Lessons learned: drive-through COVID-19 clinic testing during an adaptive epidemic response and a point-of-care test assessment of a computer-read rapid lateral flow immunoassay with fluorescence-based detection.

Background. The COVID-19 pandemic demonstrated a need for robust SARS-CoV-2 test evaluation infrastructure to underpin biosecurity and protect the population during a pandemic health emergency.Gap statement. The first generation of rapid antigen tests was less accurate than molecular methods due to their inherent sensitivity and specificity shortfalls, compounded by the consequences of self-testing. This created a need for more accurate point-of-care SARS-CoV-2 detection methods.Aim. Here we present the lessons-learned during the COVID-19 emergency response in Western Australia including the detailed set-up, evaluation and operation of rapid antigen test in a state-run drive-through sample collection service during the COVID-19 pandemic after the strict border shutdown ended.Methods. We report a conformity assessment of a novel, second-generation rapid antigen test (Virulizer) comprising a technician-operated rapid lateral flow immunoassay with fluorescence-based detection.Results. The Virulizer rapid antigen test demonstrated up to 100% sensitivity (95% CI: 61.0-100%), 91.94% specificity (95% CI: 82.5-96.5%) and 92.65% accuracy when compared to a commercial PCR assay method. Wide confidence intervals in our series reflect the limits of small sample size. Nevertheless, the Virulizer assay performance was well-suited to point-of-care screening for SARS-CoV-2 in a drive-through clinic setting.Conclusion. The adaptive evaluation process necessary under changing pandemic conditions enabled assessment of a simple sample collection and point-of-care testing process, and showed how this system could be rapidly deployed for SARS-CoV-2 testing, including to regional and remote settings.

SARS-CoV-2 Seropositivity in Nursing Home Staff and Residents during the First SARS-CoV-2 Wave in Flanders, Belgium.

(1) Background: early in the COVID-19 pandemic, reverse transcription polymerase chain reaction (RT-PCR) testing was limited. Assessing seroprevalence helps understand prevalence and reinfection risk. However, such data are lacking for the first epidemic wave in Belgian nursing homes. Therefore, we assessed SARS-CoV-2 seroprevalence and cumulative RT-PCR positivity in Belgian nursing homes and evaluated reinfection risk. (2) Methods: we performed a cross-sectional study in nine nursing homes in April and May 2020. Odds ratios (ORs) were calculated to compare the odds of (re)infection between seropositive and seronegative participants. (3) Results: seroprevalence was 21% (95% CI: 18-23): 22% (95% CI: 18-25) in residents and 20% (95% CI: 17-24) in staff. By 20 May 2020, cumulative RT-PCR positivity was 16% (95% CI: 13-21) in residents and 8% (95% CI: 6-12) in staff. ORs for (re)infection in seropositive (compared to seronegative) residents and staff were 0.22 (95% CI: 0.06-0.72) and 3.15 (95% CI: 1.56-6.63), respectively. (4) Conclusion: during the first wave, RT-PCR test programmes underestimated the number of COVID-19 cases. The reinfection rate in residents was 3%, indicating protection, while it was 21% in staff, potentially due to less cautious health behaviour. Future outbreaks should use both RT-PCR and serological testing for complementary insights into transmission dynamics.

Evaluation of COVID-19 rapid antigen test against polymerase chain reaction test in immunocompromised patients.

On the 11th of March 2020, the world faced a new global pandemic, COVID-19 which is a disease caused by the novel coronavirus, it had multiple devastating outcomes on multiple sectors along with significant rates of mortality. These challenges encouraged the development of multiple testing methods, as well as anti-viral medications such as Molnupiravir, as well as evaluating the efficacy of available medications against it, like; Azithromycin, Ritonavir and Hydroxychloroquine. Vaccination against COVID-19 forged into a significant challenge, few months ensuing the first case of SARS-CoV-2, which was diagnosed in December 2019, in Wuhan-China, thus, multiple vaccines were approved for use around the world to combat this pandemic. Our study includes a sample of 556 oncology patients at Augusta Victoria Hospital in Jerusalem, all patients were tested using Panbio rapid antigen test and Allplex PCR Assay. The main objective was to study the sensitivity and specificity of Rapid antigen test, which contributes to a faster isolation call and management of infected patients, thus decreasing the risk on spread to other patients and health care. Patients were categorized based on two factors: Ct range and age group and studying their possible effect on false-negative results. Patients with Ct value less than 20, had the highest detection rate which is consistent with other studies in the literature. The sensitivity and specificity of Panbio Rapid Antigen testing were of 69.9% and 100%, respectively. A correlation between age group and false negative results could not be made, but a correlation between Ct value and false negative result was noticed, Ct value was directly related to false negative results. P-value of 0.007 indicated that results were statistically significant where PCR test is considered more sensitive compared to rapid antigen test.

