Brain Tissue Cysts in Infected Mice with RH-Strain of Toxoplasma gondii and Evaluation of BAG1 and SAG1 Genes Expression.

BACKGROUND: Toxoplasma gondii is an obligate intracellular parasite that infects humans at high prevalence rates. The virulent RH strain of T. gondii is generally considered to have lost its cyst forming capacity. This study performed to obtain tissue cysts in mice infected with tachyzoites of RH strain treated with sulfadiazine (SDZ). It provides the opportunity to analyze the conversion of tachyzoite to bradyzoite stage of the RH strain, followed by stage-specific gene-expression analyzing. METHODS: Two groups of Swiss-Webster and BALB/c mice were infected subcutaneously with 10(4) tachyzoites of T. gondii, RH strain and given SDZ (300 mg/l) with NaHCO3 (5 g(-1)) in drinking water from day1 to day 14 post infection (p.i). The infected mice were sacrificed on day 50 post infection. Their brains were removed and the numbers of tissue cysts were microscopically counted. Total RNA was extracted from brains and cDNA synthesis was carried out. Finally, RT-PCR (Reverse transcription PCR) was used to detect the expression of bradyzoite (BAG1) and tachyzoite (SAG1) specific genes during tachyzoite / bradyzoite stage conversion. RESULTS: Sixty five percent of all infected mice were survived. Cysts were detectable in mice brain (45%) on day 50 p.i. Also RT-PCR of the brain samples was positive for SAG1 and BAG1. CONCLUSION: It seems that conversion of tachyzoites to bradyzoites in brain of mice undergoing SDZ was not completed until 50 days after inoculation.

Activity of Superoxide Dismutase (SOD) Enzyme in the Excretory-Secretory Products of Fasciola hepatica and F. gigantica Parasites.

BACKGROUND: The aim of this study was to compare superoxide dismutase (SOD) activity in Fasciola hepatica and Fasciola gigantica parasites. MATERIALS: F. gigantica and F. hepatica helminths were collected from abattoir and cultured in buffer media for 4 h at 37 °C. Excretory-Secretory (ES) products were collected, centrifuged and stored at -20(°)C. E-S protein concentration was measured by Bradford method and SOD activity was detected using RANSOD kit (Randox Lab. Crumlin, UK). Statistical t-test was conducted for analysis of results. RESULTS: Protein concentration for F. hepatica and F. gigantica were obtained 7.293 ug/ml and 19.65 ug/ml respectively and SOD activity as 0.721 U/ml and 1.189 U/ml, in that order. ES protein concentration of two species was significantly different (P<0.05), however the difference of SOD activity of two species was not significant. CONCLUSION: Two species of Fasciola have comparable SOD biochemical defense enzyme and can help us explain the parasite survival in host tissue.

Use of Single-enzyme PCR-restriction Digestion Barcode Targeting the Internal Transcribed Spacers (ITS rDNA) to Identify Dermatophyte Species.

BACKGROUND: Dermatophytes are the most common causative agents of superficial mycoses. Species identification of these fungi is important from therapeutic and epidemiological point of wive. Traditional approaches for identification of dermatophytes at the species level, relying on macroscopic and microscopic features of the colonies, usually are time-consuming and unreliable in many circumstances. Recently a broad varieties of rapid and accurate DNA-based techniques were successfuly utilized for species delineation of dermatophytes. METHODS: The ITS1-5.8S-ITS2 region of rDNA from various reference strains of dermatophyte species were amplified using the universal fungal primers ITS1 and ITS4.The PCR products were digested by a single restriction enzyme, MvaI. The enzyme was evaluated in both in silico and practical PCR-RFLP assay to find the exact differentiating restriction profiles for each species. To validate the standardized PCR-RFLP system, all tested strains were subjected to sequencing and sequence analysis. RESULTS: The obtained RFLP patterns were specific for many species including T. interdigitale, T. rubrum, T. violaceum, M. persicolor, M. audouinii, M. nanum (A. obtusum) and E. floccosum but were similar for some closely related species such as M. canis / M. ferrugineum. Sequencing of the ITS1-5.8S-ITS2 fragment from all type strains affirmed the RFLP findings. CONCLUSION: It was practically revealed that the ITS-PCR followed by MvaI-RFLP is a useful and reliable schema for identification and differentiation of several pathogenic species and can be used for rapid screening of even closely related species of dermatophytes in clinical and epidemiological settings.

