Major alterations in the mononuclear phagocyte landscape associated with COVID-19 severity.

Dendritic cells (DCs) and monocytes are crucial mediators of innate and adaptive immune responses during viral infection, but misdirected responses by these cells may contribute to immunopathology. Here, we performed high-dimensional flow cytometry-analysis focusing on mononuclear phagocyte (MNP) lineages in SARS-CoV-2-infected patients with moderate and severe COVID-19. We provide a deep and comprehensive map of the MNP landscape in COVID-19. A redistribution of monocyte subsets toward intermediate monocytes and a general decrease in circulating DCs was observed in response to infection. Severe disease coincided with the appearance of monocytic myeloid-derived suppressor cell-like cells and a higher frequency of pre-DC2. Furthermore, phenotypic alterations in MNPs, and their late precursors, were cell-lineage-specific and associated either with the general response against SARS-CoV-2 or COVID-19 severity. This included an interferon-imprint in DC1s observed in all patients and a decreased expression of the coinhibitory molecule CD200R in pre-DCs, DC2s, and DC3 subsets of severely sick patients. Finally, unsupervised analysis revealed that the MNP profile, alone, pointed to a cluster of COVID-19 nonsurvivors. This study provides a reference for the MNP response to SARS-CoV-2 infection and unravels mononuclear phagocyte dysregulations associated with severe COVID-19.

High-dimensional profiling reveals phenotypic heterogeneity and disease-specific alterations of granulocytes in COVID-19.

Since the outset of the COVID-19 pandemic, increasing evidence suggests that the innate immune responses play an important role in the disease development. A dysregulated inflammatory state has been proposed as a key driver of clinical complications in COVID-19, with a potential detrimental role of granulocytes. However, a comprehensive phenotypic description of circulating granulocytes in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected patients is lacking. In this study, we used high-dimensional flow cytometry for granulocyte immunophenotyping in peripheral blood collected from COVID-19 patients during acute and convalescent phases. Severe COVID-19 was associated with increased levels of both mature and immature neutrophils, and decreased counts of eosinophils and basophils. Distinct immunotypes were evident in COVID-19 patients, with altered expression of several receptors involved in activation, adhesion, and migration of granulocytes (e.g., CD62L, CD11a/b, CD69, CD63, CXCR4). Paired sampling revealed recovery and phenotypic restoration of the granulocytic signature in the convalescent phase. The identified granulocyte immunotypes correlated with distinct sets of soluble inflammatory markers, supporting pathophysiologic relevance. Furthermore, clinical features, including multiorgan dysfunction and respiratory function, could be predicted using combined laboratory measurements and immunophenotyping. This study provides a comprehensive granulocyte characterization in COVID-19 and reveals specific immunotypes with potential predictive value for key clinical features associated with COVID-19.

COVID-19-specific metabolic imprint yields insights into multiorgan system perturbations.

Corona disease 2019 (COVID-19) affects multiple organ systems. Recent studies have indicated perturbations in the circulating metabolome linked to COVID-19 severity. However, several questions pertain with respect to the metabolome in COVID-19. We performed an in-depth assessment of 1129 unique metabolites in 27 hospitalized COVID-19 patients and integrated results with large-scale proteomic and immunology data to capture multiorgan system perturbations. More than half of the detected metabolic alterations in COVID-19 were driven by patient-specific confounding factors ranging from comorbidities to xenobiotic substances. Systematically adjusting for this, a COVID-19-specific metabolic imprint was defined which, over time, underwent a switch in response to severe acute respiratory syndrome coronavirus-2 seroconversion. Integration of the COVID-19 metabolome with clinical, cellular, molecular, and immunological severity scales further revealed a network of metabolic trajectories aligned with multiple pathways for immune activation, and organ damage including neurological inflammation and damage. Altogether, this resource refines our understanding of the multiorgan system perturbations in severe COVID-19 patients.

Targeted plasma proteomics reveals signatures discriminating COVID-19 from sepsis with pneumonia.