Should SARS-CoV-2 serological testing be used in the decision to deliver a COVID-19 vaccine booster? A pro-con assessment.

Anti-SARS-CoV-2 vaccination has saved millions of lives in the past few years. To maintain a high level of protection, particularly in at-risk populations, booster doses are recommended to counter the waning of circulating antibody levels over time and the continuous emergence of immune escape variants of concern (VOCs). As anti-spike serology is now widely available, it may be considered a useful tool to identify individuals needing an additional vaccine dose, i.e., to screen certain populations to identify those whose plasma antibody levels are too low to provide protection. However, no recommendations are currently available on this topic. We reviewed the relevant supporting and opposing arguments, including areas of uncertainty, and concluded that in most populations, spike serology should not be used to decide about the administration of a booster dose. The main counterarguments are as follows: correlates of protection are imperfectly characterised, essentially owing to the emergence of VOCs; spike serology has an intrinsic inability to comprehensively reflect the whole immune memory; and booster vaccines are now VOC-adapted, while the commonly available commercial serological assays explore antibodies against the original virus.

Nanobody-based strategy for rapid and accurate pathogen detection: A case of COVID-19 testing.

Antibody pairs-based immunoassay platforms served as essential and effective tools in the field of pathogen detection. However, the cumbersome preparation and limited detection sensitivity of antibody pairs challenge in establishment of a highly sensitive detection platform. In this study, using COVID-19 testing as a case, we utilized readily accessible nanobodies as detection antibodies and further proposed an accurate design concept with a more scientific and efficient screening strategy to obtain ultrasensitive antibody pairs. We employed nanobodies capable of binding different antigenic epitopes of the nucleocapsid (NP) or receptor-binding domain (RBD) antigens sandwich as substitutes for monoclonal antibodies (mAbs) sandwich in fast detection formats and utilized time-resolved fluorescence (TRF) microspheres as the signal probe. Consequently, we developed a multi-epitope nanobody sandwich-based fluorescence lateral flow immunoassay (FLFA) strip. Our results suggest that the NP antigen had a detection limit of 12.01pg/mL, while the RBD antigen had a limit of 6.51 pg/mL using our FLFA strip. Based on double mAb sandwiches, the values presented herein demonstrated 4 to 32-fold enhancements in sensitivity, and 32 to 256-fold enhancements compared to commercially available antigen lateral flow assay kits. Furthermore, we demonstrated the excellent characteristics of the proposed test strip, including its specificity, stability, accuracy, and repeatability, which underscores its the prospective utility. Indeed, these findings indicate that our established screening strategy along with the multi-epitope nanobody sandwich mode provides an optimized strategy in the field of pathogen detection.

Performance evaluation of microfluidic microplate-based fluorescent ELISA for qualitative detection of SARS-CoV-2-specific IgG and IgM.

