Molecular Characterization of Acanthamoeba Isolates from Surface Resting Waters in Northwest Iran.

BACKGROUND: Acanthamoeba is an opportunistic amphizoic protozoan found in different fresh water sources. The aim of this study was to identify and characterize Acanthamoeba isolates from surface resting waters, in Northwest Iran. METHODS: Samples were collected from twenty-two different areas, between May and Sep 2014. After filtration, samples were cultivated on non-nutrient agar. The extracted DNAs were amplified and sequenced using partial 18S rRNA in order to genotype and phylogenetic analyses. RESULTS: Thirty-four (68%) out of 50 collected samples were positive for free-living amoebae based on both culture and morphological characterizations but 28 samples were identified as Acanthamoeba spp. by PCR. Sequentially, one isolate was identified as A. lenticulata, (T5) (AN: KP940443, identity 99.7%-100%, and divergence 0.3%) whilst other sequenced isolates identified Acanthamoeba spp. (AN: KP940444-45) as very similar to A. rhysodes and A. royreba with identity 100% and divergence 0%. CONCLUSION: Surface resting waters in Northwest Iran, were potentially contaminated with pathogenic amphizoic protozoan. Further studies will be required to determine other Acanthamoeba species and genotypes in the region.

Molecular Detection and Genotyping of Toxoplasma gondii in Chicken, Beef, and Lamb Meat Consumed in Northwestern Iran.

BACKGROUND: Toxoplasma gondii, the obligate, intracellular parasite afflicts human in diverse ways such as ingestion of tissue cysts in undercooked meat. The aim of this study was to assess the prevalence and genotyping of Toxoplasma isolated from meat samples being consumed in East Azerbaijan, Northwest of Iran. METHODS: Overall, 150 samples including chicken, beef and lamb meat were collected from retailers in different regions of Tabriz, northwestern Iran during April to September 2014. Molecular detection was done by amplifying B1 gene and T. gondii surface antigen 2 (SAG2) loci. For genotyping of T. gondii, restriction fragment length polymorphism (RFLP) was performed using Sau3AI and HhaI restriction enzymes. Finally, three positive isolate from each host was sequenced to evaluate and phylogenetic analysis. RESULTS: Overall, 26 (17.33%) samples were positive for T. gondii including 4 (8%) isolate from chicken, 8 (16%) isolates from cattle and 14 (28%) isolates from sheep. According to the RFLP patterns, sequencing and phylogenetic results, all of the samples were identified as genotype I. CONCLUSION: The results indicated a potential risk of transmission of the disease through the consumption of infected meat. This is particularly important especially for pregnant women and immuno-compromised patients and this suggests that the training on the prevention of infection is necessary.

Molecular characterization and sequence analysis of Echinococcus granulosus from sheep isolates in East Azerbaijan province, northwest of Iran.

Echinococcus granulosus as an etiologic agent of hydatid cyst is one of the most important zoonotic helminthes in the world that causing enormous economic and health losses. The aim of this study was to evaluate genotype of E. granulosus isolated from sheep using mitochondrial cytochrome c oxidase subunit 1 (cox1) gene and sequencing method in East Azerbaijan province, northwest of Iran. Nineteen sheep hydatid cyst samples were collected. Genomic DNA was extracted from protoscoleces using commercial DNA extraction kit. Mitochondrial cox1 region was amplified by polymerase chain reaction (PCR) and all isolates were sequenced. Afterward, sequences were analyzed for determination of genotypes by related software. G1 (94.73 %) and G3 (5.27 %) genotypes were identified from the isolates which out of 19 hydatid cysts, 17 samples were G1B, 1 sample G1D and the other one had G3 genotype. Results of this study indicate that common sheep strain (G1); especially G1B is the dominant subtype of E. granulosus in East Azerbaijan province.

The correlation between Toxoplasma gondii infection and Parkinson's disease: a case-control study.

Toxoplasma gondii is an obligate intracellular parasite that infects all nucleate cells of vertebrates. Human infected by vertical transmission and also using raw or undercooked meat or food and water that contaminated with mature oocysts. Parkinson's disease as neurodegenerative disease affects people above 60 years. Due to high prevalence of toxoplasmosis in Iran and evidence about effects of T. gondii on neurodegenerative diseases, this study has been conducted to investigate possible correlation between Toxoplasma and Parkinson's disease in Iran. Seventy five Parkinson's patients and equal healthy volunteers were enrolled. After obtaining informed consent and sociodemographic features, 5 ml blood sample were collected and then anti-Toxoplasma IgG and IgM levels were examined by ELISA method. Data was analyzed with Chi-squre and Fisher's test by usig stata 11 software. Binary logistic regression was used for multivariate analysis in assessing the correlation between toxoplasmosis and Parkinson. Eighty five percent of Parkinson's group and 90.3 % of control group were positive for anti-Toxoplasma IgG antibody. In this investigation no statically differences were observed between groups and age, gender, residency and using raw or undercooked meat. There is no significant association between IgG positive titer and Parkinson's disease. However, statistically significant association was found between Parkinson and keeping cat (P = 0.03) as well as the using of undercooked egg (P = 0.004). Although there is high level of anti-Toxoplasma IgG antibody in Parkinson's patients which reflects chronic Toxoplasma infection; we couldn't detect any statistical association between T. gondii infection and Parkinson's disease.

