Browsing by Subject "REAL-TIME PCR"

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    Comparing efficiency between conventional and molecular methods for detecting Legionella pneumophilia in dental unit waterline systems
    (HARD) Islam, A; Güvenir, M; Süer, K; Çetiner, S; Sanlidag, T
    The aim of our study was to detect the prevalence of Legionella pneumophilia (L. pneumophilia) in DUWL S using standard culture technique (SCT) and the real-time polymerase chain reaction (PCR) method in order to assess the risk of L. pneumophilia contamination within a dental setting. A total of 65 water samples were collected from 16 dental units and one cold water supply system from all clinical departments. L. pneumophilia could not be detected in any of the water samples using the standard SCT (0%), whereas L. pneumophilia was detected using real time PCR in three (4.6%) water samples collected from the tap system. Following the detection of L. pneumophilia, the tap systems were disinfected with surface disinfectant and water samples were recollected. The recollected water samples following disinfection were negative for L. pneumophilia once analyzed using culture and real time PCR technique. Although the culture method using BCYE media is the 'gold standard' for the detection of L. pneumophilia; Real Time PCR analysis may also be a quick, useful, and sensitive method for the detection of L. pneumophilia in order to control and prevent possible infections that may arise in the dental setting.
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    Investigation of Dientamoeba fragilis Prevalence and Evaluation of Sociodemographic and Clinical Features in Patients with Gastrointestinal Symptoms
    (SPRINGER INT PUBL AG) Aykur, M; Kurt, CC; Erdogan, DD; Avci, CB; Vardar, R; Aydemir, S; Girginkardesler, N; Gündüz, C; Dagci, H
    BackgroundDientamoeba fragilis is a protozoan parasite of the human gastrointestinal tract and still controversial in association with gastrointestinal symptoms.PurposeWe present cross-sectional study of the prevalence of D. fragilis, and sociodemographic and clinical features in the patients with gastrointestinal symptoms.MethodsA total of 490 fecal specimens were collected from outpatients with gastrointestinal symptoms in the Department of Parasitology, Faculty of Medicine, Ege University and Celal Bayar University, Turkey. Fecal specimens were examined with microscopy and inoculated in Robinson medium. D. fragilis-positive samples were examined for the presence of other intestinal parasites using enzyme immunoassay. Real-time PCR analysis was performed on all samples.ResultsOf the 490 stool specimens examined by real-time PCR, 59 patients were positive for D. fragilis infection with prevalence rate of 12.04%. Forty-four of positive patients (74.5%) were found to be infected with only D. fragilis, while 23.7% were co-infected with Blastocystis and 1.7% were co-infected with Rotavirus. No statistically significant difference was found in all the examined patients in terms of D. fragilis positivity for all sociodemographic parameters. Loose stool consistency was associated with the presence of D. fragilis, with 18.3% (P=0.001). When the clinical symptoms of all the patients participating in this study were examined, diarrhea was statistically more significant in patients with the presence of D. fragilis (16.3%; P=0.001). The rate of diarrhea in D. fragilis-positive patients (84.09%; P=0.0005) was higher than that of D. fragilis-negative patients and it was statistically significant.ConclusionThis study is important for assessing the prevalence of D. fragilis and its association with other factors in symptomatic patients in a large sample group in Turkey, as well as investigating the relationship of identified symptoms with the D. fragilis pathogenicity. It is suggested that D. fragilis in this case is not a commensal parasite but a pathogenic parasite and that the most common clinical symptom is diarrhea.
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    Investigation of BRAF mutation analysis with different technical platforms in metastatic melanoma
    (ELSEVIER GMBH, URBAN & FISCHER VERLAG) Sener, E; Yildirim, P; Tan, A; Gokoz, O; Tezel, GG
    In metastatic melanoma, the detection of somatic mutations in the BRAF gene is crucial regarding patient selection for targeted therapy. Several screening methods have been developed to identify BRAF gene mutations. In this study, our objective was to evaluate the detection of the BRAF V600 mutations using two molecular methods, real-time polymerase chain (real-time PCR) assay and pyrosequencing, and immunohistochemistry (IHC), and to compare the results of these different technical platforms. This study included 98 patients diagnosed with metastatic melanoma at the Hacettepe University, Department of Pathology between 2002 and 2014. BRAF mutation analysis was tested with real-time PCR, pyrosequencing and IHC methods. The results of all three tests were compared with a reference test, and the sensitivity, specificity rates and kappa coefficient values were analysed for each test. We successfully analysed BRAF mutations using all three methods in 92 patients. According to our findings, the pyrosequencing method had the highest kappa value regarding the determination of BRAF V600 mutations. The kappa values were at almost perfect agreement levels in pyrosequencing and realtime PCR assay (kappa coefficient for pyrosequencing = 0.895 (95% CI: 0.795-0.995); kappa coefficient for real-time PCR=0.871 (95% CI: 0.761-0.981). The kappa value was at a substantial agreement level in the IHC analysis (kappa coefficient = 0.776 (95% CI: 0.629-0.923). According to our results, we found that real-time PCR and pyrosequencing methods were equally excellent in determination of BRAF V600 mutations. The IHC method, which is commonly used in routine pathology practice, can also be safely used as a screening test for determination of BRAF V600 mutations. (C) 2017 Elsevier GmbH. All rights reserved.
