Detection of rifampin resistance in Mycobacterium tuberculosis strains by RNA/RNA mismatch method [Mycobacterium tuberculosis i·zolatlarinda ri·fampi·n di·renci·ni·n RNA/RNA uyumsuz bi·rleşme (mismatch) yöntemi· i·le saptanmasi]

PubMed ID: 12838673Rifampin is one of the most potent antituberculosis drugs and therefore, rifampin resistance leads to high clinical relapse rates. Detection of rifampin resistance could be an indication of multidrug resistance. In the recent years several molecular methods have been developed to evaluate the mutations in rpoB gene for the detection of rifampin resistance. The aim of the present study was to evaluate the performance of the RNA/RNA Mismatch Assay for detection of the mutations in the rpoB gene in 20 M.tuberculosis isolates which were determined as resistant to rifampin by agar proportion method. While RNA/RNA Mismatch Assay detected the mutations in the rpoB region in 16 of 20 (80%) M.tuberculosis isolates, the remaining four isolates yielded no band pattern indicating resistance. However, there may be situations where interpretation of the results is difficult in RNA/RNA Mismatch Assay which is already cheaper than DNA sequencing and other molecular methods. In conclusion, if the RNA/RNA Mismatch Assay can be optimized, it can be used for the rapid detection of rifampin resistance

Genotyping of rifampin-resistant Mycobacterium tuberculosis isolates by mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) analysis [Ri·fampi·si·ne di·rençli· Mycobacterium tuberculosis i·zolatlarinin MIRU-VNTR (mycobacterial interspersed repetitive unit-variable number tandem repeat) yöntemi·yle genoti·plendi· rmesi·]

PubMed ID: 17933249Molecular typing methods have greatly enhanced our understanding on epidemiology of tuberculosis and allowed us to identify outbreaks and intertransmission within populations. Recently, a set of 12 variable-number tandem repeat (VNTR), designated mycobacterial interspersed repetitive units (MIRU), has been described as being useful for the typing of M.tuberculosis. In this study, 26 rifampin (RIF) resistant M.tuberculosis isolates with known IS6110-RFLP patterns obtained from 26 different patients in Aegean Region were typed by MIRU-VNTR and the data were compared with IS6110-RFLP results. The results showed that in most isolates the clustering on the basis of IS6110 RFLP typing and that on the basis of MIRU-VNTR typing were in agreement. It was also determined that the loci including MIRU 16, MIRU 40, MIRU 26, MIRU 10, MIRU 04 and MIRU 31, respectively, have the highest allelic diversities and discriminatory power. In conclusion, since the discrimination level of conventional MIRU-VNTR including 12 loci might be variable, by the use of additional loci which present high degree of allelic differentiation, this method would be reliable for the epidemiologic studies

Investigation of the virulence genes in methicillin-resistant Staphylococcus aureus strains isolated from biomaterial surfaces [Biyomalzeme yüzeylerinden izole edilen metisiline dirençli Staphylococcus aureus suşlar?nda virülar?s genlerinin araşt?r?lmas?]