Serum selenium, zinc, and copper in early diagnosed patients with pemphigus vulgaris.

BACKGROUND: Pemphigus vulgaris is a life threatening, blistering skin disease. It is an autoimmune abnormality. Due to involvement of oral cavity and pharynx, patients are at risk of nutrients deficiency. The aim of this study was to evaluate the status of selenium, copper, and zinc in these patients. METHODS: In a case-control study, 43 newly diagnosed pemphigus vulgaris patients were compared with 58 healthy people from 2009 to 2010. The severity of the disease was estimated according to Harman's scores. Serum selenium was measured with atomic absorption but serum zinc and copper concentrations were determined spectrophotometrically. Data were compared with independent t test. Correlations were evaluated by Pearson correlation test. RESULTS: Both groups were the same based on sex, age, and weight and body mass index. The mean duration of disease was 5.6 month. The oral and skin severities were 1.79 and 2.3 respectively, based on Harman's scores. Serum selenium of pemphigus patients was significantly less than that of healthy people (P<0.001). Serum copper was negatively correlated with duration of disease in males (P=0.02, r=-0.5). CONCLUSIONS: Pemphigus vulgaris negatively affects on serum selenium, copper and zinc. It seems that serum selenium, copper and zinc decrease as the disease lasts longer.

Evidence for augmented oxidative stress in the subjects with type 1 diabetes and their siblings: a possible preventive role for antioxidants.

BACKGROUND/OBJECTIVES: Oxidative stress (OS) is thought to be involved in both development of type 1 diabetes (T1D) and its further complications. In this study, certain biomarkers of OS were compared among the subjects with T1D, their non-diabetic siblings and unrelated healthy controls. SUBJECTS/METHODS: Known cases of T1D from both sexes aged 5-25 years were enrolled in a case-control study (n(1)=60). There were two control groups; non-diabetic siblings (n(2)=60) and unrelated apparently healthy subjects (n(3)=60). Anthropometric, dietary and laboratory assessments were done. RESULTS: There was no significant difference in dietary data among the groups. Total antioxidant capacity was significantly lower in T1D than both related and unrelated controls (1.6 ± 0.05, 1.7 ± 0.05 and 1.8 ± 0.06 mmol BSA equivalent/l, respectively, P=0.044). Both T1D subjects and their siblings showed lower glutathione peroxidase (GSH-px) levels (median (interquartile range): 22.2 (28.6), 29.9 (23) and 41.8 (73.6) U/ml, respectively, P=0.006). On the contrary, superoxide dismutase concentrations were significantly higher in T1D group and the siblings than unrelated healthy controls (243 (45.3), 157.8 (176.9) and 27.9 (8.7) U/l, respectively, P<0.001). Serum concentrations of GSH correlated with energy intake in the siblings (r=0.521, P<0.001) and unrelated controls (r=0.268, P=0.042) but not in T1D group. The associations remained significant after controlling for blood glucose (r=0.437, P=0.001 and r=0.420, P=0.011, respectively) in both the groups. CONCLUSION: Augmented OS in the siblings may indicate an increased requirement for antioxidants in genetically diabetes-prone subjects.

Vitamin D status and the predictors of circulating T helper 1-type immunoglobulin levels in Iranian subjects with type 1 diabetes and their siblings: a case-control study.

