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Background: Trimethyltin (TMT) is a toxic agent that causes oxidative stress, a laboratory model for inducing hippocampal injuries. Pentoxifylline (PTX) inhibits phosphodiesterase, inflammation and oxidative stress. This study evaluated the neuroprotective effects of PTX on injuries induced by TMT in the hippocampus. Methods: Sixty male Wistar rats were divided into five groups of 12 each. Group 1 received normal saline while Group 2 received a single dose of TMT (8 mg/kg). The other four groups received TMT at 8 mg/kg plus 60, 100 or 120 mg/kg PTX twice daily for six consecutive days. The rats’ working and reference memory were investigated, using radial arm maze tasks. At the end of the experiments, the rats’ brains were removed and processed for histological study of the hippocampus. Results: The TMT treatment prolonged the four baited arms tasks (P

Background: Chronic lymphocytic leukemia (CLL) is one of the most common hematologic malignancy in adults worldwide. This cancer has a poor prognosis at different stages. So, the identification of new biomarkers is important for diagnosis of B-CLL. Considering the oncogenic role of APRIL molecule in this leukemia as well as the regulatory role of miRNAs in different signaling pathways, the present study evaluated the miRNAs targeting APRIL gene in B-CLL. Methods: The miRNAs were predicted and selected using bioinformatics algorithms. A total of 80 plasma samples were subjected to RNA extraction and synthesis of cDNA. The expressions levels of predicted miRNAs and APRIL gene in plasma of B-CLL patients and healthy individuals were assessed by Real time PCR analysis. ROC analysis was performed to investigate the role predicted miRNAs as novel biomarkers in diagnosis of B-CLL. Results: The results of the prediction showed that miR-145-5p and miR-185-5p target the APRIL gene. The expression level of APRIL gene was strikingly higher in plasma of B-CLL patients than in the healthy individuals (102, P= 0.001). On the other hand, expression levels of miR-145-5p and miR-185-5p were strikingly lower in B-CLL patients than in the healthy individuals (0.07, P= 0.001) (0.29, P= 0.001). Also, ROC curve analyses demonstrated that miR-145-5p and miR-185-5p are specific and sensitive and may serve as new biomarkers for the detection of B-CLL. (AUC; 0.95, sensitivity; %90) (AUC; 0.87, sensitivity; %63). Conclusion: These data suggest that miR-145-5p and miR-185-5p target the APRIL gene and might have a role in diagnosis of B-CLL. Therefore, these two miRNAs can be served as a novel and potential biomarker for detection of B-CLL.

Background T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive and malignant neoplasm that arises from the hematopoietic T-cell precursors. Inactivation of FBXW7 gene is frequently observed in T-cell acute lymphoblastic leukemia, suggesting a significant tumor-suppressive role for FBXW7 in the pathobiology of this leukemia. Considering the role of microRNAs in cell proliferation and regulation of apoptosis, the aim of this study was to identify novel oncogenic microRNAs that suppress FBXW7 in patients with T-ALL. Patients and Methods The expression levels of two bioinformatically predicted microRNAs - miR-32 and miR-107 were compared in patients with T-ALL and a control group. A total of 80 plasma samples were subjected to RNA extraction, and the microRNA expression profiles were assessed by the RT-qPCR. The expression level of miR-103 was used as the endogenous reference for normalization of quantitative data. Results The plasma levels of miR-32 and miR-107 in patients with T-ALL were significantly higher (5.65, P < 0.001) and lower (0.432, P = 0.002), respectively. On the other hand, the expression levels of FBXW7 gene were significantly downregulated by -76.9 fold in T-ALL patients (P < 0.001). The results of the ROC curve analysis indicated that overexpression of miR-32 might be used to distinguish T-ALL patients with reasonable sensitivity and specificity. Conclusion miR-32 is considered as a novel oncomir that targets FBXW7 and might have a role in the etiology or progression of T-ALL. Furthermore, miR-32 can potentially serve as a non-invasive biomarker for detection of T-ALL.

