The emergence of plasmid-encoded oxacillinase and carbapenemase among uropathogenic Escherichia coli (UPEC) isolated from hospitalized patients in the North of Iran.

Carbapenemase enzyme production is responsible for resistance to carbapenem among Gram-negative bacteria. This study aimed to detect common carbapenemase and oxacilinase genes among uropathogenic E. coli (UPEC) isolated from hospitalized patients in Rasht, north of Iran. In the present study, from 2000 urine samples, 263 UPEC strains were isolated from inpatients with urinary tract infections (UTI) in 2020. The Kirby-Bauer disk diffusion susceptibility test was used to determine the sensitivity or resistance of isolates to antimicrobial compounds. The double-disk test confirmed extended-spectrum ß lactamase (ESBL) production phenotypically, and the presence and distribution of genes encoding carbapenemase and oxacilinase were investigated using polymerase chain reaction (PCR). Based on the findings, 13/263 isolates (8 ESBL and five non-ESBL) showed a non-susceptible phenotype to at least one of the studied carbapenem group antibiotics, and 121 (46%) isolates were ESBL-producers. PCR for oxacilinase and carbapenemase genes was done on all 126 isolates, including ESBL-positive and carbapenem-resistant strains, in which 10 (7.9%) and 25 (19.8%) isolates harbored OXA-1 and IMP genes, respectively. Also, OXA-2, OXA-10, OXA-48, VIM, and NDM genes were not found in any studied isolates. IMP and OXA-1 genes among carbapenemase-producing isolates indicate the possible spread of antibiotic-resistant strains. Hence, identification and control of ESBL and carbapenemase-producing strains, although with almost low frequency due to plasmid genes encoding carbapenemase, is essential for infection control.

Phenotypic and genotypic characterization of metallo-ß-lactamase producing Pseudomonas aeruginosa isolated from burn patients.

Pseudomonas aeruginosa is an opportunistic pathogen associated with many nosocomial infections. This study aimed to detect blaIMP and blaVIM genes and their common subtypes, including bla IMP-1, bla IMP-2, bla VIM-1, and bla VIM-2, among imipenem-resistant Pseudomonas aeruginosa strains. In this study, 117 P. aeruginosa strains were isolated from clinical samples of burn wound patients in Velayat hospital, Rasht, Iran, between 2018 and 2019. These isolates were tested for antimicrobial susceptibilities by disk diffusion and Metallo-ß-Lactamase (MßL) activity. The polymerase chain reaction (PCR) test was applied to detect MßLs encoding genes in MßL-producing strains. The resistance rates were as follows: Tobramycin (59%), Gentamicin (57%), Piperacillin (52%), Ciprofloxacin (51%), Ceftazidime (32%), and Amikacin (26%). Among 27 (23%) imipenem-resistant isolates, 13 (48%) produced the MßL enzyme. PCR results of imipenem-resistant isolates showed that five and four isolates contained the blaVIM (4 blaVIM1, 2 blaVIM2) and blaIMP (4 blaIMP1, 2 blaIMP2) genes, respectively. In addition some of isolates had more than one gene. In this study, 48% of imipenem-resistant strains produced the MßL enzyme. Therefore, systematic surveillance to detect MßL-producing bacteria and rational prescription and use of carbapenems could be helpful to prevent the spread of carbapenem resistance.

Does Conjugation of Silver Nanoparticles with Thiosemicarbazide Increase Their Antibacterial Properties?

The opportunistic pathogen, Pseudomonas aeruginosa, uses different mechanisms as well as biofilm production to acquire antibiotic resistance. The polysaccharide synthesis locus (psl) genes play an important role in P. aeruginosa biofilm formation. Therefore, targeting the expression of psl genes can be a suitable strategy to prevent the formation of biofilms by antibiotic-resistant strains. Today, advances in nanotechnology provide a novel potential strategy to combat antibiotic-resistant bacteria. In this study, the silver nanoparticles (Ag NPs) synthesized using a chemical co-precipitation method and, after conjugation with thiosemicarbazide, their effect on the biofilm-forming ability are studied in P. aeruginosa isolates. Chemical properties of synthesized nanoparticles were determined by scanning and transmission electron microscopy, Fourier transform infrared spectroscopy, diffuse reflectance spectroscopy, ultraviolet-visible spectroscopy, X-ray diffraction, and energy dispersive X-ray spectroscopy. The results confirmed the spherical/cubic morphology, solution stability, and good dispersion of Ag@Glu-TSC NPs with an average size of 40-60 nm. In addition, minimum inhibitory concentration values of functionalized Ag NPs were at least twofold lower than the Ag NPs (alone). The quantitative PCR data analysis showed a decrease in the expression of the pslA gene in the presence of Ag@Glu-TSC NPs, up to 60%, which was associated with a reduction of biofilm formation compared to control. In conclusion, the Ag@Glu-TSC NPs can be considered a new inhibitor of biofilm production in antibiotic-resistant bacteria.

