The Rapid Antigen Detection Test for SARS-CoV-2 Underestimates the Identification of COVID-19 Positive Cases and Compromises the Diagnosis of the SARS-CoV-2 (K417N/T, E484K, and N501Y) Variants

Abstract

Timely detection of severe acute respiratory syndrome due to coronavirus 2 (SARS-CoV-2) by reverse transcription quantitative polymerase chain reaction (RT-qPCR) has been the gold- strategy for identifying positive cases during the current pandemic. However, faster and less expensive methodologies are also applied for the massive diagnosis of COVID-19. In this way, the rapid antigen test (RAT) is widely used. However, it is necessary to evaluate its detection efficiency considering the current pandemic context with the circulation of new viral variants. In this study, we evaluated the sensitivity and specificity of RAT (SD BIOSENSOR, South Korea), widely used for testing and SARS-CoV-2 diagnosis in Santiago of Chile. The RAT showed a 90% (amplification range of 20 ≤ Cq <25) and 10% (amplification range of 25 ≤ Cq <30) of positive SARS-CoV-2 cases identified previously by RT-qPCR. Importantly, a 0% detection was obtained for samples within a Cq value>30. In SARS-CoV-2 variant detection, RAT had a 42.8% detection sensitivity in samples with RT-qPCR amplification range 20 ≤ Cq <25 containing the single nucleotide polymorphisms (SNP) K417N/T, N501Y and E484K, associated with beta or gamma SARS-CoV-2 variants. This study alerts for the special attention that must be paid for the use of RAT at a massive diagnosis level, especially in the current scenario of appearance of several new SARS-CoV-2 variants which could generate false negatives and the compromise of possible viral outbreaks.

Keywords: COVID-19 diagnosis; COVID-19 false negative rapid test for COVID-19 diagnosis; SARS-CoV-2 detection; pandemic control strategies; rapid antigen test.

Figure 1

Sensitivity evaluation of the rapid antigen test (RAT; SD BIOSENSOR, South Korea) at different ranges of Cq values for ancestral strain of SARS-CoV-2 positive samples diagnosed by RT-qPCR. SARS-CoV-2 detection by RAT from RT-qPCR positive samples with Cq value between 20 ≤ Cq <25, 25 ≤ Cq <30, 30 ≤ Cq <35, and Cq≥35, respectively. All samples were positive by RT-qPCR. Table shows: the RFU (relative fluorescence units) and Cq value for the viral ORF1ab probe (n = 10 samples per Cq range).

Figure 2

SARS-CoV-2 Variant detection by RAT. RAT detection of SARS-CoV-2 variants which contain K417N/T, E484K, and N501Y mutations, from RT-qPCR positive samples with Cq value between 20≥Cq <25, 25≥Cq <30, 30≥Cq <35. All samples were positive by RT-qPCR. Table shows: the RFU (relative fluorescence units) and Cq value for the viral ORF1ab probe (n = 16).

Figure 3

Summary of the results obtained for the analysis of the 55 nasopharyngeal swab samples (NPSs) evaluated by Rapid Antigen Test (RAT). The 55 NPSs were diagnosed as positive for SARS-CoV-2 using the ORF1ab probe following a one-step strategy. From them, 40 NPSs showed no mutations for SARS-CoV-2 (black arrow) (Group 1). By contrast, in the other 15 positive samples we identified the SARS-CoV-2 variants K417N/T, E484K, and N501Y (gray arrows) (Group 2). Each one of these groups were used to evaluate the sensitivity of the Rapid Antigen Test (RAT) according to the Cq ranges previously obtained by RT-qPCR. The group 1 showed a 90% of positive diagnosis using the RAT. However, the group 2 showed only a 42.8% of positive diagnosis. Importantly, all of them were grouped in the 20 ≤ Cq <25 interval.