ABSTRACT
Pseudomonas aeruginosa isolates have acquired resistance to antibiotics such as novel beta-lactams. The aim of this study was to investigate the bla[PER-1], bla[VEB-1], and bla[PSE-1] genes among isolates of P. aeruginosa among intensive care unit [ICU] patients. Sixty-five isolates were collected. The antibiotic susceptibility testing and combined disk tests were performed to detect the isolates producing extended spectrum beta-lactamases [ESBLs] among ceftazidime-resistant isolates. Polymerase chain reaction [PCR] amplification of blaPER-1, blaVEB-1, and blaPSE-1 genes was conducted. Ten [15.3%] isolates were ESBL-positive, of which 40% [n = 4] belonged to males and 60% [n = 6] were collected from females. Moreover, two and one isolates harbored blaPER-1 and blaVEB-1 genes, respectively
ABSTRACT
The aim of this study was to analyze antimicrobial resistance patterns and their encoding genes and genotypic diversity of Acinetobacter baumannii isolated from burn patients in Tehran, Iran. The presence of extended-spectrum beta-lactamase- and bla[OXA]-encoding genes among 37 multidrug resistant [MDR] A. baumannii strains isolated from patients hospitalized in a teaching hospital in Tehran was evaluated. Susceptibility to 7 antibiotics was tested by disk agar diffusion and to polymyxin B and colistin was tested by E-test, according to CLSI guidelines. All isolates were then analyzed by PCR for the presence of bla[IMP],bla[VIM], bla[SIM] bla[OXA-23], bla[OXA-24], and bla[OXA-58]-like carbapenemase genes, and bla[OXA-51] like, bla[TEM],bla[SHV], bla[PER], bla[VEB], and bla[GIM] genes. Genotyping of A. baumannii strains was performed by repetitive sequence-based [REP]-PCR and cluster analysis of REP-PCR profiles. A. baumannii isolates were assigned to international clones by multiplex PCR sequence group analysis. Twenty-five A. baumannii isolates were classified as MDR, and 12 were classified as extensively drug resistant. All isolates were susceptible to colistin and polymyxin B. Eighty-one percent of the isolates was resistant to imipenem or meropenem and harbored at least one or both of the bla[OXA-23]-like or bla[OXA-24]-like carbapenemase genes. Co-existence of different resistance genes was found among carbapenem-resistant isolates. Multiplex PCR sequence group analysis most commonly assigned A. baumannii isolates to international clones I [18/37; 48.6%] and II [18/37; 48.6%]. An alarming increase in resistance to carbapenems and the spread of bla[OXA-23]-like and/or bla[OXA-24]-like carbapenemase genes was observed among A. baumannii strains belonging to clonal lineages I and II, isolated from burn patients in Tehran
ABSTRACT
OBJECTIVE@#To assess the quality of expressed MSP-2 and also to confirm the immune response against different domains of these proteins.@*METHODS@#Mice were immunized with a schizont extract to stimulate the immune system to make antibodies against different antigens of the late stage parasite including production of antibodies against different domains of Plasmodium falciparum (P. falciparum) MSP-2. B lymphocytes of immunized mice were extracted from the spleen and the fusion was performed using NS-1 myeloma cells and the hybridoma cells were assayed by ELISA either with a schizont extract or different domains of MSP-2 and/or by IFAT with whole schizont preparation. Fusion of NS-1 and spleen cells was performed. The positive hybrids were cloned and ELISA was applied against different dilutions. The positive clones were transferred to a small tissue culture flask and after developing they were assayed against schizont extract and the different MSP-2 domains. The positive clones were expanded to large (75 cm(2)) flask and cultured under the same conditions, checking them using both ELISA and IFAT and the positive cells were frozen as soon as possible.@*RESULTS@#A total number of 7 fusions including 26 plates (2 496 wells) were performed, of which 1 336 hybrids were produced and the overall efficiency (1 336/2496 × 100) was about 53%. ELISA was performed to detect the positive hybrids against crude schizont extract by which the highest frequency to crude schizont extract was found for the supernatant of the hybrids produced in fusion number 3 (66 out of 315 hybrids). The supernatant of both B5 and F1 hybridoma cells were more positive against domain 2 of the MSP-2 recombinant protein in Western blotting test. Western blotting results also showed that different domains of the MSP-2 recombinant protein and also the MSP-2 of the P. falciparum parasite were recognized by some of the positive clones and also immune sera.@*CONCLUSIONS@#Bringing together all the results of this study it has been confirmed that some clones have recognized both schizont extract and different domains of the MSP-2 recombinant protein and therefore confirming the quality of the MSP-2 domains.
