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1.  Prevalence of Trichinella spp. Infections in Hunted Wild Boars in Northern Iran 
Iranian Journal of Public Health  2017;46(12):1712-1719.
Trichinellosis is an important and neglected foodborne zoonotic infectious disease in worldwide. The most human outbreaks in recent years have been related to consumption of wild boar meat. This cross-sectional study determined the prevalence of Trichinella spp. infections in hunted wild boars in northern Iran.
Thirty-five hunted wild boars were subjected in this study in 2015. All samples were examined by conventional artificial digestion method to detect of muscle larvae. Genomic DNA was extracted by phenol-chloroform method from isolated larvae. To identify the Trichinella species, a PCR-based method was applied using the internal transcribed spacer 2 (ITS2) and mitochondrial small-subunit ribosomal RNA (rRNA) gene sequences.
The overall prevalence of Trichinella spp. infection was 5.7% (2/35, 95%CI= 0–13.4). The mean larval burdens in two positive samples were 0.05 and 6 larvae per gr tissue muscle, respectively. The PCR reaction, using specific primers, yielded two 367 bp and 195 bp bands on agarose gel for ITS 2 and rrnS, respectively.
There is a hidden burden of Trichinella spp. infection in wild boar population in Iran. Moreover, T. britovi is the prevalent species circulating in wild boars of Iran. Therefore, education of the hunters and other consumers should be performed about the risk of consumption of raw or undercooked meat and meat products from wild boars.
PMCID: PMC5734972
Wild boar; Meat; Human trichinellosis; Iran
2.  Syphacia obvelata: A New Hope to Induction of Intestinal Immunological Tolerance in C57BL/6 Mice 
The ability of nematodes to manipulate the immune system of their host towards a Th2 and T regulatory responses has been proposed to suppress the inflammatory response. Clinical trials have proposed a useful effect of helminth infections on improvement of inflammatory disorders. In this study, we investigated the immunomodulatory effect of Syphacia obvelata infection to induce intestinal tolerance in C57BL/6 mice. Mice were infected through the cagemates with self-infected BALB/c mice. Four weeks post-infection, expression levels of IFN-γ, TNF-α, IL-17, and IL-10 were assessed in the supernatant of mesenteric lymph node (MLN) culture. Foxp3+Treg were measured in MLN cells by flow cytometry. In the S. obvelata-infected group, the percentage of Tregs (5.2±0.4) was significantly higher than the control (3.6±0.5) (P<0.05). The levels of IL-10 (55.3±2.2 vs 35.2±3.2), IL-17 (52.9±3.8 vs 41±1.8), IFN-γ (44.8±4.8 vs 22.3±2.3) and TNF-α (71.1±5.8 vs 60.1±3.3) were significantly increased in infected mice compared to the control group (P<0.05). The above results showed the potential effects of S. obvelata to induce intestinal tolerance. Therefore, it seems that S. obvelata may increase the immunological suppressive function in the intestinal tract.
PMCID: PMC5594727  PMID: 28877578
Syphacia obvelata; T regulatory cell; Foxp3; C57BL/6; inflammatory disorder; intestinal tolerance
3.  Prevalence, Clinical Manifestations and Genotyping of Cryptosporidium Spp. in Patients with Gastrointestinal Illnesses in Western Iran 
Iranian Journal of Parasitology  2017;12(2):169-176.
Cryptosporidium species are recognized as important gastrointestinal pathogens. This study was conducted to identify the prevalence, clinical manifestations and genotyping of Cryptosporidium spp. in patients with gastrointestinal illnesses (GIs) in western Iran.
Overall, 1301 fecal samples were collected from patients with GIs referred to the 12 clinical laboratories in Nahavand County, west of Iran. Modified Ziehl-Neelsen staining method was used to identify the oocysts. DNA was extracted from positive samples and Cryptosporidium spp. were characterized by Nested PCR and sequence analysis of the 60-kDa glycoprotein (gp60) gene. Data analysis was performed using SPSS ver. 16.
Prevalence of cryptosporidiosis was 1.3% (17/1301). Cryptosporidium infection was significantly associated with vomiting and nausea (P=0.001, OR=0.013; CI 95%=0.004– 0.044), abdominal pain (P=0.018, OR=0.073; CI 95%=0.008– 0.633) and diarrhea (P=0.001, OR=0.092; CI 95%=0.023– 0.362). Of the 17 isolates typed, 11 belonged to the C. parvum IId subtype family (subtypes IIdA26G1 and IIdA20G1) and six belonged to the C. parvum IIa subtype family (subtypes IIaA15G2R1 and IIaA16G3R1). There was no significant difference between sub-type families IIa and IId in occurrence of clinical symptoms (P= 0.75).
