Prolonged Unfrozen Storage and Repeated Freeze-Thawing of SARS-CoV-2 Patient Samples Have Minor Effects on SARS-CoV-2 Detectability by RT-PCR

Reliable transportation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) patient samples from a swabbing station to a diagnostics facility is essential for accurate results. Therefore, cooling or freezing the samples is recommended in case of longer transportation times. In this study, SARS-CoV-2 detectability by RT-PCR was assessed after prolonged unfrozen storage or repetitive freeze-thawing of SARS-CoV-2 samples. SARS-CoV-2-positive patient swabs stored in viral transport medium were exposed to different temperatures (4 degrees C, 25 degrees C, and 35 degrees C) and to repetitive freeze-thawing, to assess the effect of storage conditions on RT-PCR detection. SARS-CoV-2 RNA was still reliably detected by RT-PCR after 21 days of storage in viral transport medium, even when the samples had been stored at 35 degrees C. The maximum observed change in cycle threshold value per day was 0.046 (+/- 0.019) at 35 degrees C, and the maximum observed change in cycle threshold value per freeze-thaw cycle per day was 0.197 (+/- 0.06). Compared with storage at 4 degrees C, viral RNA levels deviated little but significantly when stored at 25 degrees C or 35 degrees C, or after repeated freeze-thawing. The results of this study indicate that viral RNA levels are relatively stable at higher temperatures and repetitive freeze-thawing.