Exploring virulence factors of Helicobacter pylori isolated from gastric biopsy.

BACKGROUND: Helicobacter pylori (H. pylori) colonizes human gastric mucosa and is classified as class one carcinogenic bacteria. In this regard, this study aimed to detect major virulence factors in H. pylori strains recovered from gastric biopsy in patients referred to Aras Clinique in Ardabil, northwest of Iran (2019-2021). MATERIALS AND METHODS: In this descriptive-cross sectional study, 287 dyspeptic patients were included. For bacterial isolation, gastric biopsy specimens (n=287) were taken from gastric antrum, then aseptically were cultured on the selective medium and incubated at 37C in microaerophilic conditions for 3-5 days. RESULTS: 25.18% of all (n = 70) patients were found to be infected with H. pylori upon endoscopy. Of them, 9 patients (12.857%) and 2 patients (2.875%) had peptic ulcer disease and gastric cancer respectively. According to the different patterns of virulence factors, 57 virutypes were identified in which oipA-vacAs1-vacAm2 (3, 4.28% n =) and oipA-vacAs1-vacAs2-vacAm2 (3, 4.28% n =) were the most common patterns. The simultaneous presence of vacAS2, vacAm2 and hopQ2 genes was observed in both patients with gastric cancer. OipA (n = 562.5%), VacAs1 (n = 6.75%), VacAs2 (n = 6.75%), and VacAm2 (n = 787.5%) were found to be the most prevalent virulence factor. CONCLUSION: According previous studies, it is confirmed that the cagPAI gene cluster and vacA gene alleles are strongly correlated with gastritis and gastrointestinal tract adenocarcinomas. Our study indicated that 50% of the indigenous strains of H. pylori harbor these oncogenic genes and they are hypervirulent.

Characterization of disinfectant susceptibility profiles among clinical isolates of Acinetobacter baumannii in Ardabil, Iran.

Antimicrobial disinfectants have been extensively used to control hospital-acquired infections worldwide. Prolonged exposure to bacteria could promote resistance to antimicrobial disinfectants. This study evaluated the antimicrobial activity of four commonly used disinfectants; triclosan, chlorhexidine digluconate, benzalkonium chloride, and formaldehyde against Acinetobacter baumannii clinical isolates. This study also determined the prevalence and association of efflux pumps encoding genes qacE, qacED1, emrA, and aceI with tolerance to disinfectants. A total of 100 A. baumannii isolates were included in the current study. The antimicrobial disinfectants' minimum inhibitory concentration (MIC) was determined using an agar dilution method. Genes involved in resistance to disinfectants were investigated by PCR method. The benzalkonium chloride MICs ranged between 32 and 128 µg mL-1, chlorhexidine digluconate 8-64 µg mL-1, triclosan 1-32 µg mL-1, and formaldehyde 128 µg mL-1. Overall, the highest MIC90 value was identified for formaldehyde (128 µg mL-1), followed by benzalkonium chloride and chlorhexidine digluconate (64 µg mL-1, each one) and triclosan (4 µg mL-1). In the present study, the qacE, qacED1, emrA, and aceI genes were found in 91%, 55%, 100%, and 88% of isolates, respectively. The qacG gene was not identified in our A. baumannii isolates. The qacED1 gene was associated with higher MICs for all disinfectants tested (P < 0.05), while the qacE and aceI genes were associated with higher MICs for benzalkonium chloride and chlorhexidine. This study indicated that triclosan is the most effective disinfectant against A. baumannii isolates.

Molecular identification of multiple drug resistance (MDR) strain of Mycobacterium tuberculosis.

