Genotyping of toxoplasma gondii isolates from soil samples in Tehran, Iran

The protozoan parasite Toxoplasma gondii can infect any warm blooded nucleated cells. One of the ways for human infection is ingestion of oocysts directly from soil or via infected fruits or vegetables. To survey the potential role of T. gondii oocyst in soil samples, the present study was conducted in Tehran City, Iran. A total of 150 soil samples were collected around rubbish dumps, children's play ground, parks and public places. Oocysts recovery was performed by sodium nitrate flotation method on soil samples. For molecular detection, PCR reaction targeting B1 gene was performed and then, the positive results were confirmed using repetitive 529 bp DNA fragment in other PCR reaction. Finally, the positive samples were genotyped at the SAG2 locus. Toxoplasma DNA was found in 13 soil samples. After genotyping and RFLP analysis in SAG2 locus, nine positive samples were revealed type III, one positive sample was type I whereas three samples revealed mixed infection [type, I and III]. The predominant genotype in Tehran soil samples is type III

Brain tissue cysts in infected mice with RH-strain of Toxoplasma gondii and evaluation of BAG1 and SAG1 genes expression

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. Two groups of Swiss-Webster and BALB/c mice were infected subcutaneously with 10[4] tachyzoites of T. gondiiy RH strain and given SDZ [300 mg/l] with NaHCO3 [5 g[-1]] in drinking water from day 1 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 [BAG[1]] and tachyzoite [SAG[1]] specific genes during tachyzoite / bradyzoite stage conversion. 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. It seems that conversion of tachyzoites to bradyzoites in brain of mice undergoing SDZ was not completed until 50 days after inoculation

rate of occupational exposure to patients specimen among personnel of medical diagnostic laboratories in Birjand City

Having knowledge about the potential risks and the usage of safety equipment in laboratories can decrease the risk of occupational exposure. The aim of this study was to evaluate the predisposing factors for occupational exposure and to assess the usage of safety equipment among personnel of medical diagnostic laboratories in Birjand. In this descriptive analytic study, all staff of laboratories was assessed by using a questionnaire including demographic data, type of accidental exposures and the use of protective equipment. Using SPSS software, we analyzed the data. Of 110, 84 [76%] have at least one accidental exposure to patients' specimens in that 55% of accidents are related to sample preparation step. In 82% of contacts, carelessness is the underlying cause of exposure. Gown, glove, fume hood and mouth mask are routinely used by 97%, 48%, 34%, 1% of personnel, respectively. Nearly all of [97%] personnel were vaccinated against hepatitis B and 78% of them have performed routine blood test for detecting any infections. The results of current study show that accidental exposures to patients' samples are common among personnel of medical diagnostic laboratories. The level of preventive education and the rate of safety equipment usage are low; therefore, we recommend planning of some training sessions to persuade the personnel for using safety equipment

Production and evaluation of Toxoplasma gondii recombinant surface antigen 1 [SAG1] for serodiagnosis of acute and chronic toxoplasma infection in human sera

The assays currently available for the detection of specific anti-Toxoplasma antibodies may vary in their abilities to detect serum immunoglobulins, due to the Lack of a purified standardized antigen. The aim of this study was evaluation the recombinant Toxoplasma gondii SAG1 antigen for the serodiagnosis of acute and chronic toxoplasmosis. This study describes an ELISA using recombinant SAG1 for detection of IgM and IgG antibodies against Toxoplasma gondii in human sera. Genomic DNA of T. gondii [RH Strain] was isolated and PCR reaction was performed. Recovered DNA was cloned into PTZ57R cloning vector. The recombinant plasmid was detected by restriction analysis. The SAG1 gene was subcloned in the pET- 28a expression vector. Protein production was then induced with 1 mM isopropyl-D - thiogalactopyranoside [IPTG]. A total of 204 sera were tested using a commercial IgG and IgM ELISA kit [Trinity, USA] as gold standard prior to testing them with the recombinant antigen. Tested sera were divided into the following groups:[a] The 74 T. gondii IgG positive [b] 70 T.gondii IgM positive [c] 60 sera who had no serological evidence of toxoplasmosis as negative sera.To determine the specificity of the test, we used other parasitic diseases including echinococusis [N=5], malaria [N=14], leishmaniasis [N=7],fasciolasis [N=4], sterengyloidiasis [N=1]. Sensitivity and specificity of the generated recombinant IgG ELISA in comparison with commercial ELISA [Com ELISA] were 93% and 95%, and the sensitivity and specificity of the generated recombinant IgM ELISA were 87% and 95% respectively. The results acquired here show that this antigen is useful for diagnostic purposes and could be replaced by lysed, whole cell antigens for diagnosis of chronic toxoplasmosis