PubMed ID: 18444560Staphylococci are the most important agents of nosocomial infections originating from biomaterials. The aim of this study was to investigate the presence of virulence genes and their phenotypic expressions in 11 methicillin-resistant Staphylococcus aureus strains isolated from the surfaces of clinically used biomaterials of 48 thorasic intensive-care unit patients. By the use of specific primers, the presence of genes encoding the attachment and biofilm production (icaA, icaC, bap), methicillin resistance (mecA), enterotoxins A-E (sea, seb, sec, sed, see), toxic shock syndrome toxin (tst), exfoliative toxins A and B (eta and etb), alpha- and beta-hemolysins (hla and hlb), staphylococcal exotoxin-like protein-1 (set1), proteases (sspA, sspB, aur, serine proteaz gene), lipase (geh) and the regulatory genes (sarA and agrCA) were investigated by polymerase chain reaction (PCR). The phenotypic properties of the isolates such as biofilm formation, antibiotic susceptibility, extracellular protease and lipase production were also evaluated. None of the isolates were found to be biofilm and/or slime producers, however, all strains were found to have icaA gene which is responsible for biofilm formation. Nevertheless the presence of icaC and bap genes that are also responsible for biofilm formation were not detected. All the strains have had mecA gene and were resistant to oxacillin, penicilin G and gentamicin, while 10 were also resistant to erythromycin and nine were also resistant to ofloxacin. The isolates were susceptible to vancomycin, teicoplanin and co-trimoxazole. Screening of toxin and regulatory genes revealed that all the strains harboured sea, set1, hla, hlb and sarA genes. The phenotypic tests for the determination of extracellular protease production revealed that all the strains formed very weak zones on skim milk and milk agar plates, and yielded negative results on casein agar plates. Furthermore, all strains were found to harbour sspA, sspB, aur and serine protease genes. Tween 20, Tween 80 and tributyrin containing media were used to detect lipase production and all strains gave late-positive results (on the third day of incubation), although they all lacked for lipase gene (geh). As a result, S.aureus strains isolated from biomaterial surfaces yielded positivity for some of the tested virulence genes, of which some of them have not been expressed phenotypically. Although there were some limitations in the study, it could be concluded that the presence of these virulence genes in S.aureus strains might be considered as potential threats especially in intensive care unit patients

In vitro activity of linezolid against multidrug-resistant mycobacterium tuberculosis isolates by BACTEC MGIT 960 method

• Objective: The aim of our study was to determine susceptibility of linezolid to multi-resistant Mycobacterium tuberculosis isolates. • Material and Method: Seventy two multi drug-resistant Mycobacterium tuberculosis isolates were studied under the activity of 1 µg/ml of linezolid by the BACTEC MGIT 960 susceptibility method. • Results: Seventy two multi drug-resistant Mycobacterium tuberculosis species were determined to be susceptible to linezolid. • Conclusion: Linezolid was found to be effective to multi-resistant Mycobacterium tuberculosis in vitro

Evaluation of the genotype MTBDRplus assay for the diagnosis of tuberculosis and rapid detection of rifampin and isoniazid resistance in clinical specimens [Klinik örneklerden tüberküloz tani{dotless}si{dotless} ve hi{dotless}zli{dotless} rifampin ve izoniyazid direnci saptanmasi{dotless} için geno type MTBDRplus testinin degerlendirilmesi]

Aim: To determine the performance of the Genotype MTBDRplus assay for diagnosis of tuberculosis and rapid detection of rifampin (RIF) and isoniazid (INH) resistance in clinical specimens. Materials and methods: A total of 90 clinical specimens of 57 patients (69 sputum samples, 11 bronchoscopic aspirates, 5 bronchoalveolar lavage, 4 deep tracheal aspirate, 1 lymph aspirate) sent to the Ege University Medical Faculty, Department of Medical Microbiology, Mycobacteriology Laboratory between December 2007 and 2009 during the clinical routine were included in the study. Results: Overall 80 valid results were obtained for 90 clinical specimens (88.9%) with MTBDRplus. While 74 of 82 (90.2%) smear positive specimens gave interpretable results by MTDRplus, 2 of 8 smear negative specimens gave invalid results. The overall rates of concordance between the results of the MTBDRplus assay and those of the drug susceptibility testing for the assessment of RIF and INH resistance were 97.5% (78/80) and 98.8% (79/80), respectively. Conclusion: Although the MTBDRplus assay could be a useful tool for rapid identification of RIF- and INH-resistant Mycobacterium tuberculosis in both clinical samples and strains, the test results must always be confi rmed by culture and drug susceptibility testing. © TÜBITAK

Development of a database for tracking HIV positive/AIDS patients [HIV pozi·ti·f/aids hastalarinin tani ve i·zlemi· i·çi·n geli·şti·ri·len veri· tabani ortami]

