Your browser doesn't support javascript.
Show: 20 | 50 | 100
Results 1 - 15 de 15
Filter
Add filters

Document Type
Year range
1.
Preprint in English | medRxiv | ID: ppmedrxiv-21266786

ABSTRACT

Serological surveillance studies of infectious diseases provide population-level estimates of infection and antibody prevalence, generating crucial insight into population-level immunity, risk factors leading to infection, and effectiveness of public health measures. These studies traditionally rely on detection of pathogen-specific antibodies in samples derived from venipuncture, an expensive and logistically challenging aspect of serological surveillance. During the COVID-19 pandemic, guidelines implemented to prevent the spread of SARS-CoV-2 infection made collection of venous blood logistically difficult at a time when SARS-CoV-2 serosurveillance was urgently needed. Dried blood spots (DBS) have generated interest as an alternative to venous blood for SARS-CoV-2 serological applications due to their stability, low cost, and ease of collection; DBS samples can be self-generated via fingerprick by community members and mailed at ambient temperatures. Here, we detail the development of four DBS-based SARS-CoV-2 serological methods and demonstrate their implementation in a large serological survey of community members from 12 cities in the East Bay region of the San Francisco metropolitan area using at- home DBS collection. We find that DBS perform similarly to plasma/serum in enzyme-linked immunosorbent assays and commercial SARS-CoV-2 serological assays. In addition, we show that DBS samples can reliably detect antibody responses months post-infection and track antibody kinetics after vaccination. Implementation of DBS enabled collection of valuable serological data from our study population to investigate changes in seroprevalence over an eight-month period. Our work makes a strong argument for the implementation of DBS in serological studies, not just for SARS-CoV-2, but any situation where phlebotomy is inaccessible.

2.
Preprint in English | medRxiv | ID: ppmedrxiv-21265622

ABSTRACT

BackgroundCOVID-19 convalescent plasma (CCP) was widely used as passive immunotherapy during the first waves of SARS-CoV-2 infection in the US. However, based on observational studies and randomized controlled trials, beneficial effects of CCP were limited, and its use was virtually discontinued early in 2021, in concurrence with increased vaccination rates and availability of monoclonal antibody (mAb) therapeutics. However, as new variants of the SARS-CoV-2 spread, interest in CCP derived from vaccine-boosted CCP donors is resurging. The effect of vaccination of previously infected CCP donors on antibodies against rapidly spreading variants of concern (VOC) is still under investigation. Study Design/MethodsIn this study, paired samples from 11 CCP donors collected before and after vaccination were tested to measure binding antibodies levels and neutralization activity against the ancestral and SARS-CoV-2 variants (Wuhan-Hu-1, B.1.1.7, B.1.351, P.1, D614G, B.1.617.2, B.1.427) on the Ortho Vitros Spike Total Ig and IgG assays, the MSD V-PLEX SARS-CoV-2 Panel 6 arrays for IgG binding and ACE2 inhibition, and variant-specific Spike Reporter Viral Particle Neutralization (RVPN) assays. Results/FindingsBinding and neutralizing antibodies were significantly boosted by vaccination, with several logs higher neutralization for all the variants tested post-vaccination compared to the pre-vaccination samples, with no difference found among the individual variants. DiscussionVaccination of previously infected individuals boosts antibodies including neutralizing activity against all SARS-CoV-2 VOC, including the current spreading delta (B.1.617.2) variant. Animal model and human studies to assess clinical efficacy of vaccine boosted CCP are warranted, especially since 15-20% of current donations in the US are from previously infected vaccine-boosted donors.

3.
Preprint in English | medRxiv | ID: ppmedrxiv-21262414

ABSTRACT

SARS-CoV-2 serosurveys can estimate cumulative incidence for monitoring epidemics but require characterization of employed serological assays performance to inform testing algorithm development and interpretation of results. We conducted a multi-laboratory evaluation of 21 commercial high-throughput SARS-CoV-2 serological assays using blinded panels of 1,000 highly-characterized blood-donor specimens. Assays demonstrated a range of sensitivities (96%-63%), specificities (99%-96%) and precision (IIC 0.55-0.99). Durability of antibody detection in longitudinal samples was dependent on assay format and immunoglobulin target, with anti-spike, direct, or total Ig assays demonstrating more stable, or increasing reactivity over time than anti-nucleocapsid, indirect, or IgG assays. Assays with high sensitivity, specificity and durable antibody detection are ideal for serosurveillance. Less sensitive assays demonstrating waning reactivity are appropriate for other applications, including characterizing antibody responses after infection and vaccination, and detection of anamnestic boosting by reinfections and vaccine breakthrough infections. Assay performance must be evaluated in the context of the intended use.

