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    Association between polymorphisms in HLA-A, HLA-B, HLA-DR, and DQ genes from gastric cancer and duodenal ulcer patients and cagL among cagA-positive Helicobacter pylori strains: The first study in a Turkish population
    (Elsevier, 2020) Kocak, Banu Tufan; Saribas, Suat; Demiryas, Suleyman; Yilmaz, Erkan; Uysal, Omer; Kepil, Nuray; Demirci, Mehmet
    Colonization of the human gastric mucosa by H. pylori may cause peptic and duodenal ulcers (DUs), gastric lymphomas, and gastric cancers. The cagL gene is a component of cag T4SS and is involved in cagA translocation into host. An association between the risk of gastric cancer and the type of HLA class II (DR and/or DQ) was suggested in different populations. The aim of this study was to investigate, the clinical association of the cagL gene with host HLA alleles in H. pylori strains that were isolated from patients with gastric cancer, DU, and non-ulcer dyspepsia (NUD) and to determine the HLA allele that confers susceptibility or resistance for the risk of gastric cancer and DU development in Turkish patients. A total of 94 patients (44 gastric cancer and 50 DU patients; 58 male, 36 female; mean age, 49.6 years), and 86 individuals (50 NUD patients and 36 persons with normal gastrointestinal system [NGIS]; 30 male, 56 female; mean age, 47.3 years) were included as the patient and the control groups, respectively. CagA and cagL were determined by PCR method. DNA from peripheral blood samples was obtained by EZ-DNA extraction kit. For HLA SSO typing, LIFECODES SSO Typing kits (HLA-A, HLA-B HLA-C, HLA-DRB1 and HLA-DQA1/B1 kits) were used. The CagL/CagA positivity distribution in the groups were as follows: 42 (95.4%) gastric cancer, 46 (92%) DU and, 34 (68%) NUD and no NGIS cases. The HLA-DQA1*01 (OR: 3.82) allele was significantly different, suggesting that these individuals with H. pylori strains harbouring the CagL/CagA positivity are susceptible to the risk of gastric cancer and DU, and the HLA-DQA1*05 (OR, 0.318) allele was suggested as a protective allele for the risk of gastric cancer and DU using univariate analyses. HLA-DQA1*01 (OR, 2.21), HLA-DQB1*06 (OR, 2.67), sex (male, OR, 2.27), and CagL/CagA/(< 2) EPIYA C repeats (OR, 5.72) were detected independent risk factors that increased the risk of gastric cancer and DU using multivariate analyses. However, the HLA-DRB1*04 (OR, 0.28) allele was shown to be a protective allele, which decreased the risk of gastric cancer and DU. Gastric pathologies result from an interaction between bacterial virulence factors, host epigenetic and environmental factors, and H. pylori strain heterogeneity, such as genotypic variation among strains and variations in H. pylori populations within an individual host.
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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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    The Prevalence of Periodontal Pathogenic Bacteria in Atherosclerotic Cardiovascular Disease
    (Clin Lab Publ, 2020) Gode, Safa; Sarp, Tevhide Z.; Saribas, Suat; Ergin, Sevgi; Kasnak, Gokhan; Dinc, Harika O.; Caliskan, Reyhan
    Background: A possible link between periodontal pathogenic bacteria and atherosclerosis may exist based on the inflammatory mechanisms initiated by bacteria found in periodontal lesions. Our aim was to investigate the presence of DNA originating from T. denticola, C. rectus, T. forsythia, and P. gingivalis in the vascular tissue specimens obtained from patients who underwent surgery for arteriosclerotic vascular disease in this study. Methods: A total of 96 patients diagnosed with valvular heart disease due to atherosclerosis and 85 patients with advanced aortic valve stenosis due to rheumatic fever and had undergone aortic valve replacement were included as the study (PG) and the control groups (CG), respectively. Atheroma plaques and vascular tissue specimens were collected from PG and CG during cardiovascular surgical procedures. Revitalization of the lyophilized T. denticola, ATCC 35405; C. rectus, ATCC 33238; P. gingivalis, ATCC 33277 and T. forsythia, ATCC 43037 strains was performed according to the manufacturer's instructions. C. rectus, T. forsythia, and T. denticola DNA samples were analyzed using the one-step in-house PCR method. Results: In one (1.04%) and three (3.13%) out of 96 atherosclerotic PG tissue specimens, P. gingivalis and T. forsythia DNA were detected, respectively. No T. denticola or C. rectus DNA was found in the study specimens. Periodontal pathogenic bacteria were not observed in 85 CG tissue specimens. There was no statistically significant difference between PG and CG for the presence of P. gingivalis and T. forsythia DNA using Fischer's Exact test (p > 0.05). Conclusions: In conclusion, with the case-control studies on a small scale such as in our study, it is not possible to determine a causality relationship between periodontal pathogenic bacteria and formation of atherosclerosis. Periodontal pathogenic bacteria may not be the only factor that causes inflammatory diseases associated with atherosclerosis. Host response and inflammatory mechanisms may be affected by other factors such as ethnicity, dietary habits, nutritional availability, and lifestyle. Taken together, it is difficult to conclude a causal link between periodontal pathogenic bacteria and formation of atherosclerosis.
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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.
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    A Retrospective Analysis of Anaerobic Bacteria Isolated in 236 Cases of Pleural Empyema and their Prevalance of Antimicrobial Resistance in Turkey
    (Clin Lab Publ, 2018) Demirci, Mehmet; Gemicioglu, Bilun; Saribas, Suat; Halis, Ayse N.; Taner, Zeynep; Mamal-Torun, Muzeyyen; Karatoka, Belma
    Background: Parapneumonic effusions usually occur secondary to an infection and produce pus (empyema) that accumulates in the pleural space. We aimed to evaluate the prevalence of anerobes in patients with empyema and to assess their resistance patterns for seven antimicrobials. Methods: Pleural fluid specimens from 236 patients were inoculated on Schaedler agar. Anaerobic bacteria were identified via API 20 A. Susceptibility testing for penicillin, ampicillin + sulbactam, amoxicillin + clavulanate, cefoxitin, clindamycin, metronidazole, and imipenem were performed with the E-test. Results: There were 118 anaerobic bacterial strains detected in 66 (27.9%) of the 236 specimens. Gram-positive anaerobic cocci were detected in 54.23% and the predominant cocci were 41 Peptostreptococcus spp, (34.75%) followed by 17 P. acnes (14.41%) and 6 C. tertium (5.08%). The Gram-negative anaerobes were B. fragilis (28, 23.73%), P. melaninogenica (8, 6.78%), P. intermedia (4, 3.39%), F. nucleatum (6, 5.08%), F mortiferum (5, 4.24%), and P. asaccharolytica (3, 2.54%). All anaerobic strains were susceptible to ampicillin + sulbactam, amoxicillin + clavulanate, and imipenem. The highest MIC was found to be > 256 mu g/mL for penicillin in B. fragilis strains, 128 mu g/mL for cefoxitin in P. melaninogenica strains, 32 mu g/mL for clindamycin and 64 mu g/mL for metronidazole in P. acnes strains. Clindamycin resistance was detected in 46.6% B. fragilis, and 17.6% for P. acnes. Thirty-eight (32.2%) strains produced beta-lactamase. Conclusions: The use of antimicrobial agents for thoracic empyema should be based on the isolated pathogens and their resistance profiles. Clinicians should be aware of the wide diversity of anaerobic genera and species in cases of pleural empyema.