Dynamics of SARS-CoV-2 Seroprevalence in a Large US population Over a Period of 12 Months (preprint)

Due to a combination of asymptomatic or undiagnosed infections, the proportion of the United States population infected with SARS-CoV-2 was unclear from the beginning of the pandemic. We previously established a platform to screen for SARS-CoV-2 positivity across a representative proportion of the US population, from which we reported that almost 17 million Americans were estimated to have had undocumented infections in the Spring of 2020. Since then, vaccine rollout and prevalence of different SARS-CoV-2 variants have further altered seropositivity trends within the United States population. To explore the longitudinal impacts of the pandemic and vaccine responses on seropositivity, we re-enrolled participants from our baseline study in a 6- and 12- month follow-up study to develop a longitudinal antibody profile capable of representing seropositivity within the United States during a critical period just prior to and during the initiation of vaccine rollout. Initial measurements showed that, since July 2020, seropositivity elevated within this population from 4.8% at baseline to 36.2% and 89.3% at 6 and 12 months, respectively. We also evaluated nucleocapsid seropositivity and compared to spike seropositivity to identify trends in infection versus vaccination relative to baseline. These data serve as a window into a critical timeframe within the COVID-19 pandemic response and serve as a resource that could be used in subsequent respiratory illness outbreaks.

Immune response to COVID-19 vaccines in people with immunodeficiencies (preprint)

COVID-19 vaccine research in immune-deficient/disordered people (IDP) has primarily focused on cancer and organ transplantation populations. We followed 195 IDP with varied immune disorders and 35 healthy volunteers (HV) from April 2021-April 2022. Anti-spike IgG and angiotensin-converting enzyme 2 pseudo-neutralization were measured though six months post-dose 3. Anti-spike IgG was detected in 93% of IDP by six months post-dose 3. IgG dynamics in IDP and HV were similar, though median IgG levels for IDP were <33% of HV at all timepoints. IgG concentrations were lower against Omicron BA.1 than other variants at all timepoints. Pseudo-neutralization capacity was modestly correlated with anti-spike IgG concentration, but was especially low for Omicron BA.1. Post-vaccination adverse events were minimal. Results were largely unaffected by participants’ immunomodulatory medication and treatments. COVID-19 vaccines are safe, induce anti-spike IgG in most IDP, and should be more strongly recommended for people with immunodeficiencies.

Recent SARS-CoV-2 infection abrogates antibody and B-cell responses to booster vaccination (preprint)

SARS-CoV-2 mRNA booster vaccines provide protection from severe disease, eliciting strong immunity that is further boosted by previous infection. However, it is unclear whether these immune responses are affected by the interval between infection and vaccination. Over a two-month period, we evaluated antibody and B-cell responses to a third dose mRNA vaccine in 66 individuals with different infection histories. Uninfected and post-boost but not previously infected individuals mounted robust ancestral and variant spike-binding and neutralizing antibodies, and memory B cells. Spike-specific B-cell responses from recent infection were elevated at pre-boost but comparatively less so at 60 days post-boost compared to uninfected individuals, and these differences were linked to baseline frequencies of CD27lo B cells. Day 60 to baseline ratio of BCR signaling measured by phosphorylation of Syk was inversely correlated to days between infection and vaccination. Thus, B-cell responses to booster vaccines are impeded by recent infection.

COVID-19 redux: clinical, virologic, and immunologic evaluation of clinical rebound after nirmatrelvir/ritonavir (preprint)

Clinical rebound of COVID-19 after nirmatrelvir/ritonavir treatment has been reported. We performed clinical, virologic, and immune measurements in seven patients with symptomatic rebound, six after nirmatrelvir/ritonavir treatment and one without previous treatment. There was no evidence of severe disease or impaired antibody and T-cell responses in people with rebound symptoms.

Undiagnosed COVID-19 in households with a child with mitochondrial disease (preprint)