We evaluated the diagnostic performance of newly developed microfluidic microplate-based fluorescent ELISA for anti-SARS-CoV-2 antibody detection: the Veri-Q opti COVID-19 IgG and IgM ELISAs (hereafter, "Opti IgG/M"; MiCo BioMed, Gyeonggi-do, Republic of Korea), in comparison with conventional ELISAs. A total of 270 serum samples were analyzed, among which 90 samples were serially obtained from 25 COVID-19 patients. Another 180 samples were collected from 180 SARS-CoV-2-negative individuals. As comparative assays, we used SCoV-2 Detect IgG/M ELISA (hereafter, "InBios IgG/M"; InBios, Seattle, WA, USA) and Veri-Q COVID-19 IgG/IgM ELISA (hereafter, "Veri-Q IgG/M"; MiCo BioMed). Compared with conventional ELISAs, the Opti IgG yielded 97.1-100.0% positive percent agreement, 95.2-98.0% negative percent agreement, 96.3-97.8% total percent agreement, and kappa values of 0.90-0.94. Between the Opti IgM and the InBios IgM, the values were 93.7%, 96.6%, 95.9%, and 0.89, respectively. For the Opti IgG, sensitivities for the samples collected from 0-7, 8-14, 15-21, and ≥ 22 days after symptom onset were 40.0, 58.3, 94.1, and 100.0%, respectively. The values for the Opti IgM were 30.0, 54.2, 88.2, and 80%, respectively. The diagnostic specificities of the Opti IgG and IgM were 99.4 and 97.2%, respectively. The microfluidic microplate-based fluorescent ELISAs showed comparable diagnostic performance to conventional ELISAs for detecting anti-SARS-CoV-2 antibodies. With the combination of high throughput, a simplified workflow, and the ability to analyze reduced volumes, this new technology has great potential for improving SARS-CoV-2 serologic testing.

A single domain antibody-based Luminex assay for the detection of SARS-CoV-2 in clinical samples.

Within the past decade, single domain antibodies (sdAbs) have been recognized as unique affinity binding reagents that can be tailored for performance in a variety of immunoassay formats. Luminex MagPlex color-coded magnetic microspheres provide a high-throughput platform that enables multiplexed immunoassays. We developed a MagPlex bead-based assay for the detection of SARS-CoV-2, using sdAbs against SARS-CoV-2 nucleocapsid (N) protein in which we engineered the sdAb capture reagents to orient them on the beads. The oriented sdAbs provided an increase in sensitivity over randomly oriented sdAbs for samples of N diluted in buffer, which also translated into better detection of SARS-CoV-2 in clinical samples. We assessed the specificity of the assay by examining seasonal coronavirus clinical samples. In summary, we provide a proof-of-concept that a bead-based assay using sdAbs to detect SARS-CoV-2 is feasible and future research combining it with other sdAb-coated beads that can detect other viruses may provide a useful diagnostic tool.

[Evaluation of Viral Subgenomic RNAs and Antigen Presence in SARS-CoV-2 PCR Positive Cases]. / SARS-CoV-2 PCR Pozitif Olgularda Viral Subgenomik RNA'larin ve Antijen Varliginin Degerlendirilmesi.

Polymerase chain reaction (PCR) and antigen test (AgT) are frequently used in the diagnosis of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Routine PCR tests that detect the virus genome cannot determine whether the virus is infectious or not. However, detection of subgenomic RNA (sgRNA) produced during the replication period may indicate active viral infection. Active virus detection can offer various health and economic benefits from isolation time to treatment. Antigen tests are also considered as indicators of infectiousness since they can detect viruses above a certain load amount. The aim of this study was to use two different subgenomic RNAs and antigen test instead of genomic RNA to examine the relationship with each other and the clinic in terms of infectiousness. Evaluating the antigen test together with subgenomic RNA as an indicator of infectiousness may show the importance of this test. SARS-CoV-2 PCR positive 109 naso/oropharyngeal swab samples stored at -80 °C were included in the study. In order to confirm the PCR positivity of these samples, E gene PCR was performed and AgT, and E and N sgRNA quantitative real-time reverse transcription-PCR (RT-qPCR) detection was performed. Of the 109 SARSCoV-2 PCR positive samples, 83 (76.14%) had antigen test positivity, 88 (80.73%) had E gene sgRNA, 96 (88.07%) had N gene sgRNA and 97 (89%) had at least one sgRNA positivity.The antigen test was found positive in 77.3% of the samples in which at least one sgRNA was detected and in 66.7% of the negative samples and this difference was not statistically significant (p= 0.475). The difference between E sgRNA and AgT positivity was significant (p= 0.023). N sgRNA was positive in 98.9% of E sgRNA positive samples and 42.9% of the negative samples and this difference was statistically significant (p= 0.0001). The AgT positivity rate was found to be 98.15% (53/54) for cycle threshold (Ct) value ≤ 25, 57.14% (12/21) for Ct 25-30, and 52.94% (18/34) for Ct ≥ 30. The difference in antigen test positivity between E gRNA Ct value ≤ 25 and > 25, ≤ 29 and > 29, < 30 and ≥ 30 was statistically significant (p= 0.0001). Antigen test positivity appears to be associated with viral load and infectivity, as expected. In our study, it has been shown that sgRNAs and AgT which are indicators of infectiousness can be detected at least 10 days after the symptom period. Using these two tests together could detect infective individuals with higher accuracy and shorten the duration of hospital stay and isolation.