Protective Effect of the Viola spathulata Extract on NCX3 Gene Expression in an Animal Model of Cerebral Ischemia.

Introduction: Viola plant has been used traditionally to treat neurological disorders. We aimed at determining whether pretreatment with Viola spathulata extract can alleviate the severity of ischemic-reperfusion damages and exert its protective effects through the regulation of a sodium/calcium exchanger (NCX3) gene expression in a rat brain. Methods: Male Wistar rats were divided into two main groups: one main group for evaluating Neurologic Deficit Score (NDS) and Infarct Volume (IV) and the other group for the evaluation of NCX3 gene expression in the brain tissue. The latter group was subdivided into the intact, control (vehicle), sham, V5, and V10. The vehicle (control) subgroup received Dimethyl Sulfoxide (DMSO), and V5 and V10 subgroups received V. spathulata extract at the doses of 5 and 10 mg/kg (IP), respectively, for 7 days. After pretreatment, we carried out Middle Cerebral Artery Occlusion (MCAO) for 60 min. Results: In the V5 and V10 subgroups, NDS and IV significantly decreased. MCAO upregulated NCX3 gene expression in the core, penumbra, and subcortical regions compared with the intact subgroup. The V5 subgroup significantly downregulated the NCX3 gene expression level in the core compared with the control subgroup. The V10 subgroup showed downregulation of the NCX3 gene expression level in the core, penumbra, and subcortex compared with the control subgroup. Conclusion: V. spathulata extract may have a neuroprotective role against MCAO-induced ischemic brain damage, possibly by preventing the alteration of NCX3 gene expression level. Highlights: MCAO results Infarct Volume (IV) and Neurologic Deficit Score (NDS);MCAO upregulated NCX3 gene expression in brain tissues;Viola spathulata extract pretreatment decreased IV and NDS in brain ischemia;Viola spathulata pretreatment downregulated NCX3 gene expression in brain tissues. Plain Language Summary: Stroke is the second leading cause of death and long term disability. Recently it has been reported that herbal extracts have protective role in ischemia injury. In Iranian traditional medicine Viola plant has a long history to treat disorders such as cancer. So we designed an animal study to investigate Viola plant extract in brain ischemia injury. Viola spathulata extract was administrated to rats for seven days, then animal model of brain ischemia was operated on them and some behavioral, histological and molecular factors were analyzed. Our findings showed that Viola spathulata extract improved behavioral disability, decreased infarct volume in brain tissue, and modulate Sodium/Calcium exchanger 3 gene expression. It could be concluded that Viola spathulata has the neuroprotective effect in animal stroke model and is a good candidate for nutritional supplements, although further studies are needed.

Toward a universal influenza virus vaccine: Some cytokines may fulfill the request.

