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Updated: Oct 19, 2025
Author Spotlight: Advancements in Multiplex Detection of Respiratory Viruses
Published on: November 10, 2023
Monitoring the SARS-CoV-2 pandemic: screening algorithm with single nucleotide polymorphism detection for the rapid
Joachim Mertens1, Jasmine Coppens2, Katherine Loens2
1Department of Microbiology and National Reference Centre for Respiratory Pathogens, University Hospital Antwerp, Edegem, Antwerp, Belgium; Laboratory of Medical Microbiology, Vaccine & Infectious Disease Institute (VAXINFECTIO), University of Antwerp, Wilrijk, Antwerp, Belgium.
Objectives:
To evaluate a testing algorithm for the rapid identification of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants that includes the use of PCR-based targeted single nucleotide polymorphism (SNP) detection assays preceded by a multiplex PCR sensitive to S-Gene Target Failure (SGTF).
Methods:
PCR SNP assays targeting SARS-CoV-2 S-gene mutations ΔH69-V70, L452R, E484K, N501Y, H655Y and P681R using melting curve analysis were performed on 567 samples in which SARS-CoV-2 viral RNA was detected by a multiplex PCR. Viral whole-genome sequencing (WGS) was performed to confirm the presence of SNPs and to identify the Pangolin lineage. Additionally, 1133 SARS-CoV-2 positive samples with SGTF were further assessed by WGS to determine the presence of ΔH69-V70.
Results:
The N501Y-specific assay (n = 567) had an overall percentage agreement (OPA) of 98.5%. The ΔH69-V70-specific (n = 178) and E484K-specific (n = 401) assays had OPA of 96.6% and 99.7%, respectively. Assessment of H655Y (n = 139) yielded a 100.0% concordance when applied in the proposed algorithm. The L452R-specific (n = 67) and P681R-specific (n = 62) assays had an OPA of 98.2% and 98.1%, respectively. The proposed algorithm identified six variants of concern/interest (VOC/VOI)-Alpha (n = 149), Beta (n = 65), Gamma (n = 86), Delta (n = 49), Eta (n = 6), Kappa (n = 6)-and 205 non-VOC/VOI strains-including the variants under monitoring B.1.214.2 (n = 43) and B.1.1.318 (n = 18) and Epsilon (n = 1). An excellent concordance was observed for the identification of all SARS-CoV-2 lineages evaluated.
Conclusions:
We present a flexible testing algorithm for the rapid detection of current and emerging SARS-CoV-2 VOC/VOIs, which can be easily adapted based on the local endemicity of specific variants.
