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Nonencapsulated, nontypeable Hemophilus influenzae [NTHi] remains an important cause of acute otitis and respiratory diseases in children and adults. NTHi bacteria are one of the major causes of respiratory tract infections, including acute otitis media, cystic fibrosis, and community-acquired pneumonia among children, especially in developing countries. The bacteria can also cause chronic diseases such as chronic bronchitis and chronic obstructive pulmonary disease in the lower respiratory tract of adults. Such bacteria express several outer membrane proteins, some of which have been studied as candidates for vaccine development. Due to the lack of effective vaccines as well as the spread and prevalence of NTHi worldwide, there is an urgent need to design and develop effective vaccine candidates against these strains
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Background: The pathogenesis of nontypeable Haemophilus influenzae [NTHi] begins with adhesion to the rhinopharyngeal mucosa. Almost 38-80% of NTHi clinical isolates produce proteins that belong to the High Molecular Weight [HMW] family of adhesins, which are believed to facilitate colonization
Methods: In the present study, the prevalence of hmwA, which encodes the HMW adhesin, was determined for a collection of 32 NTHi isolates. Restriction Fragment Length Polymorphism [RFLP] was performed to advance our understanding of hmwA binding sequence diversity
Results: The results demonstrated that hmwA was detected in 61% of NTHi isolates. According to RFLP, isolates were divided into three groups
Conclusion: Based on these observations, it is hypothesized that some strains of nontypeable Haemophilus influenzae infect some specific areas more than other parts
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Background: Tuberculosis is second cause of death after HIV infection related to infectious diseases in the world. According to WHO reports, there are many risk factors for reactivation of tuberculosis infection. The aim of this study was to know if assisted reproductive techniques [ART] reactivate latent tuberculosis infection
Materials and methods: In this retrospective cross sectional descriptive study that was made to evaluate the effect of ART on reactivation of latent tuberculosis. All patients with positive PPD test, TB in the past or patients who had sequels of TB from 2001 to 2012 were included in this study. Based on questionnaire, including demographics, kind of infertility treatment and active tuberculosis, one expert physician was asking some questions about sign and symptoms of tuberculosis. If Patients had signs or symptoms of tuberculosis, they were visited by infectious disease specialist. Patients with tuberculosis were signed as reactivation of latent tuberculosis
Results: Of 770 patients, 433 [56.2%] cases were only PPD positive, and 30.3% had PPD positive and signs of TB in the past. 81 [10.5%] patients did IUI, 126 [16.4%] ICSI, 43 [5.6%] IVF, 11 [1.4%] IVF/ICSI, and 7 [0/9%] ZIFT and 3 had donor of oocyts. There were 90 [11.7%] pregnancy, 24 [3.1%] deliveries, 30 [3.9%] abortions and 13 [1.7%] ectopic pregnancies, but there was only one case of reactivated TB in this study
Conclusion: This study shows that patients with positive PPD do not need TB prophylaxis. For better result, we need a prospective study
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Background: During the recent years, significant progress has been achieved on development of novel anti-viral drugs. Natural products are assumed as the potential sources of novel anti-viral drugs; therefore, there are some previous studies reporting the anti-viral compounds from venomous animals. Based on the significant value for tracing of non-toxic anti-viral agents from natural resources, this study was aimed to investigate the anti-viral activity of some HPLC purified fractions derived from the venom of Iranian scorpion, Hemiscorpius lepturus, against human immunodeficiency virus 1 [HIV-1] and herpes simplex virus 1 [HSV-1]
Methods: H. Lepturus crude venom was subjected to reverse phase HPLC analysis to determine its active components precisely where four dominant fractions obtained at retention time of 156-160 minutes. The phospholipase A2 and hemolytic activities of the purified fractions were first evaluated. Then the anti-viral activity was measured using single cycle HIV [NL4-3] replication and HSV [KOS] plaque reduction assays