BACKGROUND AND OBJECTIVES: Isoniazid and rifampin are the first -line drugs against Mycobacterium tuberculosis. Resistance to these important drugs is a serious threat to human public health. Therefore, this study aimed at molecular detection of resistance to these valuable drugs. MATERIALS AND METHODS: In this descriptive cross-sectional study, 111 non - duplicated clinical samples including sputum and Bronchoalveolar lavage (BAL) samples were collected from patients referred to the Ardabil Health Center between 2017 and 2020. The samples were first examined by microscopic method, then their DNA was extracted using the boiling method. Specific primers and MAS-PCR method were employed for the detection resistance to isoniazid and rifampin drugs and identification of MDR strain. RESULTS: of 111 specimens, 15.3% belonged to NTM. In total, the resistance rate to isoniazid and rifampin was 17% and 27% respectively while the resistance rate to isoniazid and rifampin among NTM was 61.54% and 38.46%. CONCLUSION: In our study, the prevalence of resistance to isoniazid and rifampin among Mycobacterium tuberculosis complex(MTC) and non-tuberculous mycobacteria(NTM) was investigated using the MAS-PCR method. This work highlighted the high anti- tuberculosis resistance rate among NTM compared to MTC strains.

Outcomes of COVID-19 in immunocompromised patients: a single center experience.

Malignancy, bone marrow and organ transplantation are associated with deficient and defective immune systems. Immunocompromised patients are at risk for severe and chronic complication of COVID-19 infection. However, the pathogenesis, diagnosis and management of this comorbidity remain to be elucidated. The purpose of the present study was to describe key aspects of COVID-19 infection in immunocompromised patients. In this retrospective, cross-sectional study, lab findings and outcomes of 418 COVID-19 patients with secondary immunodeficiency disorders admitted to Taleghani Hospital in Tehran, from March 2020 to September 2022 were investigated. Of the 418 immunocompromised patients with COVID-19, 236 (56.5%) | were male and the median age of all studied patients was 56.6 ± 16.4 with range of 14 to 92 years. Totally, 198 (47.4%) of the patients died during hospitalization. Remdesivir was used for treatment of all patients. Mortality rate among patients admitted to ICU ward (86.8%) was significantly higher than non ICU admission (p < 0.001). The death rate in patients with CKD was substantially higher than other underlying disease (p < 0.001). In terms of laboratory finding, there was a significant relationship between ICU admission and worse outcome with WBC count (HR = 1.94, 95% CI = 1. 46-2.59, p < 0.001), PMN count (HR = 1.93, 95% CI = 1.452.56, p < 0.001), Hb (HR = 1.49, 95% CI = 1.042.13, p = 0.028), AST (HR = 2.55, 95% CI = 1.913.41, p < 0.001), BUN (HR = 2.56, 95% CI = 2.063.69, p < 0.001), Cr (HR = 2.63, 95% CI = 1.89-3.64, p < 0.001), Comorbidities index (HR = 1.71, 95% CI = 1.29-2.27, p < 0.001) and aging (HR = 1.91, 95% CI = 1.4-2.54, p < 0.001). Immunocompromised status increased the risk of mortality or worse outcome in patients diagnosed with COVID-19. Our finding showed outcome predicting markers in whom the waned immune system encounter new emerging disease and improved our understanding of COVID-19 virus behavior in immunocompromised individuals.

Serotyping and molecular profiles of virulence-associated genes among Klebsiella pneumoniae isolates from teaching hospitals of Ardabil, Iran: A cross-sectional study.

Background and Aims: Klebsiella pneumoniae is a Gram-negative bacterium that colonized various organs. This bacterium is associated with different community-acquired and hospital-acquired infections. The present study aims to assess the capsular serotypes and frequency of virulence-associated genes in K. pneumoniae isolates from teaching hospitals in Ardabil, Iran. Methods: From October 1, 2019, to November 31, 2021, different clinical samples were collected and K. pneumoniae isolates were diagnosed using conventional biochemical tests. The final identification of K. pneumoniae was performed through the polymerase chain reaction (PCR) method using a specific primer targeting the khe gene. The PCR method was employed to confirm the presence of virulence-associated genes and aerobactin, and the main capsular serotypes based on the specific primers. Results: Of all 100 K. pneumoniae isolates, 4% and 2% were typeable with K5 and K2 primers, respectively. In addition, entB (94%), fimH (91%), and wcaG (87%) had the highest frequency among the virulence-associated genes. 24% of K. pneumoniae isolates harbored the entB-wcaG-fimH genes simultaneously. Moreover, 50% of capsular serotype 5 harbored the ybts-mrkD-entB-wcaG-fimH genes simultaneously. Conclusion: The findings revealed that 6% of all K. pneumoniae isolates were typeable, distributed in the two serotypes K5 and K2. Most K. pneumoniae isolates were positive for multiple types of virulence genes. Identifying bacterial virulence genes aids in molecular detection, assay development, and therapeutic pathways.

