscolicidal effects were determined by 1% eosin. A 25% glucose solution had no significant protoscolicidal effect. However, a 50% glucose solution revealed higher protoscolicidal effect than 0.5% silver nitrate but weaker effect than 0.5% cetrimide; the effect was comparable with that of 20% hypertonic saline. The results showed that hypertonic glucose solution is highly effective in killing protoscolices of Echinococcus granulosus in vitro.

Application of specific monoclonal antibodies for characterization of leishmania spp. promastigotes using indirect immunofluorescent and immunoperoxidase tests

Different methods have been used for characterization of Leishmania promastigotes. Monoclonal antibodies are useful in characterization of Leishmania spp. both amastogotes and promastigotes. Comparing the characterization of Leishmania spp. promastigotes with immunoperoxidase test [Avidin-Biotin] techniques and an indirect immunofluorescent assay [IFA]. Application of specific monoclonal antibodies for characterization of different Leishmania species. Immunoperoxidase tests [Avidin-Biotin] and indirect immunofluorescent assay [IFA] were employed for characterization of different Leishmnia promastigotes from culture. Five monoclonal antibodies including LXXVIII-2E5- A8 [D2] specific for L. donovani:L. infantum, IS2-2B4 [A11] specific for L. tropica, XCIV-H2- AB [T10] specific for L. tropica, XLVI-5B8- B3 [Tl] specific for L. major, and T7 reactive to both L. major and L. tropica as well as an anti GP63 mAb were used. The best result was obtained with the dilution of 1:50 for both mAb and conjugate. One hundred percent sensitivity and specificity was achieved for characterization of Leishmania promastogotes with both methods. Conclusion: As immunoperoxidase method needs less equipments compared to IFA technique, it would be a preferred method for characterization of promastigotes

Identification and characterization of leishmania spp. in impression smears of patients with cutaneous leishmaniasis

Bakground: Monoclonal antibodies have been employed extensively for the identification of Leishmania species, development of diagnostic tests and in the characterization of defined leishmanial antigens

Objectives: Identification and characterization of Leishmania spp. directly from cutaneous lesions of infected individuals

Methods: An immunoperoxidase test [Avidin-Biotin technique] using monoclonal antibodies was used for this purpose. One hundred and fifty individuals referring to Dermatology Clinic or Parasitology and Mycology Department of Shiraz University of Medical Sciences were chosen of whom a total of 28 individuals whose smears showed a large number of amastigotes after staining with Giemsa were included in this study. Five monoclonal antibodies designated: D2 [against L. donovani], All andTIO [against L. Tropica], Tl [against L. major] and T7 [against L. tropica and L. major] were used. Amastigotes were identified by Labeled Avidin Biotin [LAB] method

Results: LAB method for identification of amastigotes in impression smears of patient lesions showed that 20 out of 28 cases [71%] were positive. Among these 12 [60%] and 7[35%] were identified as L. tropica andL. major respectively

Conclusion: The results showed that immunoperoxidase is suitable for in situ identification and characterization of Leishmania spp. at the species level