ABSTRACT
The antimicrobial effects of the extracts of different kinds of plants have been demonstrated in several studies. However, no study has been conducted so far on the synergistic effects of two herbal extracts on their germicidal effects. In this study, in addition to antibacterial effects of the aqueous, methanol or ethanol extracts of Tribulus terrestris and bitter almond on some bacteria, the synergistic effects of the extracts of these two plants were also evaluated. In this experimental study, water, methanol and ethanol extracts of seeds were screened against some bacterial strains. Seeds were extracted by percolation method. Aliquots of the extracts at variable concentrations were then incubated with different bacterial strains, and the antimicrobial activities of the extracts from seeds were determined by MIC. Three antibiotics were used as reference compounds for antibacterial activities. Seeds extract inhibited significantly the growth of the tested bacterial strains. The greatest synergistic effect of T. terrestris and bitter almond extracts is detected in methanol and aqueous extracts. Among the bacterial strains tested, Staphylococcus aureus was most susceptibility. The results showed the highest antibacterial effect in the combination of methanol extract of T. terrestris and the aqueous extract of the bitter almond
ABSTRACT
Toxoplasma gondii is an obligatory intracellular protozoan. Considering to high prevalence of this disease the best way to reduce the raised loses is prevention of human and animal infection, rapid diagnosis, differentiation between acute and chronic disease. Rhoptry protein 1 of Toxoplasma gondii is an excretory-secretory antigen that exists in the most stages of life cycle. According to specifications of excretory-secretory antigen that seems this antigen is a suitable candidate to produce recombinant vaccine and diagnostic kit. The main object of the present work was cloning rhoptry protein 1 [ROP1] gene of Toxoplasma gondii [RH] in a cloning vector for further production of rhoptry proteins. Genomic DNA was extracted by phenol-chloroform method. The ROP1 fragment was amplified by PCR. This product was approved by sequencing and was cloned between the EcoR1 and Sal1 sites of the pTZ57R/T vector. Then transformed into Escherichia coli DH5 alpha strain and screened by IPTG and X-Gal. After isolating of this gene from pTZ57R/T, it was subcloned into pET32a plasmid. The plasmid was purified and approved by electrophoresis, enzyme restriction and PCR. After isolating of this gene from pTZ57R/T, it was subcloned into pET32a plasmid. After enzyme restriction and electrophoresis a fragment about 1183bp was separated from pET32a. Recombinant plasmid of ROP1 gene was constructed and ready for future study. That seems the antigen is a suitable candidate to produce recombinant vaccine and diagnostic kit
ABSTRACT
The aim of this study was compared the efficacy of the designed primers and already published primers for detection of the exoA, oprL and algD genes by PCR assay for finding a rapid, accurate and highly sensitive and specific procedure to detect the Pseudomonas aeruginosa in the serious and fatal infections such as cystic fibrosis disease, burned individual. A total of 150 clinical specimens were inoculated in to routine and selective culture media for Pseudomonas aeruginosa isolation. Specific primers were designed by bioinformatics analysis for detection of the virulence genes exoA, oprL and algD. The available sequences of these three genes were obtained from NCBI and multiple alignments were performed to find the conserved sequences of each gene for primer designing. Both multiple alignment and primer designing steps were carried out by AlleleID software, version 7.0. Microbiological culture methods were showed that 70 Pseudomonas aeruginosa strains isolated from the 150 clinical specimens. PCR assay performed by using the designed primers shown 68, 70 and 69 positive results from 70 direct specimens for exoA, oprL and algD respectively that shown 97.2%, 100% and 98.6% sensitivity for above genes. PCR assay performed by using the already published primers shown 57, 49 and 28 positive results for above genes respectively that shown 81.5%, 70% and 40% sensitivity. The present study shows that by using the high specific primers for detection of the mentioned genes of the Pseudomonas aeruginosa. The conventional PCR assay detected the early colonization of the organism in Cystic Fibrosis patients with more sensitivity and specificity before several mounts to obtain positive culture. Indeed PCR assay with high specific primers has more sensitivity and specificity as a rapid and accurate diagnosis of the organism in other deadly infections by using the direct clinical specimens.
