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Background: The great gerbil (Rhombomys opimus), is widely distributed in Asia and is a natural reservoir for zoonotic cutaneous leishmaniasis in many endemic areas, as well as Iran. Materials and Methods: In this study, infection to Leishmania species was investigated by two methods, parasitological and molecular survey, in the small number of R. opimus collected from Jovain, a Zoonotic Cutaneous Leishmaniasis (ZCL) focus located in North East of Iran. Results: Parasitological observation showed infection in only one of five rodents. But, ITS2-Nested-PCR revealed Leishmania infection in three out of 5 gerbils, including the parasitological positive one. Based on the PCR amplified size, two cases of infections were Leishmania major and one Leishmania turanica, their sequences are accessible in GenBank. The results of sequence analysis were consistent with the results obtained based on the size of the PCR. Conclusion: These findings re-confirm the important role of R. opimus in the natural circulation of Leishmania spp and indicate the need to be concerned about the disease in the study area. [ABSTRACT FROM AUTHOR]

BACKGROUND: Updated information on the vectorial capacity of vectors is required in each malarious areas as well in Iran and its neighboring countries such as Afghanistan. The aims of this study were to investigate the potential infection of about 800 specimens collected from malarious areas of Afghanistan and Iran, and to differentiate biological forms of Anopheles stephensi. Method: Two molecular markers, 18S RNA gene subunit and AsteObp1 intron I, were used respectively for investigation Plasmodium infection and identifying the biological forms of An. stephensi. RESULTS: Plasmodium infection was detected in 4 pools of Afghanistan specimens, including An. stephensi, collected from Nangarhar. Individually examination showed infection in 5 An. stephensi (infection rate: 1.25), to P. falciparum (2), P. vivax (2) and a mix infection. Out of five infected specimens, three were intermediate forms and two were mysorensis. No infection was found in specimens collected from Iran (Chabahar County), probably due to the active malaria control program in south-east of Iran. CONCLUSION: The key role of An. stephensi, as a known Asian malaria vector, was re-emphasized in Afghanistan by the results achieved here. The fauna of vectors and the pattern of biological forms of An. stephensi are similar in both countries that urge regional investigations to provide evidence-based and applied data for decision-maker in malaria control. [ABSTRACT FROM AUTHOR]

Toxoplasmosis causes serious complications in immunocompromised and pregnant women. Serological tests for the detection of toxoplasmosis are often designed from parasitic tachyzoites antigens. The process of producing these antigens is very difficult. The purpose of this study was evaluation of T. gondii‐rGRA5 for the immunodiagnosis and molecular detection of Toxoplasma infection using enzyme‐linked immunosorbent assay (ELISA) and LAMP methods in hemodialysis patients. The GRA5 gene was successfully expressed and purified by affinity chromatography assay and evaluated by western blot. Then it was used to design an ELISA assay. A total of 260 samples were tested for anti‐Toxoplasma IgG and IgM antibodies using a commercial ELISA kit and designed ELISA kit. Finally, the LAMP method was used to evaluate the precision and reliability of the results obtained by commercial and designed ELISA kits. The consistency of the results of two methods was analyzed using the Kappa coefficient of agreement. The rGRA5 revealed higher immunoreactivity with 1:100 dilution of sera from toxoplasmosis patients. The specificity and sensitivity of the assay were 93% and 96%, respectively. According to the Kappa coefficient, there was a substantial correlation between the results of ELISA and LAMP based on rGRA5 (≈98%, p < 0.001). Also it showed that rGRA5 protein can be used as an antigenic protein for designing sero‐diagnostic tests to identify Toxoplasma infection especially in hemodialysis patients. [ABSTRACT FROM AUTHOR]

BACKGROUND፡ Leishmaniasis is a vector-borne disease caused by an intracellular protozoan parasite called Leishmania spp. Different species produce different clinical outcomes; the majority of cases are cutaneous forms. Leishmania major is one of the main causative agents of cutaneous leishmaniasis (CL). Various methods are being using to diagnose CL, including microscopic examination, culture, and molecular detection of the parasite genome. METHOD: In the current study, we tried to compare three common molecular markers, including Kinetoplast DNA (kDNA), Cytochrome b (Cyt b), and Internal transcribed space 1 (ITS1), for the detection of Leishmania major. After cultivation of standard strain of L. major MHOM/IR/75/ER in RPMI 1640, certain number of promastigotes was subjected to DNA extraction and different PCR reactions. RESULTS: The lowest number of the parasite (5 promastigotes) can be detected by kDNA-PCR, followed by Cyt b-PCR (10 promastigotes), and ITS1-PCR (50 promastigotes). CONCLUSION: In conclusion, kDNA-PCR was the most sensitive marker and may provide more reliable data in the initial screening, especially in false-negative results provided by parasitological methods due to the low number of parasites. [ABSTRACT FROM AUTHOR]

