Altered and allele-specific open chromatin landscape reveals epigenetic and genetic regulators of innate immunity in COVID-19.

SARS-CoV-2 infection causes severe COVID-19 in some patients and mild in others. Dysfunctional innate immune responses have been identified to contribute to COVID-19 severity, but the key regulators are still unknown. Here, we present an integrative single-cell multi-omics analysis of peripheral blood mononuclear cells from hospitalized and convalescent COVID-19 patients. In classical monocytes, we identified genes that were potentially regulated by differential chromatin accessibility. Then, sub-clustering and motif-enrichment analyses reveals disease condition-specific regulation by transcription factors and their targets, including an interaction between C/EBPs and a long-noncoding RNA LUCAT1, which we validated through loss-of-function experiments. Finally, we investigated genetic risk variants that exhibit allele-specific open chromatin (AsoC) in COVID-19 patients and identified a SNP rs6800484-C, which is associated with lower expression of CCR2 and may contribute to higher viral loads and higher risk of COVID-19 hospitalization. Altogether, our study highlights the diverse genetic and epigenetic regulators that contribute to COVID-19.

Pausing methotrexate prevents impairment of Omicron BA.1 and BA.2 neutralisation after COVID-19 booster vaccination.

OBJECTIVE: The level of neutralising capacity against Omicron BA.1 and BA.2 after third COVID-19 vaccination in patients on paused or continuous methotrexate (MTX) therapy is unclear. METHODS: In this observational cohort study, neutralising serum activity against SARS-CoV-2 wild-type (Wu01) and variant of concern Omicron BA.1 and BA.2 were assessed by pseudovirus neutralisation assay before, 4 and 12 weeks after mRNA booster immunisation in 50 rheumatic patients on MTX, 26 of whom paused the medication. 44 non-immunosuppressed persons (NIP) served as control group. RESULTS: While the neutralising serum activity against SARS-CoV-2 Wu01 and Omicron variants increased 67-73 fold in the NIP after booster vaccination, the serum activity in patients receiving MTX increased only 20-23 fold. Patients who continued MTX treatment during vaccination had significantly lower neutralisation against all variants at weeks 4 and 12 compared with patients who paused MTX and the control group, except for BA.2 at week 12. Patients who paused MTX reached comparably high neutralising capacities as NIP, except for Wu01 at week 12. The duration of the MTX pause after-not before-was associated with a significantly higher neutralisation capacity against all three variants, with an optimal duration at 10 days after vaccination. CONCLUSION: Patients pausing MTX after COVID-19 booster showed a similar vaccine response to NIP. Patients who continued MTX demonstrated an impaired response indicating a potentially beneficial second booster vaccination. Our data also suggest that a 1 week MTX break is sufficient if the last administration of MTX occurs 1-3 days before vaccination.

The human host response to monkeypox infection: a proteomic case series study.

The rapid rise of monkeypox (MPX) cases outside previously endemic areas prompts for a better understanding of the disease. We studied the plasma proteome of a group of MPX patients with a similar infection history and clinical manifestation typical for the current outbreak. We report that MPX in this case series is associated with a strong plasma proteomic response among nutritional and acute phase response proteins. Moreover, we report a correlation between plasma proteins and disease severity. Contrasting the MPX host response with that of COVID-19, we find a range of similarities, but also important differences. For instance, CFHR1 is induced in COVID-19, but suppressed in MPX, reflecting the different roles of the complement system in the two infectious diseases. Of note, the spatial overlap in response proteins suggested that a COVID-19 biomarker panel assay could be repurposed for MPX. Applying a targeted protein panel assay provided encouraging results and distinguished MPX cases from healthy controls. Hence, our results provide a first proteomic characterization of the MPX human host response and encourage further research on protein-panel assays in emerging infectious diseases.

Humoral immunity to SARS-CoV-2 elicited by combination COVID-19 vaccination regimens.

The SARS-CoV-2 pandemic prompted a global vaccination effort and the development of numerous COVID-19 vaccines at an unprecedented scale and pace. As a result, current COVID-19 vaccination regimens comprise diverse vaccine modalities, immunogen combinations, and dosing intervals. Here, we compare vaccine-specific antibody and memory B cell responses following two-dose mRNA, single-dose Ad26.COV.2S, and two-dose ChAdOx1, or combination ChAdOx1/mRNA vaccination. Plasma-neutralizing activity, as well as the magnitude, clonal composition, and antibody maturation of the RBD-specific memory B cell compartments, showed substantial differences between the vaccination regimens. While individual monoclonal antibodies derived from memory B cells exhibited similar binding affinities and neutralizing potency against Wuhan-Hu-1 SARS-CoV-2, there were significant differences in epitope specificity and neutralizing breadth against viral variants of concern. Although the ChAdOx1 vaccine was inferior to mRNA and Ad26.COV.2S in several respects, biochemical and structural analyses revealed enrichment in a subgroup of memory B cell neutralizing antibodies with distinct RBD-binding properties resulting in remarkable potency and breadth.