The influenza virus annually causes widespread damages to the health and economy of the global community. Vaccination is currently the most crucial strategy in reducing the number of patients. Genetic variations, the high diversity of pandemic viruses, and zoonoses make it challenging to select suitable strains for annual vaccine production. If new pandemic viruses emerge, it will take a long time to produce a vaccine according to the new strains. In the present study, intending to develop a universal influenza vaccine, new bicistronic DNA vaccines were developed that expressed NP or NPm antigen with one of modified IL-18/ IL-17A/ IL-22 cytokine adjuvants. NPm is a mutant form of the antigen that has the ability for cytoplasmic accumulation. In order to investigate and differentiate the role of each of the components of Th1, Th2, Th17, and Treg cellular immune systems in the performance of vaccines, Treg competent and Treg suppressed mouse groups were used. Mice were vaccinated with Foxp3-FC immunogen to produce Treg suppressed mouse groups. The potential of the vaccines to stimulate the immune system was assessed by IFN-γ/IL-17A Dual FluoroSpot. The vaccine's ability to induce humoral immune response was determined by measuring IgG1, IgG2a, and IgA-specific antibodies against the antigen. Kinetics of Th1, Th2, and Th17 cellular immune responses after vaccination, were assessed by evaluating the expression changes of IL-17A, IFN-γ, IL-18, IL-22, IL-4, and IL-2 cytokines by semi-quantitative real-time RT-PCR. To assess the vaccines' ability to induce heterosubtypic immunity, challenge tests with homologous and heterologous viruses were performed and then the virus titer was measured in the lungs of animals. Evaluation of the data obtained from this study showed that the DNA-vaccines coding NPm have more ability to induces a potent cross-cellular immune response and protective immunity than DNA-vaccines coding NP. Although the use of IL-18/ IL-17A/ IL-22 genetic adjuvants enhanced immune responses and protective immunity, Administration of NPm in combination with modified IL-18 (Igk-mIL18-IgFC) induced the most effective immunity in Treg competent mice group.

Epidemiology, laboratory diagnosis and clinical aspects of fungal pulmonary infections in 384 patients hospitalized in pulmonary units in Guilan province, Iran.

BACKGROUND AND OBJECTIVES: The respiratory tract is the most common site for developing fungal infections. People who have a weakened immune system are more susceptible to respiratory system involvement with fungi. Fungal infections of the respiratory tract are largely unrecognized and their true burden is elusive. Therefore, the aim of the current study was to evaluate the clinical spectrum, demographic characteristics, risk factors, and etiology of fungal respiratory infections in 384 patients hospitalized in pulmonary units of Razi hospital, Guilan province, Iran. MATERIALS AND METHODS: A total of 384 lung specimens (192 Bronchoalveolar lavages (BAL) and 192 sputa) were obtained from patients who met the inclusion criteria. All samples were analyzed by direct microscopy and culture. Fungal identification was accomplished by internal transcribed spacer (ITS) and beta-tubulin sequencing. Also, in patients suspected to invasive pulmonary aspergillosis BAL specimens were tested for galactomannan (GM) antigen. According to the host factors (clinical symptoms, radiology findings and predisposing factors which were defined as inclusion criteria), and the positive results in direct examination, culture and serology (GM for aspergillosis) the infection was confirmed. RESULTS: Fungal respiratory infection was confirmed in 137 cases (35.67%) including 86 (62.77%) males and 51 (37.23%) females and the highest prevalence of infection was found in the age group of 46-72 years (n=75, 54.74%). Cough (n=129, 94.16%), dyspnea (n=111, 81.02%), purulent sputum (n=85, 62.04%) and weight loss (n=77, 56.2%) were the predominant symptoms. Tuberculosis (n=34, 24.81%), taking chemotherapy regimen (n=30, 21.89%) and diabetes mellitus (n=27, 19.70%) were the predominant underlying conditions. Candida albicans (37.22%) and Candida tropicalis (21.89%) represent the two most commonly isolated species in the current study. Furthermore, according to revised EORTC/MSG (2008) definitions for invasive fungal infections, from 5 cases of pulmonary aspergillosis, 2 (40%) cases of probable invasive pulmonary aspergillosis (IPA) and 3 (60%) cases of possible IPA were diagnosed. CONCLUSION: According to the results of this study, infected infants with congenital CMV infection could identify at early stage by testing Guthrie cards (within 21 days of life). Furthermore, since there is a lack of CMV knowledge in our population, educating and effective counseling by obstetricians/gynecologists to the pregnant women are recommended.

Fungal Isolates of the Respiratory Tract in Symptomatic Patients Hospitalized in Pulmonary Units: A Mycological and Molecular Epidemiologic Study.

