Development of Detection Antibody Targeting the Linear Epitope in SARS-CoV-2 Nucleocapsid Protein with Ultra-High Sensitivity.

The COVID-19 pandemic caused by SARS-CoV-2 highlighted the importance of reliable detection methods for disease control and surveillance. Optimizing detection antibodies by rational screening antigens would improve the sensitivity and specificity of antibody-based detection methods such as colloidal gold immunochromatography. In this study, we screened three peptide antigens with conserved sequences in the N protein of SARS-CoV-2 using bioinformatical and structural biological analyses. Antibodies that specifically recognize these peptides were prepared. The epitope of the peptide that had the highest binding affinity with its antibody was located on the surface of the N protein, which was favorable for antibody binding. Using the optimal antibody that can recognize this epitope, we developed colloidal gold immunochromatography, which can detect the N protein at 10 pg/mL. Importantly, this antibody could effectively recognize both the natural peptide antigen and mutated peptide antigen in the N protein, showing the feasibility of being applied in the large-scale population testing of SARS-CoV-2. Our study provides a platform with reference significance for the rational screening of detection antibodies with high sensitivity, specificity, and reliability for SARS-CoV-2 and other pathogens.

Clinical accuracy of instrument-based SARS-CoV-2 antigen diagnostic tests: a systematic review and meta-analysis.

BACKGROUND: During the COVID-19 pandemic, antigen diagnostic tests were frequently used for screening, triage, and diagnosis. Novel instrument-based antigen tests (iAg tests) hold the promise of outperforming their instrument-free, visually-read counterparts. Here, we provide a systematic review and meta-analysis of the SARS-CoV-2 iAg tests' clinical accuracy. METHODS: We systematically searched MEDLINE (via PubMed), Web of Science, medRxiv, and bioRxiv for articles published before November 7th, 2022, evaluating the accuracy of iAg tests for SARS-CoV-2 detection. We performed a random effects meta-analysis to estimate sensitivity and specificity and used the QUADAS-2 tool to assess study quality and risk of bias. Sub-group analysis was conducted based on Ct value range, IFU-conformity, age, symptom presence and duration, and the variant of concern. RESULTS: We screened the titles and abstracts of 20,431 articles and included 114 publications that fulfilled the inclusion criteria. Additionally, we incorporated three articles sourced from the FIND website, totaling 117 studies encompassing 95,181 individuals, which evaluated the clinical accuracy of 24 commercial COVID-19 iAg tests. The studies varied in risk of bias but showed high applicability. Of 24 iAg tests from 99 studies assessed in the meta-analysis, the pooled sensitivity and specificity compared to molecular testing of a paired NP swab sample were 76.7% (95% CI 73.5 to 79.7) and 98.4% (95% CI 98.0 to 98.7), respectively. Higher sensitivity was noted in individuals with high viral load (99.6% [95% CI 96.8 to 100] at Ct-level ≤ 20) and within the first week of symptom onset (84.6% [95% CI 78.2 to 89.3]), but did not differ between tests conducted as per manufacturer's instructions and those conducted differently, or between point-of-care and lab-based testing. CONCLUSION: Overall, iAg tests have a high pooled specificity but a moderate pooled sensitivity, according to our analysis. The pooled sensitivity increases with lower Ct-values (a proxy for viral load), or within the first week of symptom onset, enabling reliable identification of most COVID-19 cases and highlighting the importance of context in test selection. The study underscores the need for careful evaluation considering performance variations and operational features of iAg tests.

Estimating SARS-CoV-2 infection probabilities with serological data and a Bayesian mixture model.

