Severe Acute Respiratory Syndrome Coronavirus 2 Infection History and Antibody Response to 3 Coronavirus Disease 2019 Messenger RNA Vaccine Doses.

BACKGROUND: Data on antibody kinetics are limited among individuals previously infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). From a cohort of healthcare personnel and other frontline workers in 6 US states, we assessed antibody waning after messenger RNA (mRNA) dose 2 and response to dose 3 according to SARS-CoV-2 infection history. METHODS: Participants submitted sera every 3 months, after SARS-CoV-2 infection, and after each mRNA vaccine dose. Sera were tested for antibodies and reported as area under the serial dilution curve (AUC). Changes in AUC values over time were compared using a linear mixed model. RESULTS: Analysis included 388 participants who received dose 3 by November 2021. There were 3 comparison groups: vaccine only with no known prior SARS-CoV-2 infection (n = 224); infection prior to dose 1 (n = 123); and infection after dose 2 and before dose 3 (n = 41). The interval from dose 2 and dose 3 was approximately 8 months. After dose 3, antibody levels rose 2.5-fold (95% confidence interval [CI] = 2.2-3.0) in group 2 and 2.9-fold (95% CI = 2.6-3.3) in group 1. Those infected within 90 days before dose 3 (and median 233 days [interquartile range, 213-246] after dose 2) did not increase significantly after dose 3. CONCLUSIONS: A third dose of mRNA vaccine typically elicited a robust humoral immune response among those with primary vaccination regardless of SARS-CoV-2 infection >3 months prior to boosting. Those with infection <3 months prior to boosting did not have a significant increase in antibody concentrations in response to a booster.

Competent immune responses to SARS-CoV-2 variants in older adults following two doses of mRNA vaccination.

Aging is associated with a reduced magnitude of primary immune responses to vaccination. mRNA-based SARS-CoV-2 vaccines have shown efficacy in older adults but virus variant escape is still unclear. Here we analyze humoral and cellular immunity against an early-pandemic viral isolate and compare that to the P.1 (Gamma) and B.1.617.2 (Delta) variants in two cohorts (<50 and >55 age) of mRNA vaccine recipients. We further measure neutralizing antibody titers for B.1.617.1 (Kappa) and B.1.595, with the latter SARS-CoV-2 isolate bearing the spike mutation E484Q. Robust humoral immunity is measured following second vaccination, and older vaccinees manifest cellular immunity comparable to the adult group against early-pandemic SARS-CoV-2 and more recent variants. More specifically, the older cohort has lower neutralizing capacity at 7-14 days following the second dose but equilibrates with the younger cohort after 2-3 months. While long-term vaccination responses remain to be determined, our results implicate vaccine-induced protection in older adults against SARS-CoV-2 variants and inform thinking about boost vaccination.

Cutting Edge: T Cell Responses to B.1.1.529 (Omicron) SARS-CoV-2 Variant Induced by COVID-19 Infection and/or mRNA Vaccination Are Largely Preserved.

Several studies have demonstrated that the SARS-CoV-2 variant-of-concern B.1.1.529 (Omicron) exhibits a high degree of escape from Ab neutralization. Therefore, it is critical to determine how well the second line of adaptive immunity, T cell memory, performs against Omicron. To this purpose, we analyzed a human cohort (n = 327 subjects) of two- or three-dose mRNA vaccine recipients and COVID-19 postinfection subjects. We report that T cell responses against Omicron were largely preserved. IFN-γ-producing T cell responses remained equivalent to the response against the ancestral strain (WA1/2020), with some (∼20%) loss in IL-2 single or IL-2+IFN-γ+ polyfunctional responses. Three-dose vaccinated participants had similar responses to Omicron relative to post-COVID-19 participants and exhibited responses significantly higher than those receiving two mRNA vaccine doses. These results provide further evidence that a three-dose vaccine regimen benefits the induction of optimal functional T cell immune memory.

Sensitive, smartphone-based SARS-CoV-2 detection from clinical saline gargle samples.

Saliva specimens have drawn interest for diagnosing respiratory viral infections due to their ease of collection and decreased risk to healthcare providers. However, rapid and sensitive immunoassays have not yet been satisfactorily demonstrated for such specimens due to their viscosity and low viral loads. Using paper microfluidic chips and a smartphone-based fluorescence microscope, we developed a highly sensitive, low-cost immunofluorescence particulometric SARS-CoV-2 assay from clinical saline gargle samples. We demonstrated the limit of detection of 10 ag/µL. With easy-to-collect saline gargle samples, our clinical sensitivity, specificity, and accuracy were 100%, 86%, and 93%, respectively, for n = 27 human subjects with n = 13 RT-qPCR positives.