BackgroundThe impact of the COVID-19 pandemic on medically fragile populations, who are at higher risk of severe illness and sequelae, has not been well characterized. Viral infection is a major cause of morbidity in children with mitochondrial disease (MtD), and the COVID-19 pandemic represents an opportunity to study this vulnerable population. MethodsA convenience sampling cross-sectional serology study was conducted (October 2020 to June 2021) in households (N = 20) containing a child with MtD (N = 22). Samples (N = 83) were collected in the home using a microsampling apparatus and shipped to investigators. Antibodies against SARS-CoV-2 nucleocapsid (IgG), spike protein (IgG, IgM, IgA), and receptor binding domain (IgG, IgM, IgA) were determined by enzyme linked immunosorbent assay. ResultsWhile only 4.8% of participants were clinically diagnosed for SARS-CoV-2 infection, 75.9% of study participants were seropositive for SARS-CoV-2 antibodies. Most samples were IgM positive for spike or RBD (70%), indicating that infection was recent. This translated to all 20 families showing evidence of infection in at least one household member. For the children with MtD, 91% had antibodies against SARS-CoV-2 and had not experienced any adverse outcomes at the time of assessment. For children with recent infections (IgM+ only), serologic data suggest household members as a source. ConclusionsCOVID-19 was highly prevalent and undiagnosed in households with a child with MtD through the 2020-2021 winter wave of the pandemic. In this first major wave, children with MtD tolerated SARS-CoV-2 infection well, potentially due to household adherence to CDC recommendations for risk mitigation. FundingThis study was funded by the Intramural Research Program of the National Institutes of Health (HG200381-03). Clinical trial numberNCT04419870

Robust, persistent adaptive immune responses to SARS-CoV-2 in the oropharyngeal lymphoid tissue of children (preprint)

SARS-CoV-2 infection triggers adaptive immune responses from both T and B cells. However, most studies focus on peripheral blood, which may not fully reflect immune responses in lymphoid tissues at the site of infection. To evaluate both local and systemic adaptive immune responses to SARS-CoV-2, we collected peripheral blood, tonsils, and adenoids from 110 children undergoing tonsillectomy/adenoidectomy during the COVID-19 pandemic and found 24 with evidence of prior SARS-CoV-2 infection, including detectable neutralizing antibodies against multiple viral variants. We identified SARS-CoV-2-specific germinal center (GC) and memory B cells; single cell BCR sequencing showed that these virus-specific B cells were class-switched and somatically hypermutated, with overlapping clones in the adenoids and tonsils. Oropharyngeal tissues from COVID-19-convalescent children showed persistent expansion of GC and anti-viral lymphocyte populations associated with an IFN-γ-type response, with particularly prominent changes in the adenoids, as well as evidence of persistent viral RNA in both tonsil and adenoid tissues of many participants. Our results show robust, tissue-specific adaptive immune responses to SARS-CoV-2 in the upper respiratory tract of children weeks to months after acute infection, providing evidence of persistent localized immunity to this respiratory virus.

SARS-CoV-2 Seroprevalence and Drug Use in Trauma Patients from Six Sites in the United States (preprint)

In comparison to the general patient population, trauma patients show higher level detections of bloodborne infectious diseases, such as Hepatitis and Human Immunodeficiency Virus. In comparison to bloodborne pathogens, the prevalence of respiratory infections such as SARS-CoV-2 and how that relates with other variables, such as drug usage and trauma type, is currently unknown in trauma populations. Here, we evaluated SARS-CoV-2 seropositivity and antibody isotype profile in 2,542 trauma patients from six Level-1 trauma centers between April and October of 2020 during the first wave of the COVID-19 pandemic. We found that the seroprevalence in trauma victims 18-44 years old (9.79%, 95% confidence interval/CI: 8.33 11.47) was much higher in comparison to older patients (45-69 years old: 6.03%, 4.59-5.88; 70+ years old: 4.33%, 2.54 - 7.20). Black/African American (9.54%, 7.77 - 11.65) and Hispanic/Latino patients (14.95%, 11.80 - 18.75) also had higher seroprevalence in comparison, respectively, to White (5.72%, 4.62 7.05) and Non-Latino patients (6.55%, 5.57 - 7.69). More than half (55.54%) of those tested for drug toxicology had at least one drug present in their system. Those that tested positive for narcotics or sedatives had a significant negative correlation with seropositivity, while those on anti-depressants trended positive. These findings represent an important consideration for both the patients and first responders that treat trauma patients facing potential risk of respiratory infectious diseases like SARS-CoV-2.