BACKGROUND: COVID-19 remains a major public health challenge, requiring the development of tools to improve diagnosis and inform therapeutic decisions. As dysregulated inflammation and coagulation responses have been implicated in the pathophysiology of COVID-19 and sepsis, we studied their plasma proteome profiles to delineate similarities from specific features. METHODS: We measured 276 plasma proteins involved in Inflammation, organ damage, immune response and coagulation in healthy controls, COVID-19 patients during acute and convalescence phase, and sepsis patients; the latter included (i) community-acquired pneumonia (CAP) caused by Influenza, (ii) bacterial CAP, (iii) non-pneumonia sepsis, and (iv) septic shock patients. RESULTS: We identified a core response to infection consisting of 42 proteins altered in both COVID-19 and sepsis, although higher levels of cytokine storm-associated proteins were evident in sepsis. Furthermore, microbiologic etiology and clinical endotypes were linked to unique signatures. Finally, through machine learning, we identified biomarkers, such as TRIM21, PTN and CASP8, that accurately differentiated COVID-19 from CAP-sepsis with higher accuracy than standard clinical markers. CONCLUSIONS: This study extends the understanding of host responses underlying sepsis and COVID-19, indicating varying disease mechanisms with unique signatures. These diagnostic and severity signatures are candidates for the development of personalized management of COVID-19 and sepsis.

The Karolinska KI/K COVID-19 Immune Atlas: An open resource for immunological research and educational purposes.

The Karolinska KI/K COVID-19 Immune Atlas project was conceptualized in March 2020 as a part of the academic research response to the developing SARS-CoV-2 pandemic. The aim was to rapidly provide a curated dataset covering the acute immune response towards SARS-CoV-2 infection in humans, as it occurred during the first wave. The Immune Atlas was built as an open resource for broad research and educational purposes. It contains a presentation of the response evoked by different immune and inflammatory cells in defined naïve patient-groups as they presented with moderate and severe COVID-19 disease. The present Resource Article describes how the Karolinska KI/K COVID-19 Immune Atlas allow scientists, students, and other interested parties to freely explore the nature of the immune response towards human SARS-CoV-2 infection in an online setting.

Allergy development is associated with consumption of breastmilk with a reduced microbial richness in the first month of life.

BACKGROUND: Early colonization with a diverse microbiota seems to play a crucial role for appropriate immune maturation during childhood. Breastmilk microbiota is one important source of microbes for the infant, transferred together with maternal IgA antibodies. We previously observed that allergy development during childhood was associated with aberrant IgA responses to the gut microbiota already at 1 month of age, when the IgA antibodies are predominantly maternally derived in breastfed infants. OBJECTIVE: To determine the microbial composition and IgA-coated bacteria in breastmilk in relation to allergy development in children participating in an intervention trial with pre- and post-natal Lactobacillus reuteri supplementation. METHODS: A combination of flow cytometric cell sorting and 16S rRNA gene sequencing was used to characterize the bacterial recognition patterns by IgA in breastmilk samples collected one month post-partum from 40 mothers whose children did or did not develop allergic and asthmatic symptoms during the first 7 years of age. RESULTS: The milk fed to children developing allergic manifestations had significantly lower bacterial richness, when compared to the milk given to children that remained healthy. Probiotic treatment influenced the breastmilk microbiota composition. However, the proportions of IgA-coated bacteria, the total bacterial load and the patterns of IgA-coating were similar in breastmilk between mothers of healthy children and those developing allergies. CONCLUSION: Consumption of breastmilk with a reduced microbial richness in the first month of life may play an important role in allergy development during childhood.

MAMI: a birth cohort focused on maternal-infant microbiota during early life.

BACKGROUND: Early microbial colonization is a relevant aspect in human health. Altered microbial colonization patterns have been linked to an increased risk of non-communicable diseases (NCDs). Advances in understanding host-microbe interactions highlight the pivotal role of maternal microbiota on infant health programming. This birth cohort is aimed to characterize the maternal microbes transferred to neonates during the first 1000 days of life, as well as to identify the potential host and environmental factors, such as gestational age, mode of delivery, maternal/infant diet, and exposure to antibiotics, which affect early microbial colonization. METHODS: MAMI is a prospective mother-infant birth cohort in the Spanish-Mediterranean area. Mothers were enrolled at the end of pregnancy and families were follow-up during the first years of life. Maternal-infant biological samples were collected at several time points from birth to 24 months of life. Clinical and anthropometric characteristics and dietary information is available. Specific qPCR and 16S rRNA gene sequencing as well as short chain fatty acid (SCFAs) profile would be obtained. Multivariable models will be used to identy associations between microbiota and clinical and anthropometric data controlling for confounders. MAMI would contribute to a better understanding of the interaction between diet, microbiota and host response in early life health programming, enabling new applications in the field of personalized nutrition and medicine. TRIAL REGISTRATION: The study is registered on the ClinicalTrial.gov platform NCT03552939. (June 12, 2018).