Background Myeloproliferative neoplasms (MPN) are heterogeneous diseases that classified by the presence of Philadelphia chromosome into Philadelphia chromosome negative (Ph-neg) and positive (Ph-pos) myeloproliferative neoplasms. In ph-neg group A somatic point mutation (c.1849G>T) in the JAK2 gene, part of the JAK2-STAT signal-transduction pathway, causes substitution of phenylalanine for valine (V617F) in the JAK2 protein and has been identified. This mutation was seen in PV by 65% to 97% and ET (30–57%) and primary myelofibrosis (35–95%). Highly sensitive methods have been used to determine the presence of the JAK2V617F mutation instead of direct sequencing. We aimed to assess JAK2 exon14 mutations by high-resolution melting (HRM) analysis, which allows variation screening in compare to other method for detecting mutation. Methods The mutation analysis included 45 individuals who were subjected for diagnosis of ph-neg MPN. Genomic DNA was isolated and different methods are performed. Results PCR RFLP, ARMS PCR and HRM method has a detection sensitivity comparable with conventional methods (Qiagen) to identify the mutations and sequencing. Conclusions For HRM analysis is cost-effective and beside that it is enzyme independence method also this method able to show amount of the mutant allele carried in samples and it’s helpful for treatments follow-up and determining MRD for them.

Background: Timely and sensitive diagnosis of gastric cancer is crucial for efficient treatment and survival of the patients. microRNAs have been considered as diagnostic biomarkers in different type of cancers including gastric cancer. In the present study, the expression profile of four microRNAs, miR-103, miR-107, miR-194 and miR-210 were evaluated in patients with intestinal-type of gastric cancer (IGC) in order to assess their diagnosis utility as noninvasive biomarkers. Methods: A total number of 100 plasma samples from patients with gastric cancer and healthy controls were obtained and total RNA was extracted using a commercial monophasic solution of phenol and guanidium thiocyanate. Reverse transcription (RT) reactions were performed by specific stem-loop RT primers and M-MuLV RT-enzyme. The expression patterns of microRNAs were assessed using reverse transcription quantitative real-time PCR (RT-qPCR) method and the expression of SNORD47 RNA was used as the reference for normalization. Results: The results indicate that the plasma levels of miR-107, miR-194, and miR-210 were significantly lower in patients. Receiver operating characteristic (ROC) curve analysis showed that the patients could be distinguished from healthy individuals at the cutoff levels of 0.504, 0.266, and 0.394 of miR-107, miR-194, and miR-210, respectively. On the other hand, the expression levels of these miRNAs were not significantly different in different clinicopathological stages of the disease. Conclusion: These findings suggest that the plasma levels of miR-107, miR-194 and miR-210 were downregulated in patients with ICG and propose these molecules as potential non-invasive biomarkers for detection of IGC.

Background: Coronavirus disease 2019 (COVID-19) is an infectious disease caused by a newly discovered coronavirus. Understanding the spreading routes of SARS-CoV-2 is crucial for patient management and defining biosafety strategies for public and health care workers. Objectives: In the current study, the virus shedding in the upper respiratory as well as blood, stool, and urine specimens of infected patients, was examined using quantitative real-time PCR assay (qRT-PCR). Methods: The samples of 50 positive patients with high, moderate, and low virus titers in their respiratory specimens were collected. All specimens were subjected to RNA extraction and RT-qPCR assay using two different internally-controlled test systems. Results: The results indicated that no positive results were observed in the urine samples of all patients. The viral genome was diagnosed in 5% of blood and 3.3% of rectal swab samples. The Cq (Cycle of quantification)-values of positive results on the stool and blood samples were always higher than the Cq-value of the respiratory specimen of the same patient. Conclusions: SARS-CoV-2 is mainly detected in the respiratory samples, and the virus is not detectable in the urine. The importance of viremia and the existence of the virus in feces in virus spread in the human population needs further investigation.

