SARS-COV-2 infections in inborn errors of immunity: A single center study.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a single-stranded RNA virus that causes coronavirus disease 2019 (COVID-19). One of the main topics of conversation in these past months in the world of immunology has been the issue of how patients with immune defects will fare if they contract this infection. To date there has been limited data on larger cohorts of patients with Inborn Errors of Immunity (IEI) diagnosed with COVID-19. Here, we review the data of COVID-19 infections in a single center cohort of 113 patients from the Mount Sinai Immunodeficiency program, who had 132 infections between January 2020 and June 2022. This included 56 males and 57 females, age range 2 - 84 (median 42). The mortality rate was 3%. Comparison between admitted patients revealed a significantly increased risk of hospitalization amongst the unvaccinated patients, 4% vaccinated vs 40% unvaccinated; odds ratio 15.0 (95% CI 4.2 - 53.4; p <0.00001). Additionally, COVID anti-spike antibody levels, determined in 36 of these patients post vaccination and before infection, were highly variable.

Novel clinical and immunological features associated with persistent post-acute sequelae of COVID-19 after six months of follow-up: a pilot study.

BACKGROUND: Currently, there is scant information regarding the features associated to the persistence of post-COVID-19 syndrome, which is the main aim of the present study. METHODS: A cohort study of 102 COVID-19 patients was conducted. The post-COVID-19 symptoms were assessed by a standardised questionnaire. Lymphocyte immunophenotyping was performed by flow cytometry and chemokines/cytokines, neutrophil extracellular traps, the tripartite motif 63, anti-cellular, and anti-SARS-CoV-2 IgG antibodies were addressed in serum. The primary outcome was the persistence of post-COVID-19 syndrome after six months follow-up. RESULTS: Thirteen patients (12.7%) developed the primary outcome and had a more frequent history of post-COVID-19 syndrome 3 months after infection onset (p = .044), increased levels of IL-1α (p = .011) and IP-10 (p = .037) and increased CD57 expression in CD8+ T cells (p = .003). There was a trend towards higher levels of IFN-γ (p = .051), IL-1ß (p = .062) and IL-6 (p = .087). The history of post COVID-19 in the previous 3 months, obesity, baseline serum MIP-1α and IP-10, and CD57 expression in CD8+ T cells were independently associated with the persistence of post-COVID-19 syndrome. CONCLUSION: Our data suggest an important relationship between a pro-inflammatory state mediated through metabolic pathways related to obesity and increased cellular senescence as a key element in the persistence of post-COVID-19 syndrome at six months of follow-up.

SARS-COV-2 infections in inborn errors of immunity: A single center study

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a single-stranded RNA virus that causes coronavirus disease 2019 (COVID-19). One of the main topics of conversation in these past months in the world of immunology has been the issue of how patients with immune defects will fare if they contract this infection. To date there has been limited data on larger cohorts of patients with Inborn Errors of Immunity (IEI) diagnosed with COVID-19. Here, we review the data of COVID-19 infections in a single center cohort of 113 patients from the Mount Sinai Immunodeficiency program, who had 132 infections between January 2020 and June 2022. This included 56 males and 57 females, age range 2 - 84 (median 42). The mortality rate was 3%. Comparison between admitted patients revealed a significantly increased risk of hospitalization amongst the unvaccinated patients, 4% vaccinated vs 40% unvaccinated;odds ratio 15.0 (95% CI 4.2 – 53.4;p <0.00001). Additionally, COVID anti-spike antibody levels, determined in 36 of these patients post vaccination and before infection, were highly variable.

Clinical and immunological features associated to the development of a sustained immune humoral response in COVID-19 patients: Results from a cohort study.

