Utvrđivanje delecije H69/V70 u SARS-CoV-2 virusu upotrebom TaqPath reagensa

master's thesis

• Title (english): Determination of H69/V70 deletion in SARS-CoV-2 virus using TaqPath reagents
• Author: Lucija Novosel
• Mentor: Marija Grdić Rajković (mentor)
• Mentor: Ivana Ćelap (mentor)
• Committee member: Marija Grdić Rajković (predsjednik povjerenstva)
• Committee member: Ivana Ćelap (član povjerenstva)
• Committee member: Dubravka Vitali Čepo (član povjerenstva)
• Granter: University of Zagreb; Faculty of Pharmacy and Biochemistry; (Department of medical biochemistry and haematology); Zagreb
• Defense date and country: 2022-11-29, Croatia
• Scientific / art field, discipline and subdiscipline: BIOMEDICINE AND HEALTHCARE; Pharmacy; Pharmacy

Abstract

Virus SARS-CoV-2 velikom se brzinom proširio cijelim svijetom 2020. godine te uzrokovao globalnu pandemiju. Podložan je brojnim mutacijama koje na različite načine utječu na njegova svojstva kao i na rezultate molekularne analize RT-PCR kojom se potvrđuje infekcija ovim virusom. Cilj ovog diplomskog rada bio je usporediti rezultate RT-PCR testiranja uzoraka nazofaringealnih briseva na prisutnost virusa SARS-CoV-2 korištenjem komercijalnog test paketa koji detektira prisutnost više virusnih gena (ORF1ab, N i S protein) s rezultatima sekvenciranja različitih uzoraka kod kojih je RT-PCR metodom dokazana i kod kojih nije dokazana prisutnost S gena. Za provođenje testiranja korišten je TaqPathTM COVID-19 CE-IVD RT-PCR Kit, a sekvenciranje je provedeno NGS metodom u laboratoriju Eurofins Genomics Europe. Rezultati su potvrdili postojanje delecije H69/V70 kod svih uzoraka kod kojih RT-PCR metodom nije detektiran S gen. Utvrđeno je da postojanje delecije H69/V70 na S genu onemogućuje njegovu detekciju. S obzirom na to da se određuje prisutnost druga dva gena, lažno negativni rezultati za S gen ne znače odsustvo virusa nego ukazuju na potencijalno prisustvo ove mutacije u soju virusa koji kruži u populaciji.

Abstract (english)

The SARS-CoV-2 virus spread rapidly throughout the world and caused a global pandemic in 2020. The virus is susceptible to numerous mutations that affect to its properties in different ways. Mutations also affect to the results of the RT-PCR molecular analysis which is used to confirm virus infection. The aim of this study was to compare RT-PCR results of nasopharyngeal swab samples which were analyzed to detect the presence of the SARS-CoV-2 virus using a commercial test package that detects the presence of several viral genes (ORF1ab, N and S protein) with different sequencing results of samples where the presence of S protein was confirmed and samples where the presence of S protein was not confirmed using RT-PCR. The TaqPathTM COVID-19 CE-IVD RT-PCR Kit was used for testing and the sequencing was performed using the NGS method in the Eurofins Genomics Europe laboratory. The results confirmed deletion H69/V70 in every sample where the S gene was not detected using RT-PCR It was found that H69/V70 S gene deletion prevents S gene to be detected while PCR testing. False-negative results for S gene presence should not indicate the absence of the virus because there are two other gens which presence is detected. But negative S gene results can indicate the potential presence of this virus mutation and its spreading through population.

• Keywords: SARS-CoV-2; virus; delecija H69/V70; RT-PCR; sekvenciranje nove generacije
• Keywords (english): SARS-CoV-2; virus; deletion H69/V70; RT-PCR; next generation sequencing
• Language: croatian
• URN:NBN: urn:nbn:hr:163:774936
• Study programme: Title: Pharmacy; Study programme type: university; Study level: integrated undergraduate and graduate; Academic / professional title: magistar/magistra farmacije (magistar/magistra farmacije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2022-11-29 11:29:49