Diagnostic Performance Evaluation of the GXT96 X3 Extraction System with the FluoroType® SARS-CoV-2 varID Q Assay for Detection and Variant Identification of SARS-CoV-2

Background: The continued evolution of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) created ongoing challenges for molecular diagnostics and variant surveillance. Reliable assays capable of maintaining diagnostic sensitivity across emerging variants while providing rapid variant information remain essential for clinical management and public health monitoring. This study evaluated the performance of the GXT96 X3 extraction kit in combination with the FluoroType® SARS-CoV-2 varID Q version 1.0 assay (Hain LifeScience SA (Pty) Ltd, South Africa) for the detection, semi-quantitative assessment, and variant characterization of SARS-CoV-2. Methods: A total of 220 samples were evaluated, including residual nasopharyngeal clinical specimens (n = 183), reference materials, and cultured SARS-CoV-2 virus dilutions. Residual specimens collected during multiple COVID-19 waves in South Africa (wild type, Beta, Delta, and Omicron) were compared against standard-of-care (SOC) molecular assays used for routine diagnosis. RNA extraction was performed using the automated GXT96 X3 platform, followed by amplification on the FluoroCycler® XT using the FluoroType® SARS-CoV-2 varID Q assay targeting RdRp and N genes, with additional spike gene mutation detection for variant identification. Diagnostic accuracy, agreement (Cohen’s kappa), precision, linearity, and limit of detection (LoD) were assessed. Results: The assay demonstrated a sensitivity of 98.4% (95% CI: 94.2–99.8) and specificity of 100% (95% CI: 95.9–100.0) compared with SOC assays, with an overall agreement of κ = 0.981. Precision analysis showed acceptable reproducibility with standard deviation ≤1.49 and coefficient of variation ≤3.83%. Regression analysis demonstrated strong linearity across dilution series (R² = 0.9882 for RdRp and 0.994 for N genes). The LoD was ≤100 copies/mL for the RdRp gene and 250 copies/mL for the N gene. Variant-associated spike mutations detected by the assay corresponded broadly with epidemiological wave patterns observed in South Africa. Conclusions: The GXT96 X3 extraction platform combined with the FluoroType® SARS-CoV-2 varID Q assay demonstrated high diagnostic accuracy, reproducibility, and reliable SARS-CoV-2 detection across a range of viral loads. The assay additionally provides rapid mutation-based variant information, supporting its utility for routine diagnostic testing and complementary variant surveillance.