Prevalence and mechanisms of ciprofloxacin resistance in Escherichia coli isolated from hospitalized patients, healthy carriers, and wastewaters in Iran.

BACKGROUND: This study was aimed to evaluate the prevalence and molecular characteristics of ciprofloxacin resistance among 346 Escherichia coli isolates collected from clinical specimens (n = 82), healthy children (n = 176), municipal wastewater (n = 34), hospital wastewater (n = 33), poultry slaughterhouse wastewater (n = 12) and livestock (n = 9) slaughterhouse wastewater in Iran. METHODS: Ciprofloxacin minimum inhibitory concentration (MIC) was determined by agar dilution assay. Phylogroups and plasmid-mediated quinolone resistance (PMQR) genes were identified using PCR. Mutations in gyrA, gyrB, parC, and parE genes and amino acid alterations were screened through sequencing assay. The effect of efflux pump inhibitor (PAßN) on ciprofloxacin MICs in ciprofloxacin-resistant isolates was investigated using the microdilution method. RESULTS: In total, 28.03% of E. coli isolates were phenotypically resistant to ciprofloxacin. Based on sources of isolation, 64.63%, 51.51%, 33.33%, 14.70%, 10.22% and 8.33% of isolates from clinical specimens, hospital wastewater, livestock wastewater, municipal wastewater, healthy children and poultry wastewater were ciprofloxacin-resistant, respectively. Eighty-one point eighty-one percent (Ser-83 → Leu + Asp-87 → Asn; 78.78% and Ser-83 → Leu only; 3.03% (of ciprofloxacin-resistant E. coli isolates showed missense mutation in GyrA subunit of DNA gyrase, while no amino-acid substitution was noted in the GyrB subunit. DNA sequence analyses of the ParC and ParE subunits of topoisomerase IV exhibited amino-acid changes in 30.30% (Ser-80 → Ile + Glu-84 → Val; 18.18%, Ser-80 → Ile only; 9.10% and Glu-84 → Val only; 3.03%0 (and 15.38% (Ser-458 → Ala) of ciprofloxacin-resistant E. coli isolates, respectively. The PMQR genes, aac(6')-Ib-cr, qnrS, qnrB, oqxA, oqxB, and qepA were detected in 43.29%, 74.22%, 9.27%, 14.43%, 30.92% and 1.03% of ciprofloxacin-resistant isolates, respectively. No isolate was found to be positive for qnrA and qnrD genes. In isolates harboring the OqxA/B efflux pump, the MIC of ciprofloxacin was reduced twofold in the presence of PAßN, as an efflux pump inhibitor. The phylogroups B2 (48.45%) and A (20.65%) were the most predominant groups identified in ciprofloxacin-resistant isolates. CONCLUSIONS: This study proved the high incidence of ciprofloxacin-resistant E. coli isolates in both clinical and non-clinical settings in Iran. Chromosomal gene mutations and PMQR genes were identified in ciprofloxacin resistance among E. coli population.

Prosthetic knee joint infection caused by α-hemolytic Streptococcus species: a case report.

BACKGROUND: Knee arthroplasty is an orthopedic surgical procedure in which a damaged joint is replaced with an artificial one. It is estimated that 1-2% of knee arthroplasties will encounter infection over their lifetime. Although α-hemolytic Streptococcus species play an important role in prosthetic joint infection, they are less common than staphylococcal species. CASE PRESENTATION: In this report, a 50-year-old Iranian woman was diagnosed with prosthetic knee joint infection based on clinical, radiological, and laboratory findings. She was diabetic and had undergone a left total knee arthroplasty, which, 18 months after the surgery, presented pain, erythema, and edema in that knee. The primary culture of knee aspirate was positive for α-hemolytic Streptococcus species, but following antibiotic medication, culture was negative. The primary antibiotic regime was vancomycin and meropenem, which was changed to cefepime for the management of the infection based on the results of antimicrobial susceptibility testing. CONCLUSIONS: This report indicated the clinical presentation and management of the patient with prosthetic joint infection in which the patient recovered without any severe complications or surgical intervention.