The individual results of SARS-CoV-2 serological tests measured after the first pandemic wave of 2020 cannot be directly interpreted as a probability of having been infected. Plus, these results are usually returned as a binary or ternary variable, relying on predefined cut-offs. We propose a Bayesian mixture model to estimate individual infection probabilities, based on 81,797 continuous anti-spike IgG tests from Euroimmun collected in France after the first wave. This approach used serological results as a continuous variable, and was therefore not based on diagnostic cut-offs. Cumulative incidence, which is necessary to compute infection probabilities, was estimated according to age and administrative region. In France, we found that a "negative" or a "positive" test, as classified by the manufacturer, could correspond to a probability of infection as high as 61.8% or as low as 67.7%, respectively. "Indeterminate" tests encompassed probabilities of infection ranging from 10.8 to 96.6%. Our model estimated tailored individual probabilities of SARS-CoV-2 infection based on age, region, and serological result. It can be applied in other contexts, if estimates of cumulative incidence are available.

Taqman PACMAN: a simple molecular approach for positive rapid antigen test confirmation during periods of low prevalence.

Antigen-based rapid diagnostic tests (Ag-RDTs) were widely deployed to enhance SARS-CoV-2 testing capacity during the COVID-19 pandemic. Consistent with national guidance for low prevalence settings, positive Ag-RDTs were confirmed using nucleic acid amplification tests (NAATs) to avoid false positive results. However, increasing demands for positive Ag-RDT confirmation competed with other testing priorities in clinical laboratories. This work hypothesized that real-time RT-PCR without nucleic acid extraction (NAE) would be sufficiently sensitive to support positive Ag-RDT confirmation. Ag-RDT and NAAT results from community-based asymptomatic testing sites prior to the omicron variant wave were compared to calculate the weekly false positive rate (FPR) and false detection rate (FDR). Real-time RT-PCR was compared with and without NAE using 752 specimens previously tested positive for SARS-CoV-2 using commercial NAATs and 344 specimens from Ag-RDT-positive individuals. The impact of SARS-CoV-2 prevalence on laboratory resources required to sustain Ag-RDT confirmation was modeled for the RT-PCR with and without NAE. Overall, FPR was low [0.07% (222/330,763)] in asymptomatic testing sites, but FDR was high [30.7% (222/724)]. When RT-PCR was compared with and without NAE, 100% concordance was obtained with NAAT-positive specimens, including those from Ag-RDT-positive individuals. NAE-free RT-PCR significantly reduced time to results, human resources, and overall costs. A 30.7% FDR reaffirms the need for NAAT-based confirmation of positive Ag-RDT results during low SARS-CoV-2 prevalence. NAE-free RT-PCR was shown to be a simple and cost-sparing NAAT-based solution for positive Ag-RDT confirmation, and its implementation supported data-driven broader Ag-RDT deployment into communities, workplaces, and households. IMPORTANCE: Rapid antigen testing for SARS-CoV-2 was widely deployed during the COVID-19 pandemic. In settings of low prevalence, national guidance recommends that positive antigen test results be confirmed with molecular testing. Given the high testing burden on clinical laboratories during the COVID-19 pandemic, the high volume of positive antigen tests submitted for confirmatory testing posed challenges for laboratory workflow. This study demonstrated that a simple PCR method without prior nucleic acid purification is an accurate and cost-effective solution for positive rapid antigen test confirmation. Implementing this method allowed molecular confirmatory testing for positive antigen tests to be sustained as antigen testing was expanded into large populations such as workplaces, schools, and households.

Accelerated Development of a COVID-19 Lateral Flow Test in an Academic Setting: Lessons Learned.