Background: Cutaneous leishmaniasis caused by Leishmania species (L. spp) is one of the most important parasitic diseases in humans. To gain information on the metabolite variations and biochemical pathways between L. spp, we used the comparative metabolome of metacyclic promastigotes in the Iranian isolates of L. major and L. tropica by proton nuclear magnetic resonance (¹H-NMR). Methods: L. tropica and L. major were collected from three areas of Iran, namely Gonbad, Mashhad, and Bam, between 2017 and 2018, and were cultured. The metacyclic promastigote of each species was separated, and cell metabolites were extracted. ¹H-NMR spectroscopy was applied, and the data were processed using ProMatab in MATLAB (version 7.8.0.347). Multivariate statistical analyses, including the principal component analysis and the orthogonal projections to latent structures discriminant analysis, were performed to identify the discriminative metabolites between the two L. spp. Metabolites with variable influences in projection values of more than one and a P value of less than 0.05 were marked as significant differences. Results: A set of metabolites were detected, and 24 significantly differentially expressed metabolites were found between the metacyclic forms of L. major and L. tropica isolates. The top differential metabolites were methionine, aspartate, betaine, and acetylcarnitine, which were increased more in L. tropica than L. major (P<0.005), whereas asparagine, 3-hydroxybutyrate, L-proline, and kynurenine were increased significantly in L. major (P<0.01). The significantly altered metabolites were involved in eight metabolic pathways. Conclusion: Metabolomics, as an invaluable technique, yielded significant metabolites, and their biochemical pathways related to the metacyclic promastigotes of L. major and L. tropica. The findings offer greater insights into parasite biology and how pathogens adapt to their hosts. [ABSTRACT FROM AUTHOR]

Aim: we aimed to evaluate somatic mutations of CTNNA1 in DGC patients. Background: Diffuse gastric cancer (DGC) is a major type of gastric cancer where most cases are sporadic diffuse gastric cancer (SDGC). It has been shown that mutations in CTNNA1 are responsible for some cases of hereditary diffuse gastric cancer (HDGC). Methods: In the present work, 48 formalin-fixed paraffin-embedded tissues, including samples of 38 SDGC and 10 HDGC patients were examined through Sanger sequencing approach on PCR products amplified from 18 exons and boundaries of intron/exon of CTNNA1 gene. Results: We revealed 9 novel somatic mutations in CTNNA1 gene in patients with HDGC and SDGC, from which one variant was intronic. Eight patients had at least one disease-causing mutation (16.6%). Most of the patients were in the III stage of cancer (50%). Except for one patient, histological type of the rest of mutation-harboring patients was signet ring cell carcinoma, and only one HDGC patient had CTNNA1 mutation. Conclusion: Our study showed several novel variants in the CTNNA1 gene in Iranian sporadic and hereditary DGC patients, and implies that the CTNNA1 gene mutations could be involved in the pathogenesis of DGC, either hereditary or in sporadic cases. [ABSTRACT FROM AUTHOR]

Background: Cystic echinococcosis can cause severe disease and probable death in humans. Epitopes of its antigens play a key role in the sensitivity and specificity of immunodiagnostic tests. Methods: Epitope prediction software programs predict the most antigenic linear B-cell epitopes of AgB (8 kD), Ag5, and Ag95. Six such epitopes were predicted and connected by "Gly-Ser" linker and synthesized. The purity of the concentrated recombinant multi-epitope protein was assessed by 15% SDS-PAGE. Overall, 186 serum samples were collected from the Loghman Hakim Hospital and different laboratories, Tehran, Iran, from July 2016 to February 2017. Patients infected with hepatic hydatid cysts, patients infected by other parasites and viruses, and healthy individuals were used to detect the anti-CE IgG using recombinant multi-epitope protein. Results: Forty-one samples out of 43 cases of hydatidosis were diagnosed correctly as positive, and two were negative. In addition, six negative cases of healthy individual group were diagnosed as positive and negative with rMEP-ELISA and the commercial kit, respectively. Therefore, these six samples were considered as false positive using our method. In addition, a diagnostic sensitivity of 95.3% (95% CI, 84.19% to 99.43%) and a specificity of 95.0% (95% CI, 89.43% to 98.14%) were obtained using optimum cutoff value (0.20). The sensitivity and specificity of the commercial kit was 100%. Conclusion: Our findings showed high diagnostic accuracy of the ELISA test using the developed recombinant protein, which encourages the use of this recombinant multi-epitope protein for rapid serological diagnosis of hydatidosis. [ABSTRACT FROM AUTHOR]