Improved hygiene and avoidance of contact with animals and contaminated soil should be advocated to reduce the occurrence of Cryptosporidium infections, especially in children.
PMCID: PMC5527026  PMID: 28761476
Cryptosporidiosis; Clinical manifestations; Genotyping; Gastrointestinal illnesses; Iran
4.  Molecular Identification of Leishmania Species in a Re-Emerged Focus of Cutaneous Leishmaniasis in Varamin District, Iran 
Cutaneous leishmaniasis (CL) is one of the most important neglected tropical diseases and a major public health challenge in Iran caused by Leishmania spp and transmitted by phlebotomine sand flies. The number of CL cases has shown an increasing pattern all over the country, including the district of Varamin, southeast of Tehran, Iran. This study aimed to identify the Leishmania spp isolated from CL patients using molecular methods in Varamin during 2012–2013.
Exudate materials collected from the swollen edge of the skin lesions of 44 parasitological positive CL patients by disposable lancet. They were referred to Varamin Health Center by physician. The samples were subjected to molecular method for Leishmania species identification.
The digestion pattern of restriction enzyme revealed that 37 (84.1%) CL patients were infected with L. major and 7 (15.9%) were infected with L. tropica. They were mostly male than female. More than half of the patients (58%) had multiple lesions, and they were mostly observed on extremities, 34.1% on legs and 29.5% on hands. Lesions were mostly of wet ulcerative type.
Dominancy of L. major provides more evidence that Varamin District probably could be considered as Zoonotic Cutaneous Leishmaniasis (ZCL) areas. More investigation on other epidemiological aspects of disease is needed.
PMCID: PMC5629294
Cutaneous leishmaniasis; PCR-RFLP; Leishmania tropica; Leishmania major; Iran
5.  Frequency of Cutaneous Leishmaniasis and Species Identification in Suspected Individuals from Golestan Province, Northern Iran in 2014 
Iranian Journal of Public Health  2016;45(10):1348-1354.
Leishmaniasis is a zoonotic disease caused by species of protozoa of the genus Leishmania. In recent years, incidence of cutaneous leishmaniasis has increasing trend in Golestan Province, North of Iran. The aim of the present study was to identify the frequency of cutaneous leishmaniasis using PCR-RFLP in patients referred to Kalaleh Health Center, during 2013–14.
This descriptive cross-sectional study was conducted on 70 individuals with suspected cutaneous leishmaniasis that referred to health center of Kalaleh County, Golestan Province, Northern Iran, from Sep 2013 to Nov 2014. Samples of cutaneous lesions were examined microscopically. DNA was extracted from all of the positive smears and PCR was done on ITS-1 gene. RFLP was performed using HaeIII enzyme for species identification.
Totally, 38 out of the 70 (54.3%) suspected individuals including 22 males (57.9%) were found positive by microscopic examination. All of microscopically positive samples were confirmed to be positive for Leishmania DNA (approximately 340 bp bands were detected). RFLP revealed 140 bp and 200 bp bands (approximate size), indicative of L. major.
The detected species of studied region was L. major. Cutaneous leishmaniasis has high prevalence in Kalaleh County, thus more studies on leishmaniasis in the animal reservoirs, comparison of homology of animal and human isolates and a survey regarding natural infection of vectors in this region is highly recommended.
PMCID: PMC5149499  PMID: 27957442
Leishmania major; ITS-1; Cutaneous leishmaniasis; Iran
In Iran, both Plasmodium vivax and P. falciparum malaria have been detected, but P. vivax is the predominant species. Point mutations in dihydrofolate reductase (dhfr) gene in both Plasmodia are the major mechanisms of pyrimethamine resistance. From April 2007 to June 2009, a total of 134 blood samples in two endemic areas of southern Iran were collected from patients infected with P. vivax and P. falciparum. The isolates were analyzed for P. vivax dihydrofolate reductase (pvdhfr) and P. falciparum dihydrofolate reductase (pfdhfr) point mutations using various PCR-based methods. The majority of the isolates (72.9%) had wild type amino acids at five codons of pvdhfr. Amongst mutant isolates, the most common pvdhfr alleles were double mutant in 58 and 117 amino acids (58R-117N). Triple mutation in 57, 58, and 117 amino acids (57L/58R/117N) was identified for the first time in the pvdhfr gene of Iranian P. vivax isolates. All the P. falciparumsamples analyzed (n = 16) possessed a double mutant pfdhfrallele (59R/108N) and retained a wild-type mutation at position 51. This may be attributed to the fact that the falciparum malaria patients were treated using sulfadoxine-pyrimethamine (SP) in Iran. The presence of mutant haplotypes in P. vivax is worrying, but has not yet reached an alarming threshold regarding drugs such as SP. The results of this study reinforce the importance of performing a molecular surveillance by means of a continuous chemoresistance assessment.
