Hybrid immunity from SARS-CoV-2 infection and vaccination in Canadian adults: cohort study (preprint)

BackgroundFew national-level studies have evaluated the impact of "hybrid" immunity (vaccination coupled with recovery from infection) from the Omicron variants of SARS-CoV-2. MethodsFrom May 2020 to December 2022, we conducted serial assessments (each of [~]4000-9000 adults) examining SARS-CoV-2 antibodies within a mostly representative Canadian cohort drawn from a national online polling platform. Adults, most of whom were vaccinated, reported viral test-confirmed infections and mailed self-collected dried blood spots to a central lab. Samples underwent highly sensitive and specific antibody assays to spike and nucleocapsid protein antigens, the latter triggered only by infection. We estimated cumulative SARS-CoV-2 incidence prior to the Omicron period and during the BA.1/1.1 and BA.2/5 waves. We assessed changes in antibody levels and in age-specific active immunity levels. ResultsSpike levels were higher in infected than in uninfected adults, regardless of vaccination doses. Among adults vaccinated at least thrice and infected more than six months earlier, spike levels fell notably and continuously for the nine months post-vaccination. By contrast, among adults infected within six months, spike levels declined gradually. Declines were similar by sex, age group, and ethnicity. Recent vaccination attenuated declines in spike levels from older infections. In a convenience sample, spike antibody and cellular responses were correlated. Near the end of 2022, about 35% of adults above age 60 had their last vaccine dose more than six months ago, and about 25% remained uninfected. The cumulative incidence of SARS-CoV-2 infection rose from 13% (95% CI 11-14%) before omicron to 78% (76-80%) by December 2022, equating to 25 million infected adults cumulatively. However, the COVID-19 weekly death rate during the BA.2/5 waves was less than half of that during the BA.1/1.1 wave, implying a protective role for hybrid immunity. ConclusionsStrategies to maintain population-level hybrid immunity require up-to-date vaccination coverage, including among those recovering from infection. Population-based, self-collected dried blood spots are a practicable biological surveillance platform. FundingFunding was provided by the COVID-19 Immunity Task Force, Canadian Institutes of Health Research, Pfizer Global Medical Grants, and St. Michaels Hospital Foundation. PJ and ACG are funded by the Canada Research Chairs Program.

Omicron Breakthrough Infection Elicits Superior Humoral and Mucosal Immune Responses to SARS-CoV-2 Variants than an Intramuscular Booster Dose (preprint)

Our understanding of the quality of cellular and humoral immunity conferred by COVID-19 vaccination alone versus vaccination plus SARS-CoV-2 breakthrough (BT) infection remains incomplete. While the current (2023) SARS-CoV-2 immune landscape of Canadians is complex, in late 2021 most Canadians had either just received a third dose of COVID-19 vaccine, or had received their two dose primary series and then experienced an Omicron BT. Herein we took advantage of this coincident timing to contrast cellular and humoral immunity conferred by three doses of vaccine versus two doses plus BT. Our results show that mild BT infection induces cell-mediated immune responses to variants comparable to an intramuscular vaccine booster dose. In contrast, BT subjects had higher salivary IgG and IgA levels against the Omicron Spike and enhanced reactivity to the ancestral Spike for the IgA isotype, which also reacted with SARS-CoV-1. Serum neutralizing antibody levels against the ancestral strain and the variants were also higher after BT infection. Our results support the need for mucosal vaccines to emulate the enhanced mucosal and humoral immunity induced by Omicron without exposing individuals to the risks associated with SARS-CoV-2 infection.

