Implementation of an in-house real-time reverse transcription-PCR assay to detect the emerging SARS-CoV-2 N501Y variants

The SARS-CoV-2 pandemic has been associated with the emergence of several variants with a mutated spike glycoprotein that are of substantial concern regarding their transmissibility and ability to evade immune responses. This warrants implementing strategies for their detection and surveillance. We have set up an in-house one-step real-time reverse transcription-PCR (qPCR) assay that specifically detects SARS-CoV-2 spike N501Y variants. Our assay was positive for all 6 patients found spike N501Y-positive by genome sequencing. Ten cDNA samples for each of the 10 Marseille variants identified by genome sequencing and three nasopharyngeal samples of a spike N501Y-negative variant (Marseille-4) that predominates locally tested negative. All negative controls among which 5 SARS-CoV-2-negative nasopharyngeal samples tested negative. First use in the setting of diagnosis on 51 nasopharyngeal samples from SARS-CoV-2-positive but Marseille-4-negative patients showed positivity in 5 cases further confirmed by sequencing as from spike N501Y variant-infected patients. Thus, our in-house qPCR system was found reliable for the detection of the N501Y substitution and allowed estimating preliminarily that spike N501Y variant prevalence was 4% among SARS-CoV-2 diagnoses since January 2020.