Differential detection of Entamoeba histolytica, Entamoeba dispar and Entamoeba moshkovskii in faecal samples using nested multiplex PCR in west of Iran.

This study aimed to determine the prevalence of Entamoeba histolytica, Entamoeba dispar and Entamoeba moshkovskii (collectively referred to as Entamoeba complex), using microscopic and molecular methods in Kurdistan Province, northwest of Iran. The relationship between positive Entamoeba species and clinical symptoms was also investigated. Eight positive Entamoeba complex, as well as four Entamoeba complex-like isolates, were detected by microscopic stool examination. DNA was extracted from all positive and from 55 randomly selected negative stool samples. PCR was performed using species-specific 18S rRNA primers for the Entamoeba complex. All positive PCR samples were sequenced. In total, 14 (1.01%) out of 1383 isolates, i.e. 12 microscopy-positive and Entamoeba complex-like isolates and two out of 55 microscopy-negative isolates, were identified via PCR and sequencing. Overall, 0.58% (8/1383) of the isolates were E. dispar, 0.14% (2/1383) E. histolytica, 0.07% (1/1383) E. moshkovskii and 0.22% (3/1383) were mixed of E. histolytica and E. dispar. Based on our findings, the prevalence of E. dispar is greater than that of E. histoltyica. On the other hand, a case of E. moshkovskii was reported for the first time in this region. It seems that some gastrointestinal symptoms may be attributed to Entamoeba species.

Zoonotic Transmission of Cryptosporidium and Microsporidia in Individuals of the Kurdistan Province, West of Iran.

Cryptosporidium species and microsporidia, which can cause zoonotic intestinal infections in humans, have become an emerging public health concern. It seems that the identification and genotyping of these parasites are necessary for the prevention, control, and establishment of appropriate treatment. This study aimed to evaluate the distribution and zoonotic transmission routes of Cryptosporidium species and microsporidia to humans referred to medical laboratories of Kurdistan Province, Iran. A total of 1,383 stool samples were collected and investigated. Cryptosporidium spp. and microsporidia were detected using microscopic methods (i.e., formol-ether concentration, Ziehl-Neelsen staining, and modified trichrome staining methods). DNA was extracted from positive samples, and specific fragments of the Cryptosporidium GP60 gene and microsporidia SSU rRNA gene were amplified. Furthermore, positive samples were sequenced for genotype identification and bioinformatics analysis. Based on the microscopic analysis of 1,383 stool samples, 5 (0.36%) and 6 (0.43%) samples were considered positive for Cryptosporidium oocysts and microsporidia spores, respectively. Molecular analysis of positive samples identified the isolates as Cryptosporidium parvum and Enterocytozoon bieneusi. According to comparative phylogenetics, cryptosporidiosis and microsporidiosis may occur via zoonotic transmission in this region. Therefore, proper control and health education are strongly recommended to prevent zoonotic diseases.

Molecular evidence for zoonotic transmission of Blastocystis subtypes in Kurdistan province, West of Iran

Blastocystis sp. is one of the most prevalent human parasites with a vast variety of non-human hosts. The aim of the present study was to determine the subtype distribution of Blastocystis in humans and trace the route of transmission by molecular data and phylogenetic analysis. Stool samples were collected from patients who referred to 14 medical laboratories in Kurdistan, Iran. All the samples were examined using the direct wet mount and formalinether concentration techniques. DNA extraction was carried out for 30 microscopically positive isolates and 33 negative samples. DNA amplification and subtype identification were also performed using the barcoding method and sequencing techniques. Of 1383 stool samples, 239 (17.3%) were infected with Blastocystis sp. Out of the 24 sequenced isolates, two (8.3%), six (25%), and 16 (66.6 %) belonged to the ST1, ST2, and ST3 subtypes, respectively. Bioinformatics analysis indicated that all the isolates were genetically similar to animal isolates. Blastocystis sp. was very common and ST1, ST2, and ST3 subtypes were prevalent in the study population. Bioinformatics analysis suggests that zoonotic transmission plays an important role in Blastocystis sp. distribution in Kurdistan province.

Seroepidemiological Study of Toxocariasis in Children Aged 6-14 Year Old in Sanandaj, Western Iran.

BACKGROUND: Toxocariasis is a disease caused by Toxocara nematodes and occurs from consuming their eggs. The main hosts of these worms are dogs and cats. The disease in humans becomes a visceral larva migrans (VLM). This descriptive cross-sectional study was conducted to determine the prevalence of toxocariasis in children aged 6-14 years. METHODS: This cross-sectional descriptive study was conducted from Jun 1 2016 to Dec 1 2017 in Sanandaj, west of Iran. A total of 182 serum samples were collected from children age 6 to14 yr referred to medical diagnostic laboratories. Demographic data (age, sex, and parents' literacy status), clinical signs (cough, headache, fever, abdominal pain), and the history of contact with dogs and cats was collected by a questionnaire. The presence of anti-Toxocara IgG antibody was detected by T. canis IgG ELISA (IBL, Germany) kit. RESULTS: Of 182 subjects, 97 (53.3%) were male and 85 (46.7%) female. The average age was 9.2 years. Antibodies against T. canis were positive in three cases (1.65%) of all the studied subjects. CONCLUSIONS: The results showed a low prevalence of toxocariasis in children studied.