Immunogenicity of COVID-19 vaccines and their effect on the HIV reservoir in older people with HIV (preprint)

Older individuals and people with HIV (PWH) were prioritized for COVID-19 vaccination, yet comprehensive studies of the immunogenicity of these vaccines and their effects on HIV reservoirs are not available. We followed 68 PWH aged 55 and older and 23 age-matched HIV-negative individuals for 48 weeks from the first vaccine dose, after the total of three doses. All PWH were on antiretroviral therapy (cART) and had different immune status, including immune responders (IR), immune non-responders (INR), and PWH with low-level viremia (LLV). We measured total and neutralizing Ab responses to SARS-CoV-2 spike and RBD in sera, total anti-spike Abs in saliva, frequency of anti-RBD/NTD B cells, changes in frequency of anti-spike, HIV gag/nef-specific T cells, and HIV reservoirs in peripheral CD4+ T cells. The resulting datasets were used to create a mathematical model for within-host immunization. Various regimens of BNT162b2, mRNA-1273, and ChAdOx1 vaccines elicited equally strong anti-spike IgG responses in PWH and HIV-negative participants in serum and saliva at all timepoints. These responses had similar kinetics in both cohorts and peaked at 4 weeks post-booster (third dose), while half-lives of plasma IgG also dramatically increased post-booster in both groups. Salivary spike IgA responses were low, especially in INRs. PWH had diminished live virus neutralizing titers after two vaccine doses which were 'rescued' after a booster. Anti-spike T cell immunity was enhanced in IRs even in comparison to HIV-negative participants, suggesting Th1 imprinting from HIV, while in INRs it was the lowest. Increased frequency of viral 'blips' in PWH were seen post-vaccination, but vaccines did not affect the size of the intact HIV reservoir in CD4+ T cells in most PWH, except in LLVs. Thus, older PWH require three doses of COVID-19 vaccine to maximize neutralizing responses against SARS-CoV-2, although vaccines may increase HIV reservoirs in PWH with persistent viremia.

Omicron variant BA.1, BA.5, BQ.1.1, and XBB.1.5 Neutralizing Antibodies Following BNT162b2 BA.4/5 versus mRNA-1273 BA.1 Bivalent Vaccination (preprint)

Neutralization of Omicron subvariants by different bivalent vaccines have not been well evaluated. This study characterized neutralization against Omicron subvariants in 98 individuals receiving dialysis or with a kidney transplant receiving the BNT162b2 (BA.4/BA.5) or mRNA-1273 (BA.1) bivalent COVID-19 vaccine. Neutralization against Omicron BA.1, BA.5, BQ.1.1, and XBB.1.5 increased by 8-fold one month following bivalent vaccination. In comparison to wild-type (D614G), neutralizing antibodies against Omicron-specific variants were 7.3-fold lower against BA.1, 8.3-fold lower against BA.5, 45.8-fold lower against BQ.1.1, and 48.2-fold lower against XBB.1.5. Viral neutralization was not significantly different by bivalent vaccine type for wild-type (D614G) (P=0.48), BA.1 (P=0.21), BA.5 (P=0.07), BQ.1.1 (P=0.10), nor XBB.1.5 (P=0.10). Hybrid immunity conferred higher neutralizing antibodies against all Omicron subvariants. Given that both BNT162b2 (BA.4/BA.5) and mRNA-1273 (BA.1) induced similar neutralization against all Omicron subvariants, this suggests that bivalent vaccines confer protection even when they are antigenically divergent from the circulating variant.

Protection from Omicron infection in residents of nursing and retirement homes in Ontario, Canada (preprint)