Results: The H. lepturus crude venom inhibited HIV replication by 73% at the concentration of 200 micro g/ml, while it did not show significant anti-HSV activity. It also inhibited the cell-free viral particles in a virucidal assay, while it showed no toxicity for the target cells in a proliferation assay. The four HPLC fractions purified from H. lepturus inhibited HIV with IC50 of 20 micro g/ml
Conclusion: H. lepturus venom contains components with considerable anti-HIV activity insofar as it has virucidal activity that offers a novel therapeutic approach against HIV infection. Our results suggest a promising pilot for anti-HIV drug discovery with H. lepturus scorpion venom
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Haemophilus influenzae is a bacterium that can act as a pathogen of human respiratory tract. Its infections have been traditionally treated with antibiotics, and then the use of conjugate vaccines has been very successful in prevention of infectious diseases. These vaccines consisted of capsular polysaccharide of H. influenzae with an immunogenic protein. The aim of this study was molecular analysis of oapA gene in Iranian H. influenzae strains as a conserve immunogenic protein for vaccine candidate. Clinical H. influenzae strains were collected from Milad Hospital in Tehran, and cultured on chocolate agar. After diagnostic test, biochemical tests were performed for distinguishing strains biotype. The DNA extracted by boiling method and specific PCR reactions were designed for oapA gene molecular analysis. Finally sequencing of PCR products was performed for confirming our results. The results of PCR exhibited that 79% of clinical samples had 95-99% similarity to NCBI sequences of H. influenzae oapA gene, and the remaining 21% had a 125 nucleotide deletion and 94-96% similarity to NCBI sequences. Also this study showed that all strains having 125 nucleotide deletion [NontypeableH. influenzaeand H. influenzae type b] had 95-99% similarity to NCBI sequences. The results of determining the biotype revealed that the studied strains were related to different biotypes. The results of this study suggested that use of oapA gene protein for production of vaccine against H. influenzae type b be further considered
Subject(s)
Haemophilus Vaccines , Polymerase Chain ReactionABSTRACT
The incidence of extended-spectrum beta- lactamase- producing bacteria has been increased worldwide. The most common cause of resistance to extended-Spectrum cephalosporins in Klebsiella pneumoniae is the production of extended-spectrum beta lactamases [ESBLs]. In the past decade, CTX-M enzymes have become the most prevalent ESBLs in Europe, Canada and Asia. The aim of this study was to find the prevalence of ESBL-producing K .pneumoniae and molecular detection of CTX-M group in these bacteria. In the descriptive study, 100 K. pneumoniae isolates were detected bystandard biochemical tests between April 2012 and September 2012, in Besat and Imam Reza hospitals of Tehran, Iran. Susceptibility to antimicrobial agents was tested for 10 antibiotics by the disk agar diffusion [DAD] method. Also, ESBL production was screened by combined disk diffusionas recommended by the Clinical and Laboratory Standards Institute [CLSI]. Then, Screened isolates were assayed by PCR for detection of CTX-M-1 group genes. Of 100 K. pneumonia isolates, 26 isolates produced ESBLs, and also 42 isolates were CTXM- 1 producer using PCR method. According to the differences between the results of phenotypic and genotypic tests, it seems that molecular detection of drug-resistance genes is necessary. Further investigations are needed to determine the epidemiology of ESBL producing K. pneumoniae in Iran
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Application of adjuvants with microbial origins is a recently highlighted approach in the vaccinology trials. Archaeosomes are among these microbial compounds with both adjuvant and liposomal activities and features. In the present study, recombinant HBsAg encapsulated into Methanobrevibacter smithii [M. smithii] archaeosomes. Balb/c mice immunized with this compound and humoral and cytokine secretion pattern of immunized models analyzed. Frequency of IFN-gamma secreting cells in the HBsAg-containing archaeosomes group was significantly higher than HBsAg and HBsAg+C/IFA groups [p
Subject(s)
Animals, Laboratory , Hepatitis B Surface Antigens , Immunity, Humoral , T-Lymphocytes, Helper-Inducer , Immunity, Cellular , Archaea , Mice, Inbred BALB CABSTRACT