Mapping a Pandemic: SARS-CoV-2 Seropositivity in the United States (preprint)

Asymptomatic SARS-CoV-2 infection and delayed implementation of diagnostics have led to poorly defined viral prevalence rates. To address this, we analyzed seropositivity in US adults who have not previously been diagnosed with COVID-19. Individuals with characteristics that reflect the US population (n = 11,382) and who had not previously been diagnosed with COVID-19 were selected by quota sampling from 241,424 volunteers (ClinicalTrials.gov NCT04334954). Enrolled participants provided medical, geographic, demographic, and socioeconomic information and 9,028 blood samples. The majority (88.7%) of samples were collected between May 10th and July 31st, 2020. Samples were analyzed via ELISA for anti-Spike and anti-RBD antibodies. Estimation of seroprevalence was performed by using a weighted analysis to reflect the US population. We detected an undiagnosed seropositivity rate of 4.6% (95% CI: 2.6 - 6.5%). There was distinct regional variability, with heightened seropositivity in locations of early outbreaks. Subgroup analysis demonstrated that the highest estimated undiagnosed seropositivity within groups was detected in younger participants (ages 18-45, 5.9%), females (5.5%), Black/African American (14.2%), Hispanic (6.1%), and Urban residents (5.3%), and lower undiagnosed seropositivity in those with chronic diseases. During the first wave of infection over the spring/summer of 2020 an estimate of 4.6% of adults had a prior undiagnosed SARS-CoV-2 infection. These data indicate that there were 4.8 (95% CI: 2.8-6.8) undiagnosed cases for every diagnosed case of COVID-19 during this same time period in the United States, and an estimated 16.8 million undiagnosed cases by mid-July 2020.

D614G Spike Variant Does Not Alter IgG, IgM, or IgA Spike Seroassay Performance (preprint)

Emergence of a new variant of spike protein (D614G) with increased infectivity and transmissibility has prompted many to analyze the potential role of this variant in the SARS-CoV-2 pandemic. When a new variant emerges, there is a concern regarding whether an individual exposed to one variant of a virus will have cross-reactive immune memory to the second variant. Accordingly, we analyzed the serologic reactivity of D614 (original) and G614 variant spike proteins. We found that antibodies from a high-incidence population in New York City reacted both toward the original D614 spike and the G614 spike variant. These data suggest that patients who have been exposed to either SARS-CoV-2 variant have humoral immunity that can respond against both variants. This is an important finding both for SARS-CoV-2 disease biology and for potential antibody-based therapeutics.

Serologic cross-reactivity of SARS-CoV-2 with endemic and seasonal Betacoronaviruses (preprint)

In order to properly understand the spread of SARS-CoV-2 infection and development of humoral immunity, researchers have evaluated the presence of serum antibodies of people worldwide experiencing the pandemic. These studies rely on the use of recombinant proteins from the viral genome in order to identify serum antibodies that recognize SARS-CoV-2 epitopes. Here, we discuss the cross-reactivity potential of SARS-CoV-2 antibodies with the full spike proteins of four other Betacoronaviruses that cause disease in humans, MERS-CoV, SARS-CoV, HCoV-OC43, and HCoV-HKU1. Using enzyme-linked immunosorbent assays (ELISAs), we detected the potential cross-reactivity of antibodies against SARS-CoV-2 towards the four other coronaviruses, with the strongest cross-recognition between SARS-CoV-2 and SARS /MERS-CoV antibodies, as expected based on sequence homology of their respective spike proteins. Further analysis of cross-reactivity could provide informative data that could lead to intelligently designed pan-coronavirus therapeutics or vaccines.

Standardization of enzyme-linked immunosorbent assays for serosurveys of the SARS-CoV-2 pandemic using clinical and at-home blood sampling (preprint)

The extent of SARS-CoV-2 infection throughout the United States population is currently unknown. High quality serology is a key tool to understanding the spread of infection, immunity against the virus, and correlates of protection. Limited validation and testing of serology assays used for serosurveys can lead to unreliable or misleading data, and clinical testing using such unvalidated assays can lead to medically costly diagnostic errors and improperly informed public health decisions. Estimating prevalence and clinical decision making is highly dependent on specificity. Here, we present an optimized ELISA-based serology protocol from antigen production to data analysis. This protocol defines thresholds for IgG and IgM for determination of seropositivity with estimated specificity well above 99%. Validation was performed using both traditionally collected serum and dried blood on mail-in blood sampling kits, using archival (pre-2019) negative controls and known PCR-diagnosed positive patient controls. Minimal cross-reactivity was observed for the spike proteins of MERS, SARS1, OC43 and HKU1 viruses and no cross reactivity was observed with anti-influenza A H1N1 HAI titer during validation. This strategy is highly specific and is designed to provide good estimates of seroprevalence of SARS-CoV-2 seropositivity in a population, providing specific and reliable data from serosurveys and clinical testing which can be used to better evaluate and understand SARS-CoV-2 immunity and correlates of protection.