The Elecsys® Anti-SARS-CoV-2 and Elecsys® Anti-SARS-CoV-2 S antibody assays: Differentiating between vaccination and infection, and assessing long-term performance.

INTRODUCTION: Serological surveillance is useful for assessing SARS-CoV-2 immunity in populations. To effectively study the presence and persistence of antibodies, it is necessary to distinguish between persons with past infection, and persons who only received vaccination. Knowledge of the duration of antibody persistence is essential for correct interpretation of surveillance results. METHODS: Starting in April 2020, waning of SARS-CoV-2 antibodies was studied in a longitudinal cohort study of 495 SARS-CoV-2 antibody-positive Dutch blood donors, not pre-selected by PCR testing or disease severity. Additionally, in May 2021, a sample of donors representative for the Dutch population was tested for antibodies against the SARS-CoV-2 spike (S) protein, using the Wantai Ab ELISA and the Elecsys® Anti-SARS-CoV-2 S assay; and for antibodies against the nucleocapsid protein, which indicate past infection, using the Elecsys® Anti-SARS-CoV-2 assay. RESULTS: The anti-S response in donors that were infected in April or May 2020 remained positive in 100% of donors in the Elecsys® Anti-SARS-CoV-2 S assay one year after infection, after which follow up of waning was no longer possible because of large scale vaccination. The anti-nucleocapsid response results were still positive in approximately 80% of donors two years after infection. In May 2021, 51% of the donors showed anti-S reactivity and 16.8% tested positive for anti-nucleocapsid antibodies. CONCLUSION: Infection with SARS-CoV-2 resulted in spike and nucleocapsid antibody levels still detectable in the majority of donors 1-2 years after infection. In May 2021, 51% of donors were vaccinated and 16.8% had had an infection. Thus, both Elecsys® SARS-CoV-2 antibody assays can be used to reliably assess the vaccination and infection status of individuals.

Development of a unit conversion tool for five quantitative anti-spike assays and agreement analysis of three qualitative anti-nucleocapsid assays for SARS-CoV-2.

Commercially available assays for measuring severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) anti-spike (S) or anti-nucleocapsid (N) antibodies differ in units, making results comparisons challenging. This study aimed to develop conversion equations between five quantitative anti-S antibody tests and to assess the agreement over time between three qualitative anti-N antibody tests. Blood samples from 24 216 vaccinated healthcare workers in Hiroshima Prefecture, Japan, were analyzed for anti-S antibodies using five quantitative tests (Abbott, Fujirebio, Ortho, Sysmex, Roche) and for anti-N antibodies using three qualitative tests (Abbott, Sysmex, Roche). Geometric mean regression was performed to establish equations for converting measured values between the five quantitative tests. Fleiss κ statistic was used to assess the agreement between the three qualitative tests. A strong correlation (Pearson's coefficient r > 0.9) was found for each pair of the five quantitative tests measuring anti-S antibodies, enabling the development of equations to convert values between each pair. Using these equations, which are based on the original output unit of each test, values obtained from one test can be transformed to be equivalent to the corresponding values in another test. For the three tests for anti-N antibodies, the agreement was substantial in the total sample (Fleiss' κ, 0.74) and moderate among those with self-reported past coronavirus disease 2019 (COVID-19) infection (Fleiss' κ, 0.39). The agreement decreased with time after infection. Reduced agreement between anti-N antibodies tests over time suggests caution in comparing seroepidemiological studies of COVID-19 exposure based on anti-N antibodies measurement. The findings could help improve antibody measurement systems and inform public health decision-makers.