In vivo susceptibility of plasmodium vivax to chloroquine in Southeastern Iran

Plasmodium vivax is the predominant species causes of malaria with about 90% total annual reported malaria in Iran. This study conducted to determine the susceptibility of Plasmodium vivax isolates to chloroquine in Sistan and Balochistan Province, southeastern Iran. A total 270 subjects with symptomatic malaria and confirmed P. vivax infection completed the designed 28-day in vivo study. The thick and thin film blood smears were screened for malaria parasites by microscopy. The nested PCR was applied using the Plasmodium 18 subunit ribosomal ribonucleic [Ssr RNA] genes for detecting mixed infections and diagnosis of parasites in the samples with low parasite on days 0, 5, 6, 7, and 28. P. vivax was cleared in 15%, 50%, 95%, and 100% of patients on days 1, 2, 3, 4 respectively by microscopy assessment. Six patients were exhibited specific P. vivax band in nested PCR on day 5. No recurrence was observed on days 7, 14 and 28. Mean [ +/- standard deviation] parasite clearance time was 2.41 [ +/- 0.8] days. P. vivax is still susceptible to chloroquine in Southeastern Iran. This finding is compatible with results of neighboring countries Pakistan and Afghanistan

Rapid detection of Toxoplasma gondii antigen in experimentally infected mice by dot- ELISA

Toxoplasmosis is a worldwide endemic disease. In congenitally infected infants and AIDS patients, toxoplasmosis causes high rates of morbidity and mortality. In these cases antibody detection is difficult; so detection of parasite or its components could be useful tool for early detection and following treatment of the infection. Sixty-three BALB/c mice were injected intra-peritoneal with 5x10[3] tachyzoites of Toxoplasma gondii RH strain, nine mice were sacrificed daily for 7 days. Fourteen mice were injected with phosphate buffer saline as control group. Dot-ELISA was performed for detection of T.gondii antigen in mice sera and capture -ELISA was done as golden standard assay too. Toxoplasma gondii antigen was detected from day 2 in mice sera; 22% of mice sera on day 2, 33% on day 3,77% on day 4 and 100% on day 5 till their death on day 7 had shown antigenemia by dot -ELISA, no positive result was detected in control mice by dot-ELISA. Dot-ELISA is a sensitive method for diagnosis of T. gondii infection in the animal model; also, this technique is more rapid and easy to perform method in comparison with capture-ELISA

Seroprevalence of Toxoplasma gondii infection in dogs in Tehran, Iran

Toxoplasma gondii infects a wide range of animals; felines are definitive hosts and other animals including the dogs are intermediate hosts. The aim of this study was to determine the seroprevalence of T. gondii infection in dogs in Tehran, capital of Iran and to investigate possible associated risk factors. Three hundreds ninety six serum samples were collected during 2007-8 from the dogs. Collected samples were tested using an indirect fluorescent antibody test [IFAT] in dilutions of 1:16 and more. All procedures were carried out in Shahrekord University, Iran. All the data were analyzed using SPSS software, qui square test with confidence interval of 0.95. From evaluated samples, 89 [22.47%] were positive in titers of at least 1:16. further evaluations in other dilutions showed positive results in dilutions of maximum 1:16, 1:32, 1:64, 1:128 and 1:256 in 38, 29, 15, 2 and 5 dogs respectively. Investigation of the role of risk factors showed no sex predisposition while infection rate was significantly higher in dogs older than one year old. Living places were of significant importance; infection rate was significantly higher in stray or guard dogs in compare with household dogs [P<0.05]. Relatively high seroprevalence of T. gondii infection in dogs in Tehran shows high environmental contamination. It is recommended that the dogs with suspected clinical signs be tested for T. gondii infection

Toxoplasma infection in schizophrenia patients: a comparative study with control group

Schizophrenia is a serious, chronic, and often debilitating neuropsychiatric disorder. Its causes are still poorly understood. Besides genetic and non-genetic [environmental] factors are thought to be important as the cause of the structural and functional deficits that characterize schizophrenia. This study aimed to compare Toxoplasma gondii infection between schizophrenia patients and non-schizophrenia individuals as control group. A case-control study was designed in Tehran, Iran during 2009-2010. Sixty-two patients with schizophrenia and 62 non-schizophrenia volunteers were selected. To ascertain a possible relationship between T. gondii infection and schizophrenia, anti-Toxoplasma IgG antibodies were detected by indirect-ELISA. Data were statistically analyzed by chi- square at a confidence level of 99%. The sero-positivity rate among patients with schizophrenia [67.7%] was significantly higher than control group [37.1] [P <0. 01]. A significant correlation between Toxoplasma infection and schizophrenia might be expected

Evaluation of confounder in toxoplasmosis indirect fluorescent antibody assay

The IFA test is one of the most usual methods for detecting anti-Toxoplasma antibodies, although it has not any unique standardization. It seems that the microscopic judgment of results is an important confounder in IFA test. Therefore, we conducted the present study to clarify the role of microscopic observer, and other confounders on the test. Eighty sera were collected from patients suspicious to toxoplasmosis for detection IgG anti-T. gondii by this test. Samples were examined against different series of antigens, IgG anti-human conjugates, and observes. There were no significant differences between the two series of antigens and conjugates. For the observers groups the kappa coefficient of the test results in the experts group [0.97, 0.94- 1.00] were significantly higher than the less experienced observers [0.77, 0.68-0.87]. We recommend the IFA test to be performed only in reference laboratories and by laboratory technicians that have enough experience for this test. Otherwise, we suggest the substitution of this test with other tests like ELISA for the diagnosis and epidemiological studies