Haps protein plays central role in initial interaction of nontypeable Haemophilus influenzae [NTHi] with human respiratory epithelial cells. While other surface-exposed proteins of NTHi are highly variable, The HapS domain is highly conserved among H. influenzae strains. Recent studies demonstrated that HapS adhesive activity resides within the C-terminal 311 amino acids of the protein and also they showed that the C-terminal 311 amino acids of HapS [C-Haps] are capable of eliciting a protective immune response against NTHi colonization. The pET24a-chaps plasmid harboring c-haps sequence from NTHi PTCC1766 was constructed. The amino acid sequences of C-Haps of this study was aligned with C-Haps of three NTHi strains [N187, TN106, P860295] and antigenicity plot of studied rC-Haps was done bioinformatic software. The pET24a-chaps expression was conducted in E.coli BL21 [D3E] and its expression was confirmed by SDS-PAGE and Western blotting methods. The rC-Haps was purified via immobilized metal affinity chromatography. Amino acid sequence alignment of rCHaps sequence of current study and rC-Haps from the NTHi strains N187, TN106, P860295 showed more than%97 identity. Antigenicity plot identified 9 common highly antigenic domains that were located exactly in conserved regions among 4 different NTHi strains. Due to presence of highly conserved antigenic epitopes among C-Haps of NTHi PTCC1766 and other NTHi strains, rC-Haps of current study could be theoretically a vaccine candidate against NTHi strains of different geographical areas
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Nontypeable Haemophilus influenzae [NTHi] is a common cause of respiratory tract disease and initiates infection by colonization in nasopharynx. The Haemophilus influenzae [H. influenzae] Hap adhesin is an auto transporter protein that promotes initial interaction with human epithelial cells. Hap protein contains a 110 kDa internal passenger domain called "HapS" and a 45 kDa Cterminal translocator domain called "Hapbeta". Hap adhesive activity has been recently reported to be connected to its Cell Binding Domain [CBD] which resides within the 311 C-terminal residues of the internal passenger domain of the protein. Furthermore, immunization with this CBD protein has been shown to prevent bacterial nasopharynx colonization in animal models. To provide enough amounts of pure HapS protein for vaccine studies, we sought to develop a highly optimized system to overexpress and purify the protein in large quantities. To this end, pET24alpha-cbd plasmid harboring cbd sequence from NTHi ATCC49766 was constructed and its expression was optimized by testing various expression parameters such as growth media, induction temperature, IPTG inducer concentration, induction stage and duration. SDS-PAGE and Western-blotting were used for protein analysis and confirmation and eventually the expressed protein was easily purified via immobilized metal affinity chromatography [IMAC] using Ni-NTA columns. The highest expression level of target protein was achieved when CBD expressing E. coli BL21 [DE3] cells were grown at 37°C in 2xTY medium with 1.0 mM IPTG at mid-log phase [OD[600 nm] equal to 0.6] for 5 hrs. Amino acid sequence alignment of expressed CBD protein with 3 previously published CBD amino acid sequences were more than%97 identical and antigenicity plot analysis further revealed 9 antigenic domains which appeared to be well conserved among different analyzed CBD sequences. Due to the presence of high similarity among CBD from NTHi ATCC49766 and other NTHi strains, CBD protein expressed here sounds to be theoretically ideal as a universal candidate for being used in vaccine studies against NTHi strains of various geographical areas. Further investigations to corroborate the potency of this protein as a vaccine candidate are under process
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Staphylococcus aureus [S. aureus] is a major nosocomial pathogen and the infection with this organism in human is increasing due to the spread of antibiotic resistant strains. One of the resistance mechanisms of 5. aureus comprises modification in binding proteins to penicillin. Vaccine strategy may be useful in controlling the infections induced by this organism. This study aimed at developing and producing the recombinant protein PBP2a as a vaccine candidate and evaluating the related humoral immune response in a murine model. A 242 bp fragment of mecA gene was amplified by PCR from S. aureus COL strain and then cloned into prokaryotic expression vector pET-24a. For expression of recombinant protein, pET24alpha-mec cplasmid was transformed into competent E.coll BL2l [DE3] cells. Recombinant protein was over expressed with 1 mM isopropythio-beta-D-galctoside [IPTG] and purified using Ni-NTA agarose. SDS-PAGE and western blotting were carried out to confirm protein expression. For immunization of