Comparison between Nasal and Nasopharyngeal Swabs for SARS-CoV-2 Rapid Antigen Detection in an Asymptomatic Population, and Direct Confirmation by RT-PCR from the Residual Buffer.

Containment measures employed during the COVID-19 pandemic included prompt recognition of cases, isolation, and contact tracing. Bilateral nasal (NA) swabs applied to a commercial antigen-based rapid diagnostic test (Ag-RDT) offer a simpler and more comfortable alternative to nasopharyngeal (NP) collection; however, little is known about the sensitivity of this method in an asymptomatic population. Participants in community-based asymptomatic testing sites were screened for SARS-CoV-2 using an Ag-RDT with NP sampling. Positive individuals returned for confirmatory molecular testing and consented to repeating the Ag-RDT using a bilateral NA swab for comparison. Residual test buffer (RTB) from Ag-RDTs was subjected to real-time reverse transcription-PCR (RT-PCR). Of 123,617 asymptomatic individuals, 197 NP Ag-RDT-positive participants were included, with 175 confirmed positive by RT-PCR. Of these cases, 154 were identified from the NA swab collection with Ag-RDT, with a sensitivity of 88.0% compared to the NP swab collection. Stratifying results by RT-PCR cycle threshold demonstrated that sensitivity of the nasal collection method varied based on the cycle threshold (CT) value of the paired RT-PCR sample. RT-PCR testing on the RTB from the Ag-RDT using NP and NA swab collections resulted in 100.0% and 98.7% sensitivity, respectively. NA swabs provide an adequate alternative to NP swab collection for use with Ag-RDT, with the recognition that the test is most sensitive in specimens with high viral loads. With the high sensitivity of RT-PCR testing on RTB from Ag-RDT, a more streamlined approach to confirmatory testing is possible without recollection or use of paired collections strategies. IMPORTANCE Nasal swabbing for SARS-CoV-2 (COVID-19) comes with many benefits but is slightly less sensitive than traditional nasopharyngeal swabbing; however, confirmatory lab-based testing could be performed directly from the residual buffer from either sample type.

Avoiding False-Positive SARS-CoV-2 Rapid Antigen Test Results with Point-of-Care Molecular Testing on Residual Test Buffer.

Antigen-based rapid diagnostic tests (Ag-RDTs) have been widely used for the detection of SARS-CoV-2 during the coronavirus disease 2019 (COVID-19) pandemic. In settings of low disease prevalence, such as asymptomatic community testing, national guidelines recommend confirmation of positive Ag-RDT results with a nucleic acid amplification test (NAAT). This often requires patients to be recalled for repeat specimen recollection and subsequent testing in reference laboratories. This project assessed the use of a point-of-care molecular NAAT for SARS-CoV-2 detection (i.e., ID NOW), which was performed on-site at a volunteer-led asymptomatic community testing site on the residual test buffer (RTB) from positive Ag-RDTs. The ID NOW NAAT assay was performed on RTB from two Ag-RDTs: the Abbott Panbio and BTNX Rapid Response assays. Results of ID NOW were compared to real-time RT-PCR at a reference laboratory. Along with investigations into the clinical performance of ID NOW on RTB, analytical specificity was assessed with a panel of various respiratory organisms. Of the Ag-RDTs results evaluated, all 354 Ag-RDTs results characterized as true positives by RT-PCR were accurately identified with ID NOW testing of RTB. No SARS-CoV-2 detections by ID NOW were observed from 10 specimens characterized as false-positive Ag-RDTs, or from contrived specimens with various respiratory organisms. The use of on-site molecular testing on RTB provides a suitable option for rapid confirmatory testing of positive Ag-RDTs, thereby obviating the need for specimen recollection for molecular testing at local reference laboratories. IMPORTANCE During the COVID-19 pandemic, rapid antigen tests have been widely used for the detection of SARS-CoV-2. These simple devices allow rapid test results. However, false-positive results may occur. As such, individuals with positive rapid tests often must return to testing centers to have a second swab collected, which is then transported to a specialized laboratory for confirmation using molecular tests. As an alternative to requiring a repeat visit and a prolonged turn-around time for result confirmation, this project evaluated whether the leftover material from rapid antigen tests could be confirmed directly on a portable point-of-care molecular instrument. Using this approach, molecular confirmation of positive antigen tests could be performed in less than 15 min, and the results were equivalent to laboratory-based confirmation. This procedure eliminates the need for individuals to return to testing centers following a positive rapid antigen test and ensures accurate antigen test results through on-site confirmation.

Generation of False-Positive SARS-CoV-2 Antigen Results with Testing Conditions outside Manufacturer Recommendations: A Scientific Approach to Pandemic Misinformation.

