Structural inequalities in the pathway to COVID-19 immunity (preprint)

Disease outbreaks often highlight existing inequalities and injustices within society. The COVID 19 pandemic has underscored long-existing health inequalities, both within countries and between the Global North and South. These disparities have been observed throughout the pandemic, from disparities in the severity and impact of the initial waves of cases to disparities in who was most protected during the roll-out of vaccination. As the Delta variant surges in many countries, structural inequalities shape the trajectory of the pandemic and exacerbate existing health disparities. In the age of vaccination, the double burden of disparities in both exposure to infection and vaccination coverage intersect to determine the current and future patterns of infection, immunity, and mortality. It is important to consider the ways in which these disparities, with overlapping but distinct drivers, interact to determine population-level immunity and the burden of COVID 19 in different communities. Individuals or communities can experience different pathways to immunity, whether through infection, vaccination, or both. Using San Francisco as a case study, we show how a seroepidemiological approach can illuminate disparities in the pathway to immunity.

SARS-CoV-2 serology across scales: a framework for unbiased seroprevalence estimation incorporating antibody kinetics and epidemic recency (preprint)

Serosurveys are a key resource for measuring SARS-CoV-2 cumulative incidence. A growing body of evidence suggests that asymptomatic and mild infections (together making up over 95% of all infections) are associated with lower antibody titers than severe infections. Antibody levels also peak a few weeks after infection and decay gradually. We developed a statistical approach to produce adjusted estimates of seroprevalence from raw serosurvey results that account for these sources of spectrum bias. We incorporate data on antibody responses on multiple assays from a post-infection longitudinal cohort, along with epidemic time series to account for the timing of a serosurvey relative to how recently individuals may have been infected. We applied this method to produce adjusted seroprevalence estimates from five large-scale SARS-CoV-2 serosurveys across different settings and study designs. We identify substantial differences between reported and adjusted estimates of over two-fold in the results of some surveys, and provide a tool for practitioners to generate adjusted estimates with pre-set or custom parameter values. While unprecedented efforts have been launched to generate SARS-CoV-2 seroprevalence estimates over this past year, interpretation of results from these studies requires properly accounting for both population-level epidemiologic context and individual-level immune dynamics.

multiSero: open multiplex-ELISA platform for analyzing antibody responses to SARS-CoV-2 infection (preprint)

Serology has provided valuable diagnostic and epidemiological data on antibody responses to SARS-CoV-2 in diverse patient cohorts. Deployment of high content, multiplex serology platforms across the world, including in low and medium income countries, can accelerate longitudinal epidemiological surveys. Here we report multiSero, an open platform to enable multiplex serology with up to 48 antigens in a 96-well format. The platform consists of three components: ELISA-array of printed proteins, a commercial or home-built plate reader, and modular python software for automated analysis (pysero). We validate the platform by comparing antibody titers against the SARS-CoV-2 Spike, receptor binding domain (RBD), and nucleocapsid (N) in 114 sera from COVID-19 positive individuals and 87 pre-pandemic COVID-19 negative sera. We report data with both a commercial plate reader and an inexpensive, open plate reader (nautilus). Receiver operating characteristic (ROC) analysis of classification with single antigens shows that Spike and RBD classify positive and negative sera with the highest sensitivity at a given specificity. The platform distinguished positive sera from negative sera when the reactivity of the sera was equivalent to the binding of 1 ng mL-1 RBD-specific monoclonal antibody. We developed normalization and classification methods to pool antibody responses from multiple antigens and multiple experiments. Our results demonstrate a performant and accessible pipeline for multiplexed ELISA ready for multiple applications, including serosurveillance, identification of viral proteins that elicit antibody responses, differential diagnosis of circulating pathogens, and immune responses to vaccines.

SARS-CoV-2 antibody magnitude and detectability are driven by disease severity, timing, and assay (preprint)

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.

Long-Term SARS-CoV-2-Specific Immune and Inflammatory Responses Across a Clinically Diverse Cohort of Individuals Recovering from COVID-19 (preprint)

A detailed understanding of long-term SARS-CoV-2-specific T cell responses and their relationship to humoral immunity and markers of inflammation in diverse groups of individuals representing the spectrum of COVID-19 illness and recovery is urgently needed. Data are also lacking as to whether and how adaptive immune and inflammatory responses differ in individuals that experience persistent symptomatic sequelae months following acute infection compared to those with complete, rapid recovery. We measured SARS-CoV-2-specific T cell responses, soluble markers of inflammation, and antibody levels and neutralization capacity longitudinally up to 9 months following infection in a diverse group of 70 individuals with PCR-confirmed SARS-CoV-2 infection. The participants had varying degrees of initial disease severity and were enrolled in the northern California Long-term Impact of Infection with Novel Coronavirus (LIINC) cohort. Adaptive T cell responses remained remarkably stable in all participants across disease severity during the entire study interval. Whereas the magnitude of the early CD4+ T cell immune response is determined by the severity of initial infection (participants requiring hospitalization or intensive care), pre-existing lung disease was significantly associated with higher long-term SARS-CoV2-specific CD8+ T cell responses, independent of initial disease severity or age. Neutralizing antibody levels were strongly correlated with SARS-CoV-2-specific CD4+ T but not CD8+ T cell responses. Importantly, we did not identify substantial differences in long-term virus-specific T cell or antibody responses between participants with and without COVID-19-related symptoms that persist months after initial infection.

Universal PCR and antibody testing demonstrate little to no transmission of SARS-CoV-2 in a rural community (preprint)

BackgroundThe absence of systematic surveillance for SARS-CoV-2 has curtailed accurate appraisal of transmission intensity. Our objective was to perform case detection of an entire rural community to quantify SARS-CoV-2 transmission using PCR and antibody testing. MethodsWe conducted a cross-sectional survey of the prevalence and cumulative incidence of SARSCoV-2 infection in the rural town of Bolinas, California (population 1,620), four weeks following shelter-in-place orders. Residents and county essential workers were tested between April 20th - 24th, 2020. Prevalence by PCR and seroprevalence combining data from two forms of antibody testing were performed in parallel (Abbott ARCHITECT IgG to nucleocapsid protein and in-house IgG ELISA to the receptor binding domain). ResultsOf 1,891 participants, 1,312 were confirmed Bolinas residents (>80% community ascertainment). Zero participants were PCR positive. Assuming 80% sensitivity, it would have been unlikely to observe these results (p< 0.05) if there were > 3 active infections in the community. Based on antibody results, estimated prevalence of prior infection was 0.16% (95% CrI: 0.02%, 0.46%). Seroprevalence estimates using only one of the two tests would have been higher, with greater uncertainty. The positive predictive value (PPV) of a positive result on both tests was 99.11% (95% CrI: 95.75%, 99.94%), compared to PPV 44.19%-63.32% (95% CrI range 3.25%-98.64%) if only one test was utilized. ConclusionsFour weeks following shelter-in-place, active and prior SARS-CoV-2 infection in a rural Northern California community was extremely rare. In this low prevalence setting, use of two antibody tests increased the PPV and precision of seroprevalence estimates.