4.
Preprint in English | bioRxiv | ID: ppbiorxiv-458520

ABSTRACT

Early in the SARS-CoV-2 pandemic, there was a high level of optimism based on observational studies and small controlled trials that treating hospitalized patients with convalescent plasma from COVID-19 survivors (CCP) would be an important immunotherapy. However, as more data from controlled trials became available, the results became disappointing, with at best moderate evidence of efficacy when CCP with high titers of neutralizing antibodies was used early in infection. To better understand the potential therapeutic efficacy of CCP, and to further validate SARS-CoV-2 infection of macaques as a reliable animal model for testing such strategies, we inoculated 12 adult rhesus macaques with SARS-CoV-2 by intratracheal and intranasal routes. One day later, 8 animals were infused with pooled human CCP with a high titer of neutralizing antibodies (RVPN NT50 value of 3,003), while 4 control animals received normal human plasma. Animals were monitored for 7 days. Animals treated with CCP had detectable levels of antiviral antibodies after infusion. In comparison to the control animals, they had similar levels of virus replication in the upper and lower respiratory tract, but had significantly reduced interstitial pneumonia, as measured by comprehensive lung histology. By highlighting strengths and weaknesses, data of this study can help to further optimize nonhuman primate models to provide proof-of-concept of intervention strategies, and guide the future use of convalescent plasma against SARS-CoV-2 and potentially other newly emerging respiratory viruses. Author summaryThe results of treating SARS-CoV-2 infected hospitalized patients with COVID-19 convalescent plasma (CCP), collected from survivors of natural infection, have been disappointing. The available data from various studies indicate at best moderate clinical benefits only when CCP with high titer of neutralizing antibodies was infused early in infection. The macaque model of SARS-CoV-2 infection can be useful to gain further insights in the value of CCP therapy. In this study, animals were infected with SARS-CoV-2 and the next day, were infused with pooled human convalescent plasma, selected to have a very high titer of neutralizing antibodies. While administration of CCP did not result in a detectable reduction in virus replication in the respiratory tract, it significantly reduced lung inflammation. These data, combined with the results of monoclonal antibody studies, emphasize the need to use products with high titers of neutralizing antibodies, and guide the future development of CCP-based therapies.

5.
Preprint in English | medRxiv | ID: ppmedrxiv-21255576

ABSTRACT

IntroductionThe REDS-IV-P Epidemiology, Surveillance and Preparedness of the Novel SARS-CoV-2 Epidemic (RESPONSE) seroprevalence study conducted monthly cross-sectional testing for SARS-CoV-2 antibodies on blood donors in six U.S. metropolitan regions to estimate the extent of SARS-COV-2 infections over time. Study Design/MethodsDuring March-August 2020, approximately [≥]1,000 serum specimens were collected monthly from each region and tested for SARS-CoV-2 antibodies using a well-validated algorithm. Regional seroprevalence estimates were weighted based on demographic differences with the general population. Seroprevalence was compared with reported COVID-19 case rates over time. Results/FindingsFor all regions, seroprevalence was <1.0% in March 2020. New York experienced the biggest increase (peak seroprevalence, 15.8 % in May). All other regions experienced modest increases in seroprevalence(1-2% in May-June to 2-4% in July-August). Seroprevalence was higher in younger, non-Hispanic Black, and Hispanic donors. Temporal increases in donor seroprevalence correlated with reported case rates in each region. In August, 1.3-5.6 estimated cumulative infections (based on seroprevalence data) per COVID-19 case reported to CDC. ConclusionIncreases in seroprevalence were found in all regions, with the largest increase in New York. Seroprevalence was higher in non-Hispanic Black and Hispanic blood donors than in non-Hispanic White blood donors. SARS-CoV-2 antibody testing of blood donor samples can be used to estimate the seroprevalence in the general population by region and demographic group. The methods derived from the RESPONSE seroprevalence study served as the basis for expanding SARS-CoV-2 seroprevalence surveillance to all 50 states and Puerto Rico. SummarySARS-CoV-2 serosurveillance data from blood donors in 6 US regions were used to estimate population weighted seroprevalence. Seroprevelance rates were higher in case rates. The study was expanded to a national donor serosurveillance program. DisclaimerThe content is solely the responsibility of the authors and does not represent the policy of the National Institutes of Health or the Department of Health and Human Services. Any specific brandnames included in this manuscript are for identification purposes only and are not intended to represent an endorsement by CDC. The findings and conclusions in this report are those of the authorsand do not necessarily represent the official position of the Centers of Disease Control and Prevention.