Objectives: To identify factors that contribute to protection from infection with the Omicron variant of SARS-CoV-2 in older adults in nursing and retirement homes. Design: Longitudinal cohort study with retrospective analysis of infection risk. Setting and Participants: 997 residents of nursing and retirement homes from Ontario, Canada, in the COVID-in-LTC study. Methods: Residents with three mRNA dose vaccinations were included in the study. SARS-CoV-2 infection was determined by positive nasopharyngeal PCR test and/or circulating anti-nucleocapsid IgG antibodies. Cumulative probability of Omicron infection after recent COVID-19 was assessed by log-rank test of Kaplan-Meier curves. Cox regression was used to assess risk of Omicron infection by age, sex, mRNA vaccine combination, whether individuals received a fourth dose, as well as recent COVID-19. Results: 171 residents (17.2%) had a presumed Omicron variant SARS-CoV-2 infection between December 15, 2021 (local start of the first Omicron wave) and May 3, 2022. Risk of Omicron infection was not different by age [hazard ratio (95% confidence interval): 1.01 (0.99-1.02)], or in women compared to men [0.97 (0.70-1.34)], but infection risk decreased 47% with three vaccine doses of mRNA-1273 (Moderna) compared to BNT162b2 (Pfizer) [0.53 (0.31-0.90)], 81% with any fourth mRNA vaccine dose [0.19 (0.12-0.30)], and 48% with SARS-CoV-2 infection in the 3 months prior to beginning of the Omicron wave [0.52, (0.27-0.99)]. Conclusions and Implications: Vaccine type (i.e., mRNA-1273/Spikevax vs BNT162b2/Cominarty), any fourth vaccine dose, and hybrid immunity from recent COVID-19, were protective against infection with the Omicron variant. These data emphasize the importance of vaccine type, and number of vaccine doses, in maintenance of protective immunity and reduction of risk of Omicron variant breakthrough infection. These findings promote continued public health efforts to support vaccination programs and monitor vaccine immunogenicity in older adults.

Safety and Efficacy of Preventative COVID Vaccines: The StopCoV Study (preprint)

Background: To partially immunize more persons against COVID-19 during a time of limited vaccine availability, Canadian public health officials recommended extending the vaccine dose interval and brand mixing. Impact on the antibody response among the older ambulatory population was unclear. Methods: Decentralized prospective cohort study with self-report of adverse events and collection of dried blood spots. Data is presented for 1193 (93%) of the 911 older (aged >70 years) and 375 younger (30-50 years) recruits. Findings: Local and systemic reactivity rates were high but short-lived, particularly in the younger cohort and with mRNA-1273 vaccine. After a single COVID-19 vaccine, 84% younger but only 46% older participants had positive IgG antibodies to both spike protein and receptor binding domain (RBD) antigens, increasing to 100/98% with the second dose respectively. In multivariable linear regression model, lower normalized IgG RBD antibody ratios two weeks after the second dose were statistically associated with older age, male gender, cancer diagnosis, lower body weight, BNT162b2 relative to mRNA-1273 and longer dose intervals. Antibody ratios in both cohorts declined 12 weeks post second vaccine dose. Interpretation: We report success of a decentralized serology study. Antibody responses were higher in the younger than older cohort and were greater for those with at least one mRNA-1273 dose. The immunity threshold is unknown but correlations between binding and neutralizing antibodies are strongly positive. Trends with time and at breakthrough infection will inform vaccine booster strategies. Funding: Supported by the Public Health Agency of Canada and the University Health Network Foundation.

Immunogenicity of convalescent and vaccinated sera against clinical isolates of ancestral SARS-CoV-2, beta, delta, and omicron variants (preprint)

The omicron variant of concern (VOC) of SARS-CoV-2 was first reported in November 2021 in Botswana and South Africa. Omicron variant has evolved multiple mutations within the spike protein and the receptor binding domain (RBD), raising concerns of increased antibody evasion. Here, we isolated infectious omicron from a clinical specimen obtained in Canada. The neutralizing activity of sera from 65 coronavirus disease (COVID-19) vaccine recipients and convalescent individuals against clinical isolates of ancestral SARS-CoV-2, beta, delta, and omicron VOCs was assessed. Convalescent sera from unvaccinated individuals infected by the ancestral virus during the first wave of COVID-19 in Canada (July, 2020) demonstrated reduced neutralization against beta, delta and omicron VOCs. Convalescent sera from unvaccinated individuals infected by the delta variant (May-June, 2021) neutralized omicron to significantly lower levels compared to the delta variant. Sera from individuals that received three doses of the Pfizer or Moderna vaccines demonstrated reduced neutralization of both delta and omicron variants relative to ancestral SARS-CoV-2. Sera from individuals that were naturally infected with ancestral SARS-CoV-2 and subsequently received two doses of the Pfizer vaccine induced significantly higher neutralizing antibody levels against ancestral virus and all VOCs. Importantly, infection alone, either with ancestral SARS-CoV-2 or the delta variant was not sufficient to induce high neutralizing antibody titers against omicron. This data will inform current booster vaccination strategies and we highlight the need for additional studies to identify longevity of immunity against SARS-CoV-2 and optimal neutralizing antibody levels that are necessary to prevent infection and/or severe COVID-19.