PMCID: PMC4804553  PMID: 27007559
Plasmodium vivax; Plasmodium falciparum; Pyrimethamine; Point mutations and drug resistance
7.  An experimental model of colitis induced by dextran sulfate sodium from acute progresses to chronicity in C57BL/6: correlation between conditions of mice and the environment  
To induce acute colitis progresses to chronicity in C57BL/6 mice by dextran sulfate sodium.
Murine models are essential tools to understand IBD pathogenesis. Among different types of chemically induced colitis models, the dextran sulfate sodium (DSS)-induced colitis model is the most common model of IBD, due to its simplicity.
Patients and methods:
Male C57BL/6 mice 6–8 weeks old, were collected and matched by age with controls. C57BL/6 mice treated with 2 cycles of 3.5% DSS for 4 days and 4 days of pure water between each cycle. After that, mice were sacrificed and the entire colon was removed. Small sections of the colon were fixed in formaldehyde, embedded in paraffin and sectioned with a microtome. Sections were stained with hematoxylin eosin to analyses the degree of inflammation.
After the first cycle oral administration of DSS, mice with severe and visible rectal bleeding and diarrhea entered into the acute phase. After day 4-5, bleeding and diarrhea were improved and mice entered into the chronic phase with peak levels of weight loss. Macroscopically, the inflammation was predominantly located in the distal colon. Microscopically, examination of the distal colon sections showed a decrease number of goblet cells, loss of crypts, signs of surface epithelial regeneration and moderate to severe infiltration of inflammatory cells in the mucosa.
In order to achieve an experimental colitis model, our protocol is recommended for future therapies in IBD experimental modeling.
PMCID: PMC4702041  PMID: 26744614
Inflammatory bowel disease; Murine models; DSS; C57BL/6
8.  Prevalence and Genetic Characterization of Cryptosporidium Spp. In Diarrheic Children from Gonbad Kavoos City, Iran 
Iranian Journal of Parasitology  2015;10(3):441-447.
Background: Cryptosporidium is an intestinal protozean parasite causing waterborne and foodborne outbreaks of diarrheal diseases. The present study was performed in order to find prevalence and subtypes of Cryptosporidium among children with diarrhea in Gonbad Kavoos City, Northern Iran.
Methods: Diarrheic samples were collected from 547 children. The initial parasitological diagnosis was made based on detection of oocysts using the modified Ziehl-Neelsen acid-fast staining method. The positive microscopically samples were selected for sequence analysis of partial 60 kDa glycoprotein (gp60) gene.
Results: Out of 547 collected samples, 27 (4.94%) were positive for Cryptosporidium oocysts. Fifteen from 27 positive samples successfully amplified in PCR. Sequences analysis of gp60 gene in 15 Cryptosporidium isolates revealed that all of them (100%) were C. parvum. The results showed three subtypes of IIa subtype family (7 cases) including IIaA16G2R1, IIaA17G1R1, IIaA22G3R1 and one subtype of IId subtype family (8 cases). The most common allele was IId A17G1d (53.3%).
Conclusion: The predominance of zoonotic subtype families of C. parvum species (IIa, IId) in the present study is in concordance with previous studies in Iran and emphasizes the significance of zoonotic transmission of cryptosporidiosis in the country.
PMCID: PMC4662744  PMID: 26622299
Cryptosporidium; Subtypes; Gp60 gene; Children; Iran
9.  Cloning and Sequence Analysis of Recombinant Plasmodium vivax Merozoite Surface Protein 1 (PvMSP-142 kDa) In pTZ57R/T Vector 
Iranian Journal of Parasitology  2015;10(2):197-205.
Carboxy-terminal 42 kDa region of Plasmodium vivax merozoite surface protein-1 is considered as an important antigen in blood stage. Since, this region has been observed to be polymorphic among isolates of P. vivax, it is significant to survey on different regions of this antigen in various areas of the world.
In the present study, the genetic diversity of cloned PvMSP-142 kDa gene from an Iranian patient is analyzed. Parasite DNA was extracted from a P. vivax - infected patient in Iran. The region of PvMSP-142 kDa was amplified by PCR, cloned into pTZ57R/T vector and then sequenced.
