Influence of Different Inactivation Methods on Severe Acute Respiratory Syndrome Coronavirus 2 RNA Copy Number.

The outbreak of coronavirus disease 2019 (COVID-19) has spread across the world and was characterized as a pandemic. To protect medical laboratory personnel from infection, most laboratories inactivate the virus causing COVID-19, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), in clinical samples before testing. However, the effect of inactivation on the detection results remains unknown. Here, we used a digital PCR assay to determine the absolute SARS-CoV-2 RNA copy number in 63 nasopharyngeal swab samples and assess the effect of inactivation methods on viral RNA copy number. Viral inactivation was performed by three different methods: (i) incubation with the TRlzol LS reagent for 10 min at room temperature, (ii) heating in a water bath at 56 degrees C for 30 min, and (iii) high-temperature treatment, including autoclaving at 121 degrees C for 20 min, boiling at 100 degrees C for 20 min, and heating at 80 degrees C for 20 min. Compared to the amount of RNA in the original sample, TRlzol treatment destroyed 47.54% of the nucleocapsid protein (N) gene and 39.85% of open reading frame (ORF) lab. For samples treated at 56 degrees C for 30 min, the copy number of the N gene and ORF lab was reduced by 48.55% and 56.40%, respectively. The viral RNA copy number dropped by 50 to 66% after heating at 80 degrees C for 20 min. Nearly no viral RNA was detected after autoclaving at 121 degrees C or boiling at 100 degrees C for 20 min. These results indicate that inactivation reduced the quantity of detectable viral RNA and may cause false-negative results, especially in weakly positive cases. Thus, use of the TRlzol reagent rather than heat inactivation is recommended for sample inactivation, as the TRlzol reagent had the least effect on the RNA copy number among the tested methods.