ConspectusThe COVID-19 pandemic further demonstrated the need for usable, reliable, and cost-effective point-of-care diagnostics that can be broadly deployed, ideally for self-testing at home. Antigen tests using more-detectable reporter labels (usually at the cost of reader complexity) achieve better diagnostic sensitivity, supporting the value of higher-analytical-sensitivity reporter technologies in lateral flow.We developed a new approach to simple, inexpensive lateral flow assays (LFAs) of great sensitivity, based on the glow stick peroxyoxalate chemistry widely used in emergency settings and in children's toys. At the peak of the COVID-19 pandemic, we had the opportunity to participate in the pandemic-driven NIH Rapid Acceleration of Diagnostics (RADx) initiative aiming to develop a deployable lateral flow diagnostic for SARS-CoV-2 nucleoprotein based on our novel glow stick-inspired light-emitting reporter technology. During this project, we screened more than 250 antibody pairs for analytical sensitivity and specificity directly in LFA format, using recombinant nucleoprotein and then gamma-irradiated virions spiked into negative nasal swab extracts. Membranes and other LFA materials and swabs and extraction reagent components also were screened and selected. Optimization of conjugate preparation and spraying as well as pretreatment/conditioning of the sample pad led to the final optimized LFA strip. Technology development also included optimization of excitation liquid enclosed in disposable droppers, design of a custom cartridge and smartphone-based reader, and app development, even a prototype reader usable with any mobile phone. Excellent preclinical performance was first demonstrated with contrived samples and then with leftover clinical samples. Moving beyond traditional academic focus areas, we were able to establish a quality management system (QMS), produce large numbers of customized LFA cassettes by contract injection molding, build in-house facilities to assemble and store thousands of complete tests for verification and validation and usability studies, and source kitting/packaging services and quality standard reagents and build partnerships for clinical translation, regulatory guidance, scale up, and market deployment. We were not able to bring this early stage technology to the point of commercialization within the limited time and resources available, but we did achieve strong proof-of-concept and advance translational aspects of the platform including initial high-performance LFAs, reading by the iPhone app using only a $2 plastic dark box with no lens, and convenient, usable excitation liquid packaging in droppers manufacturable in very large numbers.In this Account, we aim to provide a concise overview of our 18-month sprint toward the practical development of a deployable antigen lateral flow assay under pandemic conditions and the challenges and successes experienced by our team. We highlight what it takes to coach a technically savvy but commercially inexperienced academic team through the accelerated translation of an early stage technology into a useful product. Finally, we provide a guided tutorial and workflow to empower others interested in the rapid development of translatable LFAs.

SARS-CoV-2 Viral Shedding and Rapid Antigen Test Performance - Respiratory Virus Transmission Network, November 2022-May 2023.

As population immunity to SARS-CoV-2 evolves and new variants emerge, the role and accuracy of antigen tests remain active questions. To describe recent test performance, the detection of SARS-CoV-2 by antigen testing was compared with that by reverse transcription-polymerase chain reaction (RT-PCR) and viral culture testing during November 2022-May 2023. Participants who were enrolled in a household transmission study completed daily symptom diaries and collected two nasal swabs (tested for SARS-CoV-2 via RT-PCR, culture, and antigen tests) each day for 10 days after enrollment. Among participants with SARS-CoV-2 infection, the percentages of positive antigen, RT-PCR, and culture results were calculated each day from the onset of symptoms or, in asymptomatic persons, from the date of the first positive test result. Antigen test sensitivity was calculated using RT-PCR and viral culture as references. The peak percentage of positive antigen (59.0%) and RT-PCR (83.0%) results occurred 3 days after onset, and the peak percentage of positive culture results (52%) occurred 2 days after onset. The sensitivity of antigen tests was 47% (95% CI = 44%-50%) and 80% (95% CI = 76%-85%) using RT-PCR and culture, respectively, as references. Clinicians should be aware of the lower sensitivity of antigen testing compared with RT-PCR, which might lead to false-negative results. This finding has implications for timely initiation of SARS-CoV-2 antiviral treatment, when early diagnosis is essential; clinicians should consider RT-PCR for persons for whom antiviral treatment is recommended. Persons in the community who are at high risk for severe COVID-19 illness and eligible for antiviral treatment should seek testing from health care providers with the goal of obtaining a more sensitive diagnostic test than antigen tests (i.e., an RT-PCR test).

SARS-CoV-2 seroprevalence among children in Greece during Omicron variant period.