Background: Until now, most of the research addressing long-term humoral responses in coronavirus disease 2019 (COVID-19) had only evaluated the serum titers of anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) IgGs, without the assessment of the baseline antiviral clinical and immune profile, which is the aim of this study and may be the key factor leading to a broad and sustained antibody response. Methods: We included 103 patients with COVID-19. When the patients sought medical attention (baseline), a blood sample was drawn to perform immunophenotype of lymphocytes by flow cytometry. The patients were assessed 15 days after baseline and then every month until the third month, followed by a last visit 6 months after recruitment. We evaluated the anti-SARS-COV-2 IgG at all time points, and the serum levels of cytokines, chemokines, anti-cellular (AC) antibodies and neutrophil extracellular traps were also assessed during the follow-up. The primary outcome of the study was the presence of a sustained immune humoral response, defined as an anti-SARS-CoV-2 IgG titer >4.99 arbitrary units/mL in at least two consecutive measures. We used generalized lineal models to assess the features associated with this outcome and to assess the effect of the changes in the cytokines and chemokines throughout time on the development of a sustained humoral immune response. Results: At baseline the features associated to a sustained immune humoral response were the diagnosis of critical disease, absolute number of lymphocytes, serum IP-10, IL-4, IL-2, regulatory T cells, CD8+ T cells, and positive AC antibodies. Critical illness and the positivity of AC antibodies were associated with a sustained humoral immune response after 3 months, whilst critical illness and serum IL-13 were the explanatory variables after 6 months. Conclusion: A sustained immune humoral response is strongly related to critical COVID-19, which is characterized by the presence of AC antibodies, quantitative abnormalities in the T cell compartment, and the serum cytokines and chemokines during acute infection and throughout time.

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.

Household air pollution from wood-burning cookstoves and C-reactive protein among women in rural Honduras.

Household air pollution from solid fuel combustion was estimated to cause 2.31 million deaths worldwide in 2019; cardiovascular disease is a substantial contributor to the global burden. We evaluated the cross-sectional association between household air pollution (24-h gravimetric kitchen and personal particulate matter (PM2.5) and black carbon (BC)) and C-reactive protein (CRP) measured in dried blood spots among 107 women in rural Honduras using wood-burning traditional or Justa (an engineered combustion chamber) stoves. A suite of 6 additional markers of systemic injury and inflammation were considered in secondary analyses. We adjusted for potential confounders and assessed effect modification of several cardiovascular-disease risk factors. The median (25th, 75th percentiles) 24-h-average personal PM2.5 concentration was 115 µg/m3 (65,154 µg/m3) for traditional stove users and 52 µg/m3 (39, 81 µg/m3) for Justa stove users; kitchen PM2.5 and BC had similar patterns. Higher concentrations of PM2.5 and BC were associated with higher levels of CRP (e.g., a 25% increase in personal PM2.5 was associated with a 10.5% increase in CRP [95% CI: 1.2-20.6]). In secondary analyses, results were generally consistent with a null association. Evidence for effect modification between pollutant measures and four different cardiovascular risk factors (e.g., high blood pressure) was inconsistent. These results support the growing evidence linking household air pollution and cardiovascular disease.

Circulating B10 regulatory cells are decreased in severe and critical COVID-19.

The contribution of B cells in COVID-19 pathogenesis, beyond the production of specific antibodies against SARS-CoV-2, is still not well understood. Since one of their most relevant functional roles includes their immune-suppressive mechanisms, we decided to evaluate one of the most recognized human B regulatory subpopulations: the IL-10+ B10 cells, during COVID-19 onset. After stimulation of PBMCs for IL-10 induction, we employed multiparametric flow cytometry to determine B10 frequencies in severe and critical COVID-19 patients and then correlated those with clinical and laboratory parameters. Compared with healthy individuals, we detected a significant reduction in the B10 subset in both patient groups, which correlates with some inflammatory parameters that define the disease severity. This evidence suggests an aberrant role of B10 cells in immune responses against SARS-CoV-2 that needs to be further explained.

Artificial Intelligence and Democratization of the Use of Lung Ultrasound in COVID-19: On the Feasibility of Automatic Calculation of Lung Ultrasound Score

During the COVID-19 pandemic, lung ultrasound has been revealed as a powerful technique for diagnosis and follow-up of pneumonia, the principal complication of SARS-CoV-2 infection. Nevertheless, being a relatively new and unknown technique, the lack of trained personnel has limited its application worldwide. Computer-aided diagnosis could possibly help to reduce the learning curve for less experienced physicians, and to extend such a new technique such as lung ultrasound more quickly. This work presents the preliminary results of the ULTRACOV (Ultrasound in Coronavirus disease) study, aimed to explore the feasibility of a real-time image processing algorithm for automatic calculation of the lung ultrasound score (LUS). A total of 28 patients positive on COVID-19 were recruited and scanned in 12 thorax zones following the lung score protocol, saving a 3 s video at each probe position. Those videos were evaluated by an experienced physician and by a custom developed automated detection algorithm, looking for A-Lines, B-Lines, consolidations, and pleural effusions. The agreement between the findings of the expert and the algorithm was 88.0% for B-Lines, 93.4% for consolidations and 99.7% for pleural effusion detection, and 72.8% for the individual video score. The standard deviation of the patient lung score difference between the expert and the algorithm was ±2.2 points over 36. The exam average time with the ULTRACOV prototype was 5.3 min, while with a conventional scanner was 12.6 min. Conclusion: A good agreement between the algorithm output and an experienced physician was observed, which is a first step on the feasibility of developing a real-time aided-diagnosis lung ultrasound equipment. Additionally, the examination time was reduced to less than half with regard to a conventional ultrasound exam. Acquiring a complete lung ultrasound exam within a few minutes is possible using fairly simple ultrasound machines that are enhanced with artificial intelligence, such as the one we propose. This step is critical to democratize the use of lung ultrasound in these difficult times.