A "Made-in-Canada" serology solution for profiling humoral immune responses to SARS-CoV-2 infection and vaccination (preprint)

BACKGROUND: Testing for antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been instrumental in detecting previous exposures and analyzing vaccine-elicited immune responses. Here, we describe a scalable "Made-in-Canada" solution that can detect and quantify SARS-CoV-2 antibodies, discriminate between natural infection- and vaccination-induced responses, and assess antibody-mediated inhibition of the spike-angiotensin converting enzyme 2 (ACE2) interaction. METHODS: We developed a set of methods and reagents to detect SARS-CoV-2 antibodies by enzyme-linked immunosorbent assay (ELISA). The main assays focus on the parallel detection of immunoglobulin (Ig)Gs against the spike trimer, its receptor binding domain (RBD), and the nucleocapsid (N) protein. These antigens are complemented by a detection antibody (human anti-IgG fused to horseradish peroxidase (HRP)) and a positive control reference antibody (recombinant IgG against the RBD), permitting intra- and inter-laboratory comparisons. Using this toolkit and commercial reagents, we optimized automated ELISAs on two different high throughput platforms to measure antibody responses to SARS-CoV-2 antigens. The assays were calibrated to a reference standard from the World Health Organization. We also automated a surrogate neutralization (sn)ELISA that measures inhibition of ACE2-Spike or -RBD interactions by antibodies using biotinylated ACE2. RESULTS: Our individual IgG-based ELISAs measure antibody levels in single-point measurements in reference to a standard antibody curve to accurately distinguish non-infected and infected individuals (area under the curve > 0.96 for each assay). Positivity thresholds can be established in individual assays using precision-recall analysis (e.g., by fixing the false positive rate), or more stringently, by scoring against the distribution of the means of negative samples across multiple assays performed over several months. For seroprevalence assessment (in a non-vaccinated cohort), classifying a sample as positive if antibodies were detected for at least 2 of the 3 antigens provided the highest specificity. In vaccinated cohorts, increases in anti-spike and -RBD (but not -N) antibodies are observed. Here, we present detailed protocols to perform these assays using either serum/plasma or dried blood spots both manually and on two automated platforms, and to express the results in international units to facilitate data harmonization and inter-study comparisons. We also demonstrate that the snELISA can be performed automatically at single points, increasing the scalability of this functional assay for large seroprevalence studies. INTERPRETATION: The ability to measure antibodies to three viral antigens and identify neutralizing antibodies capable of disrupting spike-ACE2 interactions in high-throughput assays enables large-scale analyses of humoral immune responses to SARS-CoV-2 infection and vaccination. The "Made-in-Canada" set of protein reagents, produced at the National Research Council of Canada are publicly available to enable the up-scaling of standardized serological assays, permitting nationwide data comparison and aggregation.

Neutralizing antibody responses to SARS-CoV-2 variants in vaccinated Ontario long-term care home residents and workers (preprint)

Prioritizing Ontarios long-term care home (LTCH) residents for vaccination against severe acute respiratory syndrome coronavirus 2 has drastically reduced their disease burden; however, recent LTCH outbreaks of variants of concern (VOCs) have raised questions regarding their immune responses. In 198 residents, mRNA vaccine dose 1 elicited partial spike and receptor binding domain antibody responses, while the second elicited a response at least equivalent to convalescent individuals in most residents. Residents administered mRNA-1273 (Moderna) mounted stronger total and neutralizing antibody responses than those administered BNT162b2 (Pfizer-BioNTech). Two to four weeks after dose 2, residents (n = 119, median age 88) produced 4.8-6.3-fold fewer neutralizing antibodies than staff (n = 78; median age 47) against wild-type (with D614G) pseudotyped lentivirus, and residents administered BNT162b2 produced 3.89-fold fewer neutralizing antibodies than those who received mRNA-1273. These effects were exacerbated upon serum challenge with pseudotyped VOC spike, with up to 7.94-fold reductions in B.1.351 (Beta) neutralization. Cumulatively, weaker vaccine stimulation, age/comorbidities, and the VOC produced an [~]130-fold reduction in apparent neutralization titers in LTCH residents and 37.9% of BNT162b2-vaccinated residents had undetectable neutralizing antibodies to B.1.351. Continued immune response surveillance and additional vaccine doses may be required in this population with known vulnerabilities.