Nonproductive exposure of PBMCs to SARS-CoV-2 induces cell-intrinsic innate immune responses.

Cell-intrinsic responses mounted in PBMCs during mild and severe COVID-19 differ quantitatively and qualitatively. Whether they are triggered by signals emitted by productively infected cells of the respiratory tract or result from physical interaction with virus particles remains unclear. Here, we analyzed susceptibility and expression profiles of PBMCs from healthy donors upon ex vivo exposure to SARS-CoV and SARS-CoV-2. In line with the absence of detectable ACE2 receptor expression, human PBMCs were refractory to productive infection. RT-PCR experiments and single-cell RNA sequencing revealed JAK/STAT-dependent induction of interferon-stimulated genes (ISGs) but not proinflammatory cytokines. This SARS-CoV-2-specific response was most pronounced in monocytes. SARS-CoV-2-RNA-positive monocytes displayed a lower ISG signature as compared to bystander cells of the identical culture. This suggests a preferential invasion of cells with a low ISG baseline profile or delivery of a SARS-CoV-2-specific sensing antagonist upon efficient particle internalization. Together, nonproductive physical interaction of PBMCs with SARS-CoV-2- and, to a much lesser extent, SARS-CoV particles stimulate JAK/STAT-dependent, monocyte-accentuated innate immune responses that resemble those detected in vivo in patients with mild COVID-19.

ELF5 is a potential respiratory epithelial cell-specific risk gene for severe COVID-19.

Despite two years of intense global research activity, host genetic factors that predispose to a poorer prognosis of COVID-19 infection remain poorly understood. Here, we prioritise eight robust (e.g., ELF5) or suggestive but unreported (e.g., RAB2A) candidate protein mediators of COVID-19 outcomes by integrating results from the COVID-19 Host Genetics Initiative with population-based plasma proteomics using statistical colocalisation. The transcription factor ELF5 (ELF5) shows robust and directionally consistent associations across different outcome definitions, including a >4-fold higher risk (odds ratio: 4.88; 95%-CI: 2.47-9.63; p-value < 5.0 × 10-6) for severe COVID-19 per 1 s.d. higher genetically predicted plasma ELF5. We show that ELF5 is specifically expressed in epithelial cells of the respiratory system, such as secretory and alveolar type 2 cells, using single-cell RNA sequencing and immunohistochemistry. These cells are also likely targets of SARS-CoV-2 by colocalisation with key host factors, including ACE2 and TMPRSS2. In summary, large-scale human genetic studies together with gene expression at single-cell resolution highlight ELF5 as a risk gene for severe COVID-19, supporting a role of epithelial cells of the respiratory system in the adverse host response to SARS-CoV-2.

SARS-CoV-2 Omicron sublineages exhibit distinct antibody escape patterns.

SARS-CoV-2 neutralizing antibodies play a critical role in COVID-19 prevention and treatment but are challenged by viral evolution and the emergence of novel escape variants. Importantly, the recently identified Omicron sublineages BA.2.12.1 and BA.4/5 are rapidly becoming predominant in various countries. By determining polyclonal serum activity of 50 convalescent or vaccinated individuals against BA.1, BA.1.1, BA.2, BA.2.12.1, and BA.4/5, we reveal a further reduction in BA.4/5 susceptibility to vaccinee sera. Most notably, delineation of sensitivity to an extended 163-antibody panel demonstrates pronounced antigenic differences with distinct escape patterns among Omicron sublineages. Antigenic distance and/or higher resistance may therefore favor immune-escape-mediated BA.4/5 expansion after the first Omicron wave. Finally, while most clinical-stage monoclonal antibodies are inactive against Omicron sublineages, we identify promising antibodies with high pan-SARS-CoV-2 neutralizing potency. Our study provides a detailed understanding of Omicron-sublineage antibody escape that can inform on effective strategies against COVID-19.

Early and Rapid Identification of COVID-19 Patients with Neutralizing Type I Interferon Auto-antibodies.

PURPOSE: Six to 19% of critically ill COVID-19 patients display circulating auto-antibodies against type I interferons (IFN-AABs). Here, we establish a clinically applicable strategy for early identification of IFN-AAB-positive patients for potential subsequent clinical interventions. METHODS: We analyzed sera of 430 COVID-19 patients from four hospitals for presence of IFN-AABs by ELISA. Binding specificity and neutralizing activity were evaluated via competition assay and virus-infection-based neutralization assay. We defined clinical parameters associated with IFN-AAB positivity. In a subgroup of critically ill patients, we analyzed effects of therapeutic plasma exchange (TPE) on the levels of IFN-AABs, SARS-CoV-2 antibodies and clinical outcome. RESULTS: The prevalence of neutralizing AABs to IFN-α and IFN-ω in COVID-19 patients from all cohorts was 4.2% (18/430), while being undetectable in an uninfected control cohort. Neutralizing IFN-AABs were detectable exclusively in critically affected (max. WHO score 6-8), predominantly male (83%) patients (7.6%, 18/237 for IFN-α-AABs and 4.6%, 11/237 for IFN-ω-AABs in 237 patients with critical COVID-19). IFN-AABs were present early post-symptom onset and at the peak of disease. Fever and oxygen requirement at hospital admission co-presented with neutralizing IFN-AAB positivity. IFN-AABs were associated with lower probability of survival (7.7% versus 80.9% in patients without IFN-AABs). TPE reduced levels of IFN-AABs in three of five patients and may increase survival of IFN-AAB-positive patients compared to those not undergoing TPE. CONCLUSION: IFN-AABs may serve as early biomarker for the development of severe COVID-19. We propose to implement routine screening of hospitalized COVID-19 patients for rapid identification of patients with IFN-AABs who most likely benefit from specific therapies.