Background: Chronic lymphocytic leukemia (CLL) is one of the most common hematologic malignancy in adults worldwide. This cancer has a poor prognosis at different stages. So, the identification of new biomarkers is important for early diagnosis of B-CLL. Considering the oncogenic role of APRIL molecule in this leukemia as well as the regulatory role of microRNAs (miRNAs) in different signaling pathways, the present study evaluated the miRNAs targeting APRIL gene in B-CLL.Methods: The miRNAs were predicted and selected using bioinformatics algorithms. A total of 80 plasma samples (40 samples of healthy individuals and 40 samples of B-CLL patients) were subjected to RNA extraction and synthesis of cDNA. The expressions levels of predicted miRNAs and APRIL gene in plasma of B-CLL patients and healthy individuals were assessed by Real time PCR analysis. ROC analysis was performed to investigate the role predicted miRNAs as novel biomarkers in diagnosis of B-CLL. Results: The results of the prediction showed that miR-145-5p and miR-185-5p target the APRIL gene. The expression level of APRIL gene was strikingly higher in plasma of B-CLL patients than in the healthy individuals (102, P= 0.001). On the other hand, expression levels of miR-145-5p and miR-185-5p were strikingly lower in B-CLL patients than in the healthy individuals (0.07, P= 0.001) (0.29, P= 0.001). Also, ROC curve analyses demonstrated that miR-145-5p and miR-185-5p are specific and sensitive and may serve as new biomarkers for the early detection of B-CLL. Conclusions: These data suggest that the study miRNAs may have a role in B-CLL development and progression. Moreover, miR-145-5p and miR-185-5p can be served as a novel and potential biomarker in the diagnosis of B-CLL.

Background T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive and malignant neoplasm that arises from the hematopoietic T-cell precursors. Inactivation of FBXW7 gene is frequently observed in T-cell acute lymphoblastic leukemia, suggesting a significant tumor-suppressive role for FBXW7 in the pathobiology of this leukemia. Considering the role of microRNAs in cell proliferation and regulation of apoptosis, the aim of this study was to identify novel oncogenic microRNAs that suppress FBXW7 in patients with T-ALL. Patients and Methods The expression levels of two bioinformatically predicted microRNAs – miR-32 and miR-107 were compared in patients with T-ALL and a control group. A total of 80 plasma samples were subjected to RNA extraction, and the microRNA expression profiles were assessed by the RT-qPCR. The expression level of miR-103 was used as the endogenous reference for normalization of quantitative data. Results The plasma levels of miR-32 and miR-107 in patients with T-ALL were significantly higher (5.65, P < 0.001) and lower (0.432, P = 0.002), respectively. On the other hand, the expression levels of FBXW7 gene were significantly downregulated by –76.9 fold in T-ALL patients (P < 0.001). The results of the ROC curve analysis indicated that overexpression of miR-32 might be used to distinguish T-ALL patients with reasonable sensitivity and specificity. Conclusion miR-32 is considered as a novel oncomir that targets FBXW7 and might have a role in the etiology or progression of T-ALL. Furthermore, miR-32 can potentially serve as a non-invasive biomarker for detection of T-ALL.

Hydatidosisis a parasitic disease caused by the larval stage of Echinococcus granulosus with different genotypes, and major complications in vital organs such as liver, lungs and, brain. Also, this parasite can infect animals and cause economic damages. Recently, some investigations indicated that the genetic variation of the parasite affects the antigenic, immunogenic and pathogenic features. Therefore, present study conducted to genotyping of the E. granulosus larva based on mitochondrial cox1 gene in livestock in the endemic areas of Markazi province, Iran. In this study, 49 hydatid cysts samples collected from 36 sheep, 11 goats and 2 cattle from different slaughterhouses of Markazi province in central part of Iran, 2017. The mitochondrial cox1 gene was amplified and genotyping were accomplished using sequence analysis. The sequencing analysis indicated that the main genotype G1 (61%) and G3 (37%) were identified. Also, one of the samples shows similarity with the G2 (2%) genotype. The results showed the statistically significant differences between the genotypes in different livestock (P