Minding the margins: Evaluating the impact of COVID-19 among Latinx and Black communities with optimal qualitative serological assessment tools.

COVID-19 disproportionately affected minorities, while research barriers to engage underserved communities persist. Serological studies reveal infection and vaccination histories within these communities, however lack of consensus on downstream evaluation methods impede meta-analyses and dampen the broader public health impact. To reveal the impact of COVID-19 and vaccine uptake among diverse communities and to develop rigorous serological downstream evaluation methods, we engaged racial and ethnic minorities in Massachusetts in a cross-sectional study (April-July 2022), screened blood and saliva for SARS-CoV-2 and human endemic coronavirus (hCoV) antibodies by bead-based multiplex assay and point-of-care (POC) test and developed across-plate normalization and classification boundary methods for optimal qualitative serological assessments. Among 290 participants, 91.4% reported receiving at least one dose of a COVID-19 vaccine, while 41.7% reported past SARS-CoV-2 infections, which was confirmed by POC- and multiplex-based saliva and blood IgG seroprevalences. We found significant differences in antigen-specific IgA and IgG antibody outcomes and indication of cross-reactivity with hCoV OC43. Finally, 26.5% of participants reported lingering COVID-19 symptoms, mostly middle-aged Latinas. Hence, prolonged COVID-19 symptoms were common among our underserved population and require public health attention, despite high COVID-19 vaccine uptake. Saliva served as a less-invasive sample-type for IgG-based serosurveys and hCoV cross-reactivity needed to be evaluated for reliable SARS-CoV-2 serosurvey results. The use of the developed rigorous downstream qualitative serological assessment methods will help standardize serosurvey outcomes and meta-analyses for future serosurveys beyond SARS-CoV-2.

Robust Regression Techniques for Multiple Method Comparison and Transformation.

A generalization of Passing-Bablok regression is proposed for comparing multiple measurement methods simultaneously. Possible applications include assay migration studies or interlaboratory trials. When comparing only two methods, the method boils down to the usual Passing-Bablok estimator. It is close in spirit to reduced major axis regression, which is, however, not robust. To obtain a robust estimator, the major axis is replaced by the (hyper-)spherical median axis. This technique has been applied to compare SARS-CoV-2 serological tests, bilirubin in neonates, and an in vitro diagnostic test using different instruments, sample preparations, and reagent lots. In addition, plots similar to the well-known Bland-Altman plots have been developed to represent the variance structure.

A safe, cost-effective, and high-throughput SARS-CoV-2 antigen capture ELISA suitable for large-scale screening in low-resource settings.

Diagnostics employing multiple modalities have been essential for controlling and managing COVID-19, caused by SARS-CoV-2. However, scaling up Reverse Transcription-Quantitative Polymerase Chain Reaction (RT-qPCR), the gold standard for SARS-CoV-2 detection, remains challenging in low and middle-income countries. Cost-effective and high-throughput alternatives like enzyme-linked immunosorbent assay (ELISA) could address this issue. We developed an in-house SARS-CoV-2 nucleocapsid capture ELISA, and validated on 271 nasopharyngeal swab samples from humans (n = 252), bovines (n = 10), and dogs (n = 9). This ELISA has a detection limit of 195 pg/100 µL of nucleocapsid protein and does not cross-react with related coronaviruses, ensuring high specificity to SARS-CoV-2. Diagnostic performance was evaluated using receiver operating characteristic curve analysis, showing a diagnostic sensitivity of 67.78 % and specificity of 100 %. Sensitivity improved to 74.32 % when excluding positive clinical samples with RT-qPCR Ct values > 25. Furthermore, inter-rater reliability analysis demonstrated substantial agreement (κ values = 0.73-0.80) with the VIRALDTECT II Multiplex RT-qPCR kit and perfect agreement with the CoVeasy™ COVID-19 rapid antigen self-test (κ values = 0.89-0.93). Our findings demonstrated that the in-house nucleocapsid capture ELISA is suitable for SARS-CoV-2 testing in humans and animals, meeting the necessary sensitivity and specificity thresholds for cost-effective, large-scale screening.

Double testing with lateral flow antigen test devices for COVID-19: does a second test in quick succession add value?