Background: The present study aimed to assess the grouping of subtypes 1-3 and 6 of Blastocystis according to the size and generation time of the parasite among the symptomatic and asymptomatic subjects. Methods: Blastocystis subtypes1-3 and 6 isolated from symptomatic and asymptomatic subjects and were cultivated in DMEM medium. In order to assess inter- and intra-subtype variation in size, all the isolates were measured using morphometric criteria. Generation time was calculated using approximately 1×104 Blastocystis, which were cultivated in DMEM, every 24h for 4 days. Results: All subtypes had 5 to 185 µm diameter range. The smallest size was attributed to ST1, followed by ST6 and ST2. ST3 showed the most variable size and phenotypes compared with the other three subtypes. Furthermore, amoeboid forms and parasite clumping were only seen in ST3-S (symptomatic subjects). Generation time analysis showed that the number of ST1 isolated from symptomatic and asymptomatic subjects peaked higher than the other subtypes. Conclusion: This is the first study discussing inter-intra-size, phenotype and generation time variations among 4 common subtypes of Blastocystis. Accordingly, ST3 was largest subtype and showed most diversities in both size and phenotype, while ST1 was smallest subtype with lowest intra-subtype variation. [ABSTRACT FROM AUTHOR]

Background: Intestinal parasitic infections are major causative agents of wildlife health complications among different parts of the world. This study aimed to investigate the gastro-intestinal parasites in feces of the zoo animals based on parasitological and morphometric criteria. Methods: One hundred fresh fecal samples were collected from 35 species of animal lived in Eram park zoo, Tehran, Central Iran during Oct 2015 to Jun 2015. All collected samples were examined by microscopic observation following direct wet mount preparation (normal saline and Lugol's iodine), formalin-ether concentration, and permanent staining. The morphometric aspects of the recovered eggs were surveyed with the aid of Camera Lucida (×400). Results: 65.7% (23/35) of zoo animal species were infected with intestinal parasites. The superfamily Trichostrongyloidea (6/16) and Strongylus sp. (16/4) were the most prevalent helminthic infections, while Blastocystis sp. (6/14), Entamoeba cyst (3/14) and Eimeria sp. (3/14) were the common protozoan parasites. For the first time, Bivitellobilharzia nairi egg was identified an elephant at Iran. Intestinal parasitic infections were apparently circulating among animals of the Eram park zoo. Conclusion: Identified parasitic infections can consider as a threatening source to visitors and workers' health that have contact with animals or their feces. Therefore, the effectual preventive strategies should be addressed to determine the risk factors, mechanisms of cross-transmission of parasite, the importance of applying the hygienic practices and well adjusting deworming programs for the animals, zoo workers and visitors. [ABSTRACT FROM AUTHOR]

Theileria lestoquardi, T. ovis, and T. annulata are recognized as major causative agents for ovine and cattle theileriosis in Iran, respectively. Recently, there have been reports on the presence of Theileria spp. (Theileria sp. OT1, Theileria sp. OT3, and Theileria sp.). In this study, 37 blood samples were collected from sheep and cattle with clinically suspected Theileria infection in the Northwest of Iran. The samples were analyzed using a light microscope. DNA samples were amplified via nested-polymerase chain reaction (PCR) of 18S rRNA gene. The amplicons were digested with HpaII, following restriction fragment length polymorphism (RFLP) and sequenced to reconfirm Theileria species. The microscopic examination indicated that 4 out of 37 (10.8%) blood samples were infected with Theileria spp. Based on the nested PCR-RFLP and sequencing data, 5.4%, 13.5%, and 27% of blood samples were infected with Theileria sp. OT3, T. ovis, and T. annulata, respectively. The pairwise distance matrix of Theileria sp. OT3 showed 99.8-100% identity and 0-0.2% divergence in comparison with the registered sequences. The present study is the first report of Theileria sp. OT3 in Iran. To the evaluate evolution of Theileria spp. and providing resultant genetic data, further research with a larger sample is necessary in the region. [ABSTRACT FROM AUTHOR]