Background: Helicobacter pylori is the main cause of stomach ulcers and gastric cancer. Hence, the diagnosis, treatment, and prevention of H. pylori infection can considerably reduce the fatality. Objectives: This study aimed to construct a dual-antigen protein by combining the antigenic regions of UreB and FlaA of H. pylori and determine its antigenicity as a promising vaccine and serodiagnosis candidate. Methods: The antigenic regions of FlaA and UreB were detected by immunological bioinformatics, amplified and joined together by polymerase chain reaction (PCR) with special primers containing linker sequences. Then, it was cloned into pET-32a and after expression and purification of the recombinant multi-epitope protein (rFlaA-UreB), its antigenicity was evaluated by immunoblotting using the sera of infected patients. Results: DNA sequencing and enzyme digestion analysis showed the rFlaA-UreB gene was successfully inserted into pET32a. The recombinant protein was produced and purified via affinity chromatography and its molecular weight was similar to what had been predicted. Moreover, data indicated that rFlaA-UreB was recognized by all patients’ sera and its sensitivity and specificity were high. Conclusions: Although the developed recombinant multi-epitope protein was very smaller and lighter than the natural forms of these two critical antigens, they all had close antigenic properties. Therefore, this recombinant protein can be an important antigen in the diagnosis and vaccination against H. pylori.

Background: Chronic infection with hepatitis B virus (HBV) is related to more than 50% of all cases of hepatocellular carcinoma (HCC). The HBx protein of the virus plays an important role in carcinogenesis through different mechanisms, including interaction with the Notch1 signaling pathway. Several microRNAs have also been shown to play essential roles in the carcinogenesis of HCC, and the molecules can be considered the novel biomarkers for the diagnosis of different types of cancer. Methods: In the present study, the expression levels of four bioinformatically predicted microRNAs, including miR-214, miR-6510, miR-5193, and miR-34a, were compared in individuals with HBV-associated HCC and controls in order to assess the diagnostic utility of these microRNAs as noninvasive biomarkers. Seventy three plasma samples were subjected to RNA extraction, and the microRNA expression profiles were assessed by RT-qPCR. The expression of miR-103 was used as the endogenous reference for the normalization of quantitative data. Results: The plasma expression of all miRNAs was significantly lower in the HCC group, but the downregulation was most marked for the newly described molecule, miR-5193 (-18.86 folds and -5.71 folds compared to healthy individuals and chronic HBV patients). Another novel microRNA, miR-6510, was downregulated by -4.23 folds (P = 0.001). The results of the ROC curve analysis indicated that the differential expression of miR-5193 could distinguish HCC patients with high sensitivity and specificity. Conclusions: The study microRNAs may have a role in HCC development and progression. In addition, miR-5193 can be potentially used as a non-invasive biomarker for the detection of HBV-induced HCC.

Background Cell-mediated immunity including T-cells (T helper and cytotoxic) plays an essential role in efficient antiviral responses against coronavirus disease-2019 (COVID-19). Therefore, in this study, we evaluated the ratio and expression of CD4 and CD8 markers in COVID-19 patients to clarify the immune characterizations of CD4 and CD8 T-cells in COVID-19 patients. Methods Peripheral blood samples of 25 COVID-19 patients and 25 normal individuals with similar age and sex as the control group were collected. White blood cells, platelets, and lymphocytes were counted and CD4 and CD8 T lymphocytes were evaluated by flow cytometry. Results The number of white blood cells, lymphocytes, and platelets were reduced significantly in COVID-19 patients (P 0.05); however, the CD8 MFI increased significantly in COVID-19 infected patients (P