6.
Preprint in English | medRxiv | ID: ppmedrxiv-21254260

ABSTRACT

BackgroundAntibody response duration following SARS-CoV-2 infection tends to be variable and depends on severity of disease and method of detection. Study design and methodsCOVID-19 convalescent plasma (CCP) from 18 donors was collected longitudinally for a maximum of 63 - 129 days following resolution of symptoms. All the samples were initially screened by the Ortho Total Ig test to confirm positivity and subsequently tested with 7 additional direct sandwich or indirect binding assays (Ortho, Roche, Abbott, Broad Institute) directed against a variety of antigen targets (S1, RBD, and NC), along with 2 neutralization assays (Broad Institute live virus PRNT and Vitalant Research Institute Pseudovirus RVPN). ResultsThe direct detection assays (Ortho Total Ig total and Roche Total Ig) showed increasing levels of antibodies over the time period, in contrast to the indirect IgG assays that showed a decline. Neutralization assays also demonstrated declining responses; the VRI RVPN pseudovirus had a greater rate of decline than the Broad PRNT live virus assay. DiscussionThese data show that in addition to variable individual responses and associations with disease severity, the detection assay chosen contributes to the heterogeneous results in antibody stability over time. Depending on the scope of the research, one assay may be preferable over another. For serosurveillance studies, direct, double Ag-sandwich assays appear to be the best choice due to their stability; in particular, algorithms that include both S1 and NC based assays can help reduce the rate of false-positivity and discriminate between natural infection and vaccine-derived seroreactivity.

7.
Preprint in English | medRxiv | ID: ppmedrxiv-21251639

ABSTRACT

Serosurveillance studies are critical for estimating SARS-CoV-2 transmission and immunity, but interpretation of results is currently limited by poorly defined variability in the performance of antibody assays to detect seroreactivity over time in individuals with different clinical presentations. We measured longitudinal antibody responses to SARS-CoV-2 in plasma samples from a diverse cohort of 128 individuals over 160 days using 14 binding and neutralization assays. For all assays, we found a consistent and strong effect of disease severity on antibody magnitude, with fever, cough, hospitalization, and oxygen requirement explaining much of this variation. We found that binding assays measuring responses to spike protein had consistently higher correlation with neutralization than those measuring responses to nucleocapsid, regardless of assay format and sample timing. However, assays varied substantially with respect to sensitivity during early convalescence and in time to seroreversion. Variations in sensitivity and durability were particularly dramatic for individuals with mild infection, who had consistently lower antibody titers and represent the majority of the infected population, with sensitivities often differing substantially from reported test characteristics (e.g., amongst commercial assays, sensitivity at 6 months ranged from 33% for ARCHITECT IgG to 98% for VITROS Total Ig). Thus, the ability to detect previous infection by SARS-CoV-2 is highly dependent on the severity of the initial infection, timing relative to infection, and the assay used. These findings have important implications for the design and interpretation of SARS-CoV-2 serosurveillance studies.

8.
Preprint in English | bioRxiv | ID: ppbiorxiv-300871

ABSTRACT

A coronavirus antigen microarray (COVAM) was constructed containing 11 SARS-CoV-2, 5 SARS-1, 5 MERS, and 12 seasonal coronavirus recombinant proteins. The array is designed to measure immunoglobulin isotype and subtype levels in serum or plasma samples against each of the individual antigens printed on the array. We probed the COVAM with COVID-19 convalescent plasma (CCP) collected from 99 donors who recovered from a PCR+ confirmed SARS-CoV-2 infection. The results were analyzed using two computational approaches, a generalized linear model (glm) and Random Forest (RF) prediction model, to classify individual specimens as either Reactive or Non-Reactive against the SARS-CoV-2 antigens. A training set of 88 pre-COVID-19 specimens (PreCoV) collected in August 2019 and102 positive specimens from SARS-CoV-2 PCR+ confirmed COVID-19 cases was used for these analyses. Results compared with an FDA emergency use authorized (EUA) SARS-CoV2 S1-based total Ig chemiluminescence immunoassay (Ortho Clinical Diagnostics VITROS(R) Anti-SARS-CoV-2 Total, CoV2T) and with a SARS-CoV-2 S1-S2 spike-based pseudovirus micro neutralization assay (SARS-CoV-2 reporter viral particle neutralization titration (RVPNT) showed high concordance between the 3 assays. Three CCP specimens that were negative by the VITROS CoV2T immunoassay were also negative by both COVAM and the RVPNT assay. Concordance between VITROS CoV2T and COVAM was 96%, VITROS CoV2T and RVPNT 93%, and RVPNT and COVAM 95%. The discordances were all weakly reactive samples near the cutoff threshold of the VITROS CoV2T immunoassay. The multiplex COVAM allows CCP to be grouped according to antibody reactivity patterns against 11 SARS-CoV-2 antigens. Unsupervised K-means analysis, via the gap statistics, as well as hierarchical clustering analysis revealed 3 main clusters with distinct reactivity intensities and patterns. These patterns were not recapitulated by adjusting the VITROS CoV2T or RVPNT assay thresholds. Plasma classified according to these reactivity patterns may be better associated with CCP treatment efficacy than antibody levels alone. The use of a SARS-CoV-2 antigen array may be useful to qualify CCP for administration as a treatment for acute COVID-19 and to interrogate vaccine immunogenicity and performance in preclinical and clinical studies to understand and recapitulate antibody responses associated with protection from infection and disease.

