Degradation of SARS-CoV-2 specific ribonucleic acid in samples for nucleic acid amplification detection

Abstract

The degradation of SARS-CoV-2 specific ribonucleic acid (RNA) was investigated by a numerical modeling approach based on nucleic acid amplification test (NAAT) results utilizing the SmartAmp technique. The precision of the measurement was verified by the relative standard deviation (RSD) of repeated measurements at each calibration point. The precision and detection limits were found to be 6% RSD (seven repeated measurements) and 94 copies/tube, respectively, at the lowest calibration point. RNA degradation curves obtained from NAAT data on four different temperatures were in good agreement with the first-order reaction model. By referring to rate constants derived from the results, the Arrhenius model was applied to predict RNA degradation behavior. If the initial RNA concentration was high enough, such as in samples taken from infected bodies, the NAAT results were expected to be positive during testing. On the other hand, if initial RNA concentrations were relatively low, such as RNA in residual viruses on environmental surfaces, special attention should be paid to avoid false-negative results. The results obtained in this study provide a practical guide for RNA sample management in the NAAT of non-human samples.

Validating the inactivation of viral pathogens with a focus on SARS-CoV-2 to safely transfer samples from high-containment laboratories

“…In this study, heat inactivation of SARS-CoV-2 causes a decrease in genome copy numbers ( Figures 2C, D , 3B, C ) or remains undetectable in RT-qPCR ( Figures 4B , 5D ) over the incubation period, validating virus inactivation. A decrease in genome copy number is expected in the treated group due to RNA degradation at 37°C over time ( Takeuchi et al., 2022 ) as reflected in Figures 2D , 3B . To the contrary, the untreated virus should show a decrease in CT value or an increase in genome copy numbers over time in cell cultures, as demonstrated in this study ( Figures 2 – 5 ).…”

Streamlined Specimen Purification for Rapid COVID-19 Diagnosis Using Positively Charged Polymer Thin Film-Coated Surfaces and Chamber Digital PCR

“…However, increasing the heating temperature did not improve the RNA capture efficiency. This may be because although high temperatures did not damage DNA structures, according to their stability with double-stranded molecules, the temperatures could have damaged RNA structures and resulted in inefficient PCR (Figure S4). We measured the capture efficiency of various RNA concentrations under the optimized RNA capture conditions.…”

The life and death of RNA across temperatures

“…RNA degradation plays an important role in diagnostic and forensic science [142] . Evidence of degradation of some mRNAs suggest that the 5′ end of an mRNA transcript degrades in dried stains faster than the 3′ end.…”