The Omicron variant is associated with increased transmissibility, but evidence about the impact of Omicron in seropositivity of children is limited. This study aims to evaluate SARS-CoV-2 seroprevalence in children during the different variants' subperiods. A prospective multicenter seroprevalence study was conducted in 7 University public hospitals in Greece from November 2021 to August 2022 (3 subperiods: November 2021-February 2022, March 2022-May 2022, June 2022-August 2022). Children from different age groups, admitted to the hospital or examined in outpatient clinics for reasons other than COVID-19 were enrolled. Neutralizing antibodies (Nabs), anti-Spike (anti-S) and anti-nucleocapsid (anti-N) SARS-CoV-2 IgG in serum were evaluated. A total of 2127 children (males:57,2%; median age:4,8years) were enrolled. Anti-N IgG seropositivity increased from 17,8% in the first sub-period to 40,7% in the second sub-period and then decreased in the third sub-period (36,7%). Anti-S IgG seropositivity appeared to have an increasing trend over the study period, starting from 34,8% and reaching 80,7%. Children aged 1-4 years old have significantly higher anti-N IgG titers compared to children aged 0-1 years old (p < 0,001). Infants have significantly lower anti-S IgG titers compared to all other age groups (p < 0,001). Immunocompromised children and infants have the lowest seropositivity for NAbs.Conclusions During the Omicron period, seropositivity significantly increased, as a result of higher transmissibility. Neonates and infants have lower antibody titers compared to other age groups, while young children aged 1-4 years old present higher antibody titers, suggesting that this age group may mount a higher antibody response. Continuous surveillance seroprevalence studies are needed in children, in order to identify the true extent of SARS-CoV-2 and guide the planning of adequate public health measures.

WHAT IS KNOWN: • Seroprevalence surveys among children may be extremely useful, in order to properly monitor the immunity, either natural or acquired, through the quantification of IgG antibodies and to plan further immunization policies. • There are variations in the seroprevalence of COVID-19 between the different periods, according to the vaccination rates, the type of circulating variant and the transmissibility of the virus. • The Omicron variant is associated with increased transmissibility, but evidence about the impact of Omicron in seropositivity of children is limited. WHAT IS NEW: • In this large multicenter seroepidemiological study, SARS-CoV-2 seroprevalence rate in children is higher during the Omicron period in comparison to the previous pandemic waves, due to the high transmissibility of the virus and the increased rates of reinfection. • Neonates and infants have lower antibody titers compared to other age groups, while young children aged 1­4 years old present higher antibody titers, indicating that the children of this age group mount a higher antibody response. • This study provides essential information about immunity and the level of protection in the pediatric population, as neutralizing antibodies were evaluated, in addition to the anti-N and anti-S IgG antibodies.

Nucleocapsid and Spike Protein-Based Anti-SARS-CoV-2 Assay Performance in the Minority and Rural Coronavirus Insights Study: Characteristics of Socioeconomically Disadvantaged Populations with Health Disparities.