Aberrant IgA responses to the gut microbiota during infancy precede asthma and allergy development.

BACKGROUND: Although a reduced gut microbiota diversity and low mucosal total IgA levels in infancy have been associated with allergy development, IgA responses to the gut microbiota have not yet been studied. OBJECTIVE: We sought to determine the proportions of IgA coating together with the characterization of the dominant bacteria, bound to IgA or not, in infant stool samples in relation to allergy development. METHODS: A combination of flow cytometric cell sorting and deep sequencing of the 16S rDNA gene was used to characterize the bacterial recognition patterns by IgA in stool samples collected at 1 and 12 months of age from children staying healthy or having allergic symptoms up to 7 years of age. RESULTS: The children with allergic manifestations, particularly asthma, during childhood had a lower proportion of IgA bound to fecal bacteria at 12 months of age compared with healthy children. These alterations cannot be attributed to differences in IgA levels or bacterial load between the 2 groups. Moreover, the bacterial targets of early IgA responses (including coating of the Bacteroides genus), as well as IgA recognition patterns, differed between healthy children and children with allergic manifestations. Altered IgA recognition patterns in children with allergy were observed already at 1 month of age, when the IgA antibodies are predominantly maternally derived in breast-fed children. CONCLUSION: An aberrant IgA responsiveness to the gut microbiota during infancy precedes asthma and allergy development, possibly indicating an impaired mucosal barrier function in allergic children.

Bugging allergy; role of pre-, pro- and synbiotics in allergy prevention.

Large-scale biodiversity loss and complex changes in social behaviors are altering human microbial ecology. This is increasingly implicated in the global rise in inflammatory diseases, most notably the "allergy epidemic" in very early life. Colonization of human ecological niches, particularly the gastrointestinal tract, is critical for normal local and systemic immune development and regulation. Disturbances in composition, diversity and timing of microbial colonization have been associated with increased allergy risk, indicating the importance of strategies to restore a dysbiotic gut microbiota in the primary prevention of allergic diseases, including the administration of probiotics, prebiotics and synbiotics. Here, we summarize and discuss findings of randomized clinical trials that have examined the effects of these microbiome-related strategies on short and long-term allergy preventative effects - including new guidelines from the World Allergy Organization which now recommend probiotics and prebiotics for allergy prevention under certain conditions. The relatively low quality evidence, limited comparative studies and large heterogeneity between studies, have collectively hampered recommendations on specific probiotic strains, specific timing and specific conditions for the most effective preventive management. At the same time the risk of using available products is low. While further research is needed before specific practice guidelines on supplement probiotics and prebiotics, it is equally important that the underlying dietary and lifestyle factors of dysbiosis are addressed at both the individual and societal levels.

SARS-CoV-2-specific humoral and cellular immunity persists through 9 months irrespective of COVID-19 severity at hospitalisation.

OBJECTIVES: Humoral and cellular immunity to SARS-CoV-2 following COVID-19 will likely contribute to protection from reinfection or severe disease. It is therefore important to characterise the initiation and persistence of adaptive immunity to SARS-CoV-2 amidst the ongoing pandemic. METHODS: Here, we conducted a longitudinal study on hospitalised moderate and severe COVID-19 patients from the acute phase of disease into convalescence at 5 and 9 months post-symptom onset. Utilising flow cytometry, serological assays as well as B cell and T cell FluoroSpot assays, we assessed the magnitude and specificity of humoral and cellular immune responses during and after human SARS-CoV-2 infection. RESULTS: During acute COVID-19, we observed an increase in germinal centre activity, a substantial expansion of antibody-secreting cells and the generation of SARS-CoV-2-neutralising antibodies. Despite gradually decreasing antibody levels, we show persistent, neutralising antibody titres as well as robust specific memory B cell responses and polyfunctional T cell responses at 5 and 9 months after symptom onset in both moderate and severe COVID-19 patients. CONCLUSION: Our findings describe the initiation and, importantly, persistence of cellular and humoral SARS-CoV-2-specific immunological memory in hospitalised COVID-19 patients long after recovery, likely contributing towards protection against reinfection.