[Allele frequency of C-482T polymorphism in apolipoprotein CIII gene in a tehranian population: Tehran lipid and glucose study]

Apoc3, an apolipoprotein, is well known as a lipolysis inhibitor. It inhibits lipolysis by both HP and LPL activity inhibition and has been studied as a factor for hypertriglyceridemia for years. C-482T polymorphism in apoc3 gene promoter has associated with hypertriglyceridemia and insulin resistance, factors associated with the metabolic syndrome association factors. Subjects were randomly selected from the Tehran Lipid and Glucose Study. A 231 bp segment of the mentioned gene was amplified by PCR and the polymorphism revealed by RFLP using the MspI restriction enzyme. Allele frequencies obtained for APOC3-482C and-482T polymorphisms were 0.653 and 0.347 respectively. Genotype frequencies were in conformity with the Hardy-Weinberg expectation. The observed genotype and allele frequencies were similar to those reported for other Caucasians samples. The data generated from this study will be of importance in the context of ongoing studies concerning the factors that influence lipid levels in Iranian populations

[Effect of potato cultivar on acrylamide formation in potato chips]

In April 2002, a research group at Stockholm University and the Swedish Food Administration announced that significant amounts of acrylamide might be formed during common heat processing of foods. Since acrylamide is classified as "probably carcinogenic to humans" by the International Agency for Research on Cancer [IARC], these findings were considered as alarming. The most important food sources of dietary acrylamide are fried potato products like chips and French fries, which are popular snacks in Iran. The aim of this study was to investigate the effects of reducing sugars and amino acids in three 'potato cultivars on acrylamide formation in potato chips produced on a laboratory scale. Potato chips samples were produced on lab-scale from three potato cultivars namely Sante, Agria and Omidbakhsh by frying at 180°C for 4.15 min. Reducing sugars [glucose and fructose] and asparagine concentration in raw potato samples were determined using HPLC. A suitable and valid method was set up for determination of acrylamide in the chips samples by gas chromatography/mass spectrometry, GC/MS. The reducing sugars and acrylamide contents were significantly different among potato chips samples produced from different cultivars [p<0.05]. The highest amount of reducing sugars was found in cultivar Sante [3513 mg/kg] followed by Agria [2111 mg/kg] and Omidbakhsh [1622 mg/kg], respectively. On the other hand, cultivar Omidbakhsh had the highest amount of asparagine [1871 mg/kg]. The highest amount of acrylamide [8825 mg/kg] was formed in the chips from cultivar Sante and the lowest amount was formed in the chips from cultivar Omidbakhsh [5112 micro g/kg]. A high content of asparagine in raw potatos was not necessarily an indication of high content of acrylamide in the produced chips. The two cultivars with higher content of reducing sugars, showed the higher potential for acry1amide formation in the chips. This indicates that the concentration of reducing sugars is a more important parameter than asparagine content in the formation of acrylamide in potato chips. The selection of proper potato cultivars with naturally lower contents of precursors of acrylamide specially reducing sugars is important factor to reduce the acrylamide formation during production of potato chips. With respect to the results obtained from this study, it is suggested that cultivar Omidbakhsh is the most suitable cultivar in Iran for the industrial production of potato chips with low levels of acrylamide

[Evaluation of TP[53] oncoprotein expression in mucoepidermoid carcinoma of salivary glands and it's relation with grading]

Mucoepidermoid carcinoma is the most common malignancy in salivary glands. This tumor has a variable biologic potential, so it was first divided into two groups: one with malignant behavior and the other with benign behavior and good prognosis. The purpose of this study was evaluation of TP[53] oncoprotein and its relation to different grades of mucoepidermoid carcinoma. This retrospective study included 22 paraffin embedded mucoepidermoid carcinoma samples stained by H and E. The samples were classified into low grade, intermediate grade and high grade. Then, new sections were made and stained by immunohistochemistry method [IHC method] for TP[53] marker. Finally, the relation between the two methods was statistically [ANOVA and Kendall test] analyzed. Seven sections of normal salivary gland tissue were also used as control group. All control cases were negative for TP[53] marker while 68.2% of mucoepidermoid carcinoma samples were positive. A significant relation was revealed between histological grade and nuclear TP[53] staining by IHC method. Parallel to increasing histologic grade of salivary gland mucoepidermoid carcinoma, TP[53] expression is also increased so that immunohistochemistry technique is helpful for determination of the biologic behavior of salivary gland mucoepidermoid corcinoma and prognosis of patients