Construction of A Synthetic Gene Encoding the Multi-Epitope of Toxoplasma gondii and Demonstration of the Relevant Recombinant Protein Production: A Vaccine Candidate.

BACKGROUND: Toxoplasma gondii is a widely-distributed parasite all over the world whose attributed severe afflicting complications in human necessitate the development of serodiagnostic tests and vaccines for it. Immunological responses to monovalent vaccines and the application of diagnostic reagents including single antigens are not optimally effective. Bioinformatics approaches were used to introduce these epitopes, predict their immunogenicity and preliminarily evaluate their potential as an effective DNA vaccine and for serodiagnostic goals. MATERIALS AND METHODS: A 3D structure of proteins was predicted by I-TASSER server, and linear and conformational B cell and T cell epitopes were predicted using the online servers. Then, the predicted epitopes were constructed and called Toxoeb, and their expression in the prokaryotic and eukaryotic cells was demonstrated using SDS-PAGE. In the next step, Western blotting with pooled sera of mice infected with T. gondii was done. RESULTS: The current in silico analysis revealed that the B cell epitopes with high immunogenicity for GRA4 protein were located in the residues 34-71, and 230-266, for GRA14 in 308-387, for SAG1 in 182-195, 261-278, and for GRA7 in residues 101-120, 160-176. The T cell epitopes were selected in overlapping regions with the B cell epitopes. The immunogenic region for GRA4 are in the residues 245-253, 50-58, and 40-54, for GRA14 in 307-315, 351-359, and 308- 322, for SAG1 261-269, and 259-267, and for GRA7 in the residues 103-112, and 167-175. The results of the western blotting showed that the expressed protein had immunogenicity. CONCLUSION: Our constructed multi-epitope of T. gondii could be considered as a candidate for diagnostic and vaccination purposes.

Effects of frequent announced parasitology quizzes on the academic achievement.

BACKGROUND: The effect of frequent examinations on the students' learning has had inconsistent results. This study aimed to assess the effectiveness of frequent announced quizzes on the learning of a representative sample of Iranian medical students. METHODS: This experimental study was conducted among 37 fifth semester medical students who had taken the course in Protozoology and Helminthology, in which the same basic information were provided about different types of protozoa and worms. Initially, in the teaching of helminthology, ten routine sessions were handled with lectures and interactive questions and answers. Then at the beginning of the protozoology topic in the beginning of all of the next 9 sessions, the students were informed that they will have a quiz at the end of each session. At the end of the semester, the total scores of quizzes were compared with the mean final scores of protozoology and helminthology using paired t and repeated measure tests. RESULTS: The mean final scores of the protozoology lesson were not significantly different from that of the helminthology (10.45 ± 2.75 vs.11.25 ± 2.56 on the scale of 20, respectively, P=0.13). There was no significant difference in the mean score of the five quizzes compared with the mean final term score of protozoology. The overall mean scores in the helminthology lesson (11.25±2.56), protozoology lesson (10.45±2.75), and the quizzes (9.16 ± 3.55) were significantly different (P <0.0001). CONCLUSION: Frequent announced quizzes were not effective on increasing the medical students' motivation and learning.

Natural oils enhance IL-10 and IFN-γ production by human PBMCs cultured with Malassezia furfur.

BACKGROUND: Malassezia furfur is a lipophilic yeast that causes skin disease. OBJECTIVE: To evaluate the level of IL-10, IFN-γ and IL-12P70 in co-incubation of peripheral blood mononuclear cells (PBMCs) with M. furfur grown in the presence of some different types of natural oils. METHODS: PBMCs were obtained from blood samples of normal volunteers. M. furfur was cultured in different culture media containing almond oil, fish oil, walnut oil, full-fat milk, and a fat-free medium; and the yeasts grown were harvested and used for co-incubation with PBMCs in vitro. The IFN-γ, IL-10, and IL-12P70 levels were measured at different time intervals using ELISA methods. RESULTS: Generally, IFN-γ and IL-10 levels in the co-incubation of yeasts with walnut oil group (WOG) and fish oil group (FOG) were higher than those in the almond oil group (AOG) and full-fat milk group (FFMG). Although the IL-12P70 was higher in groups such as AOG, FOG, and WOG; the increase was not statistically significant. CONCLUSION: The results demonstrated that the type of fat used by M. furfur in the culture media can influence the immune response and increasesIFN-γ and IL-10 levels in an early time point of the culture system.