Drug-induced liver injury due to tofacitinib: a case report.

BACKGROUND: Drug-induced liver injury is an acute or chronic liver damage in response to drugs, herbals, and any chemical compound. CASE PRESENTATION: In the present work, liver failure following the use of tofacitinib was reported. The patient was an 18-year-old iranian woman without any history of underlying disease. She complained of alopecia areata, and tofacitinib was administered for disease management. Following adherence to tofacitinib medication, partial recovery was obtained. At the time of hospitalization, the patient had a stable condition and only anorexia, jaundice, and elevation of liver enzymes were reported. During hospitalization, liver injury progressed and liver transplantation was suggested. After drug-induced liver injury diagnosis, the use of the drug was discontinued and the patient underwent supportive treatment. The patient recovered without any severe sequelae. CONCLUSIONS: Tofacitinib is a Janus kinase inhibitor that is useful in the treatment of disorders such as rheumatoid arthritis, psoriatic arthritis, and ulcerative colitis. Until now, the severe side effect of this drug has not been reported and in most cases it is used as a last resort, but here we report a rare side effect of this drug.

Molecular detection and characterization of Shigella spp. harboring extended-spectrum ß-lactamase genes in children with diarrhea in northwest Iran.

Shigellosis is one of the acute bowel infections and remains a serious public health problem in resource-poor countries. The present study aimed to survey the distribution of extended-spectrum ß-lactamase (ESBL)-producing Shigella strains isolated from patients with diarrhea in northwest Iran. In the present cross-sectional study, from January 2019 to December 2020, 1280 fecal samples were collected from children with diarrhea in Ardabil, Iran. Multiplex PCR assay was applied for the presence of ipaH, invC, wbgZ, rfpB, and rfc genes to detect Shigella spp., Shigella sonnei, Shigella dysenteriae, Shigella flexneri, and Shigella boydii, respectively. Phenotypic detection of ESBL-producing isolates was carried out using the Double Disc Test (DDT). The frequency of main ESBL encoding genes including blaCTX-M, blaSHV, and blaTEM was detected using multiplex PCR. The genetic similarity of S. sonnei isolates was determined using ERIC PCR. A total of 49 Shigella isolates (3.8%; 49/1280) including 42 (85.7%) S. sonnei, 5 (10.2%) S. flexneri, and 2 (4%) S. dysenteriae were identified. S. boydii was not detected in any fecal samples. ESBLs were produced by 10.2% of Shigella spp. including 3 S. sonnei, 1 S. flexneri, and 1 S. dysenteriae. The ESBL encoding genes include blaCTX-M and blaTEM found in 65.3% and 61.2% of isolates, respectively. blaSHV gene was not detected in any isolates. The ERIC-PCR profiles allowed the differentiation of 42 S. sonnei strains into 6 clusters. Our study revealed a high frequency of ESBL-encoding genes among Shigella spp. in northwest Iran. The high prevalence of S. sonnei harboring ESBL genes, in the present work, is the main challenge for dysentery treatment, and this concern justifies the need for effective and regular monitoring of antibiotic usage among patients.

Phenotypic and Molecular Characterization of Carbapenems Resistant Escherichia coli Isolated from Patients with Urinary Tract Infections in Ardabil Province, Iran.

