Differential chromatin accessibility in peripheral blood mononuclear cells underlies COVID-19 disease severity prior to seroconversion (preprint)

SARS-CoV-2 infection triggers profound and variable immune responses in human hosts. Chromatin remodeling has been observed in individuals severely ill or convalescing with COVID-19, but chromatin remodeling early in disease prior to anti-spike protein IgG seroconversion has not been defined. We performed the Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) and RNA-seq on peripheral blood mononuclear cells (PBMCs) from outpatients with mild or moderate symptom severity at different stages of clinical illness. Early in the disease course prior to IgG seroconversion, modifications in chromatin accessibility associate with mild or moderate symptoms are already robust and include severity-associated changes in accessibility of genes in interleukin signaling, regulation of cell differentiation and cell morphology. Furthermore, single-cell analyses revealed evolution of the chromatin accessibility landscape and transcription factor motif accessibility for individual PBMC cell types over time. The most extensive remodeling occurred in CD14+ monocytes, where sub-populations with distinct chromatin accessibility profiles were observed prior to seroconversion. Mild symptom severity is marked by upregulation classical antiviral pathways including those regulating IRF1 and IRF7, whereas in moderate disease these classical antiviral signals diminish suggesting dysregulated and less effective responses. Together, these observations offer novel insight into the epigenome of early mild SARS-CoV-2 infection and suggest that detection of chromatin remodeling in early disease may offer promise for a new class of diagnostic tools for COVID-19. 

An atlas connecting shared genetic architecture of human diseases and molecular phenotypes provides insight into COVID-19 susceptibility (preprint)

While genome-wide associations studies (GWAS) have successfully elucidated the genetic architecture of complex human traits and diseases, understanding mechanisms that lead from genetic variation to pathophysiology remains an important challenge. Methods are needed to systematically bridge this crucial gap to facilitate experimental testing of hypotheses and translation to clinical utility. Here, we leveraged cross-phenotype associations to identify traits with shared genetic architecture, using linkage disequilibrium (LD) information to accurately capture shared SNPs by proxy, and calculate significance of enrichment. This shared genetic architecture was examined across differing biological scales through incorporating data from catalogs of clinical, cellular, and molecular GWAS. We have created an interactive web database (interactive Cross-Phenotype Analysis of GWAS database (iCPAGdb); http://cpag.oit.duke.edu) to facilitate exploration and allow rapid analysis of user-uploaded GWAS summary statistics. This database revealed well-known relationships among phenotypes, as well as the generation of novel hypotheses to explain the pathophysiology of common diseases. Application of iCPAGdb to a recent GWAS of severe COVID-19 demonstrated unexpected overlap of GWAS signals between COVID-19 and human diseases, including with idiopathic pulmonary fibrosis driven by the DPP9 locus. Transcriptomics from peripheral blood of COVID-19 patients demonstrated that DPP9 was induced in SARS-CoV-2 compared to healthy controls or those with bacterial infection. Further investigation of cross-phenotype SNPs with severe COVID-19 demonstrated colocalization of the GWAS signal of the ABO locus with plasma protein levels of a reported receptor of SARS-CoV-2, CD209 (DC-SIGN), pointing to a possible mechanism whereby glycosylation of CD209 by ABO may regulate COVID-19 disease severity. Thus, connecting genetically related traits across phenotypic scales links human diseases to molecular and cellular measurements that can reveal mechanisms and lead to novel biomarkers and therapeutic approaches.

Chromatin remodeling in peripheral blood cells reflects COVID-19 symptom severity (preprint)

SARS-CoV-2 infection triggers highly variable host responses and causes varying degrees of illness in humans. We sought to harness the peripheral blood mononuclear cell (PBMC) response over the course of illness to provide insight into COVID-19 physiology. We analyzed PBMCs from subjects with variable symptom severity at different stages of clinical illness before and after IgG seroconversion to SARS-CoV-2. Prior to seroconversion, PBMC transcriptomes did not distinguish symptom severity. In contrast, changes in chromatin accessibility were associated with symptom severity. Furthermore, single-cell analyses revealed evolution of the chromatin accessibility landscape and transcription factor motif occupancy for individual PBMC cell types. The most extensive remodeling occurred in CD14+ monocytes where sub-populations with distinct chromatin accessibility profiles were associated with disease severity. Our findings indicate that pre-seroconversion chromatin remodeling in certain innate immune populations is associated with divergence in symptom severity, and the identified transcription factors, regulatory elements, and downstream pathways provide potential prognostic markers for COVID-19 subjects. One sentence summaryChromatin accessibility in immune cells from COVID-19 subjects is remodeled prior to seroconversion to reflect disease severity.

