Abstract
The pandemic caused by SARS-CoV-2 resulted in increasing demands for diagnostic tests, leading to a shortage of recommended testing materials and reagents. This study reports on the performance of self-sampled alternative swabbing material (ordinary Q-tips tested against flocked swab and rayon swab), of reagents for classical RNA extraction (phenol/guanidine-based protocol against a commercial kit), and of intercalating dye-based one-step quantitative reverse transcription real-time PCRs (RT-qPCR) compared against the gold standard hydrolysis probe-based assays for SARS-CoV-2 detection. The study found sampling with Q-tips, RNA extraction with classical protocol and intercalating dye-based RT-qPCR as a reliable and comparably sensitive strategy for detection of SARS-CoV-2—particularly valuable in the current period with a resurgent and dramatic increase in SARS-CoV-2 infections and growing shortage of diagnostic materials especially for regions limited in resources.
Highlights
The pandemic caused by SARS-CoV-2 resulted in increasing demands for diagnostic tests, leading to a shortage of recommended testing materials and reagents
Guidelines recommended the preferred use of flocked swabs stored in transport medium over dry swabs collected by s pecialists[6] and listed only kit-based RNA extraction methodologies followed by a hydrolysis probe-based reverse transcription quantitative real-time PCR (RT-qPCR) assay
To provide alternatives for molecular detection of SARS-CoV-2 infections during material shortages, we compared the following: ordinary cotton swab “Q-tips” available in supermarkets against medical sampling approaches, a classical protocol for RNA extraction against the commercially available kit for viral RNA extraction, and a cost-saving intercalating dye-based one-step, reverse transcription quantitative real-time PCR (SYBR RTqPCR) protocol against the gold standard hydrolysis probe-based one-step, reverse transcription quantitative real-time PCR for detection of SARS-CoV-2
Summary
The pandemic caused by SARS-CoV-2 resulted in increasing demands for diagnostic tests, leading to a shortage of recommended testing materials and reagents. This study reports on the performance of self-sampled alternative swabbing material (ordinary Q-tips tested against flocked swab and rayon swab), of reagents for classical RNA extraction (phenol/guanidine-based protocol against a commercial kit), and of intercalating dye-based one-step quantitative reverse transcription realtime PCRs (RT-qPCR) compared against the gold standard hydrolysis probe-based assays for SARSCoV-2 detection. Guidelines recommended the preferred use of flocked swabs stored in transport medium over dry swabs collected by s pecialists[6] and listed only kit-based RNA extraction methodologies followed by a hydrolysis probe-based RT-qPCR assay These materials and methods became the gold standard procedure for detection of SARS-CoV-2. Adaptation of diagnostic procedures to available and alternative reagents is necessary to ensure continued reliable diagnosis For this purpose, we tested self-sampled, ordinary cotton swabs “Q-tips” as an alternative to medical swabs for oropharyngeal sampling, the classical RNA extraction protocol, and a costsaving intercalating dye-based RT-qPCR assay for the detection of SARS-CoV-2
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