BACKGROUND/OBJECTIVES: We investigated if performing two lateral flow device (LFD) tests, LFD2 immediately after LFD1, could improve diagnostic sensitivity or specificity for detecting severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) antigen. STUDY DESIGN: Individuals aged ≥16 years attending UK community testing sites (February-May 2021) performed two successive LFD tests and provided a nose-and-throat sample for a polymerase chain reaction (PCR) test. Using the PCR result as the reference diagnosis, we assessed whether improvements could be achieved in sensitivity (by counting a positive result in either LFD as a positive overall test result) or specificity (by using LFD2 as confirmatory test). RESULTS: Overall, 2231 participants were included with 159 (7 %) having a positive PCR test. Of 2223 participants who completed both LFD tests, LFD results were highly concordant both with each other and with PCR tests (>97 %). The proportion of discord LFD results decreased significantly over the study period. Combined LFD usage achieved a sensitivity of 68.6 %, versus 67.1 % for either LFD individually. The specificity increased from 99.5 % to 99.8 % when using LFD2 as confirmatory test. Observed increases in sensitivity and specificity were not statistically significant. Void results were recorded for 31 (1.4 %) LFD1s, 19 (0.9 %) LFD2s and 6 (0.3 %) combined LFD tests. CONCLUSIONS: LFD tests were highly reproducible even when they were performed by untrained users following only written instructions and without supervision. While performing two LFD tests of the same type in quick succession marginally increased sensitivity or specificity, statistically significant improvements were not detected in our study.

Detection of Nucleocapsid Antibodies Associated with Primary SARS-CoV-2 Infection in Unvaccinated and Vaccinated Blood Donors.

Nucleocapsid antibody assays can be used to estimate SARS-CoV-2 infection prevalence in regions implementing spike-based COVID-19 vaccines. However, poor sensitivity of nucleocapsid antibody assays in detecting infection after vaccination has been reported. We derived a lower cutoff for identifying previous infections in a large blood donor cohort (N = 142,599) by using the Ortho VITROS Anti-SARS-CoV-2 Total-N Antibody assay, improving sensitivity while maintaining specificity >98%. We validated sensitivity in samples donated after self-reported swab-confirmed infections diagnoses. Sensitivity for first infections in unvaccinated donors was 98.1% (95% CI 98.0-98.2) and for infection after vaccination was 95.6% (95% CI 95.6-95.7) based on the standard cutoff. Regression analysis showed sensitivity was reduced in the Delta compared with Omicron period, in older donors, in asymptomatic infections, <30 days after infection, and for infection after vaccination. The standard Ortho N antibody threshold demonstrated good sensitivity, which was modestly improved with the revised cutoff.

Performance assessment of a new serological diagnostic test for COVID-19 with candidate peptides from spike and nucleocapsid viral proteins.

Numerous commercial tests for the serological diagnosis of COVID-19 have been produced in recent years. However, it is important to note that these tests exhibit significant variability in their sensitivity, specificity, and accuracy of results. Therefore, the objective of this study was to utilize bioinformatics tools to map SARS-CoV-2 peptides, with the goal of developing a new serological diagnostic test for COVID-19. Two peptides from the S protein and one from the N protein were selected and characterized in silico, chemically synthesized, and used as a serological diagnostic tool to detect IgM, IgG, and IgA anti-SARS-CoV-2 antibodies through the ELISA technique, confirmed as positive and negative samples by RT-qPCR or serology by ELISA. The results showed a sensitivity, specificity, Positive Predictive Value and Negative Predictive Value of 100% (p < 00001, 95% CI) for the proposed test. Although preliminary, this study brings proof-of-concept results that are consistent with the high-performance rates of the ELISA test when compared to other well-established methods for diagnosing COVID-19.

Canada's approach to SARS-CoV-2 sero-surveillance: Lessons learned for routine surveillance and future pandemics.

