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Challenges in the Detection of SARS-CoV-2: Evolution of the Lateral Flow Immunoassay as a Valuable Tool for Viral Diagnosis

Abstract Objectives This proof of concept study was aimed to validate the hypothesis that the time of positivization of SARS-CoV-2 self-performed rapid diagnostic tests (RDTs) may reflect the actual viral load in the specimen. Methods A SARS-CoV-2 positive sample with high viral load was diluted and concomitantly assayed with molecular assay (Xpert Xpress SARS-CoV-2) and RDT (COVID-VIRO ALL IN RDT). The (mean cycle threshold; Ct) values and RDT positivization times of these dilutions were plotted and interpolated by calculating the best fit. The parameters of this equation were then used for converting the positivization times into RDT-estimated SARS-CoV-2 Ct values in routine patient samples. Results The best fit between measured and RDT-estimated Ct values could be achieved with a 2-degree polynomial curve. The RDT-estimated Ct values exhibited high correlation (r=0.996) and excellent Deming fit (y=1.01 × x − 0.18) with measured Ct values. In 30 consecutive patients with positive RDT test, the correlation between RDT positivization time and measured Ct value was r=0.522 (p=0.003). The correlation of RDT-estimated and measured Ct values slightly improved to 0.577 (Deming fit: y=0.44 × x + 11.08), displaying a negligible bias (1.0; 95% CI, −0.2 to 2.2; p=0.105). Concordance of RDT-estimated and measured Ct values at the <20 cut-off was 80%, with 0.84 sensitivity and 0.73 specificity. Conclusions This proof of concept study demonstrates the potential feasibility of using RDTs for garnering information on viral load in patients with acute SARS-CoV-2 infection.

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Positivization time of a COVID-19 rapid antigen self-test predicts SARS-CoV-2 viral load: a proof of concept

The new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus has high infectivity, often masked by asymptomatic carriers, which allows it to spread rapidly and become a pandemic. Attempts to slow the pandemic at this stage depend on the ability to unmask asymptomatic carriers. The rapid diagnosis of active coronavirus disease 2019 (COVID-19) infection is one of the cornerstones of pandemic control, as the nasal cavity is the main gateway for SARS-CoV-2 entry and altered sense of smell is a feature of the current virus. In the present study, we therefore tested the olfactory threshold coupled with heart–lung parameters in subjects undergoing traditional molecular testing, resulting in a significantly different score between asymptomatic subjects and healthy controls. In total, 82% of asymptomatic positives showed olfactory impairment; of these, 46% had severe hyposmia and 7% had anosmia, while in the control 9% had severe hyposmia and 0% had anosmia, respectively, which agrees with heart rate, breathing rate, and blood pressure parameter variations. The olfactory test coupled with physiological parameters may help to identify asymptomatic people. In conclusion, our results suggest that most asymptomatic individuals could be unmasked by mass olfactory rapid threshold screening and then referred to traditional slower diagnostic tests.

/pdf/unmasking-the-asymptomatic-covid-19-a-nose-question-kz2varrl.pdf

Unmasking the ‘Asymptomatic’ COVID-19: A Nose Question

Abstract Background This article provides a critical literature review and pooled analysis of diagnostic accuracy of the fully-automated Siemens SARS-CoV-2 Antigen (CoV2Ag) chemiluminescent immunoassay for diagnosis of acute SARS-CoV-2 infections. Methods An electronic search was conducted in Scopus, PubMed and medRxiv using the keywords [“Siemens AND CoV2Ag”] OR [“Siemens AND SARS-CoV-2 AND antigen”] for capturing studies that investigated the accuracy of Siemens CoV2Ag for diagnosing acute SARS-CoV-2 infection against a reference SARS-CoV-2 molecular test. The retrieved information was used for constructing a 2 × 2 table and for calculating pooled diagnostic sensitivity, specificity, Summary Receiver Operating Characteristic Curve (SROC) and Agreement. This study followed the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) reporting checklist. Results Four studies totalling 1,310 respiratory samples (612 with high viral load) were finally included in our analysis. The cumulative area under the curve, accuracy, sensitivity, specificity, were 0.964 (95% CI, 0.957–0.971), 86.9% (95% CI, 84.9–88.7%), 0.79 (95% CI, 0.76–0.82) and 0.98 (95% CI, 0.96–0.99), respectively. The negative (NPV) and positive (PPV) predictive values were 0.77 (0.74–0.79) and 0.98 (95% CI, 0.96–99), respectively. The diagnostic sensitivity in samples with high viral load (i.e., Ct<29–30) was 0.95 (95% CI, 0.93–0.97). Conclusions The Siemens CoV2Ag fully-automated and high-throughput immunoassay approximates the minimum performance criteria for general SARS-CoV-2 antigen testing and displays excellent performance in samples with high viral load, thus representing a valuable screening solution for risk assessment in COVID-19 and for limiting viral spread.