Subject(s)
Cystic Fibrosis/microbiology , Pseudomonas Infections/diagnosis , Bacteriological Techniques , Pseudomonas aeruginosa/genetics , DNA Primers/genetics , Polymerase Chain Reaction/methods , Sensitivity and Specificity , Computational Biology , DNA, BacterialABSTRACT
Immune response to recombinant L7/L12, in addition to protective role, may show its importance in detection Brucellosis tests. The aim of this study was to examine antigenicity of recombinant L7/L12 from Brucella abortus by Brucellosis human sera. We amplified L7/L12 gene by polymerase chain reaction [PCR] method and sub- cloned to prokaryotic expression vector pET28a. Escherichia coli BL21-DE3-plySs was transformed with pET28a-L7/L12 and gene expression was induced by IPTG. Recombinant L7/L12 was further analyzed by Western Blot. Sera reactivity of five infected individual were further analyzed against the recombinant L7/L12 protein. The sequencing result was confirmed by Sanger method and it was the same as L7/L12 gene. Escherichia coli BL21 [DE3] pLysS was transformed with pET28a-L7/L12 and gene expression was induced by IPTG. The expressed protein was purified by affinity chromatography by Ni-NTA resin. The data also indicated that L7/L12 protein from Brucella abortus recognized by patient sera. Our data showed that recombinant L7/L12 protein can be produced by pET28a in Escherichia coli. This protein was recognized by sera in infected human as an antigen. Therefore, recombinant L7/L12 has same epitopes with natural form of this antigen. Recombinant L7/L12 also seems to be a promising antigen for protection and serologic diagnosis of Human brucellosis
ABSTRACT
Extended-spectrum beta-lactamase [ESBLs] producing bacteria are issued as a serious problem considering their ability to hydrolyze most of beta-lactam antibiotics. The outbreak of infections derived by ESBL-producing enterobacteriaceae is increasing throughout the world. Therefore, this study aims to determine a pattern of antibiotic resistance and investigate the extended-spectrum beta-lactamases production of enterobacteriaceae isolates separated from clinical specimens. In this study, 170 various strains of enterobacteriaceae isolated from clinical specimens in teaching hospitals of Arak cultured and identified applying standard methods during one year [2010-2011]. The antibiotic resistance of isolates was investigated through disk Agar diffusion according to CLSI criteria. The resistant isolates against ceftazidime and cefotaxime antibiotics were studied through the combined disk test for the final confirmation of ESBL-production. The minimum inhibitory concentration was determined through micro broth dilution. In this study, the resistance rate of various strains of enterobacteriaceae against amoxiclav, cefotaxime, ceftriaxone, ceftazidime, cefoxitin, cefotetan, meropenem and imipenem were respectively, 91.1%, 70%, 68.8%, 62.9%, 28.2%, 11.1%, 11.1% and 1.7%. Among 125 resistant enterobacteriaceae isolates against ceftazidime or cefotaxime, 108 isolates [86.4%] had ESBL-positive phenotype and 17 isolates [13.6%] had ESBLnegative phenotype. The MICs of the resistant isolates were indicated within a range of 16 to 512 micro g/ml for ceftazidime and 64 to 512 micro g/ml for cefotaxime. According to the results of this study, imipenem is the most effective antimicrobial antibiotic. On the other hand, the present study indicates that the bacteria within the family of ESBL producing enterobacteriaceae are highly prevalent among the patients. The increase in rate of such cases is often resulted by irrational antibiotic prescription. Application of new antimicrobials, limitation of the use of antimicrobial factors and increasing the utilization of infection control tools are all required in order to solve this problem.
ABSTRACT
Extensive use of quinolones has been associated with raising level of resistance, in the current, we focused on assessing the prevalence of Escherichia coli resistance to quinolones and frequency of qnrA, qnrB and qnrS in non ESBLs [extended spectrum beta-lactamases] and ESBLs producing E. coli with blaSHV and blaTEM. One hundred and fifty E. coli isolates were identified during Mar. 2007 to Apr. 2008 in Milad [Tehran] hospital. They were tested for ESBLs production as well as quinolone resistance. PCR was performed for detection of blaSHV and blaTEM as well as qnrA, B and S. Of 150 isolates, forty-two [28%] ESBLs producing and one hundred and eight [72%] non-ESBLs producing E. coli were identified. 64.2% [n= 24] of E. coli producing ESBLs and 4.62% [n= 5] of non-ESBLs E. coli were resistance to ciprofloxacin. 95.2% [n= 40] and 26.1% [n= 11] of the isolates harbored blaTEM and blaSHV, respectively. 23.8% [n= 10] had both genes. 37.5% [n= 9] and 20.8% [n= 4] of ESBLs producing E. coli were positive for qnrA and qnrB respectively. qnrS was not identified in any isolate. Our study showed high frequency of ESBLs producing E. coli as well as quinolone resistance genes [qnrA, qnrB] in Milad hospital
ABSTRACT
Brucellosis, caused by Brucella spp., is an important zoonotic disease that causes abortion and infertility in cattle and undulant fever in humans. Various studies have examined cell-free native and recombinant proteins as candidate protective antigens in animal models. Among Brucella immunogenes, antigen based on ribosomal preparation has been widely investigated. In this study, the immunogenic ribosomal protein L7/L12 gene from Brucella abortus, S19, was amplified by PCR and sub-cloned to prokaryotic expression vector pET28a. Escherichia coli BL21 [DE3] pLysS was transformed with pET28a-L7/L12 and gene expression was induced by IPTG. The expressed protein was purified by affinity chromatography with Ni-NTA resin. The concentration of purified recombinant protein calculated to 8 mg/L of initial culture. The integrity of product was confirmed by Western-blot analysis using a standard rabbit anti Brucella abortus ribosomal protein L7/L12 antibody. Sera reactivity of five infected individual were further analyzed against the recombinant ribosomal L7/L12 protein. Data indicated that recombinant ribosomal L7/L12 protein from Brucella abortus was recognized by patient sera