SETTING: In Canada's federated healthcare system, 13 provincial and territorial jurisdictions have independent responsibility to collect data to inform health policies. During the COVID-19 pandemic (2020-2023), national and regional sero-surveys mostly drew upon existing infrastructure to quickly test specimens and collect data but required cross-jurisdiction coordination and communication. INTERVENTION: There were 4 national and 7 regional general population SARS-CoV-2 sero-surveys. Survey methodologies varied by participant selection approaches, assay choices, and reporting structures. We analyzed Canadian pandemic sero-surveillance initiatives to identify key learnings to inform future pandemic planning. OUTCOMES: Over a million samples were tested for SARS-CoV-2 antibodies from 2020 to 2023 but siloed in 11 distinct datasets. Most national sero-surveys had insufficient sample size to estimate regional prevalence; differences in methodology hampered cross-regional comparisons of regional sero-surveys. Only four sero-surveys included questionnaires. Sero-surveys were not directly comparable due to different assays, sampling methodologies, and time-frames. Linkage to health records occurred in three provinces only. Dried blood spots permitted sample collection in remote populations and during stay-at-home orders. IMPLICATIONS: To provide timely, high-quality information for public health decision-making, routine sero-surveillance systems must be adaptable, flexible, and scalable. National capability planning should include consortiums for assay design and validation, defined mechanisms to improve test capacity, base documents for data linkage and material transfer across jurisdictions, and mechanisms for real-time communication of data. Lessons learned will inform incorporation of a robust sero-survey program into routine surveillance with strategic sampling and capacity to adapt and scale rapidly as a part of a comprehensive national pandemic response plan.

RéSUMé: CONTEXTE: Au Canada, où le système de santé est fédéré, les 13 juridictions provinciales et territoriales ont la responsabilité individuelle de recueillir les données qui leur permettent d'élaborer leurs politiques de santé. Lors de la pandémie de COVID-19 (2020­2023), pour réaliser les enquêtes de séroprévalence à l'échelle régionale et nationale, les autorités ont principalement utilisé l'infrastructure existante pour pouvoir analyser les échantillons et recueillir des données rapidement, mais cela a également nécessité de la communication et de la coordination entre les différentes juridictions. INTERVENTION: Au Canada, il y a eu quatre enquêtes nationales et sept enquêtes régionales sur la séroprévalence du SARS-CoV-2 dans la population générale. Les méthodologies utilisées différaient selon la méthode de sélection des participants, le choix des tests d'analyses et les structures de rapports. Nous avons analysé la façon dont ces enquêtes avaient été réalisées afin d'en dégager des éléments essentiels qui permettront de planifier pour les futures pandémies. RéSULTATS: Entre 2020 et 2023, plus d'un million d'échantillons, répartis en 11 ensembles de données distincts, ont été analysés afin de rechercher la présence d'anticorps au SARS-CoV-2. Dans la plupart des enquêtes nationales, la taille de l'échantillon était insuffisante pour pouvoir estimer la prévalence à l'échelle régionale. La disparité des méthodologies utilisées a entravé la comparaison des enquêtes régionales. Seules quatre enquêtes fournissaient les données recueillies à partir des questionnaires. Il a été impossible de comparer les enquêtes entre elles en raison de la diversité des tests d'analyse utilisés, des méthodes d'échantillonnage et de la durée des enquêtes. Seules trois provinces avaient couplé leurs données avec les archives médicales. Pour réaliser les enquêtes dans les populations éloignées et lors des périodes de confinement, la méthode d'analyse sur gouttes de sang séché a été utilisée. CONCLUSION: Afin de pouvoir fournir, en temps et en heure, des données de haute qualité pour la prise de décisions en matière de santé publique, un système de sérosurveillance continuelle doit être adaptable, modulable et évolutif. En cas de pandémie, un plan national doit prévoir des consortiums pour la conception et la validation des tests d'analyse, des moyens d'amélioration de la capacité de dépistage, des documents de base pour le couplage des données, un mode de transfert du matériel entre les différentes juridictions et des moyens pour une communication en temps réel des données. Les leçons tirées de cette analyse permettront de mettre en place un solide programme d'enquêtes de séroprévalence au sein des systèmes de sérosurveillance continuelle, et que ce programme sera accompagné d'une stratégie d'échantillonnage et d'un plan d'intervention national, rapide et complet en cas de pandémie.

A sensitive gold nanoflower-based lateral flow assay coupled with gold staining technique for the detection of SARS-CoV-2 antigen.