Disease Severity, Fever, Age, and Sex Correlate With SARS-CoV-2 Neutralizing Antibody Responses.

Clinical trials on the use of COVID-19 convalescent plasma remain inconclusive. While data on safety is increasingly available, evidence for efficacy is still sparse. Subgroup analyses hint to a dose-response relationship between convalescent plasma neutralizing antibody levels and mortality. In particular, patients with primary and secondary antibody deficiency might benefit from this approach. However, testing of neutralizing antibodies is limited to specialized biosafety level 3 laboratories and is a time- and labor-intense procedure. In this single center study of 206 COVID-19 convalescent patients, clinical data, results of commercially available ELISA testing of SARS-CoV-2 spike-IgG and -IgA, and levels of neutralizing antibodies, determined by plaque reduction neutralization testing (PRNT), were analyzed. At a medium time point of 58 days after symptom onset, only 12.6% of potential plasma donors showed high levels of neutralizing antibodies (PRNT50 ≥ 1:320). Multivariable proportional odds logistic regression analysis revealed need for hospitalization due to COVID-19 (odds ratio 6.87; p-value 0.0004) and fever (odds ratio 3.00; p-value 0.0001) as leading factors affecting levels of SARS-CoV-2 neutralizing antibody titers in convalescent plasma donors. Using penalized estimation, a predictive proportional odds logistic regression model including the most important variables hospitalization, fever, age, sex, and anosmia or dysgeusia was developed. The predictive discrimination for PRNT50 ≥ 1:320 was reasonably good with AUC: 0.86 (with 95% CI: 0.79-0.92). Combining clinical and ELISA-based pre-screening, assessment of neutralizing antibodies could be spared in 75% of potential donors with a maximal loss of 10% of true positives (PRNT50 ≥ 1:320).

Hypertension delays viral clearance and exacerbates airway hyperinflammation in patients with COVID-19.

In coronavirus disease 2019 (COVID-19), hypertension and cardiovascular diseases are major risk factors for critical disease progression. However, the underlying causes and the effects of the main anti-hypertensive therapies-angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin receptor blockers (ARBs)-remain unclear. Combining clinical data (n = 144) and single-cell sequencing data of airway samples (n = 48) with in vitro experiments, we observed a distinct inflammatory predisposition of immune cells in patients with hypertension that correlated with critical COVID-19 progression. ACEI treatment was associated with dampened COVID-19-related hyperinflammation and with increased cell intrinsic antiviral responses, whereas ARB treatment related to enhanced epithelial-immune cell interactions. Macrophages and neutrophils of patients with hypertension, in particular under ARB treatment, exhibited higher expression of the pro-inflammatory cytokines CCL3 and CCL4 and the chemokine receptor CCR1. Although the limited size of our cohort does not allow us to establish clinical efficacy, our data suggest that the clinical benefits of ACEI treatment in patients with COVID-19 who have hypertension warrant further investigation.

SARS-CoV-2-reactive T cells in healthy donors and patients with COVID-19.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused the rapidly unfolding coronavirus disease 2019 (COVID-19) pandemic1,2. Clinical manifestations of COVID-19 vary, ranging from asymptomatic infection to respiratory failure. The mechanisms that determine such variable outcomes remain unresolved. Here we investigated CD4+ T cells that are reactive against the spike glycoprotein of SARS-CoV-2 in the peripheral blood of patients with COVID-19 and SARS-CoV-2-unexposed healthy donors. We detected spike-reactive CD4+ T cells not only in 83% of patients with COVID-19 but also in 35% of healthy donors. Spike-reactive CD4+ T cells in healthy donors were primarily active against C-terminal epitopes in the spike protein, which show a higher homology to spike glycoproteins of human endemic coronaviruses, compared with N-terminal epitopes. Spike-protein-reactive T cell lines generated from SARS-CoV-2-naive healthy donors responded similarly to the C-terminal region of the spike proteins of the human endemic coronaviruses 229E and OC43, as well as that of SARS-CoV-2. This results indicate that spike-protein cross-reactive T cells are present, which were probably generated during previous encounters with endemic coronaviruses. The effect of pre-existing SARS-CoV-2 cross-reactive T cells on clinical outcomes remains to be determined in larger cohorts. However, the presence of spike-protein cross-reactive T cells in a considerable fraction of the general population may affect the dynamics of the current pandemic, and has important implications for the design and analysis of upcoming trials investigating COVID-19 vaccines.