Background & Objective: Carbapenem-resistant is Gram-negative bacteria representing a worldwide public health problem. The present study aims to survey the phenotypic and genotypic characteristics of carbapenem-resistant Escherichia coli isolates collected from hospitalized patients and outpatients in Ardabil province, Iran. Methods: Two hundred samples were collected from the patients who had already been referred to the hospitals in Ardabil, Iran, from January to June 2017. Each patient's social and demographic data were recorded in the first step. The resistance profile of all E. coli isolates against imipenem and meropenem antibiotics were determined using the Kirby-Bauer disk diffusion method. Moreover, the broth microdilution method determined the Minimum Inhibitory Concentration (MIC) of E. coli isolates to imipenem. The Carbapenem Inactivation Method (CIM) and Carba NP test were employed for screening carbapenem-resistant strains. The frequency of carbapenem-encoding genes was determined using Polymerase Chain Reaction (PCR) method. The Enterobacterial Repetitive Intergenic Consensus (ERIC)-PCR analysis was used to evaluate the genetic relatedness of E. coli isolates. Results: Out of 200 urine samples, 66% (n = 132) of the samples were collected from women. The patients' age varied from 1 month to 93 years. Results of the disk diffusion method revealed that 33% (n=66/200) of E. coli isolates were resistant to imipenem. However, imipenem resistance was detected in 37% (n = 74/200) of the E. coli isolates using broth microdilution method. All E. coli isolates were negative in CIM and Carba NP tests. Moreover, we could not detect any carbapenemase encoding genes among E. coli isolates. The ERIC-PCR method revealed the E. coli strains were classified into 39 clusters with 80% similarity. Conclusion: It appears that E. coli is the most common cause of urinary tract infection in Ardabil province.

Phenotypic and molecular characterization of extended-spectrum ß-lactamase/AmpC- and carbapenemase-producing Klebsiella pneumoniae in Iran.

BACKGROUND: The objective of the current study is to evaluate the phenotypic and molecular characterization of ESBL/AmpC- and carbapenemase-producing K. pneumoniae isolates in Iran. METHODS: From October 2018 until the end of April 2020, different clinical samples were collected and K. pneumoniae isolates were identified using conventional biochemical tests and PCR assay. Antibiotic susceptibility pattern was determined using the Kirby-Bauer disk diffusion method. Modified Hedge Test (MHT) was applied to the identification of carbapenemase-producing K. pneumoniae. ESBL and AmpC-producing K. pneumoniae were detected using Double Disc Test (DDT) and Disc Potentiation Test (DPT), respectively. The presence of carbapenemase, ESBL, and AmpC encoding genes was screened by Polymerase Chain Reaction (PCR) assay. RESULTS: A total of 100 K. pneumoniae isolates were collected. K. pneumoniae isolates had the highest resistance rate to cefazolin (66%) and cefotaxime (66%). Meropenem and amikacin with sensitivity rates of 76% and 69% were the most effective antimicrobial agents on K. pneumoniae isolates. It was found that 12 (12%), 27 (27%), and 9 (9%) K. pneumoniae isolates were positive in MHT, DDT, and DPT tests, respectively. Among the carbapenemase-encoding genes, blaOXA-48 (24%) and blaIMP (13%) genes had the highest frequency, while blaKPC and blaGIM genes were not detected among K. pneumoniae isolates. blaTEM (48%) and blaCMY (8%) genes had the highest frequency among ESBL and AmpC ß-lactamase-encoding genes, respectively. CONCLUSIONS: It is vital to adopt effective control strategies for K. pneumoniae infections and ensure rapid identification of antibiotic resistance profile.

High-level resistance to aminoglycosides and ampicillin among clinical isolates of Enterococcus species in an Iranian referral hospital.

BACKGROUND AND OBJECTIVES: Nowadays, high-level aminoglycosides and ampicillin resistant Enterococcus species are among the most common causes of nosocomial infections. The present study was conducted to determine the prevalence of high-level resistance to aminoglycosides and ampicillin among clinical isolates of Enterococcus species in Ardabil, Iran. MATERIALS AND METHODS: In this cross-sectional study, a total of 111 Enterococcus species were collected from different clinical specimens between 2013 and 2015. Enterococcus species were identified using standard phenotypic and genotypic methods. BHI agar screen and agar dilution methods were used for detection of high-level gentamicin and streptomycin resistance (HLGR and HLSR) and minimal inhibitory concentration (MIC) of ampicillin, respectively. RESULTS: Of 111 clinical isolates, 59 (53.2%) and 25 (22.5%) isolates were E. faecalis and E. faecium, respectively, based on the PCR results. Totally, 60.3% and 56.7% of isolates were HLGR and HLSR, respectively, as well as 51.35% were HLGR plus HLSR. Among HLGR isolates, 36 (61.01%), 18 (72%) and 13 (48.14%) were E. faecium, E. faecalis and non-faecalis non-faecium species, respectively. Among HLSR isolates, 33 (55.93%), 16 (64%) and 14 (51.85%) were E. faecalis, E. faecium and non-faecalis non-faecium species, respectively. All HLGR isolates contained aac(6')Ie-aph(2″)Ia gene. Overall, the prevalence of high-level ampicillin resistance among Enterococcus species was 17.1%. For E. faecalis, E. faecium and non-faecalis non-faecium species, ampicillin resistance rates were as follows: 11 (40.74%), 7 (28%) and 1 (1.69%), respectively. For aminoglycoside antibiotics, the resistance rate was significantly higher in E. faecium isolates and for ampicillin it was higher in E. faecalis isolates. CONCLUSION: The frequency of high-level aminoglycoside resistant enterococcal isolates in our hospital was high and significant ampicillin resistance was noticed. This would require routine testing of enterococcal isolates for HLAR and ampicillin susceptibility.