Mucosal Associated Invariant T (MAIT) Cell Responses Differ by Sex in COVID-19 (preprint)

Sexual dimorphisms in immune responses contribute to coronavirus disease 2019 (COVID-19) outcomes, yet the mechanisms governing this disparity remain incompletely understood. We carried out sex-balanced sampling of peripheral blood mononuclear cells from confirmed COVID-19 inpatients and outpatients, uninfected close contacts, and healthy controls for 36-color flow cytometry and single cell RNA-sequencing. Our results revealed a pronounced reduction of circulating mucosal associated invariant T (MAIT) cells in infected females. Integration of published COVID-19 airway tissue datasets implicate that this reduction represented a major wave of MAIT cell extravasation during early infection in females. Moreover, female MAIT cells possessed an immunologically active gene signature, whereas male counterparts were pro-apoptotic. Collectively, our findings uncover a female-specific protective MAIT profile, potentially shedding light on reduced COVID-19 susceptibility in females.

Multimodal Single-Cell Omics Analysis of COVID-19 Sex Differences in Human Immune Systems (preprint)

Sex differences in the risk of SARS-CoV-2 infection have been controversial and the underlying mechanisms of COVID-19 sexual dimorphism remain understudied. Here we inspected sex differences in SARS-CoV-2 positivity, hospitalization, admission to the intensive care unit (ICU), sera immune profiling, and two single-cell RNA-sequencing (snRNA-seq) profiles from nasal tissues and peripheral blood mononuclear cells (PBMCs) of COVID-19 patients with varying degrees of disease severity. Our propensity score-matching observations revealed that male individuals have a 29% increased likelihood of SARS-CoV-2 positivity, with a hazard ration (HR) 1.32 (95% confidence interval [CI] 1.18-1.48) for hospitalization and HR 1.51 (95% CI 1.24-1.84) for admission to ICU. Sera from male patients at hospital admission had decreased lymphocyte count and elevated inflammatory markers (C-reactive protein, procalcitonin, and neutrophils). We found that SARS-CoV-2 entry factors, including ACE2, TMPRSS2, FURIN and NRP1, have elevated expression in nasal squamous cells from males with moderate and severe COVID-19. Cell-cell network proximity analysis suggests possible epithelium-immune cell interactions and immune vulnerability underlying a higher mortality in males with COVID-19. Monocyte-elevated expression of Toll like receptor 7 (TLR7) and Bruton tyrosine kinase (BTK) is associated with severe outcomes in males with COVID-19. These findings provide basis for understanding immune responses underlying sex differences, and designing sex-specific targeted treatments and patient care for COVID-19.

Dysregulated transcriptional responses to SARS-CoV-2 in the periphery support novel diagnostic approaches (preprint)

In order to elucidate novel aspects of the host response to SARS-CoV-2 we performed RNA sequencing on peripheral blood samples across 77 timepoints from 46 subjects with COVID-19 and compared them to subjects with seasonal coronavirus, influenza, bacterial pneumonia, and healthy controls. Early SARS-CoV-2 infection triggers a conserved transcriptomic response in peripheral blood that is heavily interferon-driven but also marked by indicators of early B-cell activation and antibody production. Interferon responses during SARS-CoV-2 infection demonstrate unique patterns of dysregulated expression compared to other infectious and healthy states. Heterogeneous activation of coagulation and fibrinolytic pathways are present in early COVID-19, as are IL1 and JAK/STAT signaling pathways, that persist into late disease. Classifiers based on differentially expressed genes accurately distinguished SARS-CoV-2 infection from other acute illnesses (auROC 0.95). The transcriptome in peripheral blood reveals unique aspects of the immune response in COVID-19 and provides for novel biomarker-based approaches to diagnosis.