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    Evaluation of 2015-2016 MOTAKK HBV DNA and HCV RNA External Quality Assessment National Program Results
    (ANKARA MICROBIOLOGY SOC) Karatayli, E; Soydemir, E; Aksoy, ZB; Kizilpinar, M; Altay Koçak, A; Karatayli, SC; Yurdcu, E; Yildirim, U; Güriz, H; Bozdayi, G; Yurdaydin, C; Ilhan, O; Yildirim, Y; Bozdayi, AM; Oguz, AY; Baris, A; Alp, A; Aksözek, A; Sayiner, A; Karagul, A; Ordu, A; Istanbullu, A; Otlu, B; Aridogan, B; Aksu, B; Buruk, CK; Karahan, C; Güney, Ç; Toksöz, D; Yildirim, D; Çolak, D; Daglar, DE; Findik, D; Kas, E; Çaliskan, E; Zeyrek, FY; Arslan, F; Demir, F; Milletli, F; Kibar, F; Özdinçer, F; Dündar, G; Arslan, H; Agca, H; Aliskan, HE; Güdücüoglu, H; Fidan, I; Akyar, I; Afsar, I; Kaleli, I; Dönmez, I; Yanik, K; Midilli, K; Çubukçu, K; Özdemir, M; Acar, M; Yalinay, M; Kuskucu, MA; Bakici, MZ; Aydin, N; Yilmaz, N; Çeken, N; Ziyade, N; Yilmaz, N; Özgümüs, OB; Gitmisoglu, Ö; Demirgan, R; Kesli, R; Güçkan, R; Sertöz, R; Akgün, S; Aksaray, S; Tezcan, S; Kaygusuz, S; Gökahmetoglu, S; Mese, S; Bayik, SA; Akçali, S; Gürcan, S; Karsligil, T; Us, T; Özekinci, T; Pilgir, T; Aslan, U; Dinç, U; Coskun, USS; Çetinkol, Y; Keskin, Y; Ayaydin, Z; Toraman, ZA
    MOTAKK, as a national external quality control program has been launched to evaluate the molecular detection of viral infections including HBV DNA and HCV RNA in molecular microbiology diagnostic laboratories in Turkey. This program is prepared in compliance with ISO 17043:2010 (Conformity assessment general requirements for proficiency testing) standards, and aims to take the place of external quality control programs from abroad, contributing to standardization and accuracy of molecular diagnostic tests in our country. The aim of this study was to evaluate 2015 and 2016 results of the MOTAKK External Quality Control Program for HBV DNA and HCV RNA viral load. The calls were announced on the web page of MOTAKK (www.motakk.org). The quality control samples were sent to participating laboratories in 2015 and 2016. Main stocks were prepared from patients with chronic hepatitis B and C who had viral load detection with reference methods according to WHO reference materials for viral load studies to improve quality control sera. From these main stocks, samples with different viral loads were prepared from dilutions of plasma with HBV, HCV, HAV, HIV, Parvovirus B19 and CMV negative serologic markers. Quality control samples were sent to the participating laboratories along with the negative samples in the cold chain. The laboratories accomplished the related tests within 2-3 weeks and entered their results on the MOTAKK web page. These results were analysed according to ISO 13528 (Statistical methods for use in proficiency testing by interlaboratory comparison) and scoring reports were created by a software developed by MOTAKK and sent to participating labs. Each laboratory evaluated their own results in comparison with the other laboratory results, reassessed the tests via observing the distance from the mean result and the reference values. The number of laboratories participating in the HBV DNA and HCV RNA external quality control program was 70-73 in 2015-2016. Participants were able to comply with the program tools, registering, entering results and receiving the results reports problem. In HBV panel, 72.6-89.1% and 84.7-90.3% of the participant laboratories were in 1 standard deviation (SD) in 2015-2016, respectively. In HCV panel, 70.8-89.1% and 84.7-90.3% of the participant laboratories were in 1 SD in 2015-2016, respectively. A national external quality control program for HBV DNA and HCV RNA in Turkey has been prepared for the first time with this project and implemented successfully. All the data provided in the MOTAKK external quality control program final report, compensate all the data provided by the quality control program final reports from abroad; additionally, the report allows comparison of used technologies and commercial products.