experimental groups, Balb/c mice were injected subcutaneously with 20 pg of recombinant PBP2a three times with three weeks intervals. The sera of experimental groups were collected three weeks after the last immunization and then specific antibodies were evaluated by ELISA method. Successful cloning of mecA was confirmed by colony-PCR, enzymatic digestion, and sequencing. SDS-PAGE and western blot analysis showed that recombinant protein with molecular weight of 13 kDa is over expressed. In addition, high titer of specific antibody against PBP2a in vaccinated mice was developed as compared with the control group and confirmed the immunogenicity of the vaccine candidate. Results suggest that PBP2a recombinant induced specific antibodies and can be used as Staphylococcal vaccine candidate after further studies
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The aim of this research was to investigate the Cyclooxygenase-2 [COX-2] selective inhibition effect on haloperidol-induced catatonia. In this study, the effect of orally, acutely and Sub-chronically administrations of compound 11b [1-[phenyl]-5-[4-methylsulfonylphenyl]-2-ethylthioimidazole] [2, 4 and 8 mg/kg], a newly selective COX-2 inhibitor, was investigated against the haloperidol-induced catatonia phenomenon comparing to the standard drug scopolamine [1 mg/Kg] followed by microdialysis analysis of Striatum dopaminergic neurotransmission. The results showed a great potency for compound 11b in improvement of catalepsy followed by enhancing the dopaminergic neurotransmission p < 0.05. In addition, our statistical analysis showed that the protective effect of compound 11b against haloperidol-induced catatonia was both dose- and time-dependent. These findings are additional pharmacological data that suggest the effectiveness of compound 11b in treatment of schizophrenic drug overdoses and also Parkinson's disease [PD] affiliated rigidity
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HIV virions with replication capacity are needed for HIV researches, like investigating for new anti HIV agents. Here HIV-1 replication assay was optimized with HIV-1 single cycle replicable [SCR] virions to improve biological safety condition. pSPAX2, pmzNL4-3 and pMD2G plasmids were co-transfected to the HEK293T by using polyfect reagent to produce the SCR HIV-1 virions. Virions were quantified using capture ELISA P24. Different MOI of SCR virions were used for infecting of Target cells [HEK] and the load of the supernatant P24 was monitored days after infection. Single cycle replication assay [SCRA] was developed using kinetic studies data. The P24 load of the infected cells supernatant has linear relation to the beginning infectious MOI. 24 hours post infection with HIV-1 SCR virions the viral particle production was detectable. The highest load of P24 in infected cells supernatant was detected 48 hours after infection. Using this developed method, the 50 and 95 percent inhibitory concentration of [IC[95] and IC[50]] Indinavir and Nevirapine were calculated as 25nM and 50nM. In this study, using SCR HIV-1 virions the SCRA was developed. SCR HIV-1 virions are replicable only for one cycle and this improves the safety of developed assays. The accuracy of assay was examined by quantifying the anti HIV-1 potential of two commercial anti-AIDS drugs and the calculated activity for test agents was equal to previously known amounts
Subject(s)
Virus Replication , Virion , Anti-HIV Agents , Plasmids , Cell Cycle , Indinavir , NevirapineABSTRACT
Neisseria meningitidis serogroup A Polysaccharides vaccines have been available for many years, but these vaccines have many disadvantages due to their induction of T-Cell independent responses. To overcome these problems, many researches have been focused on other parts of bacterial cell component such as OMV [Outer membrane vesicle]. In this study, OMV containing PorA were extracted and evaluated by biological and immunological methods. OMV were extracted by siadat, et al method. Physicochemical properties of extracted OMV were analyzed by electron microscopy and SDS-page. The toxicity of LPS content in OMV was assayed by LAL test. The Presence of PorA was confirmed by western blot. Antibodies synthesis after immunization by OMV was evaluated using ELISA method. The content of extracted protein was 0.1 mg/ml. Size of OMV was between 50 and 150 nanometer. SDS-PAGE showed that PorA was located in 35-40 kDa. LAL test showed that the endotoxin activity was ranged in 126EU/ml which is safe for using. The ELISA test showed that the total IgG titer was elevated after first injection. The results showed that the conformation of extracted OMV was stable, and there were no progeny determinants in OMV. Also, OMV elicited high level of specific antibodies against Neisseria meningitidis serogroup A. These results indicate that the OMV can be used as a meningococcal vaccine after further investigations