Antigen-based rapid diagnostics tests (Ag-RDTs) are useful tools for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection. However, misleading demonstrations of the Abbott Panbio coronavirus disease 2019 (COVID-19) Ag-RDT on social media claimed that SARS-CoV-2 antigen could be detected in municipal water and food products. To offer a scientific rebuttal to pandemic misinformation and disinformation, this study explored the impact of using the Panbio SARS-CoV-2 assay with conditions falling outside manufacturer recommendations. Using Panbio, various water and food products, laboratory buffers, and SARS-CoV-2-negative clinical specimens were tested with and without manufacturer buffer. Additional experiments were conducted to assess the role of each Panbio buffer component (tricine, NaCl, pH, and Tween 20) as well as the impact of temperature (4°C, 20°C, and 45°C) and humidity (90%) on assay performance. Direct sample testing (without the kit buffer) resulted in false-positive signals resembling those obtained with SARS-CoV-2 positive controls tested under proper conditions. The likely explanation of these artifacts is nonspecific interactions between the SARS-CoV-2-specific conjugated and capture antibodies, as proteinase K treatment abrogated this phenomenon, and thermal shift assays showed pH-induced conformational changes under conditions promoting artifact formation. Omitting, altering, and reverse engineering the kit buffer all supported the importance of maintaining buffering capacity, ionic strength, and pH for accurate kit function. Interestingly, the Panbio assay could tolerate some extremes of temperature and humidity outside manufacturer claims. Our data support strict adherence to manufacturer instructions to avoid false-positive SARS-CoV-2 Ag-RDT reactions, otherwise resulting in anxiety, overuse of public health resources, and dissemination of misinformation. IMPORTANCE With the Panbio severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antigen test being deployed in over 120 countries worldwide, understanding conditions required for its ideal performance is critical. Recently on social media, this kit was shown to generate false positives when manufacturer recommendations were not followed. While erroneous results from improper use of a test may not be surprising to some health care professionals, understanding why false positives occur can help reduce the propagation of misinformation and provide a scientific rebuttal for these aberrant findings. This study demonstrated that the kit buffer's pH, ionic strength, and buffering capacity were critical components to ensure proper kit function and avoid generation of false-positive results. Typically, false positives arise from cross-reacting or interfering substances; however, this study demonstrated a mechanism where false positives were generated under conditions favoring nonspecific interactions between the two antibodies designed for SARS-CoV-2 antigen detection. Following the manufacturer instructions is critical for accurate test results.

Utility of the FebriDx point-of-care assay in supporting a triage algorithm for medical admissions with possible COVID-19: an observational cohort study.

OBJECTIVE: To evaluate a triage algorithm used to identify and isolate patients with suspected COVID-19 among medical patients needing admission to hospital using simple clinical criteria and the FebriDx assay. DESIGN: Retrospective observational cohort. SETTING: Large acute National Health Service hospital in London, UK. PARTICIPANTS: All medical admissions from the emergency department between 10 August 2020 and 4 November 2020 with a valid SARS-CoV-2 RT-PCR result. INTERVENTIONS: Medical admissions were triaged as likely, possible or unlikely COVID-19 based on clinical criteria. Patients triaged as possible COVID-19 underwent FebriDx lateral flow assay on capillary blood, and those positive for myxovirus resistance protein A (a host response protein) were managed as likely COVID-19. PRIMARY OUTCOME MEASURES: Diagnostic accuracy (sensitivity, specificity and predictive values) of the algorithm and the FebriDx assay using SARS-CoV-2 RT-PCR from nasopharyngeal swabs as the reference standard. RESULTS: 4.0% (136) of 3443 medical admissions had RT-PCR confirmed COVID-19. Prevalence of COVID-19 was 46% (80/175) in those triaged as likely, 4.1% (50/1225) in possible and 0.3% (6/2033) in unlikely COVID-19. Using a SARS-CoV-2 RT-PCR reference standard, clinical triage had sensitivity of 96% (95% CI 91% to 98%) and specificity of 61.5% (95% CI 59.8% to 63.1%), while the triage algorithm including FebriDx had sensitivity of 93% (95% CI 87% to 96%) and specificity of 86.4% (95% CI 85.2% to 87.5%). While 2033 patients were deemed not to require isolation using clinical criteria alone, the addition of FebriDx to clinical triage allowed a further 826 patients to be released from isolation, reducing the need for isolation rooms by 9.5 per day, 95% CI 8.9 to 10.2. Ten patients missed by the algorithm had mild or asymptomatic COVID-19. CONCLUSIONS: A triage algorithm including the FebriDx assay had good sensitivity and was useful to 'rule-out' COVID-19 among medical admissions to hospital.