INTRODUCTION: Fungal respiratory infections are being recognized with increasing frequency in parallel with an expanding population of immunocompromised patients. In most cases, colonization is the first step in the progression to pulmonary fungal infection. This study was designed to evaluate the distribution of fungal elements in the respiratory tract of symptomatic patients hospitalized in pulmonary units. METHODS: This descriptive cross-sectional study was carried out over a period of two years, from October 2017 to October 2019 in Guilan province, located in Iran's northern region. In the current study, bronchoalveolar lavage or sputum specimens were collected. All samples were analyzed by direct microscopy using KOH 10% and culture. Fungal identification was accomplished by internal transcribed spacer (ITS) and beta-tubulin sequencing. Also, in patients suspected of invasive pulmonary aspergillosis, BAL specimens were tested for galactomannan (GM) antigen. RESULTS: A total of 384 lung specimens (192 bronchoalveolar lavage (BAL) and 192 sputum samples) were obtained from symptomatic patients hospitalized in pulmonary units. Of these, 137 (35.67%) were positive in direct examination and culture. Among the 137 positive cases, most isolates were from male patients 86 (62.77%) and most of them were between 46 and 72 years. Candida albicans (37.22%) and Candida tropicalis (21.89%) represent the two most commonly isolated species in the current study. Cough (94.16%), dyspnea (81.02%), purulent sputum (62.04%) and weight loss (56.2%) were the predominant symptoms and tuberculosis (24.81%), chemotherapy (21.89%) and diabetes mellitus (19.70%) were the predominant underlying conditions. Also, 5 cases of invasive pulmonary aspergillosis and 1 case of mucormycosis were diagnosed. CONCLUSION: Candida albicans was the most common fungal species isolated from symptomatic patients hospitalized in pulmonary units. Tuberculosis, chemotherapy and diabetes mellitus were important underlying conditions for pulmonary fungal colonization and/or infection.

Curcumin-human serum albumin nanoparticles decorated with PDL1 binding peptide for targeting PDL1-expressing breast cancer cells.

In this research, programmed death ligand 1 (PDL1) binding peptide was used for targeted delivery of curcumin to high PDL1-expressing breast cancer cells. Human serum albumin-curcumin nanoparticles (HSA/Cur NP) were first prepared by desolvation method and then functionalized with PDL1 binding peptide. Peptide conjugation to HSA/Cur NPs was confirmed by Fourier transform infrared and UV-visible spectroscopy. The formation of HSA/Cur NP was characterized by transmission electron microscope and scanning electron microscope. The size and zeta potential were determined by dynamic light scattering. The average particle size of the HSA/Cur NPs and peptide-HSA/Cur NPs were 197 and 246.5 nm, respectively. Evaluation of cellular uptake showed enhanced internalization of peptide-HSA/Cur NPs in high PDL1-expressing cancer cells compared to HSA/Cur NPs. The cell viability and apoptosis determination demonstrated higher cytotoxicity of HSA/Cur NPs relative to free curcumin in breast cancer cells. Peptide conjugation to HSA/Cur NPs increased cytotoxicity significantly concerning high PDL1-expressing breast cancer cells. In conclusion, peptide-HSA/Cur NPs improved cellular uptake and cytotoxicity of HSA/Cur NPs in high PDL1-expressing breast cancer cells. These results suggest that PDL1 has potential to be used as a target for selective drug delivery and promising candidate for the treatment of PDL1-expressing breast cancer cells.

Diphtheria toxoid nanoparticles improve learning and memory impairment in animal model of Alzheimer's disease.

BACKGROUND: Alzheimer's disease (AD) is a neurodegenerative disorder involving memory. The present study aimed at evaluating the effects of encapsulated diphtheria toxoid (DT) on behavioral learning impairment, and XBP1 mRNA splicing in AD. METHODS: A DT-loaded nanoparticle (NP) carrier was prepared using the ionic gelation method. Sixty-three rats were divided into nine groups: (1) healthy, (2-4) sham, and (5-9) AD models: (5) AD was induced by intracerebroventricular injection of amyloid beta (Aß) 1-42. (6) The rats received a subcutaneous diphtheria vaccine only 28 days before Aß injection. (7) The rats received an intranasal diphtheria vaccine, in group 8, induced by administering empty chitosan NPs. 9) it was induced by administering chitosan NPs carrying DT. Morris water maze (MWM) test was used to examine the animals' learning and memory. Also, X-box binding protein 1 (XBP-1) mRNA gene splicing was studied in the hippocampus by reverse-transcription polymerase chain reaction (RT-PCR). RESULTS: For the first time, chitosan NPs were prepared with an average diameter size of 40 nm and the effectiveness of approximately 70% during DT encapsulation. In comparison with the healthy group, the AD models exhibited significant impairment of learning and memory (P < 0.05), while DT-administrated animals showed significant improvements in learning and memory impairment (P < 0.05). XBP-1 mRNA gene splicing was only detected in an untreated AD group, while encapsulated DT completely inhibited splicing. CONCLUSION: The therapeutic effects of DT chitosan NPs against learning and memory impairment were observed in this study, and XBP1 mRNA splicing was reported in the animal models.