Severity of SARS-CoV-2 infection is linked to double-negative (CD27- IgD-) B cell subset numbers.

OBJECTIVES: The role of B cells in COVID-19, beyond the production of specific antibodies against SARS-CoV-2, is still not well understood. Here, we describe the novel landscape of circulating double-negative (DN) CD27- IgD- B cells in COVID-19 patients, representing a group of atypical and neglected subpopulations of this cell lineage. METHODS: Using multiparametric flow cytometry, we determined DN B cell subset amounts from 91 COVID-19 patients, correlated those with cytokines, clinical and laboratory parameters, and segregated them by principal components analysis. RESULTS: We detected significant increments in the DN2 and DN3 B cell subsets, while we found a relevant decrease in the DN1 B cell subpopulation, according to disease severity and patient outcomes. These DN cell numbers also appeared to correlate with pro- or anti-inflammatory signatures, respectively, and contributed to the segregation of the patients into disease severity groups. CONCLUSION: This study provides insights into DN B cell subsets' potential role in immune responses against SARS-CoV-2, particularly linked to the severity of COVID-19.

Training Conditions and Emotional Impact on Spanish Olympic Swimmers and Rowers in Social Isolation Due to COVID-19. Results of a Survey

This article analyzes the results of a survey conducted in 2020 with Spanish Olympic swimmers and rowers, who were confined to their homes due to the epidemiological crisis. The questionnaire was administered between 23 April and 25 May. Responses to the questionnaire on emotional and adaptive reactions during the COVID-19 confinement (REACOVID-19) were received from 88 subjects, who represented 100% of the total population of Spanish Olympic swimmers and rowers. Through this questionnaire, they were asked about their living conditions, their daily training habits and their psychological, cognitive and emotional adaptation during the confinement. The results show the commitment of these athletes to their sports goals and their responsibility in respecting the confinement rules. Sixty-seven per cent of them stated that they had not left their homes for 96 days. In these extreme circumstances, the majority trained an average of 11 to 13 h a week and coped with the confinement with a positive attitude, in spite of the inconveniences of social isolation, the lack of equipment and technical support for training and the limitations of their physical space. The article analyzes which emotional and social factors influenced both their motivation and their hours of training.

Redefining COVID-19 Severity and Prognosis: The Role of Clinical and Immunobiotypes.

Background: Most of the explanatory and prognostic models of COVID-19 lack of a comprehensive assessment of the wide COVID-19 spectrum of abnormalities. The aim of this study was to unveil novel biological features to explain COVID-19 severity and prognosis (death and disease progression). Methods: A predictive model for COVID-19 severity in 121 patients was constructed by ordinal logistic regression calculating odds ratio (OR) with 95% confidence intervals (95% CI) for a set of clinical, immunological, metabolomic, and other biological traits. The accuracy and calibration of the model was tested with the area under the curve (AUC), Somer's D, and calibration plot. Hazard ratios with 95% CI for adverse outcomes were calculated with a Cox proportional-hazards model. Results: The explanatory variables for COVID-19 severity were the body mass index (BMI), hemoglobin, albumin, 3-Hydroxyisovaleric acid, CD8+ effector memory T cells, Th1 cells, low-density granulocytes, monocyte chemoattractant protein-1, plasma TRIM63, and circulating neutrophil extracellular traps. The model showed an outstanding performance with an optimism-adjusted AUC of 0.999, and Somer's D of 0.999. The predictive variables for adverse outcomes in COVID-19 were severe and critical disease diagnosis, BMI, lactate dehydrogenase, Troponin I, neutrophil/lymphocyte ratio, serum levels of IP-10, malic acid, 3, 4 di-hydroxybutanoic acid, citric acid, myoinositol, and cystine. Conclusions: Herein, we unveil novel immunological and metabolomic features associated with COVID-19 severity and prognosis. Our models encompass the interplay among innate and adaptive immunity, inflammation-induced muscle atrophy and hypoxia as the main drivers of COVID-19 severity.