PubMed ID: 17427558The collection of reliable data is the first step to assess the status of HIV/AIDS in a community. HIV recording systems are necessary for organizing and analyzing the patients' data. The aim of the study was to develop a database to be used to track HIV positive/AIDS patients. The database includes general demographic fields as well as specific fields such as health history, laboratory and other clinical history, current and past drug regimens (both antiretroviral and non-antiretroviral drugs). It is also possible to organize and maintain a patient database according to specific diseases, laboratory tests and/or medication treatments

Investigation of the surface properties of Staphylococcus epidermidis strains isolated from biomaterials [Bi·yomalzemelerden i·zole edi·len Staphylococcus epidermidis suşlarinin yüzey özelli·kleri·ni·n beli·rlenmesi·]

PubMed ID: 20455404The surface properties of bacteria play an important role on adhesion to the biomaterial surface. In this study, the surface properties of Staphylococcus epidermidis strains isolated from clinically used polymeric biomaterial surfaces were investigated on the basis of zeta potential, hydrophobicity and surface topography. A total of 10 S.epidermidis strains isolated from intravenous catheters (n= 5), endotracheal tubes (n= 3) and central venous catheters (n= 2) which were used in the patients of pulmonary Intensive Care Unit, Ege University Medical Faculty Hospital, were included to the study. Seven of those isolates were biofilm producers, inhabiting biofilm genes, 2 were non-biofilm producers, however, inhabiting biofilm genes, and 1 was non-biofilm producer, inhabiting no biofilm genes. Zeta potential analysis have been performed in 3 different buffers (phosphate-buffered saline, 1 mM potassium chloride and 1 mM potassium phosphate buffer) and at different pH values (pH 4.1-8.2), in order to simulate in vivo environment of the biomaterials. Hydrophobicities of the strains were examined by bacterial adhesion to hydrocarbon (BATH) test and the surface topography of biofilms and slime layers were visualized by atomic force microscopy (AFM) and scanning electron microscopy (SEM) methods. It was found that all strains have negative zeta potential values (surface charge) in all buffers and pH values. In hydrophobicity analysis, the highest value (86%) was determined for non-biofilm forming S.epidermidis strain YT-169b (endotracheal tube isolate) and the lowest hydrophobicity (2.5%) was determined for biofilm forming S.epidermidis strain YT-212 (central venous catheter isolate). Biofilm and slime layers of the strains were imaginated by AFM and SEM analysis in µm scale. SEM analysis showed that bacteria highly adhered to rough surfaces on biomaterial surfaces and the produced slime layers covered the surface of bacteria. In conclusion, elucidating the surface properties of opportunistic pathogens in different physiologic buffers will give important clues for the production of non-adhesive materials and antibacterial surfaces for those bacteria. It was also estimated that designing the surface of the biomaterial to have negative surface charge in the body and to be as smooth as possible will hamper biofilm formation

Determination of extended spectrum beta-lactamase frequency of Klebsiella pneumoniae strains isolated from urinary tract infections and typing with isoelectric focusing method

Extended spectrum beta-lactamase (ESBL) presence was investigated in Klebsiella pneumoniae strains which were the ethiological agents of urinary tract infections and typed with isoelectric focusing (IEF) method. The antibiotic sensitivities of 52 K. pneumoniae strains were determined by disc diffusion method and ESBL were found in 12 strains (23 %) by double disc synergy test. Minimum inhibitory concentration (MIC) values of ESBL (+) K. pneumoniae strains for ceftazidime (CAZ) and cefotaxime (CTX) were investigated by E-test and isoelectric points of ESBL enzymes were determined by polyacrilamide gel electrophoresis. When the strains' sensitivity patterns were investigated for 3rd generation cephalosporins and aztreonam, 6 strains had ESBL premise sign with at least one antibiotic. Band pattern which focused around pl 7.6 was determined in 11 of 12 strains by IEF method. MIC values were found 16 µg/mL and over for CAZ and 12 µg/mL and over for CTX in 5 of these strains. When pI's and antibiotic susceptibility patterns were analysed, 5 of 12 isolates had a band and susceptibility patterns suggesting SHV-2