Background

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the cause of coronavirus disease 2019 (COVID-19), is associated with respiratory-related disease and death. Assays to detect virus-specific antibodies are important to understand the prevalence of infection and the course of the immune response.

Methods

Quantitative measurements of plasma or serum antibodies to the nucleocapsid and spike proteins were analyzed using luciferase immunoprecipitation system assays in 100 cross-sectional or longitudinal samples from patients with SARS-CoV-2 infection. A subset of samples was tested both with and without heat inactivation.

Results

At >14 days after symptom onset, antibodies against SARS-CoV-2 nucleocapsid protein showed 100% sensitivity and 100% specificity, whereas antibodies to spike protein were detected with 91% sensitivity and 100% specificity. Neither antibody levels nor the rate of seropositivity were significantly reduced by heat inactivation of samples. Analysis of daily samples from 6 patients with COVID-19 showed anti-nucleocapsid and spike protein antibodies appearing between days 8 and 14 after initial symptoms. Immunocompromised patients generally had a delayed antibody response to SARS-CoV-2, compared with immunocompetent patients.

Conclusions

Antibody to the nucleocapsid protein of SARS-CoV-2 is more sensitive than spike protein antibody for detecting early infection. Analyzing heat-inactivated samples with a luciferase immunoprecipitation system assay is a safe and sensitive method for detecting SARS-CoV-2 antibodies.

Background:SARS-CoV-2, the cause of coronavirus disease 2019 (COVID-19), is associated with respiratory-related morbidity and mortality. Assays to detect virus-specific antibodies are important to understand the prevalence of infection and the course of the immune response. Methodology:Quantitative measurements of plasma or serum antibodies by luciferase immunoprecipitation assay systems (LIPS) to the nucleocapsid and spike proteins were analyzed in 100 cross-sectional or longitudinal samples from SARS-CoV-2-infected patients. A subset of samples was tested with and without heat inactivation. Results:Fifteen or more days after symptom onset, antibodies against SARS-CoV-2 nucleocapsid protein showed 100% sensitivity and 100% specificity, while antibodies to spike protein were detected with 91% sensitivity and 100% specificity. Neither antibody levels nor the rate of seropositivity were significantly reduced by heat inactivation of samples. Analysis of daily samples from six patients with COVID-19 showed anti-nucleocapsid and spike antibodies appearing between day 8 to day 14 after initial symptoms. Immunocompromised patients generally had a delayed antibody response to SARS-CoV-2 compared to immunocompetent patients. Conclusions:Antibody to the nucleocapsid protein of SARS-CoV-2 is more sensitive than spike protein antibody for detecting early infection. Analyzing heat-inactivated samples by LIPS is a safe and sensitive method for detecting SARS-CoV-2 antibodies.

Mechanistic models fit to streaming surveillance data are critical to understanding the transmission dynamics of an outbreak as it unfolds in real-time. However, transmission model parameter estimation can be imprecise, and sometimes even impossible, because surveillance data are noisy and not informative about all aspects of the mechanistic model. To partially overcome this obstacle, Bayesian models have been proposed to integrate multiple surveillance data streams. We devised a modeling framework for integrating SARS-CoV-2 diagnostics test and mortality time series data, as well as seroprevalence data from cross-sectional studies, and tested the importance of individual data streams for both inference and forecasting. Importantly, our model for incidence data accounts for changes in the total number of tests performed. We model the transmission rate, infection-to-fatality ratio, and a parameter controlling a functional relationship between the true case incidence and the fraction of positive tests as time-varying quantities and estimate changes of these parameters nonparametrically. We compare our base model against modified versions which do not use diagnostics test counts or seroprevalence data to demonstrate the utility of including these often unused data streams. We apply our Bayesian data integration method to COVID-19 surveillance data collected in Orange County, California between March 2020 and February 2021 and find that 32--72\% of the Orange County residents experienced SARS-CoV-2 infection by mid-January, 2021. Despite this high number of infections, our results suggest that the abrupt end of the winter surge in January 2021 was due to both behavioral changes and a high level of accumulated natural immunity.

Spleen tyrosine kinase (SYK) is a previously unidentified therapeutic target that inhibits neutrophil and macrophage activation in coronavirus disease 2019 (COVID-19). Fostamatinib, a SYK inhibitor, was studied in a phase 2 placebo-controlled randomized clinical trial and was associated with improvements in many secondary end points related to efficacy. Here, we used a multiomic approach to evaluate cellular and soluble immune mediator responses of patients enrolled in this trial. We demonstrated that SYK inhibition was associated with reduced neutrophil activation, increased circulation of mature neutrophils (CD10⁺CD33^-), and decreased circulation of low-density granulocytes and polymorphonuclear myeloid-derived suppressor cells (HLA-DR^-CD33⁺CD11b^-). SYK inhibition was also associated with normalization of transcriptional activity in circulating monocytes relative to healthy controls, an increase in frequency of circulating nonclassical and HLA-DR^hi classical monocyte populations, and restoration of interferon responses. Together, these data suggest that SYK inhibition may mitigate proinflammatory myeloid cellular and soluble mediator responses thought to contribute to immunopathogenesis of severe COVID-19.

Despite signs of infection-including taste loss, dry mouth and mucosal lesions such as ulcerations, enanthema and macules-the involvement of the oral cavity in coronavirus disease 2019 (COVID-19) is poorly understood. To address this, we generated and analyzed two single-cell RNA sequencing datasets of the human minor salivary glands and gingiva (9 samples, 13,824 cells), identifying 50 cell clusters. Using integrated cell normalization and annotation, we classified 34 unique cell subpopulations between glands and gingiva. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viral entry factors such as ACE2 and TMPRSS members were broadly enriched in epithelial cells of the glands and oral mucosae. Using orthogonal RNA and protein expression assessments, we confirmed SARS-CoV-2 infection in the glands and mucosae. Saliva from SARS-CoV-2-infected individuals harbored epithelial cells exhibiting ACE2 and TMPRSS expression and sustained SARS-CoV-2 infection. Acellular and cellular salivary fractions from asymptomatic individuals were found to transmit SARS-CoV-2 ex vivo. Matched nasopharyngeal and saliva samples displayed distinct viral shedding dynamics, and salivary viral burden correlated with COVID-19 symptoms, including taste loss. Upon recovery, this asymptomatic cohort exhibited sustained salivary IgG antibodies against SARS-CoV-2. Collectively, these data show that the oral cavity is an important site for SARS-CoV-2 infection and implicate saliva as a potential route of SARS-CoV-2 transmission.