Neutrophil Extracellular Traps Contribute to COVID-19 Hyperinflammation and Humoral Autoimmunity.

The coronavirus disease 2019 (COVID-19) is related to enhanced production of NETs, and autoimmune/autoinflammatory phenomena. We evaluated the proportion of low-density granulocytes (LDG) by flow cytometry, and their capacity to produce NETs was compared with that of conventional neutrophils. NETs and their protein cargo were quantified by confocal microscopy and ELISA. Antinuclear antibodies (ANA), anti-neutrophil cytoplasmic antibodies (ANCA) and the degradation capacity of NETs were addressed in serum. MILLIPLEX assay was used to assess the cytokine levels in macrophages' supernatant and serum. We found a higher proportion of LDG in severe and critical COVID-19 which correlated with severity and inflammatory markers. Severe/critical COVID-19 patients had higher plasmatic NE, LL-37 and HMGB1-DNA complexes, whilst ISG-15-DNA complexes were lower in severe patients. Sera from severe/critical COVID-19 patients had lower degradation capacity of NETs, which was reverted after adding hrDNase. Anti-NET antibodies were found in COVID-19, which correlated with ANA and ANCA positivity. NET stimuli enhanced the secretion of cytokines in macrophages. This study unveils the role of COVID-19 NETs as inducers of pro-inflammatory and autoimmune responses. The deficient degradation capacity of NETs may contribute to the accumulation of these structures and anti-NET antibodies are related to the presence of autoantibodies.

Upper respiratory tract SARS-CoV-2 RNA loads in symptomatic and asymptomatic children and adults.

OBJECTIVES: Studies comparing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA load in the upper respiratory tract (URT) between children and adults-who either presented with coronavirus disease 2019 (COVID-19) or were asymptomatic-have yielded inconsistent results. Here, we conducted a retrospective, single-centre study to address this issue. PATIENTS AND METHODS: Included were 1184 consecutive subjects (256 children and 928 adults) testing positive for SARS-CoV-2 RNA in nasopharyngeal exudates (NPs); of these, 424 (121 children and 303 adults) had COVID-19 and 760 (135 children and 625 adults) were asymptomatic close contacts of COVID-19 patients. SARS-CoV-2 RNA testing was carried out using the TaqPath COVID-19 Combo Kit (Thermo Fisher Scientific, MS, USA). The AMPLIRUN® TOTAL SARS-CoV-2 RNA Control (Vircell SA, Granada, Spain) was used for estimating SARS-CoV-2 RNA loads (in copies/mL). SARS-CoV-2 RNA loads at the time of laboratory diagnosis (single specimen/patient) were used for comparison purposes. RESULTS: Median initial SARS-CoV-2 RNA load was lower (p 0.094) in children (6.98 log10 copies/mL, range 3.0-11.7) than in adults (7.14 log10 copies/mL, range 2.2-13.4) with COVID-19. As for asymptomatic individuals, median SARS-CoV-2 RNA load was comparable (p 0.97) in children (6.20 log10 copies/mL, range 1.8-11.6) and adults (6.48 log10 copies/mL, range 1.9-11.8). Children with COVID-19 symptoms displayed SARS-CoV-2 RNA loads (6.98 log10 copies/mL, range 3.0-11.7) comparable to those of their asymptomatic counterparts (6.20 log10 copies/mL, range 1.8-11.6) (p 0.61). Meanwhile in adults, median SARS-CoV-2 RNA load was significantly higher in symptomatic (7.14 log10 copies/mL, range 2.2-13.4) than in asymptomatic subjects (6.48 log10 copies/mL, range 1.9-11.8) (p < 0.001). Overall, the observed URT SARS-CoV-2 RNA clearance rate was faster in children than in adults. CONCLUSIONS: Based on viral load data at the time of diagnosis, our results suggest that SARS-CoV-2-infected children, with or without COVID-19, may display NP viral loads of comparable magnitude to those found in their adult counterparts. However, children may have shorter viral shedding than adults.