Sequencing of cloned PvMSP-142 kDa gene clearly has a high degree of homology (95%) with reference Sal-I sequence and also with the homogeneous sequences from some studied countries (97%). Thirty eight SNPs (single nucleotide polymorphism) were identified in cloned PvMSP-142 kDa gene which the mutations had localized in the 33 kDa fragment (PvMSP-133 kDa), while there was nearly no variation in the 19 kDa fragment (PvMSP-119 kDa). 2 out of 38 mutations were found as to be novel haplotypes.
High similarity of cloned PvMSP-142 kDa gene in comparison to reference sequence and other sequences could be beneficial as a remarkable molecular marker for serological diagnostic kits of P. vivax in malarious neighboring countries of Iran and around the world.
PMCID: PMC4522295  PMID: 26246817
Plasmodium vivax; Recombinant MSP-1 42 kDa; Sequencing; Iran
10.  Potential treatment of inflammatory bowel disease: a review of helminths therapy 
An inflammatory bowel disease (IBD) is most common in highly industrialized Western countries but uncommon in less developed areas of the world where helminths are frequent. The hygiene hypothesis proposes that the recent increase in allergic and autoimmune diseases is due to modern highly hygienic life styles and medical conditions. Loss of routine exposure to parasitic helminths, as a result of increasing lifestyle-associated factors, may be one factor leading to the increased disease prevalence.
In animal models and clinical trials of IBD, gastrointestinal nematodes colonization suppresses intestinal inflammation through multiple mechanisms including induction of innate and adaptive regulatory circuits. Studies using helminths like Trichuris suis or Necator americanus showed that these helminths are safe and may be effective therapeutic approaches for the control of IBD and other immune diseases. The aim of present review was to exploring the therapeutic use of helminths for the control of IBD.
PMCID: PMC4017549  PMID: 25436093
Inflammatory bowel disease; Helminthes; Therapeutic
11.  Application of Multiplex PCR for Detection and Differentiation of Entamoeba histolytica, Entamoeba dispar and Entamoeba moshkovskii 
Iranian Journal of Parasitology  2014;9(4):466-473.
Entamoeba moshkovskii and E. dispar are impossible to differentiate microscopically from the pathogenic species E. histolytica. Multiplex polymerase chain reaction (Multiplex PCR) is a widespread molecular biology technique for amplification of multiple targets in a single PCR experiment.
For detection and differentiation of the three-microscopy indistinguishable Entamoeba species in human, multiplex PCR assay using different DNA extraction methods was studied. A conserved forward primer was derived from the middle of the small-subunit rRNA gene, and reverse primers were designed from signature sequences specific to each of these three Entamoeba species.
A 166-bp PCR product with E. histolytica DNA, a 580-bp product with E. moshkovskii DNA and a 752-bp product with E. dispar DNA were generated in a single-round and multiplex PCR reaction.
We recommend this PCR assay as an accurate, rapid, and effective diagnostic method for the detection and discrimination of these three Entamoeba species in both routine diagnosis of amoebiasis and epidemiological surveys.
PMCID: PMC4345085  PMID: 25759727
Entamoeba hitolytica; Entamoeba dispar; Entamoeba moshkovskii; DNA extraction; Multiplex PCR
12.  Genotyping of Cryptosporidium spp. in clinical samples: PCR-RFLP analysis of the TRAP-C2 gene 
The aim of the present study was to determine the species and genotypes of Cryptosporidium spp. among children with diarrhea by PCR-RFLP using the TRAP-C2 gene.
Cryptosporidium is a globally distributed protozoan parasite and one of the most common causes of infection and diarrhea in humans.
Patients and methods
Four hundred and sixty nine stool samples were collected from children less than 12 years with diarrhea who had been referred to Pediatrics Medical Centers in Gazvin provinces. The presence of Cryptosporidium oocysts was determined by Ziehl-Neelsen acid fast staining, then, genomic DNA was extracted from positive samples and nested PCR-RFLP was performed to amplify the TRAP-C2 gene.
The overall prevalence of Cryptosporidium infection in children was 2.5 %. Results of nested PCR amplification showed that of 12 positive children samples, 10 (83.3%) were belonged to C. parvum, followed by C. hominis in 1 (8.3%) and mixed infection in 1 isolate (8.3%).
This study showed that Cryptosporidium parvum (the zoonotic genotypes) is more prevalent than other Cryptosporidium species in children from this area. This suggests that zoonotic transmission is the main mode of transmission of Cryptosporidium infection in Iran.
PMCID: PMC4017402  PMID: 24834152
Cryptosporidium; Genotypes; TRAP-C2 gene

Results 1-12 (12)