Quantitative real-time PCR (qPCR) assay is accepted as the method of choice for monitoring human cytomegalovirus (HCMV) infection in hematopoietic stem cell transplant recipients, but the high cost of commercial kits has hampered its use in many developing countries. In this study, an affordable in-house qPCR was used to manage HCMV infection in pediatric patients and the diagnostic value of this method was compared with the conventional pp65 antigenemia assay. A total number of 1179 samples from 82 recipients were used in this study, and the effect of some potential risk factors on HCMV reactivation was evaluated. The qPCR was able to detect HCMV reactivation earlier and with higher sensitivity than antigenemia assay. Forty-six episodes of reactivation were detected in 39 patients, of which all were detected by the qPCR assay, while only 21 episodes were diagnosed by antigenemia. The DNAemia level of 1284 IU/ml plasma was defined as the optimal cutoff value for starting pre-emptive therapy. It was shown that the acute GVHD severity and the relationship of donor and recipient are the most significant risk factors for HCMV reactivation. The data suggest that the antigenemia method for monitoring HCMV reactivation could be substituted by the qPCR assay.

Background: Helicobacter pylori is a common cause of chronic infections worldwide. Helicobacter pylori urease is a multimeric enzyme with a molecular weight of 530 kDa, which is collected from two separate subunits of UreA (26.5 kDa) and UreB (61.7 kDa). Objectives: This study aimed to evaluate the antigenic properties of UreB recombinant protein as a vaccine candidate for the infected sera of humans and mice. Methods: In this study, the highly antigenic region of UreB gene (609 bp) was detected, using immunological bioinformatics for epitope mapping, amplified by polymerase chain reaction (PCR). Afterwards, the antigen was cloned, using a pBSK cloning vector and was inserted into a pET-32a expression vector; the target protein was then expressed and purified. Eventually, antigenicity was examined by immunoblotting on the sera of humans infected with H. pylori UreB recombinant protein. Results: The sequencing results of PCR were indicative of UreB gene cloning into the recombinant plasmid. Production of the protein was induced by isopropyl β-D thiogalactopyranoside (IPTG), and the expressed protein was purified via dialysis, using the Ni-NTA kit. In addition, the recombinant protein with a molecular weight of 42 kDa was recognized by antibodies in Western blotting. Conclusions: The evaluation of antibodies was indicative of high antigenic properties for the immunogenic fragment, predicted by immunological bioinformatics. Therefore, UreB recombinant protein might be a proper antigen for the development of vaccines against H. pylori and other diagnostic kits.

Introduction Non-small cell lung cancer (NSCLC) is the most common subtype of lung cancer. One of the signal transduction pathways related to NSCLC is Unfolded Protein Response (UPR), which is mainly regulated by GRP78 ( HSPA5 , Gene ID: 3309). The aim of this study was to employ bioinformatics tools to predict microRNAs (miRNAs) affecting GRP78 expression, experimentally validate interaction of these miRNAs with GRP78 and also evaluating the expression correlation of GRP78 and its predicted miRNAs in clinical samples. Materials and methods Various software were used to predict miRNAs that simultaneously target all upstream and downstream components of GRP78 in the UPR, as well as the main components of PI3K/AKT, MAPK, ErbB and calcium pathways. For experimental analysis, 36 pairs of Formalin-Fixed Paraffin-Embedded (FFPE) lung tumor and non-tumor tissue samples were obtained. Additionally, A549 and QU-DB lung cancer cell lines were used for expression determination of GRP78 and its predicted targeting miRNAs. We also employed a luciferase assay to evaluate interactions between candidate miRNAs with the 3′-UTR of GRP78. Results hsa-miR-495 and hsa-miR-199-5p were chosen based on several criteria including thermodynamic binding features of miRNAs to the target transcripts, number of recognition sites, and conservation of binding sites within the 3′-UTR of GRP78. RT-qPCR data revealed a significant up-regulation of GRP78 (3.87 times, P = 0.002) and down-regulation of miR-199a-5p (0.13 times, P = 0.0001) and miR-495 (0.085 times, P = 0.0001) in tumor samples. Luciferase assay confirmed an interaction of hsa-miR-199a-5p and hsa-miR-495 with the 3′-UTR of GRP78 transcript. In addition, over-expression and competitive inhibition of the aforementioned miRNAs, significantly altered the expression of GRP78 and spliced XBP1 level. Conclusion Our data revealed a significant up-regulation of GRP78 and a concomitant down-regulation of miR-495 and miR-199a-5p in NSCLC. Accordingly, our data suggest a causative role for miR-199-5p and miR-495 in tumorgenesis of lung and probably other cancer types.