BACKGROUND: COVID-19 has had a devastating impact on Black, Hispanic, and other underserved, disadvantaged populations. Here anti-SARS-CoV-2 tests are characterized in disadvantaged patients to examine equivalence in US populations. METHODS: Underserved participant adults (age > 18 years) were enrolled before the availability of SARS-CoV-2 vaccines in Federal Qualified Health Centers in California, Florida, Louisiana, Illinois, and Ohio and contributed samples to the Minority and Rural Coronavirus Insights Study (MRCIS). A subset coined the MRCIS SARS-CoV-2 Antibody Cohort of 2365 participants was tested with the Roche Anti-SARS-CoV-2 assay (Cobas e601). Five hundred ninety-five of these were also tested with the Ortho Clinical Diagnostics VITROS Anti-SARS-CoV-2 IgG assay (VITROS-5600); 1770 were also tested with the Abbott ARCHITECT SARS-CoV-2 IgG assay (ARCHITECT-2000). Assay-specific cutoffs classified negative/positive results. RESULTS: Eight point four percent (199/2365) of the MRCIS SARS-CoV-2 Antibody Cohort was SARS-CoV-2 RNA positive at enrollment. Agreement between the Ortho/Roche and the Abbott/Roche antibody testing did not vary by enrollment RNA status. The Ortho (anti-spike protein) vs Roche (anti-nucleocapsid protein) comparison agreed substantially: kappa = 0.63 (95% CI: 0.57-0.69); overall agreement, 83%. However, agreement was even better for the Abbott vs Roche assays (both anti-nucleocapsid protein tests): kappa = 0.85 (95% CI: 0.81-0.87); overall agreement, 95%. Anti-SARS-CoV-2 comparisons stratified by demographic criteria demonstrated no significant variability in agreement by sex, race/ethnicity, or age. CONCLUSIONS: Analytical agreement is 96.4% for anti-spike-protein vs anti-nucleocapsid-protein comparisons. Physiologically, seroreversion of anti-nucleocapsid reactivity after infection occurred in the disadvantaged population similarly to general populations. No anti-SARS-CoV-2 assays included demonstrated a clinically significant difference due to the demographics of the disadvantaged MRCIS SARS-CoV-2 Antibody Cohort.

COVID-19 treatment guidelines panel: Coronavirus diseases 2019 (COVID-19) treatment guidelines

The COVID-19 Treatment Guidelines were developed to provide clinicians with guidance on caring for patients with COVID-19. Because clinical information about the optimal management of COVID-19 is evolving quickly, these Guidelines are updated frequently to reflect newly published data and other authoritative information.

Diagnostic Accuracy of SARS-CoV-2 Rapid Antigen Testing in a Pediatric Population: Children are Not Little Adults.

BACKGROUND: Information on the use of antigen-based SARS-CoV-2 rapid antigen tests (RAT) in children is limited. RATs have been used more frequently, because they are easily applicable, inexpensive, and can be easily performed at home without the need for special equipment. This study was designed to assign the diagnostic test accuracy of the SARS-CoV-2 RAT in daily clinical practice in children. METHODS: One thousand forty-two pediatric patients (aged 1 month - 18 years) who presented to the pediatric COVID-19 outpatient clinic of our hospital between January 2021 and June 2022 and met the inclusion criteria were included in this study. Nasopharyngeal samples were taken from the patients at the same visit, first for reverse transcription polymerase chain reaction (RT-PCR) and then for RAT. RESULTS: The data of all patients with RT-PCR positivity (n = 314) and additionally 14 patients with RAT positivity were analyzed in depth. The overall sensitivity and specificity were 62.1% (95% CI: 56.4 - 67.4) and 98% (95% CI: 96.7 - 98.9), respectively. The positive predictive value (PPV) and the negative predictive value (NPV) in this pediatric study were 93.3% and 85.7% (95% CI: 88.7 - 96.1 and 83.1 - 87.9), respectively. Considering the Ct values, which are indirect indicators of viral load, it was observed that the sensitivity of the rapid antigen test increased at low Ct values. The sensitivity increased to 75.1% (95% CI: 67.9 - 81.1) in patients with a Ct value of < 25. The specificity was 92.7% (95% CI: 90.7 - 94.3), PPV was 67.8% (95% CI: 60.7 - 67.8) and the NPV was 94.7% (95% CI: 93.0 - 96.1) in patients with a Ct value < 25. When the patients were evaluated according to their symptomatic/asymptomatic status, the difference between the diagnostic performance of the RAT test was found to be statistically significant (p = 0.006). CONCLUSIONS: In our study, it was found that the sensitivity of RATs in pediatric patients was lower than in adults. Our results also showed that children are not small adults, and the sensitivity of the test was higher, especially in symptomatic patients and patients with high viral load. To obtain more accurate results, we believe that performing the test in the first 3 days of symptoms will give more accurate results.

