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    Comparison of new and classical point mutations associated with clarithromycin resistance in Helicobacter pylori strains isolated from dyspeptic patients and their effects on phenotypic clarithromycin resistance
    (Microbiology Soc, 2019) Kocazeybek, Bekir; Sakli, Merve Kutlu; Yuksel, Pelin; Demirci, Mehmet; Caliskan, Reyhan; Sarp, Tevhide Ziver; Saribas, Suat
    Purpose. We aimed to investigate the presence of three recently identified point mutations (A2115G, G2141A and A2144T) of the 23 S rRNA gene and compare them with the three most frequently encountered point mutations (A2142G, A2142C and A2143G) in Helicobacter pylori strains in Turkey. Methodology. A total of 63 patients (mean 47.08 +/- 12.27) were included. The E-test method (for clarithromycin) was used for the clarithromycin antimicrobial susceptibility test of isolated H. pylori strains. Real-time PCR was used to detect the point mutations. Results. A total of 24 out of 63 H. pylori strains (38.1%) were detected as clarithromycin resistant (>0.5 mg l(-1)). The new A2115G (n:6, 25%), A2144T (n: 7, 29.1%) and G2141A, 8 (n: 8, 33.3%) mutations and the classical A2142G (n: 8, 33.3%) and A2143G (n: 11, 45.8%) point mutations were detected in the 24 clarithromycin-resistant strains. The A2144T point mutation had the highest median MIC value (3 mg l(-1)) amongst the new mutations, but the classical mutations (A2142G and A2143G) had the highest median MIC values (256 mg l(-1)) overall. The presence of the A2115G (OR: 31.66), A2144T (OR: 36.92) or G2141A (OR: 28.16) mutations increased the likelihood of clarithromycin resistance in H. pylori strains by 31.66-, 36.92- and 28.16-fold (ORs), respectively, according to the binary logistic regression analysis. Conclusion. We concluded that classical mutations of the 23 S rRNA gene resulted in higher clarithromycin MIC values than new mutations. These new point mutations caused moderate elevations in the MIC values of clarithromycin-resistant H. pylori strains.
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    Diagnostic performance of the RT-qPCR method targeting 85B mRNA in the diagnosis of pulmonary Mycobacterium tuberculosis infection
    (Elsevier Science London, 2018) Demirci, Mehmet; Saribas, Suat; Ozer, Nigar; Toprak, Sezer; Caglar, Emel; Ortakoylu, Gonenc; Yuksel, Pelin
    Background: Several nucleic acid amplification techniques (IS6110, 16S rRNA, and 85B mRNA) were developed for the rapid, direct detection of Mycobacterium tuberculosis. We aimed to assess the diagnostic performance of 85B mRNA-based RT-qPCR by comparing with the real-time PCR COBAS TagMan MTB Kit while using the BACTEC MGIT 960 method as the gold standard. Methods: 60 patients with confirmed pulmonary TB and 60 individuals without TB were included as the study and control groups, respectively. Sputum specimens were cultured using LJ and BACTEC MGIT 960 systems. Extracted DNA was used for COBAS PCR in a CONAS TagMan 48 analyzer. 85B mRNA detection was performed by RT-qPCR. Results: The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of COBAS TagMan MTB Test were detected as 93.3%, 83.3%, 84.8%, 92.6%, and 88.3%, respectively. The same diagnostic parameters of RT-qPCR were: 98.3%, 95.0%, 95.2%, 98.3%, and 96.7%, respectively. According to the binary logistic regression analysis, RT-qPCR (OR: 19,924, p < 0.001) was identified as the more optimal test. Conclusion: RT-qPCR targeting the 85B gene of M. tuberculosis seems to be a more useful and rapid technique than DNA-based methods for detecting live M. tuberculosis bacilli from sputum specimens. (C) 2018 The Authors. Published by Elsevier Limited on behalf of King Saud Bin Abdulaziz University for Health Sciences.
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    Problems encountered in conventional HIV 1/2 Algorithms: lack of necessity for immunoblot assays to confirm repeated ELISA reactive results
    (Makerere Univ, Fac Med, 2018) Yuksel, Pelin; Saribas, Suat; Kuskucu, Mert; Mutcali, Sibel Islak; Kosan, Erdogan; Habip, Zafer; Demirci, Mehmet
    Background: The use of conventional (serologically based) HIV 1/2 diagnostic algorithms has become controversial in recent years. Objectives: Sera from patients who underwent verification tests were evaluated because repeated ELISA-reactive results demonstrated a HIV1+HIV2 positive band pattern. Methods: The line immunoassay (LIA) test was used for repeated HIV enzyme immunoassays (EIA)-reactive sera in patients at three centers. The Bio-Rad Geenius T HIV 1/2 and the HIV-1 RNA tests were used. HIV-1 and RNA HIV-2 were investigated using PCR. Results: LIA was used to evaluate 3,224 out of 10,591 samples with repeated ELISA reactivity (30%). We found that 32 (1%) of the sera, along with HIV1 bands and HIV2 gp36 bands, were positive. Only 28 of the 32 verified serum samples with gp36 bands were repeated, and no gp36 band positivity was detected using the Bio-Rad Geenius T HIV-1/2 confirmatory assay in these serum samples. The HIV-2 proviral DNAs were also negative. Therefore, we excluded the possibility of HIV1+2 co-infection. All samples from the 32 patients were positive for HIV-1 RNA. Conclusion: Our findings highlight the need to exclude confirmatory tests like the LIA test from the current diagnostic HIV algorithm and replace it with rapid HIV-1 and HIV-2 confirmatory immunochromotographic tests.