High incidence of type III secretion system associated virulence factors (exoenzymes) in Pseudomonas aeruginosa isolated from Iranian burn patients.

OBJECTIVE: The present study aimed to determine the prevalence of virulence factors and antimicrobial resistance profile of Pseudomonas aeruginosa strains isolated from Iranian burn patients. RESULTS: This cross-sectional study performed on 100 P. aeruginosa isolates which were recovered from burn wound specimens in 2014-2015. All presumptive isolates were identified by standard microbiologic tests. Antimicrobial susceptibility test was carried out by disk diffusion method. The presence of virulence genes was determined by PCR method. Antibiotic susceptibility results revealed that the isolates were mostly susceptible to amikacin (61%), ceftazidime (60%), and imipenem (55%). Moreover, 59% of the isolates were multi-drug resistance (MDR). The most prevalent MDR pattern was aminoglycosides-penicillins-fluoroquinolones-carbapenems (15%). The presence of exoT, exoY, exoS and exoU genes was detected in 100%, 100%, 59%, and 41% of the tested isolates, respectively. Results points out the pattern of MDR and genetic diversity of type III secretion system among P. aeruginosa strains isolated from the burn population. Overall, the association of MDR and the presence of the specific virulence genes can be a predictive marker for the persistence of these isolates in the hospitals and subsequently a worse clinical condition for the affected patients.

Effect of Amino Acid Substitutions on Biological Activity of Antimicrobial Peptide: Design, Recombinant Production, and Biological Activity.

Recently, antimicrobial peptides have been introduced as potent antibiotics with a wide range of antimicrobial activities. They have also exhibited other biological activities, including anti-inflammatory, growth stimulating, and anti-cancer activities. In this study, an analog of Magainin II was designed and produced as a recombinant fusion protein. The designed sequence contained 24 amino acid residues (P24), in which Lys, His, Ser residues were substituted with Arg and also, hydrophobic Phe was replaced with Trp. Recombinant production of P24 in Escherichia coli (E. coli) BL21 using pTYB21, containing chitin binding domain and intein sequence at the N-terminus of the peptide gene, resulted in 1 µg mL-1 product from culture. Chitin column chromatography, followed by online peptide cleavage with thiol reducing agent was applied to purify the peptide. Antimicrobial activity was evaluated using five bacteria strains including Staphylococcus aureus, Enterococcus faecalis, Klebsiella pneumonia, E. coli, and Pseudomonas aeruginosa. Designed AMP exhibited promising antimicrobial activities with low minimum inhibitory concentration, in the range of 64-256 µg/mL. P24 showed potent antimicrobial activity preferably against Gram-positive bacteria, and more potent than pexiganan as a successful Magainin II analog for topical infections. In general, further modification can be applied to improve its therapeutic index.

Association study of copy number variation in BMP8A gene with the risk of ankylosing spondylitis in Iranian population.

BACKGROUND: Copy number variation (CNV) of DNA segments has been considered as an important component of genetic variation, affecting the quality and quantity of gene expression. Bone morphogenic protein 8A (BMP8A) has been reported to function in bone formation. With respect to the bone and joint complications in ankylosing spondylitis (AS), this investigation aimed to study the role of BMP8A gene CNV in impressing the gene expression as well as the disease risk. METHODS: A total of 900 individuals, including 450 patients with AS and 450 healthy controls were enrolled. The copy numbers of BMP8A gene were detected by TaqMan real-time polymerase chain reaction (PCR) method. BMP8A messenger RNA (mRNA) transcript level in peripheral blood mononuclear cells (PBMCs) was also measured by SYBR Green real-time gene expression PCR method. RESULTS: No significant association of BMP8A copy number was detected with the risk of AS. BMP8A mRNA expression level was significantly downregulated in patients compared with controls. mRNA expression level of BMP8A in both AS patients with and without syndesmophyte was significantly lower than the healthy control group. There was no correlation between the mRNA expression level of BMP8A and both demographic and clinical data of the patients. CONCLUSIONS: Although BMP8A gene expression was downregulated in patients with AS, its copy number could not affect the transcript level of BMP8A gene in PBMCs and was not associated with susceptibility to AS in Iranian population. BMP8a may take into account as an indicator of bone formation process in AS, but it seems that mechanisms other than CNV may regulate this protein.