Implementing SARS-CoV-2 antigen testing scale-up in Rwanda: retrospective analysis of national programme data and qualitative findings.

OBJECTIVES: Reverse transcriptase PCR is the most sensitive test for SARS-CoV-2 diagnosis. However, the scale-up of these tests in low-income and middle-income countries (LMICs) has been limited due to infrastructure and cost. Antigen rapid diagnostic tests are an alternative option for diagnosing active infection that may allow for faster, easier, less expensive and more widespread testing. We compared the implementation of antigen and PCR testing programmes in Rwanda. DESIGN: We retrospectively reviewed routinely collected PCR and antigen testing data for all reported tests conducted nationally. We administered semiquantitative surveys to healthcare workers (HCWs) involved in COVID-19 testing and care and clients receiving antigen testing. SETTING: Rwanda, November 2020-July 2021. PARTICIPANTS: National SARS-CoV-2 testing data; 49 HCWs involved in COVID-19 testing and care; 145 clients receiving antigen testing. INTERVENTIONS: None (retrospective analysis of programme data). PRIMARY AND SECONDARY OUTCOME MEASURES: Test volumes, turnaround times, feasibility and acceptability of antigen testing. RESULTS: Data from 906 204 antigen tests and 445 235 PCR tests were included. Antigen testing increased test availability and case identification compared with PCR and had a median results return time of 0 days (IQR: 0-0). In contrast, PCR testing time ranged from 1 to 18 days depending on the sample collection site/district. Both HCWs and clients indicated that antigen testing was feasible and acceptable. Some HCWs identified stockouts and limited healthcare staff as challenges. CONCLUSIONS: Antigen testing facilitated rapid expansion and decentralisation of SARS-CoV-2 testing across lower tier facilities in Rwanda, contributed to increased case identification, reduced test processing times, and was determined to be feasible and acceptable to clients and providers. Antigen testing will be an essential component of SARS-CoV-2 test and treat programmes in LMICs.

Performance of 20 rapid antigen detection tests to detect SARS-CoV-2 B.1.617.2 (Delta) and B.1.1.529 (Omicron) variants using a clinical specimen panel from January 2022, Berlin, Germany.

BackgroundThere are conflicting reports on the performance of rapid antigen detection tests (RDT) in the detection of the SARS-CoV-2 Omicron (B.1.1.529) variant; however, these tests continue to be used frequently to detect potentially contagious individuals with high viral loads.AimThe aim of this study was to investigate comparative detection of the Delta (B.1.617.2) and Omicron variants by using a selection of 20 RDT and a limited panel of pooled combined oro- and nasopharyngeal clinical Delta and Omicron specimens.MethodsWe tested 20 CE-marked RDT for their performance to detect SARS-CoV-2 Delta and Omicron by using a panel of pooled clinical specimens collected in January 2022 in Berlin, Germany.ResultsWe observed equivalent detection performance for Delta and Omicron for most RDT, and sensitivity was widely in line with our previous pre-Delta/Omicron evaluation. Some variation for individual RDT was observed either for Delta vs Omicron detection, or when compared with the previous evaluation, which may be explained both by different panel sizes resulting in different data robustness and potential limitation of batch-to-batch consistency. Additional experiments with three RDT using non-pooled routine clinical samples confirmed comparable performance to detect Delta vs Omicron. Overall, RDT that were previously positively evaluated retained good performance also for Delta and Omicron variants.ConclusionOur findings suggest that currently available RDT are sufficient for the detection of SARS-CoV-2 Delta and Omicron variants.

Binding and Avidity Signatures of Polyclonal Sera From Individuals With Different Exposure Histories to Severe Acute Respiratory Syndrome Coronavirus 2 Infection, Vaccination, and Omicron Breakthrough Infections.