The furin cleavage site in the SARS-CoV-2 spike protein is required for transmission in ferrets.

SARS-CoV-2 entry requires sequential cleavage of the spike glycoprotein at the S1/S2 and the S2' cleavage sites to mediate membrane fusion. SARS-CoV-2 has a polybasic insertion (PRRAR) at the S1/S2 cleavage site that can be cleaved by furin. Using lentiviral pseudotypes and a cell-culture-adapted SARS-CoV-2 virus with an S1/S2 deletion, we show that the polybasic insertion endows SARS-CoV-2 with a selective advantage in lung cells and primary human airway epithelial cells, but impairs replication in Vero E6, a cell line used for passaging SARS-CoV-2. Using engineered spike variants and live virus competition assays and by measuring growth kinetics, we find that the selective advantage in lung and primary human airway epithelial cells depends on the expression of the cell surface protease TMPRSS2, which enables endosome-independent virus entry by a route that avoids antiviral IFITM proteins. SARS-CoV-2 virus lacking the S1/S2 furin cleavage site was shed to lower titres from infected ferrets and was not transmitted to cohoused sentinel animals, unlike wild-type virus. Analysis of 100,000 SARS-CoV-2 sequences derived from patients and 24 human postmortem tissues showed low frequencies of naturally occurring mutants that harbour deletions at the polybasic site. Taken together, our findings reveal that the furin cleavage site is an important determinant of SARS-CoV-2 transmission.

Metabolomics analysis reveals a modified amino acid metabolism that correlates with altered oxygen homeostasis in COVID-19 patients.

We identified the main changes in serum metabolites associated with severe (n = 46) and mild (n = 19) COVID-19 patients by gas chromatography coupled to mass spectrometry. The modified metabolic profiles were associated to an altered amino acid catabolism in hypoxic conditions. Noteworthy, three α-hydroxyl acids of amino acid origin increased with disease severity and correlated with altered oxygen saturation levels and clinical markers of lung damage. We hypothesize that the enzymatic conversion of α-keto-acids to α- hydroxyl-acids helps to maintain NAD recycling in patients with altered oxygen levels, highlighting the potential relevance of amino acid supplementation during SARS-CoV-2 infection.

Targeting Viral RNA-Dependent RNA Polymerases as an Antiviral Therapy.

Progressive globalization of our society brings not only worldwide integration, it also increases and promotes our exposure to new viral pathogens with evident impacts on our global health. Especially with the emergence of SARS-CoV-2, our biomedical research infrastructure has never been more compelled to rapidly develop antiviral regimens that demonstrate improved efficacy against these pathogens. Here, we showcase 3 poignant antivirals against the lucrative target, RNA-dependent RNA polymerase (RdRP) of RNA viruses - a timely and relevant topic given the present efforts against COVID-19. While effective drug designs against RdRP are important, their benefit and potential as a standard of care truly relies on them standing out in well-designed clinical trials.

B Cell Subsets as Severity-Associated Signatures in COVID-19 Patients.

Background: SARS-CoV-2 infection represents a global health problem that has affected millions of people. The fine host immune response and its association with the disease course have not yet been fully elucidated. Consequently, we analyze circulating B cell subsets and their possible relationship with COVID-19 features and severity. Methods: Using a multiparametric flow cytometric approach, we determined B cell subsets frequencies from 52 COVID-19 patients, grouped them by hierarchical cluster analysis, and correlated their values with clinical data. Results: The frequency of CD19+ B cells is increased in severe COVID-19 compared to mild cases. Specific subset frequencies such as transitional B cell subsets increase in mild/moderate cases but decrease with the severity of the disease. Memory B compartment decreased in severe and critical cases, and antibody-secreting cells are increased according to the severity of the disease. Other non-typical subsets such as double-negative B cells also showed significant changes according to disease severity. Globally, these differences allow us to identify severity-associated patient clusters with specific altered subsets. Finally, respiratory parameters, biomarkers of inflammation, and clinical scores exhibited correlations with some of these subpopulations. Conclusions: The severity of COVID-19 is accompanied by changes in the B cell subpopulations, either immature or terminally differentiated. Furthermore, the existing relationship of B cell subset frequencies with clinical and laboratory parameters suggest that these lymphocytes could serve as potential biomarkers and even active participants in the adaptive antiviral response mounted against SARS-CoV-2.