Background: Malarious areas in Iran are close to Afghanistan and Pakistan that urge the researchers to extend their knowledge on malaria epidemiology to the neighboring countries as well. Vectorial capacity differs at species or even at population level, the first essential step is accurate identification of vectors. This study aimed to identify Anopheles species composition in selected malarious areas of Afghanistan and Iran, providing further applied data for other research in two countries. Methods: Adults Anopheles spp. were collected from four provinces in Afghanistan (Badakhshan, Herat, Kunduz, Nangarhar) by pyrethrum spray catch, hand collection methods through WHO/EMRO coordination and from Chabahar County in Iran by pyrethrum spray catch method. Identification was performed using reliable identification key. Results: Totally, 800 female Anopheles mosquitos, 400 from each country were identified at species level. Anopheles composition in Afghanistan was An. superpictus, An. stephensi and An. hyrcanus. Most prevalent species in Badakhshan and Kunduz were An. superpictus, whereas An. stephensi and An. hyrcanus were respectively found in Nangarhar and Heart. Anopheles species in Chabahar County of Iran were An. stephensi, An. fluviatilis, An. culicifacies and An. sergentii. The most prevalent species was An. stephensi. Conclusion: Current study provides a basis for future research such as detection of Plasmodium infection in collected samples which is on process by the authors, also for effective implementation of evidence-based malaria vector intervention strategies. [ABSTRACT FROM AUTHOR]

Background and Objectives: Toxoplasma gondii is an obligatory intracellular parasite which causes severe diseases in the fetus of pregnant women and immunocopmromised patients. Serological tests based on recombinant protein are one of the main diagnosis methods for the detection of specific antibodies in serum samples. Dense granule antigenic proteins derived from T. gondii (TgGRAs) are potential antigens for the development of diagnostic tools. Materials and Methods: DNA was extracted from T. gondii (RH-strain) tachyzoites and PCR reaction was done using corresponding primers for GRA5 antigen. The PCR product was purified and ligated into pTG19-t vector and then subcloned into XhoI and BamHI digested pGEX6p-1 expression vector. Recombinant plasmid was transformed into E. coli (BL21 DE3) and induced by 1mM IPTG and analyzed by 15% SDS-PAGE. Expressed protein was confirmed by western blot analysis. Results: There was no difference among the sequences of T. gondii GRA5 gene from different isolates. The recombinant plasmid pGEX-6p-1/GRA5 induced by IPTG was expressed in E. coli. It was a GST fusion protein and could react with human positive sera analyzed by western blot. Conclusion: The GRA5 gene of T. gondii isolates is highly conservative. This antigen as a recombinant protein was successfully expressed in E. coli, which showed high immunoreactivity. [ABSTRACT FROM AUTHOR]

Background: Toxocariasis is considered as an important neglected tropical disease. Although, the prevalence of Toxocara eggs in soil has previously been reported in different parts of Iran, the extent of this condition is not precisely known in Kermanshah city, west of Iran. Materials and Methods: A total of 126 soil samples were collected from different zones of Kermanshah public places during April-June 2014. The samples were examined for Toxocara spp. eggs via modified floatation method using sodium nitrate (NaNO3) and the data was analyzed using Data Analysis and Statistical Software (STATA Ver.13.1). Results: Toxocara spp. eggs were found in 17 (13.5%) out of 126 samples collected from the studied areas. There was a significant difference between contamination rate in the areas with low levels of health status and that in the areas with high levels (p=0.003). Conclusion: According to the results obtained in the present study public parks, streets, and squares of Kermanshah are contaminated with eggs of Toxocara spp. Considering these findings, establishment of a wisely planned health program for controlling helminthes in the soil and the population of the stray dogs and cats in order to reduce the distribution of parasitosis is strongly recommended. [ABSTRACT FROM AUTHOR]

Background: Toxoplasma gondii is an obligate, intracellular protozoon that develops its sexual stage in cat's intestinal epithelial cells as definitive host and develops its asexual stage in different tissues of a wide range hosts called intermediate host. The protozoon is a food-borne and worldwide parasite that can cause serious complications such as abortion in pregnant women, encephalitis, and ocular toxoplasmosis. The present study aimed to genotype T. gondii strains isolated from patients with toxoplasmic retinochoroiditis. Methods: Fifty-two blood samples were taken from patients with ocular toxoplasmosis, from July 2013 to July 2014. The specimens were collected from three ophthalmological hospitals of Tehran, Iran. After that, DNA extraction was performed using kit on separated buffy coats of serologically positive blood samples. Then PCR was done in GRA6 gene. For digestion of products, MseI endonuclease was used. Finally, some of the PCR products were sequenced. Results: All of 52 samples were found positive by serological and PCR-RFLP methods and all of isolated strains belong to type III genotype. Type III genotype has the highest prevalence in Iranian ocular toxoplasmic patients. Conclusion: T. gondii, particularly its type III should not be neglected as a cause of retinochoroiditis. [ABSTRACT FROM AUTHOR]