9.
Preprint in English | medRxiv | ID: ppmedrxiv-20184895

ABSTRACT

BACKGROUNDEfficacy of COVID-19 convalescent plasma (CCP) to treat COVID-19 is hypothesized to be associated with the concentration of neutralizing antibodies (nAb) to SARS-CoV-2. High capacity serologic assays detecting binding antibodies (bAb) have been developed, nAb assays are not adaptable to high-throughput testing. We sought to determine the effectiveness of using surrogate bAb signal-to-cutoff ratios (S/CO) in predicting nAb titers using a pseudovirus reporter viral particle neutralization (RVPN) assay. METHODSCCP donor serum collected by 3 US blood collectors was tested with a bAb assay (Ortho Clinical Diagnostics VITROS Anti-SARS-CoV-2 Total, CoV2T) and a nAb RVPN assay. CoV2T prediction effectiveness at S/CO thresholds was evaluated for RVPN nAb NT50 titers using receiver operating characteristic analysis. RESULTS753 CCPs were tested with median CoV2T S/CO of 71.2 and median NT50 of 527.5. Proportions of CCP donors with NT50 over various target nAb titers were 86% [≥]1:80, 76% [≥]1:160, and 62%[≥]1:320. Increasing CoV2Ts reduced the sensitivity to predict NT50 titers, while specificity to identify those below thresholds increased. As the targeted NT50 increased, the positive predictive value fell with reciprocal increase in negative predictive value. S/CO thresholds were thus less able to predict target NT50 titers. CONCLUSIONSelection of a clinically effective nAb titer will impact availability of CCP. Product release with CoV2T assay S/CO thresholds must balance the risk of releasing products below target nAb titers with the cost of false negatives. A two-step testing scheme may be optimal, with nAb testing on CoV2T samples with S/COs below thresholds.

10.
Preprint in English | bioRxiv | ID: ppbiorxiv-222943

ABSTRACT

A high-resolution understanding of the antibody response to SARS-CoV-2 is important for the design of effective diagnostics, vaccines and therapeutics. However, SARS-CoV-2 antibody epitopes remain largely uncharacterized, and it is unknown whether and how the response may cross-react with related viruses. Here, we use a multiplexed peptide assay ( PepSeq) to generate an epitope-resolved view of reactivity across all human coronaviruses. PepSeq accurately detects SARS-CoV-2 exposure and resolves epitopes across the Spike and Nucleocapsid proteins. Two of these represent recurrent reactivities to conserved, functionally-important sites in the Spike S2 subunit, regions that we show are also targeted for the endemic coronaviruses in pre-pandemic controls. At one of these sites, we demonstrate that the SARS-CoV-2 response strongly and recurrently cross-reacts with the endemic virus hCoV-OC43. Our analyses reveal new diagnostic and therapeutic targets, including a site at which SARS-CoV-2 may recruit common pre-existing antibodies and with the potential for broadly-neutralizing responses.

11.
Preprint in English | bioRxiv | ID: ppbiorxiv-191007

ABSTRACT

CD4 T follicular helper (Tfh) cells are important for the generation of long-lasting and specific humoral protection against viral infections. The degree to which SARS-CoV-2 infection generates Tfh cells and stimulates the germinal center response is an important question as we investigate vaccine options for the current pandemic. Here we report that, following infection with SARS-CoV-2, adult rhesus macaques exhibited transient accumulation of activated, proliferating Tfh cells in their peripheral blood on a transitory basis. The CD4 helper cell responses were skewed predominantly toward a Th1 response in blood, lung, and lymph nodes, reflective of the interferon-rich cytokine environment following infection. We also observed the generation of germinal center Tfh cells specific for the SARS-CoV-2 spike (S) and nucleocapsid (N) proteins, and a corresponding early appearance of antiviral serum IgG antibodies but delayed or absent IgA antibodies. Our data suggest that a vaccine promoting Th1-type Tfh responses that target the S protein may lead to protective immunity.