BACKGROUND: Vitamin D status has been linked to both T helper (Th)1/Th2 balance and susceptibility to type 1 diabetes (T1D). The present study aimed to evaluate vitamin D status and its relation to Th1/Th2 balance in subjects with T1D, their siblings and unrelated healthy controls during autumn and winter 2008-2009. METHODS: A case-control study was conducted on subjects with T1D (n(1) = 60) and two control groups comprising nondiabetic siblings (n(2) = 60) and unrelated healthy controls (n(3) = 60). Assessments of dietary intake, anthropometry, intact parathyroid hormone (iPTH) and 25(OH)D were performed. Serum levels of immunoglobulin (Ig)G(2) and IgE, as well as the IgG2/IgE ratio, were used to evaluate Th1/Th2 balance. Vitamin D status was defined based on circulating 25(OH)D as deficiency: <27.5 nm; insufficiency 27.5 ≤ 25(OH)D <50 nm; and sufficiency ≥50 nm. RESULTS: Vitamin D status did not differ significantly among the groups. Similarly, no significant difference in 25(OH)D, iPTH, IgG(2), IgE and IgG(2)/IgE was found. In multiple regression analysis of pooled data, PTH and body mass index were the predictors of IgG(2)/IgE. In the diabetic group, both PTH and age and, in siblings, PTH only, were the predictors of IgG(2)/IgE ratio. CONCLUSIONS: These data suggest PTH as the major predictor of immune deviance towards the Th1 response in both type 1 diabetic subjects and their siblings. Considering that the active form of vitamin D suppresses PTH production, it is hypothesised that vitamin D replenishment of just those who are genetically prone to the disease (i.e. siblings) may be regarded as a preventive measure against T1D.

Vector ecology and susceptibility in a malaria-endemic focus in southern Islamic Republic of Iran

This study aimed to carry out a malaria situation analysis, species composition and susceptibility levels of the main malaria vector, Anopheles stephensi, to different insecticides in Bashagard. A longitudinal survey was conducted in 2 randomly selected villages in Bashagard. Malaria vectors were sampled by dipping method for the larvae and hand catch, night-biting catch, total catch, and shelter pit collection for the adults. Standard WHO susceptibility tests were used for a variety of insecticides on F1 progeny of An. stephensi reared from wild-caught females. In total, 693 adult anopheline mosquitoes and 839 third and fourth-instar larvae were collected and identified. They comprised 7 species; the most abundant adult and larvae anopheline mosquito was An. Dthali [40.7% and 30.5% respectively]. An. Culicifacies[24.2%] and An. Stephensi [16.7%] were the next most common species for adult mosquitoes. An. Stephensi was fully susceptible to malathion and pyrethroid insecticides but resistant to DDT and tolerant to dieldrin

Serum zinc levels in children and adolescents with type-1 diabetes mellitus.

BACKGROUND: There have been very few studies, with contradictory results, on the zinc status of children and adolescents with type-1 diabetes mellitus. The objective of this cross-sectional study was to determine zinc status based on the serum zinc concentration in type-1 diabetic children and adolescents and compare it with that of healthy controls. METHODS: Thirty children and adolescents with type-1 diabetes mellitus, aged 6 to 18 years, and 30 age- and sex-matched healthy controls participated in the study. Serum zinc, fasting blood sugar, hemoglobin A(1c) and serum albumin were measured by flame atomic absorption spectrophotometry, enzymatic colorimetry, ion-exchange chromatography and colorimetry using bromocresol green methods, respectively. RESULTS: No statistically significant difference was found in the mean serum zinc concentration between diabetic patients and healthy controls (111.0 ± 3.1 and 107.1 ± 3.8 mg/dl respectively, P= 0.4). No correlations were found between the serum zinc levels and fasting blood sugar, hemoglobin A(1c), or the duration of the disease in the patients. CONCLUSION: The zinc levels of diabetic children and adolescents are not noticeably different compared to those of healthy controls and are independent of glycemic control and the duration of the disease.

Haemolymph Components of Infected & None Infected Lymnaea snails with Xiphidiocercariae.

