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.
CD4 T follicular helper (T
) cells are important for the generation of durable and specific humoral protection against viral infections. The degree to which SARS-CoV-2 infection generates T
cells and ...stimulates the germinal center (GC) response is an important question as we investigate vaccine induced immunity against COVID-19. Here, we report that SARS-CoV-2 infection in rhesus macaques, either infused with convalescent plasma, normal plasma, or receiving no infusion, resulted in transient accumulation of pro-inflammatory monocytes and proliferating T
cells with a T
1 profile in peripheral blood. CD4 helper cell responses skewed predominantly toward a T
1 response in blood, lung, and lymph nodes. SARS-CoV-2 Infection induced GC T
cells specific for the SARS-CoV-2 spike and nucleocapsid proteins, and a corresponding early appearance of antiviral serum IgG antibodies. Collectively, the data show induction of GC responses in a rhesus model of mild COVID-19.
IMPORTANCE: People who have been infected with or vaccinated against SARS-CoV-2 have reduced risk of subsequent infection, but the proportion of people in the US with SARS-CoV-2 antibodies from ...infection or vaccination is uncertain. OBJECTIVE: To estimate trends in SARS-CoV-2 seroprevalence related to infection and vaccination in the US population. DESIGN, SETTING, AND PARTICIPANTS: In a repeated cross-sectional study conducted each month during July 2020 through May 2021, 17 blood collection organizations with blood donations from all 50 US states; Washington, DC; and Puerto Rico were organized into 66 study-specific regions, representing a catchment of 74% of the US population. For each study region, specimens from a median of approximately 2000 blood donors were selected and tested each month; a total of 1 594 363 specimens were initially selected and tested. The final date of blood donation collection was May 31, 2021. EXPOSURE: Calendar time. MAIN OUTCOMES AND MEASURES: Proportion of persons with detectable SARS-CoV-2 spike and nucleocapsid antibodies. Seroprevalence was weighted for demographic differences between the blood donor sample and general population. Infection-induced seroprevalence was defined as the prevalence of the population with both spike and nucleocapsid antibodies. Combined infection- and vaccination-induced seroprevalence was defined as the prevalence of the population with spike antibodies. The seroprevalence estimates were compared with cumulative COVID-19 case report incidence rates. RESULTS: Among 1 443 519 specimens included, 733 052 (50.8%) were from women, 174 842 (12.1%) were from persons aged 16 to 29 years, 292 258 (20.2%) were from persons aged 65 years and older, 36 654 (2.5%) were from non-Hispanic Black persons, and 88 773 (6.1%) were from Hispanic persons. The overall infection-induced SARS-CoV-2 seroprevalence estimate increased from 3.5% (95% CI, 3.2%-3.8%) in July 2020 to 20.2% (95% CI, 19.9%-20.6%) in May 2021; the combined infection- and vaccination-induced seroprevalence estimate in May 2021 was 83.3% (95% CI, 82.9%-83.7%). By May 2021, 2.1 SARS-CoV-2 infections (95% CI, 2.0-2.1) per reported COVID-19 case were estimated to have occurred. CONCLUSIONS AND RELEVANCE: Based on a sample of blood donations in the US from July 2020 through May 2021, vaccine- and infection-induced SARS-CoV-2 seroprevalence increased over time and varied by age, race and ethnicity, and geographic region. Despite weighting to adjust for demographic differences, these findings from a national sample of blood donors may not be representative of the entire US population.
Red blood cell storage in the blood bank promotes the progressive accumulation of metabolic alterations that may ultimately impact the erythrocyte capacity to cope with oxidant stressors. However, ...the metabolic underpinnings of the capacity of RBCs to resist oxidant stress and the potential impact of donor biology on this phenotype are not known. Within the framework of the REDS-III RBC-Omics study, RBCs from 8,502 healthy blood donors were stored for 42 days and tested for their propensity to hemolyze following oxidant stress. A subset of extreme hemolyzers donated a second unit of blood, which was stored for 10, 23, and 42 days and profiled again for oxidative hemolysis and metabolomics (599 samples). Alterations of RBC energy and redox homeostasis were noted in donors with high oxidative hemolysis. RBCs from females, donors over 60 years old, donors of Asian/South Asian race-ethnicity, and RBCs stored in additive solution-3 were each independently characterized by improved antioxidant metabolism compared to, respectively, males, donors under 30 years old, Hispanic and African American race ethnicity donors, and RBCs stored in additive solution-1. Merging metabolomics data with results from an independent GWAS study on the same cohort, we identified metabolic markers of hemolysis and G6PD-deficiency, which were associated with extremes in oxidative hemolysis and dysregulation in NADPH and glutathione-dependent detoxification pathways of oxidized lipids. Donor sex, age, ethnicity, additive solution and G6PD status impact the metabolism of the stored erythrocyte and its susceptibility to hemolysis following oxidative insults.
Given the limited availability of serological testing to date, the seroprevalence of SARS-CoV-2-specific antibodies in different populations has remained unclear. Here, we report very low SARS-CoV-2 ...seroprevalence in two San Francisco Bay Area populations. Seroreactivity was 0.26% in 387 hospitalized patients admitted for non-respiratory indications and 0.1% in 1,000 blood donors in early April 2020. We additionally describe the longitudinal dynamics of immunoglobulin-G (IgG), immunoglobulin-M (IgM), and in vitro neutralizing antibody titers in COVID-19 patients. The median time to seroconversion ranged from 10.3-11.0 days for these 3 assays. Neutralizing antibodies rose in tandem with immunoglobulin titers following symptom onset, and positive percent agreement between detection of IgG and neutralizing titers was >93%. These findings emphasize the importance of using highly accurate tests for surveillance studies in low-prevalence populations, and provide evidence that seroreactivity using SARS-CoV-2 anti-nucleocapsid protein IgG and anti-spike IgM assays are generally predictive of in vitro neutralizing capacity.