Immunity to SARS-CoV-2 persists 9 months post-symptoms with an altered T cell phenotype compared to influenza A virus-specific memory (preprint)

SARS-CoV-2 induces T cell, B cell and antibody responses that are detected for several months in recovered individuals. Whether this response resembles a typical respiratory viral infection is a matter of debate. Here we followed T cell and antibody responses in 24 mainly non-hospitalized SARS-CoV-2 recovered subjects at two time points (median of 45- and 145-days post-symptom onset). Antibody responses were detected in 95% of subjects, with a strong correlation between plasma and salivary anti-S and anti-RBD IgG, as well as a correlation between circulating T follicular helper cells and the SARS-CoV-2-specific IgG response. Based on intracellular cytokine production or proliferation, CD4+ T cell responses to SARS-CoV-2 were detected in all subjects, decaying with a half-life of 5-6 months for S-specific IL-2-producing cells. CD4+ responses were largely of the T helper 1 phenotype, but with a lower ratio of IFN-{gamma} : IL-2 producing cells and a lower frequency of CD8+: CD4+ T cells compared to influenza A virus-(IAV)-specific memory responses within the same subjects. Analysis of secreted molecules also revealed a lower ratio of IFN-{gamma}: IL-2 and IFN-{gamma}: IL-6 and an altered cytotoxic profile for S- and N-specific compared to IAV-specific responses. These data suggest that the memory T-cell phenotype after a single infection with SARS-CoV-2 persists over time, with an altered cytokine and cytotoxic profile compared to long term memory to IAV within the same subjects.

The Humoral Response to the BNT162b2 Vaccine in Hemodialysis Patients (preprint)

Importance: Hemodialysis patients have an exceptionally high mortality from COVID-19 and this patient population often has a poor response to vaccinations. Randomized controlled trials for COVID-19 vaccines included few patients with kidney disease, therefore vaccine immunogenicity is uncertain in this population. Objective: Evaluate the SARS-CoV-2 antibody response in chronic hemodialysis patients following one versus two doses of BNT162b2 COVID-19 vaccination compared to health care worker controls and convalescent serum. Design: Prospective observational cohort study. Setting: Single centre study in Toronto, Ontario, Canada. Participants: 142 in-centre hemodialysis patients and 35 health care worker controls. Exposure: BNT162b2 (Pfizer-BioNTech) COVID-19 vaccine. Main Outcomes and Measures: SARS-CoV-2 IgG antibodies to the spike protein (anti-spike), receptor binding domain (anti-RBD), and nucleocapsid protein (anti-NP) were measured in 66 hemodialysis patients receiving one vaccine dose following a public health policy change, 76 patients receiving two vaccine doses, and 35 health care workers receiving two vaccine doses. Results: Detectable anti-NP suggestive of natural SARS-CoV-2 infection was detected in 15/142 (11%) of patients at baseline while only three patients had prior RT-PCR confirmed COVID-19. Two additional patients contracted COVID-19 after receiving two doses of vaccine. In patients receiving a single BNT162b2 dose, seroconversion occurred in 53/66 (80%) for anti-spike and 35/66 (55%) for anti-RBD by 28 days post dose, but only 15/66 (23%) and 4/66 (6%), respectively attained a robust response as defined by reaching the median level of anti-spike and anti-RBD in convalescent serum from COVID-19 survivors. In patients receiving two doses of BNT162b2 vaccine, seroconversion occurred in 69/72 (96%) for anti-spike and 63/72 (88%) for anti-RBD by 2 weeks following the second dose while 52/72 (72%) and 43/76 (41%) reached the median convalescent serum level of anti-spike and anti-RBD, respectively. In contrast, 35/35 (100%) of health care workers exceeded the median level of anti-spike and anti-RBD found in convalescent serum 2-4 weeks after the second dose. Conclusions and Relevance: This study confirms poor immunogenicity 28 days following a single dose of BNT162b2 vaccine in the hemodialysis population, supporting adherence to recommended vaccination schedules, and avoiding delay of the second dose in these at-risk individuals.

