Canadian SARS-CoV-2 serological survey using antenatal serum samples: a retrospective seroprevalence study.

BACKGROUND: Insufficient data on the rate and distribution of SARS-CoV-2 infection in Canada has presented a substantial challenge to the public health response to the COVID-19 pandemic. Our objective was to assess SARS-CoV-2 seroprevalence in a representative sample of pregnant people throughout Canada, across multiple time points over 2 years of the pandemic, to describe the seroprevalence and show the ability of this process to provide prevalence estimates. METHODS: This Canadian retrospective serological surveillance study used existing serological prenatal samples across 10 provinces over multiple time periods: Feb. 3-21, 2020; Aug. 24-Sept. 11, 2020; Nov. 16-Dec. 4, 2020; Nov. 15-Dec. 3, 2021; and results from the province of British Columbia during a period in which the SARS-CoV-2 B.1.1.529 (Omicron) variant was predominant, from Nov. 15, 2021, to June 11, 2022. Age and postal code administrative data allowed for comparison with concurrent polymerase chain reactivity (PCR)-positive results collected by Statistics Canada and the Canadian Surveillance of COVID-19 in Pregnancy (CANCOVID-Preg) project. RESULTS: Seropositivity in antenatal serum as early as February 2020 indicates SARS-CoV-2 transmission before the World Health Organization's declaration of the pandemic. Seroprevalence in our sample of pregnant people was 1.84 to 8.90 times higher than the recorded concurrent PCR-positive prevalence recorded among females aged 20-49 years in November-December 2020. Overall seropositivity in our sample of pregnant people was low at the end of 2020, increasing to 15% in 1 province by the end of 2021. Seroprevalence among pregnant people in BC during the Omicron period increased from 5.8% to 43% from November 2021 to June 2022. INTERPRETATION: These results indicate widespread vulnerability to SARS-CoV-2 infection before vaccine availability in Canada. During the time periods sampled, public health tracking systems were under-reporting infections, and seroprevalence results during the Omicron period indicate extensive community spread of SARS-CoV-2 infection.

Serial cross-sectional estimation of vaccine-and infection-induced SARS-CoV-2 seroprevalence in British Columbia, Canada.

BACKGROUND: The evolving proportion of the population considered immunologically naive versus primed for more efficient immune memory response to SARS-CoV-2 has implications for risk assessment. We sought to chronicle vaccine- and infection-induced seroprevalence across the first 7 waves of the COVID-19 pandemic in British Columbia, Canada. METHODS: During 8 cross-sectional serosurveys conducted between March 2020 and August 2022, we obtained anonymized residual sera from children and adults who attended an outpatient laboratory network in the Lower Mainland (Greater Vancouver and Fraser Valley). We used at least 3 immunoassays per serosurvey to detect SARS-CoV-2 spike and nucleocapsid antibodies. We assessed any seroprevalence (vaccineor infection-induced, or both), defined by positivity on any 2 assays, and infection-induced seroprevalence, also defined by dual-assay positivity but requiring both antinucleocapsid and antispike detection. We used estimates of infection-induced seroprevalence to explore underascertainment of infections by surveillance case reports. RESULTS: By January 2021, we estimated that any seroprevalence remained less than 5%, increasing with vaccine rollout to 56% by May-June 2021, 83% by September-October 2021 and 95% by March 2022. Infection-induced seroprevalence remained less than 15% through September-October 2021, increasing across Omicron waves to 42% by March 2022 and 61% by July-August 2022. By August 2022, 70%-80% of children younger than 20 years and 60%-70% of adults aged 20-59 years had been infected, but fewer than half of adults aged 60 years and older had been infected. Compared with estimates of infection-induced seroprevalence, surveillance case reports underestimated infections 12-fold between September 2021 and March 2022 and 92-fold between March 2022 and August 2022. INTERPRETATION: By August 2022, most children and adults younger than 60 years had evidence of both SARS-CoV-2 vaccination and infection. As previous evidence suggests that a history of both exposures may induce stronger, more durable hybrid immunity than either exposure alone, older adults - who have the lowest infection rates but highest risk of severe outcomes - continue to warrant prioritized vaccination.

