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Öğe Evaluation of Rapid Genotype Assay for the Identification of Gram-Positive Cocci from Blood Cultures and Detection of mecA and van Genes(Ankara Microbiology Soc, 2011) Gulhan, Baris; Atmaca, Selahattin; Ozekinci, Tuncer; Suay, AdnanRapid and accurate identification of bacterial pathogens grown in blood cultures of patients with sepsis is crucial for prompt initiation of appropriate therapy in order to decrease related morbidity and mortality rates. Although current automated blood culture systems led to a significant improvement in bacterial detection time, more rapid identification systems are still needed to optimise the establishment of treatment. Novel genotype technology which is developed for the rapid diagnosis of sepsis, is a molecular genetic assay based on DNA multiplex amplification with biotinylated primers followed by hybridization to membrane bound probes. The aim of this study was to evaluate the performance of Genotype (R) BC gram-positive test for the identification of gram-positive cocci grown in blood cultures and rapid detection of mecA and van genes. This test uses DNA.STRIP (R) technology which includes a panel of probes for identification of 17 gram-positive bacterial species and is able to determinate the methicillin and vancomycin resistance mediating genes (mecA and vanA, vanB, vanC1, vanC2/C3) simultaneously, in a single test run. A total of 55 positive blood cultures from BACTEC (TM) Plus/F (Becton Dickinson, USA) aerobic and pediatric blood culture vials were included in the study. The isolates which exhibit gram-positive coccus morphology by Gram staining were identified by Genotype (R) BC gram-positive test (Hain Life Science, Germany). All of the samples were also identified with the use of Phoenix PMIC/ID Panel (Becton Dickinson, USA) and antibiotic susceptibilities were determined. Of the 55 blood culture isolates, 17 were identified as Staphylococcus epidermidis [all were methicillin-resistant (MR)], 9 were S.aureus (one was MR), 18 were S.hominis (10 were MR), 4 were E.faecalis, 3 were E. faecium (one was vanconycin-resistant), 2 were S.saprophyticus (one was MR), 1 was S.warneri and 1 was S.haemolyticus, by Phoenix automated system. Genotype (R) BC gram-positive test results revealed consistency with Phoenix system regarding bacterial identification in 46 (83.6%) of the samples. The two bacteria identified as S.saprophyticus by the Phoenix system could not be identified by the Genotype (R) BC test since this species were not included in the identification panel of the system, however, mecA gene were detected in these two samples by Genotype (R) BC test. Genotype (R) BC test detected mecA gene in five samples which were not detected as methicillin resistant by the Phoenix system. Besides polymicrobial growth was determined in five samples by Genotype (R) BC test, but not by the automated system. One E.faecium isolate with vanA gene was correctly identified by Genotype (R) BC test. In conclusion, Genotype (R) BC gram-positive test is a fast and reliable test for the identification of the most important gram-positive pathogens and mecA and van genes directly from positive blood culture bottles. This test was also found superior than the automated Phoenix system regarding the detection of polymicrobial growth. These data indicated that, routine use of DNA strip technology-based assay would be useful for clinical diagnosis in patients with sepsis.Öğe Investigation of Antibiotic Resistance Patterns and Reduced Vancomycin Susceptibilities of Methicillin-Resistant Staphylococcus aureus Isolates: A Multi-Center Study(Ankara Microbiology Soc, 2015) Cikman, Aytekin; Aydin, Merve; Gulhan, Baris; Parlak, Mehmet; Gultepe, Bilge; Kalayci, Yildiz; Bilmen, Fulya BayindirThe aims of this study were to determine the minimum inhibitory concentration (MIC) values of vancomycin, teicoplanin, daptomycin, quinupristin/dalfopristin, linezolid, tigecycline, chloramphenicol, rifampicin, ofloxacin and tetracycline and to investigate the reduced vancomycin susceptibility among methicillin-resistant Staphylococcus aureus (MRSA) strains isolated in hospitals located in different geographical regions of Turkey. A total of 100 MRSA strains isolated from patients (of which 50% were from intensive care units) hospitalized in seven centers in Turkey [Istanbul (n= 15), Ankara (n= 15), Izmir (n= 15), Adana (n= 15), Diyarbakir (n=15), Erzincan (n= 15), Van (n= 10)], between August 201 3 August 2014, were included in the study. Fourty-three strains were isolated from blood, whereas 21 were from lower respiratory tract, 17 from wounds, eight from catheters, six from urine, four from nasal swab and one from cerebrospinal fluid samples. Methicillin resistance of the isolates was determined by using cefoxitin (30 mu g) disk with standard disk diffusion method, while the MIC values of other antibiotics were determined with E-test in accordance with the recommendations of Clinical and Laboratory Standards Institute (CLSI). MIC results obtained for quinupristin-dalfopristin (Q/D) were evaluated according to the CLSI criteria used for methicillin-susceptible S.aureus and for tigecycline according to the criteria recommended by the Food and Drug Administration for MRSA. Primarily, agar screening method (ASM) was used for determination of vancomycin-intermediate S.aureus (VISA) and heterogeneous VISA (hVISA) strains. Brain heart infusion agar containing 6 mu g/ml vanconnycin was used in ASM, and the strains with suspicion of VISA/hVISA were screened by standard E-test and macro E-test methods. All MRSA strains were susceptible to vancomycin, teicoplanin, daptomycin, Q/D and linezolid by E-test method; and their rates of susceptibility for tigecycline, chloramphenicol, rifampicin, ofloxacin and tetracycline were detected as 89%, 97%, 40%, 39% and 32%, respectively. MIC50/MIC90 values were 1.5/2 mu g/ml for vancomycin, 2/4 mu g/ml for teicoplanin, 0.19/0.38 mu g/ml for daptomycin, 0.19/0.38 mu g/ml for Q/D, 0.75/1 mu g/ml for linezolid, 0.19/0.75 mu g/ml for tigecycline, 3/6 mu g/ml for chloramphenicol, 32/32 mu g/ml for rifampicin, 32/32 mu g/ml for ofloxacin and 32/64 mu g/ml for tetracycline, respectively. For the evaluation of reduced vancomycin susceptibility, 2% (2/100) of MRSA strains were defined as VISA and 5% (5/100) as hVISA with ASM. One of those seven isolates identified as VISA/hVISA with ASM was evaluated as suspected hVISA by using both standard E-test and macro E-test methods. In conclusion, no MRSA resistant strain to vancomycin, teicoplanin, daptomycin, Q/D and linezolid was determined in our study. However tigecycline resistance (11%) was found higher than expected. As the glycopeptide resistance is increasing in the world and because of the intense use of these drugs in Turkey, the rates of vancomycin resistance among MRSA strains should be investigated periodically.