Iranian Journal of Microbiology
Year:2016 | Volume:8 | Issue:4|Papers Count:9

Background and Objectives: Pseudomonas aeruginosa (PA) is one of the most important causes of nosocomial infectionsand has an intrinsic resistance to many antibiotics. Among all the resistance-nodulation-division (RND) pumps of P. aeruginosa, MexAB-OprM is the first efflux pump found to target multiple classes of antibiotics. This study was aimed to evaluatethe expression level of genes expressing MexAB-OprM in clinical isolates of P. aeruginosa.Materials and Methods: In this study, 45 P. aeruginosa strains were isolated from patients admitted to Children's MedicalCenter Hospital, an Iranian referral hospital. Disk diffusion and Minimum Inhibitory Concentration (MIC) methods wereused for determination of the patterns of resistance to antibiotics. Real-time PCR was used to investigate the expression levelof genes of MexAB-OprM efflux pump.Results: Among 45 resistant PA isolates, the frequency of genes overexpression was as follows: MexA (n=25, 55.5%), MexB(n=24, 53.3%) and OprM (n=16, 35.5%). In addition, in 28 strains (62%) overexpression was observed in one of the studiedthree genes of MexAB-OprM efflux pump.Conclusion: In our study 28 isolates (62%) had increased expression level of efflux pumps genes, MexAB-OprM. Althoughthe efflux pumps play important roles in increasing the resistance towards different antibiotics but the role of other agentsand mechanisms in evolution of resistance should not be ignored. Since the concomitant overproduction of other Mex effluxsystems might have additive effects on antibiotic resistance, the co-expressing of a multicomponent efflux pump is recommended.On the other hand, the concomitant overproduction of two Mex pumps might have additive effects on resistance toantibiotic. Therefore co-expressing of Mex efflux systems is recommended.

Background and Objectives: Lactic acid bacteria (LAB) play important roles in processing of Sayur Asin (spontaneouslyfermented mustard). Unfortunately, information about LAB in Indonesian Sayur Asin, prepared by traditional manufactureswhich is important as baseline data for maintenance of food quality and safety, is unclear. The aim of this study was to describethe diversity and distribution of culturable lactic acid bacteria in Sayur Asin of Indonesia.Materials and Methods: Four Sayur Asin samples (fermentation liquor and fermented mustard) were collected at harvestingtimes (3-7 days after fermentation) from two traditional manufactures in Tulung Agung (TA) and Kediri (KDR), East Javaprovinces, Indonesia. LAB strains were isolated by using MRS agar method supplemented with 1% CaCO3 and characterizedmorphologically. Identification of the strains was performed basedon 16S rDNA analysis and the phylogenetic tree wasdrawn to understand the phylogenetic relationship of the collected strains.Results: Different profiles were detected in total count of the plates, salinity and pH of fermenting liquor of Sayur Asinin TA and KDR provinces. A total of 172 LAB isolates were successfully isolated and identified based on their 16S rDNAsequences. Phylogenetic analysis of 27 representative LAB strains from Sayur Asin showed that these strains belonged to 5distinct species namely Lactobacilus farciminis (N=32), L. fermentum (N=4), L. namurensis (N=15), L. plantarum (N=118)and L. parafarraginis (N=1). Strains D5-S-2013 and B4-S-2013 showed a close phylogenetic relationship with L. compostiand L. paralimentarius, respectively where as the sequence had slightly lower similarity of lower than 99%, suggesting thatthey may be classified into novel species and need further investigation due to exhibition of significant differences in theirnucleotide sequences. Lactobacillus plantarum was found being dominant in all sayur asin samples.Conclusion: Lactobacilli were recognized as the major group of lactic acid bacteria in Sayur Asin including 5 known and2 novel candidate species. The distribution of LAB species was associated with the manufactures where Sayur Asin is produced.

Background and Objectives: The microbiological monitoring of the water used for haemodialysis is important especiallyfor Legionella and non-fermentative bacteria since patients with end stage renal disease (ESRD) are suffering from deterioratedfunction of immune system.Materials and Methods: A total 50 water and dialysate samples were weekly collected over a period of 10 weeksfrom 5 sites. Total and faecal coliforms were determined by utilizing the most probable number (MPN) method.For isolation of Legionella, water samples were inoculated on a BCYE medium. DNA extraction was performedand was used to amplify 16S rRNA gene of Legionella species. Airborne bacteria were sampled using a single stageAndersen air sampler.Results: Out of total 50 water samples, 24 samples had bacterial contamination. The highest rate of Legionella contaminationwas observed in the storage tank (67 cfu/ml). Legionella was not isolated from the dialysate effluent samples. The highestrate of total bacterial count was related to the dialysate effluent and the maximum total count of coliforms was related tothe reverse osmosis. The isolated bacteria were Gram-negative bacilli (mostly Pseudomonas isolates), Gram-positive cocci(mostly Micrococcus spp.) and Gram-positive bacilli (mostly Bacillus spp.). Six samples were contaminated with coliforms.No faecal coliform was isolated from the samples.Conclusion: These results indicated that dialysis machine is an important source of contaminations such as Staphylococcus, Pseudomonas and Legionella. Therefore an efficient prevention program is needed to eliminate bacterial contamination ofdialysis water system. Moreover, in haemodialysis centres, periodic surveillance programs for microbiological qualificationcan lead to a better planning for disinfection of haemodialysis water systems.