An enhanced lateral flow assay (LFA) is presented for rapid and highly sensitive detection of acute respiratory syndrome coronavirus-2 (SARS-CoV-2) antigens with gold nanoflowers (Au NFs) as signaling markers and gold enhancement to amplify the signal intensities. First, the effect of the morphology of gold nanomaterials on the sensitivity of LFA detection was investigated. The results showed that Au NFs prepared by the seed growth method showed a 5-fold higher detection sensitivity than gold nanoparticles (Au NPs) of the same particle size, which may benefit from the higher extinction coefficient and larger specific surface area of Au NFs. Under the optimized experimental conditions, the Au NFs-based LFA exhibited a detection limit (LOD) of 25 pg mL-1 for N protein using 135 nm Au NFs as the signaling probes. The signal was further amplified by using a gold enhancement strategy, and the LOD for the detection of N protein achieved was 5 pg mL-1. The established LFA also exhibited good repeatability and stability and showed applicability in the diagnosis of SARS-CoV-2 infection.

Diagnostic Accuracy of the Abbott BinaxNOW COVID-19 Antigen Card Test, Puerto Rico.

BACKGROUND: The COVID-19 pandemic underscored the need for rapid and accurate diagnostic tools. In August 2020, the Abbott BinaxNOW COVID-19 Antigen Card test became available as a timely and affordable alternative for SARS-CoV-2 molecular testing, but its performance may vary due to factors including timing and symptomatology. This study evaluates BinaxNOW diagnostic performance in diverse epidemiological contexts. METHODS: Using RT-PCR as reference, we assessed performance of the BinaxNOW COVID-19 test for SARS-CoV-2 detection in anterior nasal swabs from participants of two studies in Puerto Rico from December 2020 to May 2023. Test performance was assessed by days post symptom onset, collection strategy, vaccination status, symptomatology, repeated testing, and RT-PCR cycle threshold (Ct) values. RESULTS: BinaxNOW demonstrated an overall sensitivity of 84.1% and specificity of 98.8%. Sensitivity peaked within 1-6 days after symptom onset (93.2%) and was higher for symptomatic (86.3%) than asymptomatic (67.3%) participants. Sensitivity declined over the course of infection, dropping from 96.3% in the initial test to 48.4% in testing performed 7-14 days later. BinaxNOW showed 99.5% sensitivity in participants with low Ct values (≤ 25) but lower sensitivity (18.2%) for participants with higher Cts (36-40). CONCLUSIONS: BinaxNOW demonstrated high sensitivity and specificity, particularly in early-stage infections and symptomatic participants. In situations where test sensitivity is crucial for clinical decision-making, nucleic acid amplification tests are preferred. These findings highlight the importance of considering clinical and epidemiological context when interpreting test results and emphasize the need for ongoing research to adapt testing strategies to emerging SARS-CoV-2 variants.

Clinical Evaluation of the Accuracy of the Panbio™ COVID-19/Flu A&B Rapid Panel: A Combination Antigen Rapid Diagnostic Test for the Omicron Variant and Influenza A Virus.

It is difficult to differentiate between coronavirus disease 2019 (COVID-19) and influenza based on the symptoms. In the present study, a newly developed antigen rapid diagnostic test (Ag-RDT) called Panbio™ COVID-19/Flu A&B that can simultaneously detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza A/B virus was evaluated. Its accuracy was evaluated using 235 pairs of nasopharyngeal samples collected from patients with respiratory symptoms and fever (>37.5°C). Reverse transcription polymerase chain reaction was used as a reference method to evaluate the accuracy of the SARS-CoV-2 detection. We confirmed the accuracy of the developed Ag-RDT against the Omicron variant where the sensitivity and specificity were 94.8% and 100%, respectively. In addition, to identify the influenza A virus, a noninferiority test was conducted using a commercial Ag-RDT, which has a sensitivity and specificity in comparison with viral culture of 94.8% and 98.4%, respectively. The positive and negative predictive values for influenza A virus were 98.5% and 98.1%, respectively, for the Panbio COVID-19/Flu A&B test. The evaluation of this newly developed Ag-RDT using clinical samples suggests that it has a high efficacy in clinical settings.