Subject(s)
Animals , Female , Humans , Mice , Antibodies, Monoclonal , Allergy and Immunology , Antibodies, Protozoan , Allergy and Immunology , Antigens, Protozoan , Chemistry , Allergy and Immunology , Immunization , Malaria , Allergy and Immunology , Parasitology , Plasmodium falciparum , Chemistry , Allergy and Immunology , Protein Structure, Tertiary , Protozoan Proteins , Chemistry , Allergy and Immunology , Schizonts , Allergy and ImmunologyABSTRACT
Background: Multiple-drug resistant Acinetobacter have widely spread in the last decades imposing a serious nosocomial source of infection. Nevertheless, little knowledge was gaimed on tracing the development of antibiotic resistance in Acinetobacter species.Objectives: Explore Acinetobacter spp. via antimicrobial susceptibility, plasmid profiles, and random amplified polymorphism DNA polymerase chain reaction (RAPD-PCR) typing.Methods: One hundred twelve Acinetobacter isolates (including 66 A. baumannii and 46 non-Acinetobacter baumannii strains) were obtained from three university hospitals. The source of infection of these isolates included blood, urine, wound, and respiratory tract. Their susceptibilities to 17 antibiotics were tested and then allAcinetobacter isolates were typed by plasmid analysis and RAPD-PCR method.Results: A. baumannii isolates revealed nine different patterns of antibiotic resistance. Of those, non-A. baumannii, were associated with plasmid and RAPD-PCR typings (p 0.05).Conclusion: There is a wide spread of multi-drug resistant Acinetobacter spp., particularly A. baumannii, in the Middle East region that can be traced efficiently by plasmid and genotyping typing of Acinetobacter. More care should be taken for tracing the development of antimicrobial resistance of Acinetobacter using precisemolecular typing techniques.Keywords: Acinetobacter baumannii, antimicrobial susceptibility, molecular typing, plasmid profiles RAPD-PCR
ABSTRACT
Pyrococcus woesei is a hyperthermophilic archaea and produces a heat stable polymerase [Pwo polymerase] that has proofreading activity. In this study, this microorganism was cultured, its DNA was extracted and the pwo gene polymerase was cloned, expressed and purified. The DNA sequence of the cloned gene was verified by sequencing. The pwo polymerase gene consists of 2,328 bps [775 amino acids with about 90 kD molecular weight]. Cloning was done by GATEWAY Cloning System and for purification of recombinant protein; His6x-Tag was added to the C-terminus of the recombinant protein. We could purify Pwo polymerase enzyme by Ni-NTA resin. PCR assay showed that Pwo polymerase activity is comparable to a commercial Pfu polymerase activity
ABSTRACT
Infections caused by Shigella re a major cause of diarrheal disease in the developing and developed countries. The present study was conducted to apply and evaluate arbitrarily primed PCR [AP-PCR] for investigation of genetic relatedness among the strains of Shigella sonnei isolated from cases of acute diarrhea in Tehran. Totally, 60 S. sonnei strains isolated from children hospitalized due to enteritis at five hospitals in Tehran during 2003 and two sporadic isolates recovered in 1984 were investigated. Molecular typing was performed by AP-PCR. Depending on the number and size of amplified DNA bands, the strains were clustered into AP-PCR profiles. All strains of S. sonnei were typeable with this approach. AP-PCR generated nine indistinguishable bands ranged from 0.35 to 2.5 kbp in all studied strains. Only a single AP-PCR pattern was observed among the S. sonnei strains recovered in 2003. Two sporadic isolates recovered in 1984 showed different AP-PCR patterns compared to recent clinical isolates. Results suggest that a very homogeneous AP-PCR cluster types might be responsible for shigellosis caused by S. sonnei in Tehran in 2003. Further molecular analysis conducted on a larger selection of isolates could confirm our findings