Discovery of lead natural products for developing pan-SARS-CoV-2 therapeutics.

The COVID-19 pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), remains a global public health crisis. The reduced efficacy of therapeutic monoclonal antibodies against emerging SARS-CoV-2 variants of concern (VOCs), such as omicron BA.5 subvariants, has underlined the need to explore a novel spectrum of antivirals that are effective against existing and evolving SARS-CoV-2 VOCs. To address the need for novel therapeutic options, we applied cell-based high-content screening to a library of natural products (NPs) obtained from plants, fungi, bacteria, and marine sponges, which represent a considerable diversity of chemical scaffolds. The antiviral effect of 373 NPs was evaluated using the mNeonGreen (mNG) reporter SARS-CoV-2 virus in a lung epithelial cell line (Calu-3). The screening identified 26 NPs with half-maximal effective concentrations (EC50) below 50 µM against mNG-SARS-CoV-2; 16 of these had EC50 values below 10 µM and three NPs (holyrine A, alotaketal C, and bafilomycin D) had EC50 values in the nanomolar range. We demonstrated the pan-SARS-CoV-2 activity of these three lead antivirals against SARS-CoV-2 highly transmissible Omicron subvariants (BA.5, BA.2 and BA.1) and highly pathogenic Delta VOCs in human Calu-3 lung cells. Notably, holyrine A, alotaketal C, and bafilomycin D, are potent nanomolar inhibitors of SARS-CoV-2 Omicron subvariants BA.5 and BA.2. The pan-SARS-CoV-2 activity of alotaketal C [protein kinase C (PKC) activator] and bafilomycin D (V-ATPase inhibitor) suggest that these two NPs are acting as host-directed antivirals (HDAs). Future research should explore whether PKC regulation impacts human susceptibility to and the severity of SARS-CoV-2 infection, and it should confirm the important role of human V-ATPase in the VOC lifecycle. Interestingly, we observed a synergistic action of bafilomycin D and N-0385 (a highly potent inhibitor of human TMPRSS2 protease) against Omicron subvariant BA.2 in human Calu-3 lung cells, which suggests that these two highly potent HDAs are targeting two different mechanisms of SARS-CoV-2 entry. Overall, our study provides insight into the potential of NPs with highly diverse chemical structures as valuable inspirational starting points for developing pan-SARS-CoV-2 therapeutics and for unravelling potential host factors and pathways regulating SARS-CoV-2 VOC infection including emerging omicron BA.5 subvariants.

Correlation of SARS-CoV-2 Viral Neutralizing Antibody Titers with Anti-Spike Antibodies and ACE-2 Inhibition among Vaccinated Individuals.

