Analytical sensitivity and efficiency comparisons of SARS-COV-2 qRT-PCR assays

Chantal B.F. Vogels; Anderson F. Brito; Anne Louise Wyllie; Joseph R Fauver; Isabel M. Ott; Chaney C. Kalinich; Mary E. Petrone; Marie-Louise Landry; Ellen F. Foxman; Nathan D. Grubaugh

When testing nasopharyngeal swabs collected prior to the COVID-19 pandemic, we found that qRT-PCR with five primer-probe sets did not result in background amplification (E-Sarbeco, RdRp-SARSr, HKU-N, HKU-ORF1, and 2019-nCoV_N1; Fig. 3 ). When using SARS-CoV-2 RNA spiked into RNA from these nasopharyngeal swabs, our results show that none of these five primer-probe sets were able to detect (Ct values <40) SARS-CoV-2 RNA at 1 (10 0 ) virus genome equivalents/μL and all could partially detect 10 (10 1 ) virus genome equivalents/μL ( Fig. 3 ). Our results suggest that the two most sensitive primer-probe sets are E-Sarbeco (Charité) and HKU-ORF1 (HKU), which each detected 6/8 (75%) of the nasopharyngeal swabs spiked with 10 virus genome equivalents/μL ( Fig. 3 ). At 100 (10 2 ) virus genome equivalents/μL, we could detect virus and differentiate between the negative samples for all replicates and primers sets, except for the RdRp-SARSr (Charité) set, which was negative (Ct values >40) for all 10 0 -10 2 genome equivalents/μL concentrations.