Molecular characterization of quinolone resistant Shigella spp. isolates from patients in Ardabil, Iran.

BACKGROUND AND OBJECTIVES: Shigella is an etiological agent of shigellosis. Antibiotic therapy has a critical role in decreasing serious complications of shigellosis. The present study aimed to determine the multi-drug resistance strains and to detect fluoroquinolone related mutations. MATERIALS AND METHODS: In this descriptive, cross sectional study, a total of 113 Shigella isolates were collected from 1280 patients admitted to Bu-Ali hospital in Ardabil province during 2015-17. Antibiotic resistance pattern of isolates was evaluated using Kirby Bauer method and finally, the MICs of ciprofloxacin were determined. In order to determine any mutations in QRDR region, parC and gyrA genes of resistant strains were amplified and sequenced. RESULTS: Shigella spp. isolates were identified using ipaH amplification and rfc and wbgz genes were used for molecular detection of S. flexneri and S. soneii, respectively. Our results showed that the predominant species in Ardabil province was S. sonnei (69.91%). Most of isolates (82%) were resistant to trimethoprim/sulfamethoxazole (TMP/SMX); 51% were nalidixic acid resistant and 4.4% were floroquinolones resistant. All examined isolates were susceptible to imipenem (100%). Mutation in gyrA and parC genes were detected in all fluoroquinolone resistant isolates (5 isolates). Although, in this study the rate of resistance to ciprofloxacin was low, but in the lack of preventive strategy it will be a major challenge of public health in future. CONCLUSION: This study provided information on the prevalence and antimicrobial susceptibility patterns of Shigella isolates in Ardabil province, Iran. Also this study showed a high-level of resistance to commonly used antibiotics among Shigella isolates.

Designing and Construction of a Cloning Vector Containing mpt64 Gene of Mycobacterium tuberculosis.

BACKGROUND: Tuberculosis caused by Mycobacterium tuberculosis (M. tuberculosis), remains as one of the leading causes of deaths worldwide, with nearly two million death cases annually. BCG (Bacille Calmette-Guerin) continues to be the most widely used vaccine in the world, but the protective immunity differs in different parts of the world. Accordingly, new strategies including DNA vaccines are essentially needed. This study was aimed to design and construct a cloning vector containing mpt64 gene of M. tuberculosis. MATERIALS AND METHODS: M. tuberculosis H37Rv was cultured on Lowenstein Jensen medium, and genomic DNA was extracted. The mpt64 gene was amplified by PCR using designed specific primers. After the digestion of mpt64 and pcDNA3.1 (+) by BamHI and EcoRI restriction enzymes, the mpt64 fragment was ligated into the digested vector using T4 DNA ligase enzyme. Then, the recombinant vector was transformed into competent Escherichia coli (E. coli) TOP10 strain. To confirm the colonies of transformed bacteria, antibiotic resistance, colony-PCR, restriction enzyme digestion and DNA sequencing were used. RESULTS: To confirm the clones, colony-PCR using mpt64 specific primers was performed and the fragment of 718 bp was observed by gel electrophoresis. Clones were also verified by restriction enzyme digestion using BamHI and EcoRI restriction enzymes and the 718 bp fragment was observed. Furthermore, results of DNA sequencing showed 100% homology with the mpt64 fragment of H37Rv in GenBank. CONCLUSION: In this study, the mpt64 fragment was successfully cloned in pcDNA3.1 (+) vector. This construct can be used in future studies as a DNA vaccine in animal models to induce immune system responses.