Preclinical evaluation of a SARS-CoV-2 mRNA vaccine PTX-COVID19-B (preprint)

Safe and effective vaccines are needed to end the COVID-19 pandemic caused by SARS-CoV-2. Here we report the preclinical development of a lipid nanoparticle (LNP) formulated SARS-CoV-2 mRNA vaccine, PTX-COVID19-B. PTX-COVID19-B was chosen among three candidates after the initial mouse vaccination results showed that it elicited the strongest neutralizing antibody response against SARS-CoV-2. Further tests in mice and hamsters indicated that PTX-COVID19-B induced robust humoral and cellular immune responses and completely protected the vaccinated animals from SARS-CoV-2 infection in the lung. Studies in hamsters also showed that PTX-COVID19-B protected the upper respiratory tract from SARS-CoV-2 infection. Mouse immune sera elicited by PTX-COVID19-B vaccination were able to neutralize SARS-CoV-2 variants of concern (VOCs), including the B.1.1.7, B.1.351 and P.1 lineages. No adverse effects were induced by PTX-COVID19-B in both mice and hamsters. These preclinical results indicate that PTX-COVID19-B is safe and effective. Based on these results, PTX-COVID19-B was authorized by Health Canada to enter clinical trials in December 2020 with a phase 1 clinical trial ongoing (ClinicalTrials.gov number: NCT04765436).

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) seroprevalence: Navigating the absence of a gold standard (preprint)

Background: Multiple anti-SARS-CoV-2 immunoassays are available, but no gold standard exists. We assessed four assays using various methodological approaches to estimate SARS-COV-2 seroprevalence during the first COVID-19 wave in Canada. Methods: This serial cross-sectional study was conducted using plasma samples from healthy blood donors between April-September 2020. Qualitative assessment of SARS-CoV-2 IgG antibodies was based on four assays: Abbott Architect SARS-Cov-2 IgG assay (target nucleocapsid) (Abbott-NP) and three in-house IgG ELISA assays (target spike glycoprotein (Spike), spike receptor binding domain (RBD), and nucleocapsid (NP)). Seroprevalence was estimated using multiple composite reference standards (CRS) and by a series of Bayesian Latent Class Models (BLCM) (using uninformative, weakly, and informative priors). Results: 8999 blood samples were tested. The Abbott-NP assay consistently estimated seroprevalence to be lower than the ELISA-based assays. Discordance between assays was common, 13 unique diagnostic phenotypes were observed. Only 32 samples (0.4%) were positive by all four assays. BLCM using uninformative priors predicted seroprevalence increased from 0.7% (95% credible interval (CrI); 0.4, 1.0%) in April/May to 0.8% (95% CrI 0.5, 1.2%) in June/July to 1.1% (95% CrI 0.7, 1.6) in August/September. Results from CRS were very similar to the BLCM. Assay characteristics varied considerably over time. Overall spike had the highest sensitivity (89.1% (95% CrI 79.2, 96.9%), while the sensitivity of the Abbott-NP assay waned from 65.3% (95% CrI 43.6, 85.0%) in April/May to 45.9% (95% CrI 27.8, 65.6) by August/September. Discussion: We found low SARS-CoV-2 seroprevalence rates at the end of the first wave and estimates derived from single assays may be biased.