Background: The Crimean-Congo hemorrhagic fever (CCHF) virus causes severe disease in humans, with a high mortality rate. Since, there is no approved vaccine or specific treatment for CCHF, an early and accurate diagnosis, as well as reliable surveillance, is essential for case management and patient improvement. Objectives: For this research, our aim was to evaluate the application of a novel SYBR Green based one-step real-time reverse-transcriptase polymerase chain reaction (rRT-PCR) assay for the in-house diagnosis of the CCHF virus. Patients and Methods: In this experimental study, the highly conserved S-region sequence of the CCHF viral genome was first adapted from GenBank, and the specific primers targeting this region were designed. Then, the viral RNA was extracted from 75 serum samples from different patients in eastern Iran. The sensitivity and specificity of the primers were also evaluated in positive serum samples previously confirmed to have the CCHF virus, by this one-step rRT-PCR assay, as well as a DNA sequencing analysis. Results: From a total of 75 suspected serum samples, 42 were confirmed to be positive for CCHF virus, with no false-positives detected by the sequencing results. After 40 amplification cycles, the melting curve analysis revealed a mean melting temperature (Tm) of 86.5 ± 0.6°C (quite different from those of the primer-dimers), and the positive samples showed only a small variation in the parameters. In all of the positive samples, the predicted length of 420 bp was confirmed by electrophoresis. Moreover, the sensitivity test showed that this assay can detect less than 20 copies of viral RNA per reaction. Conclusions: This study showed that this novel one-step rRT-PCR assay is a rapid, reliable, repeatable, specific, sensitive, and simple tool for the detection of the CCHF virus.

Background: Pseudomonas aeruginosa is an opportunistic pathogen that infects patients with cystic fibrosis, burning wounds, ophthalmic traumas and immunodeficiency. Objectives: The aim of the present study was to compare the efficiency of newly designed primer sets with some previously published primers for PCR detection of exoA, oprL and algD genes from P. aeruginosa. Materials and Methods: A total of 150 clinical specimens were inoculated into the routine and selective culture media for P. aeruginosa isolation. Specific primers were designed by bioinformatics analysis for detection of the virulence genes determinants, exoA, oprL and algD. The sequences of these three genes were obtained from NCBI and multiple alignments were performed to find the conserved sequences of each gene to design the primers. Multiple alignment and primer design steps were carried out by the AlleleID software, version 7.0. Results: Microbiological culture methods resulted 70 P. aeruginosa strains isolated from 70 of the 150 clinical specimens. The results of the PCR assay using the newly designed exoA, oprL and algD primer sets were positive in 68, 70 and 69 clinical samples which represent 97.2%, 100% and 98% sensitivity for each primer set, respectively. The PCR results using previously published exoA, oprL and algD primer sets were positive only in 57, 49 and 28 specimens that correspond to 81.5%, 70% and 40% sensitivity, respectively. Conclusions: The results of the present study showed that in comparison with previously published primer sets, P. aeruginosa infection can be diagnosed with higher sensitivity and specificity by the conventional PCR assay using the newly designed primers. It was also shown that the results of the PCR assay on clinical samples of severe infections became positive much earlier than the results of conventional culture method.

