Employment of a high-throughput functional assay to define the critical factors that influence vaccine-induced, cross-variant neutralizing antibodies for SARS-CoV-2. (preprint)

The scale and duration of neutralizing antibody responses targeting SARS-CoV-2 viral variants represents a critically important serological parameter that predicts protective immunity for COVID-19. In this study, we describe the development and employment of a new functional assay that measures neutralizing antibodies for SARS-CoV-2 and present longitudinal data illustrating the impact of age, sex and comorbidities on the kinetics and strength of vaccine-induced antibody responses for key variants in an Asian volunteer cohort. We also present an accurate quantitation of serological responses for SARS-CoV-2 that exploits a unique set of in-house, recombinant human monoclonal antibodies targeting the viral Spike and nucleocapsid proteins and demonstrate a reduction in neutralizing antibody titres across all groups six months post-vaccination. We also observe a marked reduction in the serological binding activity and neutralizing responses targeting recently newly emerged Omicron variants including XBB 1.5 and highlight a significant increase in cross-protective neutralizing antibody responses following a third dose (boost) of vaccine. These data illustrate how key virological factors such as immune escape mutations combined with host demographic factors such as age and sex of the vaccinated individual influence the strength and duration of cross-protective serological immunity for COVID-19.

Defining factors that influence vaccine-induced, cross-variant neutralizing antibodies for SARS-CoV-2 in Asians (preprint)

The scale and duration of neutralizing antibody responses targeting SARS-CoV-2 viral variants represents a critically important serological parameter that predicts protective immunity for COVID-19. In this study, we present longitudinal data illustrating the impact of age, sex and comorbidities on the kinetics and strength of vaccine-induced neutralizing antibody responses for key variants in an Asian volunteer cohort. We demonstrate a reduction in neutralizing antibody titres across all groups six months post-vaccination and show a marked reduction in the serological binding and neutralizing response targeting Omicron compared to other viral variants. We also highlight the increase in cross-protective neutralizing antibody responses against Omicron induced by a third dose (booster) of vaccine. These data illustrate how key virological factors such as immune escape mutation combined with host factors such as age and sex of the vaccinated individuals influence the strength and duration of cross-protective serological immunity for COVID-19.

Data-driven analysis of COVID-19 reveals specific severity patterns distinct from the temporal immune response (preprint)

Key immune signatures of SARS-CoV-2 infection may associate with either adverse immune reactions (severity) or simply an ongoing anti-viral response (temporality); how immune signatures contribute to severe manifestations and/or temporal progression of disease and whether longer disease duration correlates with severity remain unknown. Patient blood was comprehensively immunophenotyped via mass cytometry and multiplex cytokine arrays, leading to the identification of 327 basic subsets that were further stratified into more than 5000 immunotypes and correlated with 28 plasma cytokines. Low-density neutrophil abundance was closely correlated with hepatocyte growth factor levels, which in turn correlated with disease severity. Deep analysis also revealed additional players, namely conventional type 2 dendritic cells, natural killer T cells, plasmablasts and CD16+ monocytes, that can influence COVID-19 severity independent of temporal progression. Herein, we provide interactive network analysis and data visualization tools to facilitate data mining and hypothesis generation for elucidating COVID-19 pathogenesis.

Age and chest radiography as possible parameters for rapid triage in COVID-19 outbreak surge (preprint)

BackgroundTo evaluate the utility of age and chest radiography(CXR) in triaging COVID-19 patients for hospitalization versus isolation in non-hospital facilities, we examined how age and CXR at diagnosis were associated with clinical needs from late-January to early-April. MethodsClinical status of all COVID-19 cases was monitored for national disease surveillance. Cases were isolated in hospitals until SARS-CoV-2 RNA was undetectable on PCR. Age and CXR results on admission were analysed for association with oxygen supplementation and mechanical ventilation, the outcomes of interest.ResultsTill 4 April 2020, there were 1,481 COVID-19 cases in Singapore. Overall, 11.4% required supplemental oxygen while 4.8% required mechanical ventilation and intensive care. The respective proportions increased to 40.9% and 16.5% for cases aged ≥70 years. As a predictor of subsequent mechanical ventilation, age had an area under the receiver operator characteristic curve(AUROC) of 0.772 (95%CI:0.699-0.845). A combined criterion of either an abnormal CXR or age≥55 years had a sensitivity of 86.7% and specificity of 58.0% for the same outcome. A similar performance was observed for predicting oxygen supplementation needs.ConclusionsAge and CXR at diagnosis may be valuable in excluding severe disease, allowing safe triage for isolation in non-hospital facilities. 

Whole blood immunophenotyping uncovers immature neutrophil-to-VD2 T-cell ratio as an early prognostic marker for severe COVID-19 (preprint)

SARS-CoV-2 is the novel coronavirus responsible for the current COVID-19 pandemic. Severe complications are observed only in a small proportion of infected patients but the cellular mechanisms underlying this progression are still unknown. Comprehensive flow cytometry of whole blood samples from 54 COVID-19 patients revealed a dramatic increase in the number of immature neutrophils. This increase strongly correlated with disease severity and was associated with elevated IL-6 and IP-10 levels, two key players in the cytokine storm. The most pronounced decrease in cell counts was observed for CD8 T-cells and VD2 {gamma}{delta} T-cells, which both exhibited increased differentiation and activation. ROC analysis revealed that the count ratio of immature neutrophils to CD8 or VD2 T-cells predicts pneumonia onset (0.9071) as well as hypoxia onset (0.8908) with high sensitivity and specificity. It would thus be a useful prognostic marker for preventive patient management and improved healthcare resource management.

A SARS-CoV-2 surrogate virus neutralization test (sVNT) based on antibody-mediated blockage of ACE2-spike (RBD) protein-protein interaction (preprint)

At this critical moment of the international response to the COVID-19 outbreak, there is an urgent need for a robust serological test to detect neutralizing antibodies to SARS-CoV-2. Such a test is not only important for contact tracing, but for determining infection rate, herd immunity and predicted humoral protection. The current gold standard is a virus neuralization test (VNT) requiring live virus and a biosafety level 3 (BSL3) laboratory. On the other hand, the ELISA- or lateral flow-based assays are for the detection of binding antibodies, which does not directly correlate with their neutralizing ability. Here we report a SARS-CoV-2 surrogate virus neutralization test (sVNT) that is designed to detect total neutralizing antibodies in an isotype- and species-independent manner. Our simple and rapid test is based on antibody-mediated blockage of virus-host interaction between the ACE2 receptor protein and the receptor binding domain (RBD) of the viral spike protein. The test has been validated with two COVID-19 patient cohorts in two different countries, achieving 100% specificity and 95-100% sensitivity and is capable of differentiating antibody responses from other known human coronaviruses. Importantly, the sVNT does not require BSL3 containment, thereby making the test immediately accessible to the global community.

Potent neutralizing antibodies in the sera of convalescent COVID-19 patients are directed against conserved linear epitopes on the SARS-CoV-2 spike protein (preprint)

The ongoing SARS-CoV-2 pandemic demands rapid identification of immunogenic targets for the design of efficient vaccines and serological detection tools. In this report, using pools of overlapping linear peptides and functional assays, we present two immunodominant regions on the spike glycoprotein that were highly recognized by neutralizing antibodies in the sera of COVID-19 convalescent patients. One is highly specific to SARS-CoV-2, and the other is a potential pan-coronavirus target.