TR Dizin İndeksli Yayınlar Koleksiyonu / TR Dizin Indexed Publications Collection
Permanent URI for this collectionhttps://hdl.handle.net/20.500.14365/4
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Article Citation - WoS: 2Citation - Scopus: 2Analysis of Nucleotide Changes in Rt-Pcr Primer/Probe Binding Regions in Sars-Cov Isolates Reported From Turkey(Ankara Microbiology Soc, 2021-07-16) Demir, Ayse Banu; Bulgurcu, Alihan; Appak, Ozgur; Sayiner, Ayca ArzuThe SARS-CoV-2 virus, which caused the COVID-19 epidemic, caused more than 55 million cases and nearly 1.5 million deaths worldwide. For the microbiological diagnosis of the disease, the most valid method is detecting the presence of the viral genome by real-time reverse transcription polymerase chain reaction (rRT-PCR). However, due to the nature of the RNA viruses, frequent mutations may affect the sensitivity of the analyses made on the genetic material of the virus, such as PCR. In this study, we aimed to investigate the mutations in the primer-probe binding regions of the rRT-PCR panels used in COVID-19 diagnosis. SARS-CoV-2 whole genome sequence data (n= 194) isolated from COVID-19 cases in Turkey and uploaded on GISAID database from the centers in Istanbul (n= 78), Ankara (n= 58), Kars (n= 47), Bursa (n= 2), Adiyaman (n= 2), Erciyes (n= 1) and Kocaeli (n= 1) between March 17-September 14, 2020 were analyzed. In order to determine the nucleotide changes, SARS-CoV-2 sequences from Turkey were compared to the reference genome sequence (NC_045512.1) present in GenBank website. The constructed data set was aligned using the MAFFT program and was checked manually if the sequences were in the same frame by using the AliView program. Primer-probe binding sites of the thirteen SARS-CoV-2 rRT-PCR panels from seven different institutes (US CDC, China CDC, Charite CDC, Pasteur, HKU, Thailand, NIID) that are being used in COVID-19 diagnosis were evaluated in terms of nucleotide changes within the corresponding regions compared to the reference genome. Sequence diversities in the viral genomes were determined via positional nucleotide numerical calculator and entropy calculator modules and nucleotide and entropy changes in primer-probe binding regions for each rRT-PCR panel were examined. Among thirteen different primer-probe panels, nucleotide changes in the target regions of the seven primer-probe panels were determined. When viral sequences with nucleotide changes in the primer-probe binding regions were examined, the most common changes were observed in the China CDC N-forward primer and US CDC N3-forward primer binding regions. It is important that the kits to be used as diagnostic tests are designed specific to the regions with less nucleotide changes. Nucleotide changes may not be critical for DNA amplification for most PCR panels, but should be carefully monitored as they may affect the sensitivity of the assay. If the risk of alteration of the designed region is high, the primer - probe binding sites should be checked frequently and updated when necessary.Article Citation - WoS: 1Determination of a Sample-To Ratio To Predict True-Positivity in Blood Donor Samples Screened for Syphilis by a Chemiluminescent Immunoassay(Aves Press Ltd, 2018) Akcakanat, I. Ebru; Ozbek, Ozgen Alpay; Dogan, Yavuz; Abacioglu, Yusuf HakanPurpose: The use of Architect Syphilis TP (CMIA) in the blood bank raised the number of syphilis positive samples requiring confirmation. The aim of this study is to determine a sample-to-cutoff (s/co) ratio for CMIA predicting >= 95% of true-positive samples to reduce these samples. Methods: CMIA reactive samples (n=177) were evaluated by Western blot (WB) as the reference standard, as well as by Treponema pallidum hemagglutination (TPHA) and Rapid Plasma Reagin (RPR) tests. The s/co ratio predicting >= 95% of true-positive samples was defined as the threshold leaving >= 95% of WB confirmed samples greater than the particular value. The performances of TPHA and RPR tests were also evaluated with respect to s/co ratios of CMIA positive samples. Results: The s/co ratio 15.17 predicted a true-positive result for >= 95% of samples tested (95% confidence interval: 85.9-99.3) and reduced the number of samples requiring confirmation by 29.9%. Higher s/co ratios were correlated with the increasing number of bands on WB strips (p<0.0001, R=0.906). For the samples with s/co ratios between 3 and 15.17, the agreement of TPHA and WB test results were 90%. The lowest s/co ratio where TPHA was positive, was 3.1. Although RPR predicted > 95% of positive samples with s/co ratios > 15, its sensitivity was 47.7%. Conclusion: Higher s/co ratios can be used to define true-positivity and may indicate an active infection. TPHA may replace WB to confirm samples with s/co ratios between 3 and 15. RPR should not be used as a screening test in blood banks as it could miss almost half of the true-positive samples.