BACKGROUND: In this study the haemolymph components of infected and none infected Lymnaea gedrosiana with xiphidiocercaria larvae was compared. METHODS: Five hundred Fifty Lymnaea snails were collected from Ilam and Mazandaran provinces, Iran, during 2008-2009. The snails were transported to the lab at Tehran University of Medical Sciences and their cercarial sheddings were studied. Haemolmyphs of snails were extracted and cells were counted using haemocytometer and cell-surface carbohydrate were recognized by conjugated lectin (Lentil). Haemolymph protein concentrations were measured by Bradford protein assay method and soluble protein compositions were determined on sodium dodecyl sulphate polyacrilamide gel electrophoresis (SDS-PAGE). RESULTS: From the 550 examined Lymnaea snails for cercariae, 27 snails were infected with xiphidiocercariae. Mean of haemolymph cells (haemocyte) number were obtained 93480±2.43 (cells/ml) for none infected snails (25 snail) and 124560±2800 (cells/ml) for infected snails (25 snail). Mannose carbohydrate was recognized on haemocyte of none infected and infected snails. Mean of protein concentration of haemolymph plasma was obtained as 1354±160 µg/ml (1.4 mg/ml) for none infected snails (25 snails) and 1802±138 µg/ml (1.8 mg/ml) for infected snail (25 snails). Comparing to none infected snails, the SDS-PAGE results of haemolymph plasma of infected snails, showed an extra protein band (70 kDa). The results showed a significant difference between the amounts and the kinds of proteins in haemolymph of infected and none infected snails. CONCLUSION: This information might be useful to understand of parasite detection, adhesion, engulfment and antigen agglutination by snail.

Seroepidemiological study of human hydatidosis in meshkinshahr district, ardabil province, iran.

BACKGROUND: The aim of this study was to conduct a sero-epidemiological survey in Meshkinshahr, Ardabil Province, northwestern Iran to detect the rate of hydatidosis in the city and nearby villages. Literature shows that no such study has been conducted so far. METHODS: Overall, 670 serum samples were collected from 194 males and 476 females from patients referred to different health centers of the region. All patients filled out a questionnaire and an informed consent. Sera were analyzed using indirect-ELISA test. Ten µg /ml antigens (Antigen B derived from hydatid cyst fluid), serum dilutions of 1:500 and conjugate anti-human coombs with 1:10000 dilutions were utilized to perform the test. Data analysis was conducted using SPSS software ver. 11.5. RESULTS: The seroprevalence of human hydatidosis was 1.79% by ELISA test in the region. This rate for females was 1.68% and males 2.6%, respectively. There was no significant difference as regards all factors studied and the seropositivity. According to job, farmers and ranchmen had the highest rate of infection as 3.17%. The sero-prevalence of infection was 2.6%% in illiterate people which showed the highest rate. As regards residency, urban life showed no significant difference with rural life (1.1% vs. 2.58%). Age group of 69-90 yr old, with 4.62% as prevalence had the highest rate of positivity. CONCLUSION: Obtained sero-prevalence of hydatidosis shows more or less a resemblance to other cities of Iran, although due to the specific condition of the city we expected more rate of sero-positivity.

The effects of vitamins e and d supplementation on erythrocyte superoxide dismutase and catalase in atopic dermatitis.

BACKGROUND: Atopic dermatitis is a public health problem worldwide. Increment of reactive oxygen species (ROS) production may be one of the contributing factors of tissue damage in atopic dermatitis. The present study was designed to determine the effect of vitamins E and/or D on erythrocyte superoxide dismutase and catalase activities in patients with atopic dermatitis. METHODS: In a randomized, double blind, placebo controlled clinical trial 45 atopic dermatitis patients were divided into four groups. Each group received one of the following supplements for 60 days: group A (n=11) vitamins E and D placebos; group B (n= 12) 1600 international unit (IU) vitamin D3 plus vitamin E placebo; group C (n=11) 600 IU synthetic all-rac-α tocopherol plus vitamin D placebo; group D (n=11) 1600 IU vitamin D3 plus 600 IU synthetic all-rac-α tocopherol. Erythrocyte superoxide dismutase (SOD) and catalase activities, serum 25 (OH) D, plasma α-tocopherol were determined. The data were analyzed by analysis of variance (ANOVA) and paired t-test. RESULTS: After 60 days vitamin D and E supplementation, erythrocyte SOD activities increased in groups B, C and D (P= 0.002, P= 0.016 and P= 0.015, respectively). Erythrocyte catalase activities increased in groups B and D (P= 0.026 and P= 0.004, respectively). The increment of erythrocyte catalase activity was not significant in group C. There was a positive significant correlation between SOD activity and serum 25 (OH) D (r= 0.378, P= 0.01). CONCLUSIONS: It is concluded that vitamin D is as potent as vitamin E in increasing the activities of erythrocyte SOD and catalase in atopic dermatitis patients.