A multicenter-based study on epidemiology, antibiotic susceptibility and risk factors of toxigenic Clostridium difficile in hospitalized patients in southwestern Iran.

Clostridium difficile (recently Clostridioides difficile) is a leading cause of hospital- and antimicrobial-associated diarrhea (AAD). The present study was carried out to investigate the prevalence of toxigenic C. difficile, antibiotic resistance and its associated risk factors in Iranian hospitalized patients. This cross-sectional study was conducted from October 2017 to June 2018 in three teaching hospitals in southwestern Iran. During this period, a total of 215 non duplicated nosocomial AAD samples were collected from the hospitalized patients older than two years of age. Presumptive C. difficile isolates were identified by standard microbiologic methods and confirmed by specific PCR primers. The minimum inhibitory concentrations (MICs) were determined by the agar dilution method. PCR was carried out to determine the presence of toxin genes (tcdA, and tcdB). In all, from the 215 diarrheal samples, the frequency of C. difficile culture-positive samples was 21.4% (n = 46). Of the 46 C. difficile isolates, 43 carried both toxins, two isolates only had the tcdB gene, and one was negative for both toxins. Overall, all isolates of C. difficile were susceptible to metronidazole and vancomycin. The MIC50/MIC90 of metronidazole and vancomycin were 0.75/2 µg/mL, 0.25/0.75 µg/mL, respectively. The findings of this study show the prevalence of CDI in hospitalized patients in southwestern Iran, highlighting the importance of active surveillance of CDI in hospitals. Meanwhile, all of the tested isolates were susceptible to metronidazole and vancomycin, which encourages the use of these antibiotics as the drug of choice for initial treatment of CDI in our region.

Aptamer-based Targeted Delivery of miRNA let-7d to Gastric Cancer Cells as a Novel Anti-Tumor Therapeutic Agent.

miRNAs as one of the potential therapeutic agents have been recently considered for cancer treatment. AS1411 (aptNCL) is a DNA aptamer specifically binding to nucleolin protein on the cancer cell surface with antiproliferative effect. The aim of the study was to develop a conjugate consisting of aptNCL (as targeted delivery of therapeutic agent) and miRNA let-7d (as a tumor suppressor) using two different linking methods and also to evaluate the potential effect of the conjugates on the proliferation of gastric cancer (MKN-45) cell line compared to negative control cell line of human dermal fibroblast (HDF). Conjugation was performed covalently by SM (PEG)2 as a bifunctional crosslinker (conjugate-1) and noncovalently, using 19bp complementary sticky end sequences (conjugate-2). Nucleolin positive MKN-45 and nucleolin negative HDF cells were cultured and treated with the conjugates. Then, the changes in let-7d expression and cell proliferation were determined using Real-time PCR and MTT methods, respectively. In MKN-45 cells, the conjugates caused significant increase in let7-d uptake compared with HDF cells (P = 0.0001). The conjugate-1, likely due to its higher stability compared with the conjugate-2, led to significantly more increase in intracellular let-7d in MKN-45 cells (30 fold versus 15 fold, respectively, P = 0.0001). The conjugates revealed more potent antiproliferative effect against gastric cancer cells compared with aptNCL alone (P = 0.0001). It was found that the aptNCL-let-7d conjugate efficiently carried let-7d into the cancer cells. Also, it appears that in the setting of aptNCL-let-7d conjugate, let-7d and aptNCL moieties could cooperate and synergistically exhibit the antiproliferative effect on cancer cells.

Design, development and evaluation of microRNA-199a-5p detecting electrochemical nanobiosensor with diagnostic application in Triple Negative Breast Cancer.