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Diagnostic accuracy of Siemens SARS-CoV-2 Antigen (CoV2Ag) chemiluminescent immunoassay for diagnosing acute SARS-CoV-2 infection: a pooled analysis

This study evaluated the performances of immunoassays (LFIA and ELISA) designed for SARS-CoV-2 Antigen (Ag)-detection in nasopharyngeal (NP) and serum samples in comparison to RT-PCR. NP samples from patients with respiratory symptoms (183 RT-PCR-positive and 74 RT-PCR-negative samples) were collected from March to April and November to December 2020. Seroconversion and antigen dynamics were assessed by symptom onset and day of RT-PCR diagnosis. Serum samples from 87 COVID-19 patients were used to investigate the added value of Ag quantification, at diagnosis and during follow-up. The sensitivity of COVID-VIRO-LFIA on samples with Ct ≤ 33, considered as the contagious threshold, was 86% on NPs (CI 95%: 79–90.5) and 76% on serum samples (CI 95%: 59.4–88), with a specificity of 100%. Serum N-Ag was detected during active infection as early as day two from symptom onset, with a diagnostic sensitivity of 81.5%. Within one week of symptom onset, diagnostic sensitivity and specificity reached 90.9% (95% CI, 85.1%–94.6%) and 98.3% (95% CI, 91.1%–99.9%), respectively. Serum N-Ag concentration closely correlated with disease severity. Longitudinal analysis revealed the simultaneous increase of antibodies and decrease of N-Ag. Sensitivities of COVID-VIRO-LFIA and COV-QUANTO-ELISA tests on NP and serum samples were close to 80%. They are suitable COVID-19-laboratory diagnostic tests, particularly when blood samples are available, thus reducing the requirement for NP sampling, and subsequent PCR analysis. ELISA titers may help to identify patients at risk of poor outcomes.

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Clinical Added Value of SARS-CoV-2 Antigen Detection in Blood Samples

Coronavirus disease 2019 (COVID-19) initiated global health care challenges such as the necessity for new diagnostic tests. Diagnosis by real-time PCR remains the gold-standard method, yet economical and technical issues prohibit its use in points of care (POC) or for repetitive tests in populations. A lot of effort has been exerted in developing, using, and validating antigen-based tests (ATs). Since individual studies focus on few methodological aspects of ATs, a comparison of different tests is needed. Herein, we perform a systematic review and meta-analysis of data from articles in PubMed, medRxiv and bioRxiv. The bivariate method for meta-analysis of diagnostic tests pooling sensitivities and specificities was used. Most of the AT types for SARS-CoV-2 were lateral flow immunoassays (LFIA), fluorescence immunoassays (FIA), and chemiluminescence enzyme immunoassays (CLEIA). We identified 235 articles containing data from 220,049 individuals. All ATs using nasopharyngeal samples show better performance than those with throat saliva (72% compared to 40%). Moreover, the rapid methods LFIA and FIA show about 10% lower sensitivity compared to the laboratory-based CLEIA method (72% compared to 82%). In addition, rapid ATs show higher sensitivity in symptomatic patients compared to asymptomatic patients, suggesting that viral load is a crucial parameter for ATs performed in POCs. Finally, all methods perform with very high specificity, reaching around 99%. LFIA tests, though with moderate sensitivity, appear as the most attractive method for use in POCs and for performing seroprevalence studies.

https://www.mdpi.com/2075-4418/12/6/1388/pdf?version=1654667092

Helen Mavriki

Papers

Performance of Antigen Detection Tests for SARS-CoV-2: A Systematic Review and Meta-Analysis