Background and Objectives: Tuberculosis (TB) is a major problem in the world. Treatment and control of TB needs detectionof the Mycobacterium tuberculosis (MT) in the proper samples. While smear doesn’t have enough sensitivity, cultureand PCR are expensive, time consuming and unavailable in many centers. Recent development of a rapid TB antigen detectiontest (PrTBK) at Pasteur Institute of Iran could give a simple way for diagnosis of TB in about two hours. In this test theantigen-antibody complex will change color when gold conjugated mouse anti- rabbit antibody detects specific MT cell wallantigen in suspected samples.Materials and Methods: We evaluated the diagnostic accuracy of PrTBK for diagnosis of pulmonary TB in comparisonwith smear, culture and PCR techniques in 56 consecutive samples (47 BAL and 13 sputum samples) obtained from patientswith clinical suspicion of active TB.Results: Twentynine patients (52%) were female and seven patients were HIV positive. PrTBK was positive in 17 culturepositive and 4 culture negative samples (100% sensitivity, 89% specificity and 92% accuracy in comparison with culturemethod). In two out of four patients with negative culture who were positive for PrTBK, PCR and anti-tuberculosis drugstrial therapy responses were in favor of tuberculosis. If we take this finding into account, the accuracy of PrTBK will rise.Conclusion: High sensitivity and accuracy of PrTBK test enable us to initiate treatment on the basis of this convenient andrapid test.

Background and Objectives: Clostridium perfringens, a Gram-positive obligate anaerobic bacterium, is able to form resistantspores which are widely distributed in the environment. C. perfringens is subdivided into five types A to E based on itsfour major alpha, beta, epsilon and iota toxins. The aim of the present study was cloning and expression of C. perfringenstype D vaccine strain epsilon toxin gene.Materials and Methods: Genomic DNA was extracted and the epsilon toxin gene was amplified using Pfu DNA polymerase.The PCR product was cloned into pJET1.2/blunt cloning vector. The recombinant vector (pJETε) was sequenced usinguniversal primers. At the next step epsilon toxin gene was subcloned into pET22b (+) expression vector and transformedinto E. coli Rosetta (DE3) host strain.Results: The recombinant protein has been expressed in E. coli Rosetta (DE3) cells after subcloning of C. perfringens etxgene (1008 bp) into the expression vector.Conclusion: We concluded that E. coli Rosetta strain was suitable for the expression of recombinant C. perfringens epsilontoxin protein from pET22ε expression vector. This recombinant cell can be used for further research on recombinant vaccinedevelopment.

Background and Objectives: Biomaterials are widely used in medical devices such as urinary catheters. One of the mainproblems associated with long term using of the urinary catheters is biofilm formation on their surfaces. Many techniqueshave been presented to reduce the biofilm formation. One of the most revolutionary techniques allowing such surface fictionalizationis plasma surface modification.Materials and Methods: In this study, a glow discharge plasma (GDP) effect on Escherichia coli biofilm formation on thesurface of urinary catheter in the pressure of 1.6 × 10-1 Torr of nitrogen, discharge voltage about 1.2 kV and current of 150mA for 20 minutes has been investigated. Crystal violet binding assay and sonication method were performed in order toevaluate the amount of biofilm formation on tested biomaterials.Results: Characterization of modified surfaces by Attenuated Total Reflectance Fourier Transform Infrared Spectrometry(ATR-FTIR) and atomic force microscopy (AFM) revealed a noticeable change in hydrophobicity and roughness of cathetersurfaces achieved by nitrogen plasma. The results of crystal violet binding assay and sonication method showed that theamount of biofilm formation on modified surface was about 86% less than the pristine sample.Conclusion: Plasma surface modification can reduce the risk of infections in patients with long-term use of urinary catheters.