High-level Expression and Purification of Active Human FGF-2 inEscherichia coli by Codon and Culture Condition Optimization Mohammad Reza Soleyman 1 ; Mostafa Khalili 1 ; Behzad Khansarinejad 2 ; and Maryam Baazm 3, * 1 Department of Biotechnology, School of Medicine, Arak University of Medical Sciences, Arak, IR Iran 2 Department of Microbilogy and Immunology, School of Medicine, Arak University of Medical Sciences, Arak, IR Iran 3 Department of Anatomy, School of Medicine, Arak University of Medical Sciences, Arak, IR Iran *Corresponding Author: Maryam Baazm, Department of Anatomy, School of Medicine, Arak University of Medical Sciences, Arak, IR Iran. Tel: +98-9166621131, Fax: +98-8634173526, E-mail: Dr.Baazm@arakmu.ac.ir. Abstract Background: Basic fibroblast growth factor (bFGF) is a member of a highly conserved superfamily of proteins that are involved in cell proliferation, differentiation, and migration. Objectives: The objective of this study was to overexpress and purify the high-level active human bFGF in Escherichia coli (E. coli). Materials and Methods: This experimental study was conducted in the Islamic Republic of Iran. After codon optimization and gene synthesis, the optimized FGF-2 gene was subcloned into plasmid pET-32a. pET32-FGF-2 was transformed into E. coli BL21 for expression. The cultivation parameters were optimized to produce a high yield of FGF-2. Results: The optimal conditions were determined as follows: cultivation at 37°C in TB medium, with 1 mM isopropyl-β-D-thiogalactopyranoside (IPTG), followed by post-induction expression for 6 h. Under the abovementioned conditions, the expression volumetric productivity of FGF-2 reached 1.48 g/L. Conclusions: A fusion tag from the pET32 expression plasmid permits the recovery of the recombinant fusion FGF-2 from E. coli,without affecting its biological activity. Keywords: Fibroblast Growth Factor 2; Codon; Protein; E. coli

Background: Toxoplasma gondii is an obligatory intracellular protozoan parasite, which infects human beings. Since the current antigens used for diagnosis or vaccination are contaminated with non parasitic material in which the parasite is grown, it is tried to produce recombinant antigens to design vaccines against toxoplasmosis, or make diagnostic kits. Choosing the type of antigen to produce recombinant vaccine or diagnostic kits is considerably important. The rhoptry protein 1 is one of the excretory-secretary antigens of Toxoplasma which seems to be an appropriate candidate in production of recombinant vaccines and diagnostic kits. Objectives: The current study aimed to produce rhoptry protein 1 from Iranian strains of Toxoplasma for further investigations. Materials and Methods: Genomic DNA was isolated from tachyzoite of parasite by phenol chloroform method and gene fragment was amplified by polymerase chain reaction. The polymerase chain reaction products were ligated into restriction enzymes sites of pTZ57R/T cloning vector. The product was sub-cloned into a prokaryotic expression plasmid (pET32a). The recombinant expression vector containing rhoptry protein 1 sequence was transformed into Escherichia coli BL21 pLysS and was induced by isopropyl β-D-1-thiogalactopyranoside. The sodium dodecyl sulfate polyacrylamide gel electrophoresis and western blotting methods were used to confirm the production of protein. Results: The result showed that the desired molecular weight protein is produced. Recombinant protein was confirmed by western blot using Ni-NTA- conjugate. Conclusions: The result of this study showed that recombinant ROP1 Toxoplasma was produced successfully. In the present study, unlike other studies, the goal was to express full length of ROP1. Also this protein was produced alone not fused with other proteins.

INTRODUCTION: Prevalence of influenza A virus (Flu-A), respiratory syncytial virus (RSV), and human metapneumovirus (hMPV) was assessed in children with acute respiratory infections (ARIs). METHODS: Nasopharyngeal aspirates and throat swabs were subjected to real-time polymerase chain reaction (PCR) to detect RSV and Flu-A and to conventional PCR to detect hMPV. RESULTS: Of the 156 children assessed, 93 (59.6%) carried at least one virus, with 35.9% positive for RSV, 14.1% for hMPV, and 9.6% for Flu-A. The prevalence of co-infections was 2.6%. CONCLUSIONS: The high detection rate may reflect increased sensitivity of real-time PCR compared to traditional PCR and viral culture.