Subject(s)
Bacterial Outer Membrane Proteins , Bacterial VaccinesABSTRACT
The aim of this study was to evaluate the effects of dexamethasone on striatal dopaminergic, glutamatergic and gamma amino butyric acid [GABA] ergic neurotransmission in normal and parkinsonian rats. Dexamethasone [0.15, 0.30, 0.60 and 0.8 mg/kg] was administered to normal or parkinsonian rats [i.p.] followed by the analysis of the striatal neurotransmitters concentrations. Additionally, the effect of dexamethasone on the damaged Substantian nigra pars compata [SNc] neurons has been investigated. Dexamethasone resulted in decreased level of striatum glutamatergic-GABAergic and enhanced dopaminergic neurotransmission in normal and parkinsonian rats. In addition, acute treatment with dexamethasone did not improve the lesion at all. These findings suggest the new therapeutic mechanism of action for dexamethasone in Parkinson's disease animal model
Subject(s)
Male , Animals, Laboratory , Dexamethasone/pharmacology , Rats, Wistar , Models, Animal , Parkinson Disease/veterinary , Parkinson Disease/therapy , Dopamine , gamma-Aminobutyric Acid , Glutamic AcidABSTRACT
Neisseria meningitidis is a major cause of bacterial septicemia and meningitis. Currently, there are no vaccines to prevent disease caused by strains of N. meningitidis serogroup B, since cross-reactivity of the serogroup B capsule with human tissue has hampered efforts to develop a reliable vaccine. PilQ is an antigenically conserved outer membrane protein which is essential for meningococcal pilus expression at the cell surface. In the current study, we selected a 1095bp fragment of C-terminal of secretin pilQ and evaluated the immunogenicity of this recombinant fragment. This fragment was amplified by PCR from genomic DNA isolated from N. meningitidis serogroup B and cloned into the pET-28a expression vector. PilQ406-770 was over-expressed with IPTG and then affinity-purified by Ni2+-Sepharose resin. The recombinant PilQ406-770 was reacted with rabbit anti-N. meningitidis polyclonal antibody in western-blot analysis. Mice were immunized subcutaneously with purified rPilQ[406-770] mixed with an equal volume of Freud's adjuvant and evaluated specific serum antibody responses. Our results show pilQ[406-770] cloned in pET28a vector, while the cloning of pilQ[406-770] was confirmed by colony-PCR and enzymatic digestion. SDS-PAGE analysis showed that our constructed prokaryotic expression system pET28a- pilQ406-770-BL21 efficiently produces target recombinant protein with molecular weight of 43 kDa in the form of dissoluble inclusion body. Our results confirmed that a prokaryotic expression system for PilQ406-770 protein was successfully constructed
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Staphylococcus aureus is an important cause of serious infection in both hospital and the community. Methicillin-resistant S. aureus [MRSA] is associated with high morbidity and mortality rates with rapid development of resistance. There is a need for early and reliable detection of MRSA infection to direct antibiotic therapy, and more effectively control cross-infection. In this study, resistance to methicillin was detected by a disk diffusion method, the determination of MIC, and the PCR for mecA gene. A total of 174 S.aureus strains were isolated from different clinical specimens from three teaching Hospitals. Antibiotic susceptibility was determined by disk diffusion method, MIC for oxacillin was made by the agar dilution, and mecA gene was identified by specific primers. The prevalence of MRSA by three methods ranged from 47% to 50%, and mecA positive isolates were more resistant to all of the antibiotic tested than mecA negative isolates. All S. aureus isolates were resistant to penicillin, and susceptible to vancomycin. The results of agar dilution test indicated a low-level resistance to methicillin [MIC<64mg/l]. The distribution of MRSA isolates were uniform between three hospitals, and there were not significant differences in the presence of MRSA between isolates from different clinical specimens. The PCR method was the best test for routine detection of MRSA in the present study. An additional benefit of the mecA PCR is the potential to generate a susceptibility report, 24h earlier than the time of generation of results of conventional susceptibility testing methods