The pool of beta cell-specific CD8
T cells in type 1 diabetes (T1D) sustains an autoreactive potential despite having access to a constant source of antigen. To investigate the long-lived nature of ...these cells, we established a DNA methylation-based T cell 'multipotency index' and found that beta cell-specific CD8
T cells retained a stem-like epigenetic multipotency score. Single-cell assay for transposase-accessible chromatin using sequencing confirmed the coexistence of naive and effector-associated epigenetic programs in individual beta cell-specific CD8
T cells. Assessment of beta cell-specific CD8
T cell anatomical distribution and the establishment of stem-associated epigenetic programs revealed that self-reactive CD8
T cells isolated from murine lymphoid tissue retained developmentally plastic phenotypic and epigenetic profiles relative to the same cells isolated from the pancreas. Collectively, these data provide new insight into the longevity of beta cell-specific CD8
T cell responses and document the use of this methylation-based multipotency index for investigating human and mouse CD8
T cell differentiation.
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 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 three 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 91%. 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 three main clusters with distinct reactivity intensities and patterns. These patterns were not recapitulated by adjusting the VITROS CoV2T or RVPNT assay thresholds. Plasma classified by COVAM reactivity patterns offers potential to improve CCP therapeutic efficacy CoV2T alone. The use of a SARS-CoV-2 antigen array can qualify CCP for administration as a treatment for acute COVID-19, and interrogate vaccine immunogenicity and performance in preclinical, clinical studies, and routine vaccination to identify antibody responses predictive of protection from infection and disease.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) serosurveys can estimate cumulative incidence for monitoring epidemics, requiring assessment of serologic assays to inform testing ...algorithm development and interpretation of results. We conducted a multilaboratory evaluation of 21 commercial high-throughput SARS-CoV-2 serologic assays using blinded panels of 1,000 highly characterized specimens. Assays demonstrated a range of sensitivities (96%-63%), specificities (99%-96%), and precision (intraclass correlation coefficient 0.55-0.99). Durability of antibody detection was dependent on antigen and immunoglobulin targets; antispike and total Ig assays demonstrated more stable longitudinal reactivity than antinucleocapsid and IgG assays. Assays with high sensitivity, specificity, and durable antibody detection are ideal for serosurveillance, but assays demonstrating waning reactivity are appropriate for other applications, including correlation with neutralizing activity and detection of anamnestic boosting by reinfections. Assay performance must be evaluated in context of intended use, particularly in the context of widespread vaccination and circulation of SARS-CoV-2 variants.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, ODKLJ, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Studies have demonstrated that intensive ART alone is not capable of eradicating HIV-1, as the virus rebounds within a few weeks upon treatment interruption. Viral rebound may be induced from several ...cellular subsets; however, the majority of proviral DNA has been found in antigen experienced resting CD4+ T cells. To achieve a cure for HIV-1, eradication strategies depend upon both understanding mechanisms that drive HIV-1 persistence as well as sensitive assays to measure the frequency of infected cells after therapeutic interventions. Assays such as the quantitative viral outgrowth assay (QVOA) measure HIV-1 persistence during ART by ex vivo activation of resting CD4+ T cells to induce latency reversal; however, recent studies have shown that only a fraction of replication-competent viruses are inducible by primary mitogen stimulation. Previous studies have shown a correlation between the acquisition of effector memory phenotype and HIV-1 latency reversal in quiescent CD4+ T cell subsets that harbor the reservoir. Here, we apply our mechanistic understanding that differentiation into effector memory CD4+ T cells more effectively promotes HIV-1 latency reversal to significantly improve proviral measurements in the QVOA, termed differentiation QVOA (dQVOA), which reveals a significantly higher frequency of the inducible HIV-1 replication-competent reservoir in resting CD4+ T cells.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Persistence of HIV in people living with HIV (PWH) on suppressive antiretroviral therapy (ART) has been linked to physiological mechanisms of CD4+ T cells. Here, in the same 37 male PWH on ART we ...measure longitudinal kinetics of HIV DNA and cell turnover rates in five CD4 cell subsets: naïve (T
), stem-cell- (T
), central- (T
), transitional- (T
), and effector-memory (T
). HIV decreases in T
and T
but not in less-differentiated subsets. Cell turnover is ~10 times faster than HIV clearance in memory subsets, implying that cellular proliferation consistently creates HIV DNA. The optimal mathematical model for these integrated data sets posits HIV DNA also passages between CD4 cell subsets via cellular differentiation. Estimates are heterogeneous, but in an average participant's year ~10 (in T
and T
) and ~10
(in T
, T
, T
) proviruses are generated by proliferation while ~10
proviruses passage via cell differentiation (per million CD4). In simulations, therapies blocking proliferation and/or enhancing differentiation could reduce HIV DNA by 1-2 logs over 3 years. In summary, HIV exploits cellular proliferation and differentiation to persist during ART but clears faster in more proliferative/differentiated CD4 cell subsets and the same physiological mechanisms sustaining HIV might be temporarily modified to reduce it.
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