BACKGROUND: The number of exposures to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and to vaccine antigens affect the magnitude and avidity of the polyclonal response. METHODS: We studied binding and avidity of different antibody isotypes to the spike, the receptor-binding domain (RBD), and the nucleoprotein (NP) of wild-type (WT) and BA.1 SARS-CoV-2 in convalescent, mRNA vaccinated and/or boosted, hybrid immune individuals and in individuals with breakthrough cases during the peak of the BA.1 wave. RESULTS: We found an increase in spike-binding antibodies and antibody avidity with increasing number of exposures to infection and/or vaccination. NP antibodies were detectible in convalescent individuals and a proportion of breakthrough cases, but they displayed low avidity. Omicron breakthrough infections elicited high levels of cross-reactive antibodies between WT and BA.1 antigens in vaccinated individuals without prior infection directed against the spike and RBD. The magnitude of the antibody response and avidity correlated with neutralizing activity against WT virus. CONCLUSIONS: The magnitude and quality of the antibody response increased with the number of antigenic exposures, including breakthrough infections. However, cross-reactivity of the antibody response after BA.1 breakthroughs, was affected by the number of prior exposures.

ABO blood types and SARS-CoV-2 infection assessed using seroprevalence data in a large population-based sample: the SAPRIS-SERO multi-cohort study.

ABO blood type has been reported as a potential factor influencing SARS-CoV-2 infection, but so far mostly in studies that involved small samples, selected population and/or used PCR test results. In contrast our study aimed to assess the association between ABO blood types and SARS-CoV-2 infection using seroprevalence data (independent of whether or not individuals had symptoms or sought for testing) in a large population-based sample. Our study included 67,340 French participants to the SAPRIS-SERO multi-cohort project. Anti-SARS-CoV-2 antibodies were detected using ELISA (targeting the proteins spike (S) and nucleocapsid (NP)) and seroneutralisation (SN) tests on dried blood spots collected in May-November 2020. Non-O individuals (and especially types A and AB) were more likely to bear anti SARS-CoV-2 antibodies (ELISA-S, 2964 positive cases: ORnon-Ovs.O = 1.09[1.01-1.17], ORAvs.O = 1.08[1.00-1.17]; ELISA-S/ELISA-NP/SN, 678 triple positive cases: ORnon-Ovs.O = 1.19 [1.02-1.39], ORAvs.O = 1.19[1.01-1.41], ORABvs.O = 1.43[1.01-2.03]). Hence, our results provided additional insights into the dynamic of SARS-CoV-2 infection, highlighting a higher susceptibility of infection for individuals of blood types A and AB and a lesser risk for blood type O.

A comparison of SARS-CoV-2 rapid antigen testing with realtime RT-PCR among symptomatic and asymptomatic individuals.

BACKGROUND: Identification of SARS-CoV-2 positive patients with rapid and cost-effective test methods is the key for isolating infected individuals, interrupting the transmission chain, and thus, containment of the CoVID-19 disease. In this regard, Rapid Antigen Test (RAT) plays an important role at point of care testing but the low sensitivity attributing towards escape of positive cases is reported as a major disadvantage of RAT which led us to evaluate a RAT kit among symptomatic and asymptomatic individuals suspected of CoVID-19. METHODS: We analyzed 329 parallel nasopharyngeal swabs for RAT (Zydus Cadila, India) at the point of collection in a hospital-based facility and RealTime RT-PCR in the laboratory. The performance parameters were analyzed by evaluating the specificity, sensitivity, Negative Predictive Value (NPV), Positive Predictive Value (PPV), and Kappa coefficient. RESULTS: The sensitivity and specificity were found to be 75.17% and 98.89% respectively. Positive Predictive value was 98.25% and the negative predictive value was 82.79%. The accuracy between the two techniques was found to be 88.14% with a kappa coefficient of 0.756 (SE: 0.036 and CI at 95%: 0.686 to 0.826) with a good strength of agreement (0.61-0.80) between the two testing techniques. Among the false-negative cases, 22 (59.5%) were asymptomatic having the Cycle Threshold (Ct) range 27 to 32.9 including 12 cases with a history of close contact with the known positive cases (i.e. household contact). The remaining 15 cases (40.5%) were symptomatic having low to moderate Ct values. CONCLUSION: It is observed from the results that the false negative result for symptomatic individuals is a matter of concern as it was noted in 4 cases of our study subjects who required hospitalisation later. Also the positives among asymptomatic contacts are important from epidemiological point of view for isolation and curtailing the infection from spreading in a community. These results support the fact that RAT showing sensitivity below 80% can be used for mass screening purposes with provision for additional testing in case of false negative with symptomatic individuals. Also false-negative results should be interpreted cautiously considering the epidemiological link as well as the clinical condition of the patients.