The new sensitive diagnostic method, electrochemical nanobiosensor, is applied to cancer detection by using specific biomarkers such as miRNA. Studies have shown that for the diagnosis of Triple Negative Breast Cancer (TNBC) patients, in the early stages of the disease, miR-199a-5p as circulating miRNA can be a suitable candidate. In this study, a new electrochemical nanobiosensor for serum miR-199a-5p detection was designed by using modified glassy carbon electrode (GCE) with graphene oxide (GO) and gold nanoroad (GNR) on which a thiolated probe was immobilized. The electrochemical impedance method (EIS) was used to examine the electrochemical characteristics and behavior of nanobiosensor and for verifying the nanomaterial synthesis the scanning electron microscope (SEM) were applied, field emission scanning electron microscope (FE-SEM), UV-Vis spectrophotometry and energy dispersive spectroscopy (EDS). Optimum conditions were investigated for designed nanobiosensor and in optimal conditions, following results were obtained; the linear range of the calibration curve from 15 fM to 148 pM, the detection limit of 4.5 fM, and the standard deviation of 2.9%. The research data showed that our designed electrochemical nanobiosensor has a sensitivity and desirable precision for evaluation of miR-199a-5p that can be used to measure low concentrations of miR-199a-5p in blood serum sample.

Evaluation of the immune responses following co-administration of PilQ and type b-flagellin from Pseudomonas aeruginosa in the burn mouse model.

Considering the increased antibiotic resistance of Pseudomonas aeruginosa, the evaluation of immune response against the antigens of this bacterium seems necessary. In this study, the protective efficacy and immunological properties of P. aeruginosa recombinant PilQ (r-PilQ) and type b-flagellin (FLB) proteins was evaluated in the burn mouse model of infection. The inbred BALB/c mice were immunized with r-PilQ and FLB antigens. To investigate the type of induced immune response, sera were analyzed by ELISA for total IgG, IgG1, and IgG2a isotypes. After the final immunization, the IL-4, IFN-γ, and IL-17 cytokines level were examined in the spleen of non-challenged mice. Fifty days after lethal challenge, the survival rate and bacterial burden in the skin and other internal organs of experimental mice were assessed. The in vivo administration of r-PilQ, FLB and combined antigen resulted in a significant increase in the survival of mice (66%, 75%, and 83%, respectively) infected by the PAO1 strain of P. aeruginosa in the burn model of infection. Immunization of mice with r-PilQ and FLB mixture induced high titers of IL-4 and IL-17 cytokines compared to control groups (P < 0.05). The high titer of antisera raised against combined antigen was able to inhibit the systemic spread of the PAO1 strain from the site of infection to the internal organs. We concluded that the parallel role of IL-4 and IL-17 is necessary for elimination of the bacteria and promotion of survival in the immunized burn mice.

Molecular characterization and Functional Analysis of the PilQ380-706: a Novel Secretin Domain in Pseudomonas aeruginosa.

BACKGROUND: Type 4 pili (T4P) is an important virulence factor of Pseudomonas aeruginosa (P. aeruginosa). T4P pass the outer membrane through a large oligomeric channel made of a single PilQ protein that is most highly conserved at their C-termini. To develop a functional vaccine that can be used in clinical application, the secretin domain of the PilQ (PilQ380-706) was produced as a recombinant protein. METHODS: A 981 bp fragment of C-terminal of the pilQ secretin (pilQ1138-2118) from was designed into the prokaryotic expression vector pET28a. The presence of the pilQ1138-2118 gene in the recombinant construct (pET28a/pilQ) was assessed by double digestion and PCR. After transformation, expression of the recombinant PilQ was induced by addition of IPTG. The expressed recombinant protein was purified by a modified method using a HisTrap affinity column and finally confirmed by SDS-PAGE. The functional activities of the produced PilQ380-706 confirmed by Western blot analysis and twitching inhibition assay. RESULTS: The PCR and enzymatic digestion results showed the presence of the pilQ1138-2118 gene in the construct. The protein electrophoresis showed that the molecular weight of the recombinant PilQ380-706 is approximately 37 kDa. The Western blot analysis confirmed the specificity of specific IgG against the PilQ380-706 protein. The PilQ380-706 protein showed high biological activity in all of these standard assays. CONCLUSION: Since, the PilQ380-706 protein plays an important role in the biogenesis of pili; and thus, the primary establishment of P. aeruginosa; it seems that it can be used as a candidate vaccine or an adjuvant in the future studies.

Prevalence of group B streptococcus colonization in Iranian pregnant women: A systematic review and meta-analysis.