12.
Preprint in English | medRxiv | ID: ppmedrxiv-20105692

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has led to more than 4 million confirmed infections worldwide and over 300,000 deaths. While Remdesivir has recently received FDA emergency use authorization for treatment of SARS-CoV-2 infection, convalescent plasma (CP) with high titers of SARS-CoV-2 neutralizing antibodies (NAbs) from recovered donors remains a promising and widely accessible method to mitigate severe disease symptoms. Here, we describe the development and validation of a cell-free neutralization PCR assay using SARS-CoV-2 spike protein S1 and human ACE2 receptor-DNA conjugates. By comparing with samples collected prior to the outbreak, we confirmed that NAbs were specifically detected in COVID-19 cases. Using our unique assay, the NAb signals are detectable as early as 10 days after onset of symptoms and continue to rise, plateauing after 18 days. Notably, we showed that the use of a licensed pathogen reduction technology to inactivate potentially contaminating infectious pathogens in CP did not alter NAb signals, paving a path to safely administer effective CP therapies. The described neutralization PCR assay can serve as a qualification tool to easily identify suitable CP donors of a potentially lifesaving therapy. In addition, this assay tool is readily deployable in standard laboratories with biosafety level 2 capability, and can yield results within 2-3 hrs. This advancement can facilitate research on factors driving diverse COVID-19 disease manifestations, help evaluate the impact of various CP processing protocols on CP therapeutic efficacy and assist in accelerating vaccine efficacy assessment.

13.
Preprint in English | medRxiv | ID: ppmedrxiv-20107482

ABSTRACT

We report very low SARS-CoV-2 seroprevalence in two San Francisco Bay Area populations. Seropositivity was 0.26% in 387 hospitalized patients admitted for non-respiratory indications and 0.1% in 1,000 blood donors. We additionally describe the longitudinal dynamics of immunoglobulin-G, immunoglobulin-M, and in vitro neutralizing antibody titers in COVID-19 patients. Neutralizing antibodies rise in tandem with immunoglobulin levels following symptom onset, exhibiting median time to seroconversion within one day of each other, and there is >93% positive percent agreement between detection of immunoglobulin-G and neutralizing titers.

14.
Preprint in English | medRxiv | ID: ppmedrxiv-20092528

ABSTRACT

Serologic assays are needed to determine SARS-CoV-2 seroprevalence, but poor specificity can overestimate exposures. Here, we built a pan-human coronavirus proteome-wide programmable phage display assay (VirScan) to profile coronavirus antigens specifically enriched by 20 COVID-19 patient serum IgG. With ReScan, a new diagnostic development workflow which combines the isolation of phage expressing the most immunogenic peptides with paper-based microarrays manufactured via acoustic liquid handling, we identified 9 candidate antigens from a library of 534 SARS-CoV-2 peptides. These arrays could form the basis of a multiplexed COVID-19 serologic assay with enhanced specificity. ReScan has broad applicability for serologic assay development.

15.
Preprint in English | bioRxiv | ID: ppbiorxiv-043364

ABSTRACT

The current practice for diagnosis of COVID-19, based on SARS-CoV-2 PCR testing of pharyngeal or respiratory specimens in a symptomatic patient at high epidemiologic risk, likely underestimates the true prevalence of infection. Serologic methods can more accurately estimate the disease burden by detecting infections missed by the limited testing performed to date. Here, we describe the validation of a coronavirus antigen microarray containing immunologically significant antigens from SARS-CoV-2, in addition to SARS-CoV, MERS-CoV, common human coronavirus strains, and other common respiratory viruses. A comparison of antibody profiles detected on the array from control sera collected prior to the SARS-CoV-2 pandemic versus convalescent blood specimens from virologically confirmed COVID-19 cases demonstrates near complete discrimination of these two groups, with improved performance from use of antigen combinations that include both spike protein and nucleoprotein. This array can be used as a diagnostic tool, as an epidemiologic tool to more accurately estimate the disease burden of COVID-19, and as a research tool to correlate antibody responses with clinical outcomes.

SELECTION OF CITATIONS
SEARCH DETAIL
...