Comparison of the Modified Centers for Disease Control and Prevention 2019-Novel Coronavirus Real-Time RT-PCR Method for Detection of Infectious and Heat-Inactivated Virus on Stainless Steel

• Published in: Journal of AOAC INTERNATIONAL 2021-04, Vol.104 (4), p.867-871
• Main Authors: Brunelle, Sharon L, Bird, Patrick M, Boone, Jeremy, Nelson, Maria, Johnson, Zerlinde, Coates, Scott
• Format: Article
• Language: English
• ISSN: 1060-3271; 1944-7922
• DOI: 10.1093/jaoacint/qsab046

Infectious Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) was used in the validation of methods for detection of SARS-CoV-2 on stainless steel surfaces in the AOAC Research Institute Emergency Response Validation project. Handling infectious virus requires Biosafety Level (BSL)-3 facilities. To compare the recovery and detection of infectious and heat-inactivated (65 °C for 30 min) SARS-CoV-2 from stainless steel by the modified US Centers for Disease Control and Prevention (CDC) 2019-Novel Coronavirus Real Time Reverse Transcription Polymerase Chain Reaction (RT-PCR) Diagnostic Panel. Viral stocks were diluted in viral transport medium and deposited onto stainless steel test areas at 2 x 103 and 2 x 104 genomic copies for low and high, respectively. Test areas were sampled, and aliquots of the resulting test solutions analyzed by RT-qPCR according to the CDC method. Results were analyzed by Probability of Detection (POD) statistics. The low level, where fractional positive results (25-75%) are expected, yielded PODI = 0.80 (0.58, 0.92) for the infectious virus and PODHI = 0.15 (0.05, 0.36) for the heat-inactivated virus. The bias, dPODHI = -0.65 (-0.80, -0.35), demonstrated a statistical difference between infectious and heat-inactivated virus detection. No difference was observed at the high inoculation level. Despite the statistical difference observed, the use of the heat-inactivated virus is a viable alternative for matrix extension studies using a method comparison study design. The use of heat-inactivated SARS-CoV-2 can mitigate the need for a BSL-3 facility for matrix extension validation of alternative methods in SARS-CoV-2 studies.