COVID Symptoms, Seroprevalence, and Mortality During the First Wave of SARS-CoV-2 in Canada (preprint)

Background: Canada had substantial mortality from COVID in the first SARS-CoV-2 wave, mostly in nursing homes. Efforts to stem the pandemic (including vaccination, now being introduced) can benefit from direct understanding of the relationship of age-specific mortality to SARS-CoV-2 seroprevalence and from information on asymptomatic infection.Methods: The Action to Beat Coronavirus (Ab-C) study surveyed a reasonably representative sample of 19,994 adult Canadians about COVID symptoms and analyzed IgG antibodies against SARS-CoV-2 from self-collected dried blood spots (DBS) in 8,967 adults. A sensitive and specific chemiluminescence ELISA detected IgG to the spike trimer. We compared seroprevalence to deaths to establish infection fatality rates (IFRs) and used mortality data to estimate infection levels in nursing home residents.Findings: The best estimate (high specificity) of adult seroprevalence nationally is 1.7%, but as high as 3.5% (high sensitivity) depending on assay cutoffs. The highest prevalence was in Ontario (2.4-3.9%), the most populous province, and in younger adults aged 18-39 years (2.5-4.4%). Based on mortality, we estimated 13-16% of nursing home residents became infected. The first viral wave infected 0.54-1.08 million adult Canadians, half of whom were <40 years old. The IFR outside nursing homes was 0.20-0.40%, but the COVID death rate in nursing home residents was >70 times higher than comparably-aged adults living in the community. Seropositivity correlated with COVID symptoms, particularly during March. Asymptomatic adults constituted about a quarter of definite seropositives, with a greater proportion in the elderly.Interpretation: Canada had relatively low infection prevalence and low IFRs in the community, but not in nursing homes, during the first viral wave. The Ab-C study demonstrates the practicability of using self-collected DBS for antibody testing in national surveys to monitor the pandemic and vaccine-induced immunity in the population.Funding Statement: Pfizer Global Medical, Unity Health Foundation, Canadian COVID-19 Immunity Task ForceDeclaration of Interests: We declare no competing interests.Ethics Approval Statement: Ethics approval was provided by St. Michael's Institutional ReviewBoard. REB 20-107

Evidence for sustained mucosal and systemic antibody responses to SARS-CoV-2 antigens in COVID-19 patients (preprint)

While the antibody response to SARS-CoV-2 has been extensively studied in blood, relatively little is known about the mucosal immune response and its relationship to systemic antibody levels. Since SARS-CoV-2 initially replicates in the upper airway, the antibody response in the oral cavity is likely an important parameter that influences the course of infection. We developed enzyme linked immunosorbent assays to detect IgA and IgG antibodies to the SARS-CoV-2 spike protein (full length trimer) and its receptor binding domain (RBD) in serum (n=496) and saliva (n=90) of acute and convalescent patients with laboratory-diagnosed COVID-19 ranging from 3-115 days post-symptom onset (PSO), compared to negative controls. Anti-CoV-2 antibody responses were readily detected in serum and saliva, with peak IgG levels attained by 16-30 days PSO. Whereas anti-CoV-2 IgA antibodies rapidly decayed, IgG antibodies remained relatively stable up to 115 days PSO in both biofluids. Importantly, IgG responses in saliva and serum were correlated, suggesting that antibodies in the saliva may serve as a surrogate measure of systemic immunity.

A simple protein-based SARS-CoV-2 surrogate neutralization assay (preprint)

With the COVID-19 pandemic surpassing 12M confirmed cases and 550K deaths worldwide, defining the key components of the immune response to SARS-CoV-2 infection is critical. Of particular importance is the identification of immune correlates of infection that would support public health decision-making on treatment approaches, vaccination strategies, and convalescent plasma therapy. While ELISA-based assays to detect and quantitate antibodies to SARS-CoV-2 in patient samples have been developed, the detection of neutralizing antibodies typically requires more demanding cell-based viral assays. Here, we present and validate a safe and efficient protein-based assay for the detection of serum and plasma antibodies that block the interaction of the SARS-CoV-2 spike (S) protein receptor binding domain (RBD) with its receptor, angiotensin converting-enzyme 2 (ACE2). This test is performed on the same platform and in parallel with an enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies against the RBD and serves as a surrogate neutralization assay.