Quantitative restoration of immune defense in old animals determined by naive antigen-specific CD8 T-cell numbers.

Older humans and animals often exhibit reduced immune responses to infection and vaccination, and this often directly correlates to the numbers and frequency of naive T (Tn) cells. We found such a correlation between reduced numbers of blood CD8+ Tn cells and severe clinical outcomes of West Nile virus (WNV) in both humans naturally exposed to, and mice experimentally infected with, WNV. To examine possible causality, we sought to increase the number of CD8 Tn cells by treating C57BL/6 mice with IL-7 complexes (IL-7C, anti-IL-7 mAb bound to IL-7), shown previously to efficiently increase peripheral T-cell numbers by homeostatic proliferation. T cells underwent robust expansion following IL-7C administration to old mice increasing the number of total T cells (>fourfold) and NS4b:H-2Db -restricted antigen-specific CD8 T cells (twofold). This improved the numbers of NS4b-specific CD8 T cells detected at the peak of the response against WNV, but not survival of WNV challenge. IL-7C-treated old animals also showed no improvement in WNV-specific effector immunity (neutralizing antibody and in vivo T-cell cytotoxicity). To test quantitative limits to which CD8 Tn cell restoration could improve protective immunity, we transferred graded doses of Ag-specific precursors into old mice and showed that injection of 5400 (but not of 1800 or 600) adult naive WNV-specific CD8 T cells significantly increased survival after WNV. These results set quantitative limits to the level of Tn reconstitution necessary to improve immune defense in older organisms and are discussed in light of targets of immune reconstitution.

Smartphone-based sensitive detection of SARS-CoV-2 from saline gargle samples via flow profile analysis on a paper microfluidic chip.

Respiratory viruses, especially coronaviruses, have resulted in worldwide pandemics in the past couple of decades. Saliva-based paper microfluidic assays represent an opportunity for noninvasive and rapid screening, yet both the sample matrix and test method come with unique challenges. In this work, we demonstrated the rapid and sensitive detection of SARS-CoV-2 from saliva samples, which could be simpler and more comfortable for patients than existing methods. Furthermore, we systematically investigated the components of saliva samples that affected assay performance. Using only a smartphone, an antibody-conjugated particle suspension, and a paper microfluidic chip, we made the assay user-friendly with minimal processing. Unlike the previously established flow rate assays that depended solely on the flow rate or distance, this unique assay analyzes the flow profile to determine infection status. Particle-target immunoagglutination changed the surface tension and subsequently the capillary flow velocity profile. A smartphone camera automatically measured the flow profile using a Python script, which was not affected by ambient light variations. The limit of detection (LOD) was 1 fg/µL SARS-CoV-2 from 1% saliva samples and 10 fg/µL from simulated saline gargle samples (15% saliva and 0.9% saline). This method was highly specific as demonstrated using influenza A/H1N1. The sample-to-answer assay time was <15 min, including <1-min capillary flow time. The overall accuracy was 89% with relatively clean clinical saline gargle samples. Despite some limitations with turbid clinical samples, this method presents a potential solution for rapid mass testing techniques during any infectious disease outbreak as soon as the antibodies become available.

Direct capture and smartphone quantification of airborne SARS-CoV-2 on a paper microfluidic chip.

SARS, a new type of respiratory disease caused by SARS-CoV, was identified in 2003 with significant levels of morbidity and mortality. The recent pandemic of COVID-19, caused by SARS-CoV-2, has generated even greater extents of morbidity and mortality across the entire world. Both SARS-CoV and SARS-CoV-2 spreads through the air in the form of droplets and potentially smaller droplets (aerosols) via exhaling, coughing, and sneezing. Direct detection from such airborne droplets would be ideal for protecting general public from potential exposure before they infect individuals. However, the number of viruses in such droplets and aerosols is too low to be detected directly. A separate air sampler and enough collection time (several hours) are necessary to capture a sufficient number of viruses. In this work, we have demonstrated the direct capture of the airborne droplets on the paper microfluidic chip without the need for any other equipment. 10% human saliva samples were spiked with the known concentration of SARS-CoV-2 and sprayed to generate liquid droplets and aerosols into the air. Antibody-conjugated submicron particle suspension is then added to the paper channel, and a smartphone-based fluorescence microscope isolated and counted the immunoagglutinated particles on the paper chip. The total capture-to-assay time was <30 min, compared to several hours with the other methods. In this manner, SARS-CoV-2 could be detected directly from the air in a handheld and low-cost manner, contributing to slowing the spread of SARS-CoV-2. We can presumably adapt this technology to a wide range of other respiratory viruses.

