SARS-CoV-2 RNA identification in nasopharyngeal swabs: issues in pre-analytics

Author:

Basso Daniela12,Aita Ada12,Navaglia Filippo2,Franchin Elisa3,Fioretto Paola4,Moz Stefania12,Bozzato Dania12,Zambon Carlo-Federico12,Martin Barbara4,Dal Prà Chiara4,Crisanti Andrea35,Plebani Mario12ORCID

Affiliation:

1. Department of Medicine – DIMED , University of Padova , Padova , Italy

2. Department of Laboratory Medicine , University Hospital of Padova , Padova , Italy

3. Department of Molecular Medicine – DMM, Microbiology and Virology , University of Padova , Padova , Italy

4. Department of Medicine – DIMED, Internal Medicine 3 , University of Padova , Padova , Italy

5. Department of Life Sciences, Imperial College London , South Kensington Campus , London , UK

Abstract

Abstract Objectives The direct identification of SARS-CoV-2 RNA in nasopharyngeal swabs is recommended for diagnosing the novel COVID-19 disease. Pre-analytical determinants, such as sampling procedures, time and temperature storage conditions, might impact on the end result. Our aim was to evaluate the effects of sampling procedures, time and temperature of the primary nasopharyngeal swabs storage on real-time reverse-transcription polymerase chain reaction (rRT-PCR) results. Methods Each nasopharyngeal swab obtained from 10 hospitalized patients for COVID-19 was subdivided in 15 aliquots: five were kept at room temperature; five were refrigerated (+4 °C); five were immediately mixed with the extraction buffer and refrigerated at +4 °C. Every day and for 5 days, one aliquot per condition was analyzed (rRT-PCR) for SARS-CoV-2 gene E and RNaseP and threshold cycles (Ct) compared. To evaluate manual sampling, 70 nasopharyngeal swabs were sampled twice by two different operators and analyzed separately one from the other. Results A total of 6/10 swabs were SARS-CoV-2 positive. No significant time or storage-dependent variations were observed in SARS-CoV-2 Ct. Re-sampling of swabs with SARS-CoV-2 Ct lower than 33 resulted in highly reproducible results (CV=2.9%), while a high variability was observed when Ct values were higher than 33 (CV=10.3%). Conclusions This study demonstrates that time and temperature of nasopharyngeal swabs storage do not significantly impact on results reproducibility. However, swabs sampling is a critical step, and especially in case of low viral load, might be a potential source of diagnostic errors.

Publisher

Walter de Gruyter GmbH

Subject

Biochemistry (medical),Clinical Biochemistry,General Medicine

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