Effect of Heat inactivation on Real-Time Reverse Transcription PCR of the SARS-COV-2 Detection

ABSTRACT

Real-time reverse transcription PCR (rRT-PCR) is commonly used to diagnose SARS-CoV-2 infection. Heat inactivation prior to nucleic acid isolation may allow safe testing, while the effects of heat inactivation on SARS-CoV-2 rRT-PCR detection result need to be determined. 14 positive nasopharyngeal swab specimens were inactivated at 56{degrees}C for 30min, 56{degrees}C for 60min, 60{degrees}C for 30min, 60{degrees}C for 75min, and 100{degrees}C for 10min, then were detected by rRT-PCR. All 14 heat treated samples remained positive. Another 2 positive nasopharyngeal swab specimens were inactivated at 100{degrees}C for 10min, 100{degrees}C for 30min, and 100{degrees}C for 60min, after which the samples were isolated and detected by rRT-PCR. The range of threshold cycle (Ct) values observed when detecting ORF1a/b was 27.228-34.011 in heat-treated samples, while 25.281-34.861 in unheated samples, and the range of threshold cycle (Ct) values observed at the time of detecting N was 25.777-33.351 in heat-treated samples, while 24.1615-35.433 in unheated samples, on basis of which it showed no statistical difference otherwise a good correlation of Ct values between the heat-inactivated samples and the untreated samples. However, the 2 samples inactivated at 100{degrees}C 30min, 100{degrees}C 60min turned into negative. Heat inactivation at 56{degrees}C for 30min, 56{degrees}C for 60min, 60{degrees}C for 30min, 60{degrees}C for 75min, and 100{degrees}C for 10min shall not affect the detection results of Real-Time Reverse Transcription PCR of the SARS-COV2. Furthermore, it is recommended to inactive nasopharyngeal swab specimens 10min at 100{degrees}C before RNA extraction in consideration of efficiency and reliable results.