The Study of Total IgE Reference Range in Healthy Adults in Tehran, Iran.

BACKGROUND: IgE is an antibody class that regarded as an important factor in the pathogenesis of allergic diseases, asthma, immune responses to parasitic infection and it could be responsible for the late- phase allergic response. The objective of this study was to evaluate total IgE in healthy Iranian adults, establishment of reference range of total IgE and assess helpfulness of this value in clinical diagnosis atopic and allergic diseases. METHOD: Three hundred sixty six healthy adults from blood transfusion volunteers (18 to 60 years) were selected in this study. A specific questionnaire (including demographic factors, smoking status and …) was filled out for each person. Also, we evaluated effect of race and education on total IgE. These adults had no history of allergic disease. The total serum IgE level using a commercial enzyme immunoassay and CBC (Eosinophil count) was determined in them. RESULTS: Mean of age was 37.32± 10.93 yrs and 219 cases were males and 147 females. The geometric mean of total IgE was 20.84 IU/ml (2-373 IU/ml) (95% percentile= 250) (95% confidence interval=46.27-62.70). No differences was observed between mean of IgE log in males and females (P= NS) but mean of total IgE log in females is more than males. CONCLUSION: Normal range of serum total IgE obtained in this study could be helpful for diagnosis of IgE-dependent allergic disease, as reference ranges in Iranian healthy adults.

Effects of EPA and Vitamin E on Serum Enzymatic Antioxidants and Peroxidation Indices in Patients with Type II Diabetes Mellitus.

BACKGROUND: Diabetes mellitus is associated with chronic changes in peripheral arteries because of oxidative stress and insufficient antioxidative defense mechanism. Omega-3 fatty acid supplementation could be effective in some diabetes complications; however, polyunsaturated fatty acids may increase lipid peroxidation. This study aimed to determine whether eicosapentaenoic acid alone or in conjunction with vitamin E had differential effects on serum antioxidants and peroxidation indices. METHODS: This double-blind, placebo-controlled trial was carried out on 136 patients with type II diabetic mellitus (age 48.8±4.4 yr, BMI 27.8±1.7 kg/m(2)). The four groups of the study either received two grams of omega-3 fatty acids, 400 IU of vitamin E, a combination of the two or placebo for three months. Their serum total antioxidant capacity, enzymatic antioxidants and peroxidation indices were assessed. RESULT: Fasting serum TAC increased in EPA+E (10.7%, P< 0.001) and E groups (7.5%, P< 0.05). SOD, G-PX and G-RD increased in EPA group (7.3%, 5.1%, and 8.4%, P< 0.05, respectively). MDA and protein carbonyl decreased in EPA and E groups (respectively, 12.5%, 7.6% P< 0.05, P< 0.05; 13%, 15.3% P< 0.001, P< 0.05). After adjustment for baseline values, age, sex, BMI and duration of diagnosed diabetes, protein carbonyl decreased in EPA+E and E group (30.7%, 15.3%; P< 0.05 respectively) relative to the placebo group. CONCLUSION: EPA, by itself has a statistically significant effect on serum total antioxidant capacity, enzymatic antioxidants and peroxidation indices in diabetic patients compared to EPA+E or E alone. TRIAL REGISTRATION: ClinicalTrials.gov NCT00817622.