SARS-CoV-2 anti-spike antibody concentrations and angiotensin converting enzyme-2 (ACE-2) inhibition have been used as surrogates to live viral neutralizing antibody titers; however, validity among vaccinated individuals is unclear. We tested the correlation of these measures among vaccinated participants, and examined subgroups based on duration since vaccination and vaccine dosing intervals. We analyzed 120 samples from two-dose mRNA vaccinees without previous COVID-19. We calculated Spearman correlation coefficients between wild-type viral neutralizing antibody titers and: anti-spike (total and IgG), anti-receptor-binding-domain (RBD), and anti-N-terminal-domain (NTD) antibodies; and ACE-2 binding by RBD. We performed three secondary analyses, dichotomizing samples by the first vaccination-to-blood collection interval, second vaccination-to-blood collection interval, and by the vaccine dosing interval (all groups divided by the median), and compared correlation coefficients (Fisher's Z test). Of 120 participants, 63 (53%) were women, 91 (76%) and 29 (24%) received BNT162b2 and mRNA-1273 vaccines, respectively. Overall, live viral neutralization was correlated with anti-spike total antibody (correlation coefficient = 0.80), anti-spike IgG (0.63), anti-RBD IgG (0.62), anti-NTD IgG (0.64), and RBD ACE2 binding (0.65). Samples with long (>158 days) first vaccination-to-blood collection and long (>71 days) second vaccination-to-blood collection intervals demonstrated higher correlation coefficients, compared with short groups. When comparing cases divided by short (≤39 days) versus long vaccine dosing intervals, only correlation with RBD-ACE-2 binding inhibition was higher in the long group. Among COVID-negative mRNA vaccinees, anti-spike antibody and ACE-2 inhibition concentrations are correlated with live viral neutralizing antibody titers. Correlation was stronger among samples collected at later durations from vaccination. IMPORTANCE Live viral neutralizing antibody titers are an accepted measure of immunity; however, testing procedures are labor-intensive. COVID-19 antibody and angiotensin converting enzyme-2 (ACE-2) levels have been used as surrogates to live viral neutralizing antibody titers; however, validity among vaccinated individuals is unclear. Using samples from 120 two-dose mRNA vaccinees without previous COVID-19, we found that live viral neutralization was correlated with COVID-19 antibody and ACE2 binding levels. When grouping samples by the time interval between vaccination and sample blood collection, samples collected over 158 days after the first vaccine and over 71 days from the second vaccine demonstrated stronger correlation between live viral neutralization titers and both antibody and ACE2 levels, in comparison to those collected earlier.

Performance comparison of micro-neutralization assays based on surrogate SARS-CoV-2 and WT SARS-CoV-2 in assessing virus-neutralizing capacity of anti-SARS-CoV-2 antibodies.

We compared neutralization assays using either the wild-type severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus or surrogate neutralization markers, using characterized sera. We found the results of the neutralization assays 75 % concordant overall and 80 % concordant for samples with high antibody levels. This demonstrates that commercial surrogate SARS-CoV-2 assays offer the potential to assess anti-SARS-CoV-2 antibodies' neutralizing capacity outside CL-3 laboratory containment.

Comparative Analysis of Capillary vs Venous Blood for Serologic Detection of SARS-CoV-2 Antibodies by RPOC Lateral Flow Tests.

A comparison of rapid point-of-care serology tests using finger prick and venous blood was done on 278 participants. In a laboratory setting, immunoglobulin G (IgG) sensitivity neared 100%; however, IgG sensitivity dramatically dropped (82%) in field testing. Possible factors include finger prick volume variability, hemolysis, cassette readability, and operator training.

Evaluation of a high volume antigen test for detection of SARS-CoV-2.

Increasing transmission of SARS-CoV-2 infection in successive waves may strain the capacity of laboratories performing molecular diagnostic testing. Alternative testing approaches may offer additional diagnostic capacity. A high throughput chemiluminescent antigen assay (Ortho VITROS SARS-CoV-2 antigen test) was evaluated using both an inactivated virus preparation and prospective clinical samples (nasopharyngeal swabs in virus transport medium). The limit of detection of the assay was approximately 0.5 TCID50/ml, equivalent to a Ct value of 33. The assay was linear over a wide range. When 528 clinical samples were tested with the antigen assay, the sensitivity was 84.2% and the specificity was 100% (positive predictive value 100% and negative predictive value 97.7%). High volume antigen tests might be used to supplement molecular diagnostic testing capacity.

SARS-CoV-2 serology: Validation of high-throughput chemiluminescent immunoassay (CLIA) platforms and a field study in British Columbia.