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    Dientamoeba fragilis Infection in Patients with Gastrointestinal System Complaints
    (ANKARA MICROBIOLOGY SOC) Sivcan, E; Charyyeva, A; Ceylan, SS; Yürük, M; Erdogan, E; Sahin, I
    In this study, we aimed to investigate the incidence of Dientamoeba fragilis with different diagnostic methods in patients with gastrointestinal symptoms and determine the sensitivity and specificity of existing diagnostic methods. Fecal samples collected from 101 patients with gastrointestinal complaints (especially upper abdominal pain, abdominal and pelvic pain, nausea and vomiting, gastroenteritis and colitis, unexplained fever and diarrhea) and 20 control cases from various clinics were included in the study. Samples were first examined with native-Lugol (N-L) method and cultured in Robinson medium. All 121 stool and culture samples were stained with iron hematoxylin stain (IHS) and trichrome stain (TS) methods and examined by PCR and QPCR for D.fragilis. Among 121 stool samples 13 (10.7%), 2 (1.7%), 7 (5.7%) 13 (10.7%), and 7 (5.8%), 4 (3.3%), 2 (1.7%), 3 (2.5%) of cultured samples were determined positive with IHS, TS, PCR, QPCR respectively. Fifteen of the 121 stool samples were determined as diarrheal. All diarrheal stool samples were negative with IHS and TS. One of the diarrheal stools and 6 (4.9%) of the non-diarrheal stools were positive by PCR. All of the diarrheal stools were negative. Thirteen of the non-diarrheal stool samples (10.7%) were positive by QPCR. When the QPCR method was considered as gold standard, sensitivity and specificity values were determined as 46% and 93% in IHS, 0% and 99% in TS, 54% and 100% by PCR and sensitivity and specificity values were 67% and 96% in IHS, 33% and 98% in TS, 67% and 100% by PCR among cultured stool samples. As a result, it was determined that there was a statistically significant difference between the samples of the patients and the control groups and the sensitivity and specificity of the conventional and molecular methods (IHS, TS, PCR and QPCR) determined in this study supported the results of other compared studies. It has been determined that staining methods used for the diagnosis of D.fragilis gave false positivite or negativite results. In addition, the QPCR method is more advantageous in terms of time saving for the diagnosis and initiation of the treatment and in cases where QPCR is not available, IHS and conventional PCR methods should be used together. In our opinion, this study will contribute to the results of epidemiological and scientific studies on D.fragilis in Turkey.
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    Diagnostic accuracy of the Xpert® MTB/RIF cycle threshold level to predict smear positivity: a meta-analysis
    (INT UNION AGAINST TUBERCULOSIS LUNG DISEASE (I U A T L D)) Lange, B; Khan, P; Kalmambetova, G; Al-Darraji, HA; Alland, D; Antonenka, U; Brown, T; Balcells, ME; Blakemore, R; Denkinger, CM; Dheda, K; Hoffmann, H; Kadyrov, A; Lemaitre, N; Miller, MB; Nikolayevskyy, V; Ntinginya, EN; Ozkutuk, N; Palacios, JJ; Popowitch, EB; Porcel, JM; Teo, J; Theron, G; Kranzer, K
    SETTING: Xpert (R) MTB/RIF is the most widely used molecular assay for rapid diagnosis of tuberculosis (TB). The number of polymerase chain reaction cycles after which detectable product is generated (cycle threshold value, C-T) correlates with the bacillary burden. OBJECTIVE: To investigate the association between Xpert C-T values and smear status through a systematic review and individual-level data meta-analysis. DESIGN: Studies on the association between C-T values and smear status were included in a descriptive systematic review. Authors of studies including smear, culture and Xpert results were asked for individual-level data, and receiver operating characteristic curves were calculated. RESULTS: Of 918 citations, 10 were included in the descriptive systematic review. Fifteen data sets from studies potentially relevant for individual-level data meta-analysis provided individual-level data (7511 samples from 4447 patients); 1212 patients had positive Xpert results for at least one respiratory sample (1859 samples overall). ROC analysis revealed an area under the curve (AUC) of 0.85 (95%CI 0.82-0.87). Cut-off C-T values of 27.7 and 31.8 yielded sensitivities of 85% (95%CI 83-87) and 95% (95%C1 94-96) and specificities of 67% (95%CI 66-77) and 35% (95%CI 30-41) for smear-positive samples. CONCLUSION: Xpert CT values and smear status were strongly associated. However, diagnostic accuracy at set cut-off C-T values of 27.7 or 31.8 would not replace smear microscopy. How C-T values compare with smear microscopy in predicting infectiousness remains to be seen.