Dried blood spots are a valid alternative to venipuncture for COVID-19 antibody testing.

BACKGROUND: Serologic analysis is an important tool towards assessing the humoral response to COVID-19 infection and vaccination. Numerous serologic tests and platforms are currently available to support this line of testing. Two broad antibody testing categories are point-of-care lateral flow immunoassays and semi-quantitative immunoassays performed in clinical laboratories, which typically require blood collected from a finger-stick and a standard venipuncture blood draw, respectively. This study evaluated the use of dried blood spot (DBS) collections as a sample source for COVID-19 antibody testing using an automated clinical laboratory test system. METHODS: Two hundred and ninety-four participants in the BLAST COVID-19 seroprevalence study (NCT04349202) were recruited at the time of a scheduled blood draw to have an additional sample taken via finger stick as a DBS collection. Using the EUROIMMUN assay to assess SARS-CoV-2 anti-spike IgG status, DBS specimens were tested on 7, 14, 21, and 28 days post- collection and compared to the reference serum sample obtained from a blood draw for the BLAST COVID-19 study. RESULTS: SARS-CoV-2 anti-spike IgG status from DBS collections demonstrated high concordance with serum across all time points (7-28 days). However, the semi-quantitative value from DBS collections was lower on average than that from serum, resulting in increased uncertainty around the equivocal-to-positive analytical decision point. CONCLUSIONS: DBS collections can be substituted for venipuncture when assaying for COVID-19 IgG antibody, with samples being stable for at least 28 days at room temperature. Finger-stick sampling can therefore be advantageous for testing large populations for SARS-CoV-2 antibodies without the need for phlebotomists or immediate processing of samples. We have high confidence in serostaus determination from DBS collections, although the reduced semi-quantitative value may cause some low-level positives to fall into the equivocal or even negative range.

Performance of High Throughput SARS-CoV-2 Antigen Testing Compared to Nucleic Acid Testing.

OBJECTIVE: Independent assessment of SARS-CoV-2 antigen (COV2Ag) tests remains important as varying performance between assays is common. We assessed the performance of a new high-throughput COV2Ag test compared to SARS-CoV-2 nucleic acid amplification tests (NAAT). METHODS: A total of 347 nasopharyngeal samples collected from January to October 2021 were assessed by NAAT as part of standard-of-care testing (CDC LDT or GeneXpert System, Cepheid) and COV2Ag using the ADVIA Centaur CoV2Ag assay (Siemens Healthineers). RESULTS: Among NAAT positive specimens we found 82.4% agreement and in NAAT negative specimens we found 97.3% agreement (overall agreement 85.6%). In symptomatic persons, COV2Ag agreed with NAAT 90.0% (n = 291), and in asymptomatic persons, 62.5% (n = 56). Agreement between positive NAAT and COV2Ag increased at lower cycle threshold (Ct) values. CONCLUSION: The COV2Ag assay exceeded the World Health Organization minimum performance requirements of ≥ 80% sensitivity and ≥ 97% specificity. The COV2Ag assay is helpful for large scale screening efforts due to high-throughput and reduced wait times.