BACKGROUND: SARS-CoV-2 antibody testing is required for estimating population seroprevalence and vaccine response studies. It may also increase case identification when used as an adjunct to routine molecular testing. We performed a validation study and evaluated the use of automated high-throughput assays in a field study of COVID-19-affected care facilities. METHODS: Six automated assays were assessed: 1) DiaSorin LIAISONTM SARS-CoV-2 S1/S2 IgG; 2) Abbott ARCHITECTTM SARS-CoV-2 IgG; 3) Ortho VITROSTM Anti-SARS-CoV-2 Total; 4) VITROSTM Anti-SARS-CoV-2 IgG; 5) Siemens SARS-CoV-2 Total Assay; and 6) Roche ElecsysTM Anti-SARS-CoV-2. The validation study included 107 samples (42 known positive; 65 presumed negative). The field study included 296 samples (92 PCR positive; 204 PCR negative or not PCR tested). All samples were tested by the six assays. RESULTS: All assays had sensitivities >90% in the field study, while in the validation study, 5/6 assays were >90% sensitive and DiaSorin was 79% sensitive. Specificities and negative predictive values were >95% for all assays. Field study estimated positive predictive values at 1-10% disease prevalence were 100% for Siemens, Abbott and Roche, while DiaSorin and Ortho assays had lower PPVs at 1% prevalence, but PPVs increased at 5-10% prevalence. In the field study, addition of serology increased diagnoses by 16% compared to PCR testing alone. CONCLUSIONS: All assays evaluated in this study demonstrated high sensitivity and specificity for samples collected at least 14 days post-symptom onset, while sensitivity was variable 0-14 days after infection. The addition of serology to the outbreak investigations increased case detection by 16%.

Practical guidance for clinical laboratories for SARS-CoV-2 serology testing.

The landscape of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) diagnostic testing is rapidly evolving. While serology testing has limited diagnostic capacity for acute infection, its role in providing population-based information on positivity rates and informing evidence-based decision making for public health recommendations is increasing. With the global availability of vaccines, there is increasing pressure on clinical laboratories to provide antibody screening and result interpretation for vaccinated and non-vaccinated individuals. Here we present the most up-to-date data on SARS-CoV-2 antibody timelines, including the longevity of antibodies, and the production and detection of neutralizing antibodies. Additionally, we provide practical guidance for clinical microbiology laboratories to both verify commercial serology assays and choose appropriate testing algorithms for their local populations.

Evaluation of commercial SARS-CoV-2 serological assays in Canadian public health laboratories.

The COVID-19 pandemic has led to the influx of immunoassays for the detection of antibodies towards severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) into the global market. The Canadian Public Health Laboratory Network Serology Task Force undertook a nationwide evaluation of twelve laboratory and 6 point-of-care based commercial serological assays for the detection of SARS-CoV-2 antibodies. We determined that there was considerable variability in the performance of individual tests and that an orthogonal testing algorithm should be prioritized to maximize the accuracy and comparability of results across the country. The manual enzyme immunoassays and point-of-care tests evaluated had lower specificity and increased coefficients of variation compared to automated enzyme immunoassays platforms putting into question their utility for large-scale sero-surveillance. Overall, the data presented here provide a comprehensive approach for applying accurate serological assays for longitudinal sero-surveillance and vaccine trials while informing Canadian public health policy.

Serological survey following SARS-COV-2 outbreaks at long-term care facilities in metro Vancouver, British Columbia: Implications for outbreak management and infection control policies.

A cross-sectional serological survey was carried out in two long-term care facilities that experienced COVID-19 outbreaks in order to evaluate current clinical COVID-19 case definitions. Among individuals with a negative or no previous COVID-19 diagnostic test, myalgias, headache, and loss of appetite were associated with serological reactivity. The US CDC probable case definition was also associated with seropositivity. Public health and infection control practitioners should consider these findings for case exclusion in outbreak settings.

Real-time PCR-based SARS-CoV-2 detection in Canadian laboratories.

With emergence of pandemic COVID-19, rapid and accurate diagnostic testing is essential. This study compared laboratory-developed tests (LDTs) used for the detection of SARS-CoV-2 in Canadian hospital and public health laboratories, and some commercially available real-time RT-PCR assays. Overall, analytical sensitivities were equivalent between LDTs and most commercially available methods.