Current concepts on immunopathogenesis of hepatitis B virus infection.

Hepatitis B virus (HBV) infection is a leading cause of liver damage and hepatic inflammation. Upon infection, effective antiviral responses by CD8+ T cells, CD4+ T cells, Natural killer (NK) cells, and monocytes can lead to partial or complete eradication of the viral infection. To date, many studies have shown that the production of inhibitory cytokines such as Interleukin 10 (IL-10), Transforming growth factor beta (TGF-ß), along with dysfunction of the dendritic cells (DCs), and the absence of efficient innate immune responses could lead to T cell exhaustion, development of persistent infection, and inability to eradicate the viral infection from liver. Understanding the immunopathogenesis of the virus could be useful in providing further insights toward novel strategies in the eradication of HBV infection.

Colonization of Pneumocystis jirovecii in Patients Who Received and not Received Corticosteroids Admitted to the Intensive Care Unit: Airborne Transmission Approach.

BACKGROUND & OBJECTIVE: Pneumocystis pneumonia (PCP) is responsible for pulmonary infection in immunocompromised patients. The current study aimed at investigating the frequency of Pneumocystis colonization in patients hospitalized in the intensive care unit (ICU) and evaluating the relationship between PCP and Pneumocystis colonization. METHODS: The current cross sectional study was conducted on bronchoalveolar lavage (BAL) fluids of 100patients collected from surgery and neurosurgery ICUs with different underlying corticosteroid therapy conditions. Patients were divided into 2 groups (patients receiving and not receiving corticosteroids). Direct examination on BAL fluids was performed by the Gomori methenamine silver and Giemsa staining techniques. Additionally, 2 filtered air samples of the 2 above mentioned units were collected. A nested-PCR targeted mtLSUrRNA gene and sequencing were used to identify Pneumocystis spp. RESULTS: In direct microscopy, 31 out of 100 hospitalized patients (31%) showed positive results. Twenty-three (46%) of smear positive patients were from the group of patients receiving corticosteroid, the other 8(16%) were from the group of patients not receiving corticosteroids (P= 0.001). Pneumocystis jirovecii DNA was detected in 77 out of 100 BAL samples by nested-PCR (77%) in which 40 (52%) and 37 (48%) samples were obtained from the patients receiving and not receiving corticosteroids, respectively. Pneumocystis genome was observed in 1 of the 2 filtered air samples. CONCLUSION: A significant number of patients receiving corticosteroids were also colonized by P. jirovecii that may be predisposed to PCP or be transmitted to susceptible patients. A significant relationship was observed between the mean hospital stay and detection rate.

A study on the immune response induced by a DNA vaccine encoding Mtb32C-HBHA antigen of Mycobacterium tuberculosis.

OBJECTIVES: Tuberculosis (TB) has still remained a global health issue. One third of the world's population is infected with tuberculosis and the current BCG vaccine has low efficiency; hence, it is necessary to develop a new vaccine against TB. The aim of the current study was to evaluate the efficiency of a novel DNA vaccine encoding Mtb32C-HBHA antigen in inducing specific immune responses against Mycobacterium tuberculosis. MATERIALS AND METHODS: A DNA plasmid vaccine expressing Mtb32C-HBHA fusion protein was constructed and its ability in protein expression was examined by RT-PCR and Western blot methods. Female BALB/c mice were vaccinated with 100 µg of purified recombinant vector in an attempt to assess its immunogenicity and protective efficacy. Further, the cytokines, IFN-γ, IL-12, IL-4, IL-10, and TGF-ß were assessed. RESULTS: The levels of all the studied cytokines were significantly increased (P<0.05) compared with the control group. IFN-γ production in the group receiving DNA vaccine plus BCG was increased compared with those receiving only DNA vaccine or BCG (P<0.001). CONCLUSION: The immunogenicity of the new chimeric DNA vaccine was confirmed alone and in combination with BCG. Based on the results of the current study, the constructed DNA vaccine induced the expression of Mtb32C-HBHA fusion protein efficiently in vitro. Furthermore, high levels of the specific cytokines were induced in mice. By using this DNA vaccine as a booster after BCG, higher amounts of IFN-γ will be produced.