Background and Objectives: Acinetobacter spp. are important causes of nosocomial infections. They possess various antibioticresistance mechanisms including extended spectrum beta lactamases (ESBLs). The aim of this study was to determineantibiotic resistance profile of Acinetobacter clinical isolates especially among ESBL-producing strains and to investigatethe antimicrobial effects of oleo-gum-resin extract and essential oil of Ferula gummosa Boiss.Materials and Methods: 120 Acinetobacter strains were isolated from various clinical samples of hospitalized patients inBaqiyatallah hospital, Tehran during 2011-2012. Antibiotic susceptibility test was performed on the isolates using disk diffusionmethod. To detect and confirm the ESBL-positive isolates, phenotypic and genotypic tests were performed. Three typesof F. gummosa oleo-gum-resin extracts and essential oils were prepared and the bioactive components of F. gummosa Boissextracts were determined by GC-Mass chromatography. F. gummosa antimicrobial activity was evaluated against standardstrain of Acinetobacter baumannii (ATCC19606) as well as Acinetobacter clinical isolates using well and disk diffusionmethods. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined bybroth microdilution method.Results: 46 isolates were resistant to all tested antibiotics. All clinical isolates were resistant to cefotaxime.12.94% of theisolates were phenotypically ESBL-producing among which 94.2% carried ESBL genes (blaPER-1, blaOXA-4 and blaCTX-M)detected by PCR. Oleo-gum-resin of F. gummosa had significant antibacterial activity and alcoholic essential oil had higherinhibitory effect on Acinetobacter strains (MIC of 18.75 mg/ml).Conclusion: Ferula gummosa extract contained components with well-known antimicrobial effects.

Background and Objectives: Accurate designation of antimicrobial susceptibility pattern of the infecting microorganismsis an important crucial factor in making appropriate therapeutic decisions. Macrolide, lincosamide and streptogramin Bantibiotics are in a family, reserved as an alternative approach in treatment of resistant Gram positive cocci. Amongst them, clindamycin has been considered as the preferred agent due to its excellent pharmacokinetic properties. The inducible resistanceto clindamycin in Gram positive staphylococci and streptococci cannot be recognized by routine broth or agar basedsusceptibility tests and D-zone testing is necessary. This study is conducted to evaluate the frequency of inducible clindamycinresistance in Gram positive cocci.Materials and Methods: Using traditional culture methods, 487 isolates of staphylococcus and β-hemolytic streptococcuswere evaluated. If they were resistant to erythromycin and sensitive to clindamycin in primary antibiotic susceptibility testingby Kirby-Bauer method, they were subjected to D-zone testing to detect possible inducible clindamycin resistance.Results: Thirty three out of 172 isolates of Staphylococcus aureus and 50 out of 277 isolates of coagulase-negativestaphylococci (CoNS) were subjected for D-zone testing. Among them 13.33 and 28.50 showed inducible clindamycinresistance, respectively. There was no significant difference in inducible clindamycin resistance regarding to methicillinsusceptibility pattern. Positive D-test was observed in 17.39 and 13.33% of Group B streptococci and Streptococcus spp., respectively.Conclusion: Considerable number of isolates showed inducible clindamycin resistance in our study which falsely would bereported susceptible if D-zone testing was not performed. Thus, performing D-Zone testing is necessary to avoid misleadingresults which may cause treatment failure.

Background and Objectives: Cholera is an endemic diarrheal disease in Iran, caused by Vibrio Cholerae. The epidemiology, transmission route, environmental determinants and antimicrobial resistant pattern of cholera have been changed duringrecent years. In this study the epidemiology and antimicrobial resistance of cholera in Iran during 2013 outbreak was investigated.Materials and Methods: A retrospective, cross-sectional study was carried out using cholera national surveillance systemcollected data in 2013. Bacterial identification and antimicrobial susceptibility testing were done on 60 Vibrio cholerae isolates, serotype Inaba.Results: During July to November 2013, 256 confirmed cholera cases were diagnosed by stool culture. Two hundred andeleven out of 256 (83%) cases were imported from Afghanistan and Pakistan. The prevalent age group was 16-30 years old, 90% were male, 98.8% affected by Inaba serotype and case fatality rate was 2.7%. The results of antimicrobial susceptibilitytesting on 60 V. cholerae, serotype Inaba showed that all isolates were resistant to nalidixic acid, tetracyclin and trimethoprim-sulfamethoxazole and intermediate resistance to erythromycin but sensitive to ciprofloxacin, cefixime and ampicillin.Conclusion: Migrants from neighboring countries played a key role in cholera outbreak in Iran during 2013. The resultsof antimicrobial susceptibility testing on 60 V. cholerae, serotype Inaba showed an increasing resistance rate in comparisonwith previous years.