Immune responses to two and three doses of the BNT162b2 mRNA vaccine in adults with solid tumors.

Vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have shown high efficacy, but immunocompromised participants were excluded from controlled clinical trials. In this study, we compared immune responses to the BNT162b2 mRNA Coronavirus Disease 2019 vaccine in patients with solid tumors (n = 53) who were on active cytotoxic anti-cancer therapy to a control cohort of participants without cancer (n = 50). Neutralizing antibodies were detected in 67% of patients with cancer after the first immunization, followed by a threefold increase in median titers after the second dose. Similar patterns were observed for spike protein-specific serum antibodies and T cells, but the magnitude of each of these responses was diminished relative to the control cohort. In most patients with cancer, we detected spike receptor-binding domain and other S1-specific memory B cell subsets as potential predictors of anamnestic responses to additional immunizations. We therefore initiated a phase 1 trial for 20 cancer cohort participants of a third vaccine dose of BNT162b2 ( NCT04936997 ); primary outcomes were immune responses, with a secondary outcome of safety. At 1 week after a third immunization, 16 participants demonstrated a median threefold increase in neutralizing antibody responses, but no improvement was observed in T cell responses. Adverse events were mild. These results suggest that a third dose of BNT162b2 is safe, improves humoral immunity against SARS-CoV-2 and could be immunologically beneficial for patients with cancer on active chemotherapy.

Antihypertensive drug treatment and susceptibility to SARS-CoV-2 infection in human PSC-derived cardiomyocytes and primary endothelial cells.

The pathogenicity of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has been attributed to its ability to enter through the membrane-bound angiotensin-converting enzyme 2 (ACE2) receptor. Therefore, it has been heavily speculated that angiotensin-converting enzyme inhibitor (ACEI) or angiotensin receptor blocker (ARB) therapy may modulate SARS-CoV-2 infection. In this study, exposure of human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) and human endothelial cells (hECs) to SARS-CoV-2 identified significant differences in protein coding genes involved in immunity, viral response, and cardiomyocyte/endothelial structure. Specifically, transcriptome changes were identified in the tumor necrosis factor (TNF), interferon α/ß, and mitogen-activated protein kinase (MAPK) (hPSC-CMs) as well as nuclear factor kappa-B (NF-κB) (hECs) signaling pathways. However, pre-treatment of hPSC-CMs or hECs with two widely prescribed antihypertensive medications, losartan and lisinopril, did not affect the susceptibility of either cell type to SARS-CoV-2 infection. These findings demonstrate the toxic effects of SARS-CoV-2 in hPSC-CMs/hECs and, taken together with newly emerging multicenter trials, suggest that antihypertensive drug treatment alone does not alter SARS-CoV-2 infection.

Immune response to COVID-19 in older adults.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is the third highly pathogenic coronavirus to emerge in the human population in last two decades. SARS-CoV-2 spread from Wuhan, China, across the globe, causing an unprecedented public healthcare crisis. The virus showed remarkable age dependent pathology, with symptoms resembling common cold in most adults and children while causing more severe respiratory distress and significant mortality in older and frail humans. Even before the SARS-CoV-2 outbreak infectious diseases represented one of the major causes of death of older adults. Loss of immune function and reduced protection from infectious agents with age - immunosenescence - is a result of complex mechanisms affecting production and maintenance of immune cells as well as the initiation, maintenance and termination of properly directed immune responses. Here we briefly discuss the current knowledge on how this process affects age-dependent outcomes of SARS-CoV-2 infection.

Orthogonal SARS-CoV-2 Serological Assays Enable Surveillance of Low-Prevalence Communities and Reveal Durable Humoral Immunity.

We conducted a serological study to define correlates of immunity against SARS-CoV-2. Compared to those with mild coronavirus disease 2019 (COVID-19) cases, individuals with severe disease exhibited elevated virus-neutralizing titers and antibodies against the nucleocapsid (N) and the receptor binding domain (RBD) of the spike protein. Age and sex played lesser roles. All cases, including asymptomatic individuals, seroconverted by 2 weeks after PCR confirmation. Spike RBD and S2 and neutralizing antibodies remained detectable through 5-7 months after onset, whereas α-N titers diminished. Testing 5,882 members of the local community revealed only 1 sample with seroreactivity to both RBD and S2 that lacked neutralizing antibodies. This fidelity could not be achieved with either RBD or S2 alone. Thus, inclusion of multiple independent assays improved the accuracy of antibody tests in low-seroprevalence communities and revealed differences in antibody kinetics depending on the antigen. We conclude that neutralizing antibodies are stably produced for at least 5-7 months after SARS-CoV-2 infection.