Immunogenicity of a DNA Vaccine Encoding Ag85a-Tb10.4 Antigens from Mycobacterium Tuberculosis.

BACKGROUND: Tuberculosis is a life threatening disease that is partially prevented by BCG vaccine. Development of more effective vaccines is an urgent priority in TB control. Ag85a and Tb10.4 are the members of culture filter protein (CFP) of M. tuberculosis that have high immunogenicity. OBJECTIVE: To analyze the immunogenicity of Ag85a-Tb10.4 DNA vaccine by enzyme-linked immunosorbent assay (ELISA). METHODS: In this study a previously described plasmid DNA vaccine encoding Ag85a-Tb10.4 was used to examine its capability in the stimulation of immune responses in an animal model. Female BALB/c mice were vaccinated with 100 µg of purified recombinant vector intramuscularly 3 times at two-week intervals and the levels of five cytokines including IFN-γ, IL-12, IL-4, IL-10 and TGF-ß were measured. RESULTS: The levels of IFN-γ and IL-12 for the mice following immunization with Ag85A-Tb10.4 was significantly greater than that of the BCG and control group (p<0.05). However, there was no significant difference in the levels of IL-4, IL-10 and TGF-ß between groups. CONCLUSION: IFN-γ and IL-12 Th1 cytokines increased significantly in mice vaccinated with Ag85a-TB10.4 DNA vaccine in comparison to the control and BCG groups. Our results may serve as a groundwork for further research into the prevention and treatment of tuberculosis.

Designing and Development of a DNA Vaccine Based On Structural Proteins of Hepatitis C Virus.

BACKGROUND: Hepatitis C virus (HCV) infection is one of the most prevalent infectious diseases responsible for high morbidity and mortality worldwide. Therefore, designing new and effective therapeutics is of great importance. The aim of the current study was to construct a DNA vaccine containing structural proteins of HCV and evaluation of its expression in a eukaryotic system. METHODS: Structural proteins of HCV (core, E1, and E2) were isolated and amplified from JFH strain of HCV genotype 2a using PCR method. The PCR product was cloned into pCDNA3.1 (+) vector and finally were confirmed by restriction enzyme analysis and sequencing methods. The eukaryotic expression of the vector was confirmed by RT-PCR. RESULTS: A recombinant vector containing 2241bp fragment of HCV structural genes was constructed. The desired plasmid was sequenced and corresponded to 100% identity with the submitted sequences in GenBank. RT-PCR results indicated that the recombinant plasmid could be expressed efficiently in the eukaryotic expression system. CONCLUSION: Successful cloning of structural viral genes in pCDNA3.1 (+) vector and their expression in the eukaryotic expression system facilitates the development of new DNA vaccines against HCV. A DNA vaccine encoding core-E1-E2 antigens was designed. The desired expression vector can be used for further attempts in the development of vaccines.

Designing and Construction of a DNA Vaccine Encoding Tb10.4 Gene of Mycobacterium tuberculosis.

BACKGROUND: Tuberculosis (TB) remains as a major cause of death. Construction of a new vaccine against tuberculosis is an effective way to control it. Several vaccines against this disease have been developed. The aim of the present study was to cloning of tb10.4 gene in pcDNA3.1(+) plasmid and evaluation of its expression in eukaryotic cells. METHODS: Firstly, tb10.4 fragment was amplified by PCR and the PCR product was digested with restriction enzymes. Next, it was cloned into pcDNA3.1(+) plasmid. Following that, pcDNA3.1(+)/tb10.4 recombinant plasmid was transfected into eukaryotic cells. RESULTS: 5700 bp band for pcDNA3.1(+)/tb10.4 recombinant plasmid and 297 bp fragment for tb10.4 were observed. Cloning and transfection were successful. CONCLUSION: Successful cloning provides a basis for the development of new DNA vaccines against tuberculosis.