Toxoplasmosis and Alzheimer: can Toxoplasma gondii really be introduced as a risk factor in etiology of Alzheimer?

Alzheimer is a progressive neurological disease that results in irreversible loss of neurons and includes about two thirds of all cases of dementia. Toxoplasma gondii may be an important infectious agent involved in neurodegenerative diseases. The aim of this study was to investigate the correlation between Toxoplasma as an etiologic agent in the progress of Alzheimer's disease. This case control study was conducted on 75 Alzheimer's patients and 75 healthy volunteers. Blood samples were obtained and anti-Toxoplasma IgG and IgM tests were done by using ELISA technique. DNA was extracted from buffy coat and then GRA6 gene and SAG2 loci were amplified by PCR and nested PCR, respectively. Chi-square, Fisher's test, and binary logistic regression were used for data analysis. A percentage of 61.3 % of Alzheimer's patients and 62.6 % of healthy volunteers were positive for anti-Toxoplasma IgG but all participants were negative for anti-Toxoplasma IgM. There were no significant differences between Alzheimer's patients with their controls in terms of anti-Toxoplasma IgG antibody (P = 0.5). Due to lack of positive IgM sample, results of the molecular methods were negative by GRA6 and SAG2 fragments amplification. This result shows that, infection with T. gondii cannot be considered as a risk factor for etiology and developing Alzheimer's disease.

Latent and Asymptomatic Toxocara Infection among Young Population in Northwest Iran: The Necessity of Informing People as a Potential Health Risk.

Objectives. This study was designed to determine the frequency of anti-Toxocara antibodies in youngsters aging from 2 to 20 years in northwest Iran. Materials and Methods. 397 samples were taken randomly almost equally from four locations in Urmia, west Azerbaijan, during August 2014 to September 2015. Anti-Toxocara IgG antibody assays were done on sera by using ELISA kit (IBL, Germany). In order to prevent cross-reaction, the samples of the patients who are infected with other parasites in stool exam, especially Ascarididae family, were also excluded. SPSS 16.0 software was used for statistical analysis. Results. 12 (3%) of the serum samples were positive for anti-Toxocara IgG. According to the Chi-square analysis, risk factors such as mother's educational level, keeping dogs or cats as pets, and history of coughing were related to Toxocara infection (P < 0.05). There was no relationship between toxocariasis and gender, history of onychophagy, pica, fever, abdominal pain, and anorexia; however, we found a significant relationship between Toxocara infection and chronic coughing (P = 0.045). Conclusion. Toxocariasis in northwest Iran can be considered as a public health problem. This study may also help to increase the awareness about this infection.

The first morphometric and phylogenetic perspective on molecular epidemiology of Echinococcus granulosus sensu lato in stray dogs in a hyperendemic Middle East focus, northwestern Iran.

BACKGROUND: Hydatidosis is considered to be a neglected cyclo-zoonotic disease in Middle East countries particularly northwestern Iran which is caused by metacestode of tapeworm Echinococcus granulosus sensu lato. Human hydatidosis is a high public health priority in the area, however there is little known from a morphometric and phylogenetic perspective on molecular epidemiology of adult Echinococcus spp. in Iranian stray dogs. METHODS: 80 dogs (38 males and 42 females) were collected during June 2013 to April 2014 in northwestern Iran. The isolated parasites from each dog were distinguished by morphometric keys including small, large hook length and blade length. Subsequently, isolates were confirmed by sequencing of mitochondrial cytochrome oxidase subunit 1 gene. RESULTS: 16 (8 males and 8 females) (Prevalence 20%) out of 80 dogs were infected to genus Echinococcus. With regard to demographic factors, the frequency of parasitism in both male, female adults and their age groups showed no difference (P > 0.05). The phylogenetic analyses of cox1 sequences firmly revealed the 13 sheep strains (G1), one buffalo strain (G3), one camel strain (G6) and one mixed infection. The findings of rostellar hook morphology show an intraspecies variation range among G1 isolates. However, hook measurements in Echinococcus derived from G1 (sheep strain) were not a significant difference from those G6 and G3 strains. Six unique haplotypes were identified containing a high range of diversity (Haplotype diversity 0.873 vs. Nucleotide diversity 0.02). CONCLUSIONS: First presence of camel strain (G6) in this region seems to indicate that potential intermediate hosts play a secondary role in the maintenance of camel-dog biology. Current findings have heightened our knowledge about determination of Echinococcus prevalence, strains of taxonomy and genotypic trait of parasite in Iranian stray dogs which will also help in the development of strategies for monitoring and control of infected stray dogs in the area.