Topic Application of the Probiotic Streptococcus dentisani Improves Clinical and Microbiological Parameters Associated With Oral Health.

Streptococcus dentisani 7746, isolated from dental plaque of caries-free individuals, has been shown to have several beneficial effects in vitro which could contribute to promote oral health, including an antimicrobial activity against oral pathogens by the production of bacteriocins and a pH buffering capacity through ammonia production. Previous work has shown that S. dentisani was able to colonize the oral cavity for 2-4 weeks after application. The aim of the present work was to evaluate its clinical efficacy by a randomized, double-blind, placebo-controlled parallel group study. Fifty nine volunteers were enrolled in the study and randomly assigned to a treatment or placebo group. The treatment consisted of a bucco-adhesive gel application (2.5 109 cfu/dose) with a dental splint for 5 min every 48 h, for a period of 1 month (i.e., 14 doses). Dental plaque and saliva samples were collected at baseline, 15 and 30 days after first application, and 15 days after the end of treatment. At baseline, there was a significant correlation between S. dentisani levels and frequency of toothbrushing. Salivary flow, a major factor influencing oral health, was significantly higher in the probiotic group at day 15 compared with the placebo (4.4 and 3.4 ml/5 min, respectively). In the probiotic group, there was a decrease in the amount of dental plaque and in gingival inflammation, but no differences were observed in the placebo group. The probiotic group showed a significant increase in the levels of salivary ammonia and calcium. Finally, Illumina sequencing of plaque samples showed a beneficial shift in bacterial composition at day 30 relative to baseline, with a reduction of several cariogenic organisms and the key players in plaque formation, probably as a result of bacteriocins production. Only 58% of the participants in the probiotic group showed increased plaque levels of S. dentisani at day 30 and 71% by day 45, indicating that the benefits of S. dentisani application could be augmented by improving colonization efficiency. In conclusion, the application of S. dentisani 7746 improved several clinical and microbiological parameters associated with oral health, supporting its use as a probiotic to prevent tooth decay.

Gut Microbiota and Mucosal Immunity in the Neonate.

Gut microbiota colonization is a complex, dynamic, and step-wise process that is in constant development during the first years of life. This microbial settlement occurs in parallel with the maturation of the immune system, and alterations during this period, due to environmental and host factors, are considered to be potential determinants of health-outcomes later in life. Given that host⁻microbe interactions are mediated by the immune system response, it is important to understand the close relationship between immunity and the microbiota during birth, lactation, and early infancy. This work summarizes the evidence to date on early gut microbiota colonization, and how it influences the maturation of the infant immune system and health during the first 1000 days of life. This review will also address the influence of perinatal antibiotic intake and the importance of delivery mode and breastfeeding for an appropriate development of gut immunity.

Oral microbiome development during childhood: an ecological succession influenced by postnatal factors and associated with tooth decay.

Information on how the oral microbiome develops during early childhood and how external factors influence this ecological process is scarce. We used high-throughput sequencing to characterize bacterial composition in saliva samples collected at 3, 6, 12, 24 months and 7 years of age in 90 longitudinally followed children, for whom clinical, dietary and health data were collected. Bacterial composition patterns changed through time, starting with "early colonizers", including Streptococcus and Veillonella; other bacterial genera such as Neisseria settled after 1 or 2 years of age. Dental caries development was associated with diverging microbial composition through time. Streptococcus cristatus appeared to be associated with increased risk of developing tooth decay and its role as potential biomarker of the disease should be studied with species-specific probes. Infants born by C-section had initially skewed bacterial content compared with vaginally delivered infants, but this was recovered with age. Shorter breastfeeding habits and antibiotic treatment during the first 2 years of age were associated with a distinct bacterial composition at later age. The findings presented describe oral microbiota development as an ecological succession where altered colonization pattern during the first year of life may have long-term consequences for child´s oral and systemic health.