BACKGROUND: Group B Streptococcus (GBS) is an important pathogen in newborns and pregnant women. OBJECTIVE: The present study was carried out to estimate the prevalence of GBS colonization in pregnant women in Iran. MATERIALS AND METHODS: This systematic review and meta-analysis was based on Preferred Reporting Items for Systematic Reviews and Meta-Analysis guideline using the national databases including Society for Information Display, Magiran, Irandoc, Iran Medex, and international databases including MEDLINE, Web of Science, Scopus, PubMed, Science-Direct, Cochrane, Embase, Elton Bryson Stephens Company, Centre for Evidence-Based Medicine, Cumulative Index to Nursing and Allied Health Literature, and Google Scholar, published by 01/30/2017. The I 2 index was used to measure heterogeneity between the studies. RESULTS: In a total of 667 documents, 30 (4.49%) were selected. In this study, the prevalence of GBS colonization in 10090 Iranian pregnant women was calculated as 13.65% [confidence interval (CI): 95%: 10.56-17.45]. Based on geographic region, 24.63% [CI: 95%: 11.52-45.06] in the West and 8.75% [CI: 95%: 6.43-11.8] in the East were the highest and lowest areas in Iran, respectively, and were statistically significant (p = 0.001). Also, with regards to swapping sampling area, Vaginal with 11.96%, Vaginal and Rectal with 13.62%, and Anal and Vaginal with 25.63% were the least to the greatest, respectively, and were statistically significant (p = 0.001). CONCLUSION: Therefore, based on the recommendation of Centers for Disease Control and Prevention as reported by the Ministry of Health and Medical education, early diagnosis, and screening of high-risk women should be done at 35-37 weeks of pregnancy.

Epidemiology of Panton-Valentine Leukocidin harbouring Staphylococcus aureus in cutaneous infections from Iran: a systematic review and meta-analysis.

Panton-Valentine Leukocidin (PVL) producing Staphylococcus aureus has been associated with severity of skin infections and pathology that suggest a major role in pathogenicity. The present study aimed to determine the overall prevalence of PVL harbouring S. aureus isolates from cutaneous infections in Iran. A systematic search was performed by using Medline electronic databases (PubMed) from the papers published by Iranian authors to the end of March 2017. Ten publications which met our inclusion criteria were then selected for data extraction and analysis by Comprehensive Meta-Analysis Software. The pooled prevalence of PVL in cutaneous infections was estimated at 27.9% (95% CI: 17.9-40.6). The range of PVL positivity among S. aureus isolates obtained from cutaneous infections was from 7.4% to 55.6%. In summary, despite the emergence of multiple-drug resistant strains, it seems that the overall prevalence of PVL carrying S. aureus in Iran remains steady regardless of methicillin resistance. However, further research is required to elucidate the interplay between the risk of invasive disease and PVL, especially in Iran.

Development of a Novel Anti-Adhesive Vaccine Against Pseudomonasaeruginosa Targeting the C-terminal Disulfide Loop of the Pilin Protein.

Type IV pili (T4P) are major virulence factors of Pseudomonas aeruginosa (P. aeruginosa) that are associated with primary adhesion, biofilm formation and twitching motility. This study focuses on the introduction of a novel biologically active subunit vaccine derived from the disulfide loop (DSL) of P. aeruginosa pilin. We investigated the expression of the novel PilA in-frame with pET26b vector, which contains three domains, that each domain contains three tandem repeats. The flexible (GGGGS) and (GGGGS)3 linkers were linked between the three tandem repeats and each pilA domain, respectively. The recombinant construct (pET26b/pilA) was transformed and expressed in Escherichia coli BL21 (DE3). The reactivity of specific antiserum against PilA was assessed by ELISA method. The biological activities of this candidate vaccine were evaluated by western blotting, opsonophagocytosis and twitching inhibition assays. The pET26b/pilA plasmid was confirmed by enzymatic digestion. The purified PilA protein was confirmed by immunoblot analysis. The checkerboard titration showed that the optimal dilution of the antibody to react with antigen was 1:8. The results of opsonophagocytosis assay revealed that the antibodies raised against PilA promoted phagocytosis of the PAO1 and 6266E strains to some extent (17.5% and 16.3%, respectively), so the twitching inhibition test confirmed this result. Taken together, these are the preliminary results based on a first chimerical structure failure to induce antibodies that promote the opsonization and eradication of the pathogen. Therefore, the biological activity of the PilA protein showed that it should be introduced with other proteins or target antigens against P. aeruginosa in the future studies.