Designing and conducting in silico analysis for identifying of Echinococcus spp. with discrimination of novel haplotypes: an approach to better understanding of parasite taxonomic.

The definitive identification of Echinococcus species is currently carried out by sequencing and phylogenetic strategies. However, the application of polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) patterns is not broadly used as a result of heterogeneity traits of Echinococcus genome in different regions of the world. Therefore, designing and conducting a standardized pattern should indigenously be considered in under-studied areas. In this investigation, an in silico mapping was designed and developed for eight Echinococcus spp. on the basis of regional sequences in Iran and the world. The numbers of 60 Echinococcus isolates were collected from the liver and lungs of 15 human, 15 sheep, 15 cattle, and 15 camel cases in Semnan province, Central Iran. DNA samples were extracted and examined by polymerase chain reaction of ribosomal DNA (rDNA) internal transcribed spacer 1 (ITS1) and PCR-RFLP via Rsa1 endonuclease enzyme. Moreover, 15 amplicons of cytochrome oxidase 1 (Cox1) were directly sequenced in order to identify the strains/haplotypes. PCR-RFLP and phylogenetic analyses revealed firmly the presence of the G1 and G6 genotypes with heterogeneity (three novel haplotypes) of Cox1 gene although no other expected genotypes were found in the region. Finding shows that the identification of novel haplotypes along with discrimination of Echinococcus spp. through regional patterns can unambiguously illustrate the real taxonomic status of parasite in Central Iran.

Seroprevalence of Toxoplasma Gondii Infection among High School Girls in Ajabshir from East Azarbaijan Province, Iran.

INTRODUCTION: Toxoplasmosis is a disease parasite which can infect human and animals. The infection may be serious if is transmitted to the fetus during pregnancy. The aim of this study was to determine the prevalence of specific antibodies and the associated risk factors for toxoplasmosis in students attending high-school in Ajabshir. METHODS: In this descriptive study, 549 blood samples were collected from high school girls. The samples divided into two groups (147 and 402 samples from rural and urban schools respectively). IgG and IgM specific antibodies to Toxoplasma gondii were detected using enzyme-linked immunosorbent assay (ELISA). RESULTS: The results of study showed that from 402 urban samples, 50 (12.4) and 34(8.5) cases and from 147 rural samples, 38 (25.9) and 32 (21.8) cases were seropositive for anti-Toxoplasma IgG and IgM antibodies respectively. Of the risk factors studied, the significant association was found between T. gondii-specific antibodies with residency and age. CONCLUSION: Based on data found in our study, 87.6% of young girls from urban areas in Ajabshir did not have antibodies to Toxoplasma and this is a very important issue, because these young women were in fertile age. Therefore required Preventive and control programs especially in these cases in order to reduce the rate of disease.

Molecular and parasitological study of cryptosporidium isolates from cattle in ilam, west of iran.

BACKGROUND: Cryptosporidiosis is one of the most important parasitic infections in human and animals. This study was designed for survey on the prevalence of Cryptosporidium infection in farms of Ilam, west of Iran, using parasitology method and genotyping by Nested PCR-RFLP. METHODS: Fecal samples of 217 cattle were collected fresh and directly from the rectum of cattle. All of the samples were examined by microscopic observation after staining with modified Ziehl-Neelsen (MZN). Genomic DNA extracted by using EURx DNA kit. A Nested PCR-RFLP protocol amplifying 825 bp fragment of 18s rRNA gene conducted to differentiate species and genotyping of the isolates using SspI and VspI as restriction enzymes. RESULTS: The prevalence of Cryptosporidium infection in cattle using both methods is 3.68%. Most of the positive cattle were calves under six months. Species diagnosis carried out by digesting the secondary PCR product with SspI that C. parvum generated 3 visible bands of 448, 247 and 106 bp and digested by VspI restriction enzyme generated 2 visible bands of 628 and 104bp. In this investigation all of the positive samples were Cryptosporidium parvum. CONCLUSION: C. parvum (bovine genotype) detected in all positive cattle samples in Ilam, west of Iran. The results of the present study can help for public health care systems to prevention and management of cryptosporidiosis in cattle and the assessment of cattle cryptosporidiosis as a reservoir for the human infection.