Browsing by Author "Demir, Ayse Banu"

Now showing 1 - 2 of 2

Analysis of Nucleotide Changes in Rt-Pcr Primer/Probe Binding Regions in Sars-Cov-2 Isolates Reported from Turkey
(Ankara Microbiology Society, 2021-07-16) Sayıner, Ayça Arzu; Appak, Özgür; Demir, Ayse Banu; Bulgurcu, Alihan
The 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.
Citation - WoS: 1
Citation - Scopus: 1
Differential Effects of Choline on TLR2/4 Mediated Signaling Through Possible Regulation of Toll-Interacting Protein in Hepatocellular Carcinoma Cell Lines
(Walter de Gruyter GmbH, 2024-05-30) Barış, Elif; Demir, Ayse Banu
Objectives: Toll-like receptor (TLR) mediated inflammatory status plays an important role in development and pro- gression of hepatocellular carcinoma (HCC). Toll-interacting protein (TOLLIP) has an inhibitory effect on TLR-mediated inflammatory signalling and expression profile of TOLLIP varies between malignancies including HCC. Cholinergic anti-inflammatory pathway (CAP) is an endogenous mech- anism that controls inflammatory status via α7nicotinic acetylcholine receptors (α7nAChR). This study aims to investigate the effect of CAP-acting agent choline on TOLLIP and its related TLR-mediated inflammatory response in HCC cells with distinct differentiation stages. Methods: The expression patterns of α7nAChR, TLR2/4, TOLLIP, IL6, NFkB genes were evaluated by RT-PCR and ELISA in the presence of choline, along with the real-time cell proliferation and migration in HEP3B and SNU449 HCC cell lines. The interaction between choline and TOLLIP assessed by using in-silico analyses. Results: Choline downregulated TOLLIP in Hep3B and SNU449 cells. However, the expressions of α7nAChR, NF-κB, IL-6, TLR2 and TLR4 showed a decreased pattern in well differentiated HEP3B cells, while an increased pattern in poorly differentiated SNU449 cells. Conclusions: Choline might exert differential effects in TLR2/4-dependent signalling based on the differentiation stages of the HCC cells, suggesting its potential therapeutic effects in earlier stages of HCC which might be result of its partial modulation of TOLLIP.