Farm-dust mediated protection of childhood asthma: Mass cytometry reveals novel cellular regulation.

BACKGROUND: Farm-dust mediated asthma protection in childhood was replicated in numerous epidemiological studies. Central immune mechanisms are not fully understood. This exploratory study aimed to disentangle underlying immunological regulation of farm-dust mediated protection in peripheral blood on a single-cell level. METHODS: Single-cell protein expression of in vitro farm-dust stimulated and unstimulated cells from allergic asthmatics and healthy controls were measured using mass cytometry. Analysis of innate and adaptive cellular proportions (linear regression) and T-cell proliferation was performed. Functional marker intensity was investigated using Earth Mover's Distance and the Monte Carlo permutation test. RESULTS: Farm-dust stimulation induced cell type-specific regulation: Key-features of farm-dust stimulation comprised opposing regulation of immune-cell frequencies (downregulated innate cell populations (monocytes/DCs (p < .001), NK-cells (p < .05)) and upregulated adaptive populations (B-cells, CD4+ T-cells (both p < .05)), reduced CD4+ CD25- T-cell proliferation, and differential cell type-specific functional marker expression. Following stimulation, functional marker analysis revealed induced activation (CD25) in T-cells and NK-T-cells in both phenotypes even after correction for multiple testing. Cytotoxicity (GZMB) and inflammation (pERK1/2, pp38) related markers were reduced in T-cells exclusively in asthmatic children. Asthma-associated markers (Gata3, RORγ, and HLA-DR) were reduced in T- and innate- cell populations of asthmatics following stimulation. B-cells displayed a phenotypically independent increase of diverse functional markers upon farm-dust stimulation. CONCLUSIONS: This study mimicking in vivo environmental exposure identified a novel profile of immune-regulatory markers using mass cytometry demonstrating decreased asthma-associated markers following farm-dust stimulation. These findings may be key for further studies on asthma prevention in childhood.

Symptom trajectories in infancy for the prediction of subsequent wheeze and asthma in the BILD and PASTURE cohorts: a dynamic network analysis.

BACKGROUND: Host and environment early-life risk factors are associated with progression of wheezing symptoms over time; however, their individual contribution is relatively small. We hypothesised that the dynamic interactions of these factors with an infant's developing respiratory system are the dominant factor for subsequent wheeze and asthma. METHODS: In this dynamic network analysis we used data from term healthy infants from the Basel-Bern Infant Lung Development (BILD) cohort (435 neonates aged 0-4 weeks recruited in Switzerland between Jan 1, 1999, and Dec 31, 2012) and replicated the findings in the Protection Against Allergy Study in Rural Environments (PASTURE) cohort (498 infants aged 0-12 months recruited in Germany, Switzerland, Austria, France, and Finland between Jan 1, 2002, and Oct 31, 2006). BILD exclusion criteria for the current study were prematurity (<37 weeks), major birth defects, perinatal disease of the neonate, and incomplete follow-up period. PASTURE exclusion criteria were women younger than 18 years, a multiple pregnancy, the sibling of a child was already included in the study, the family intended to move away from the area where the study was conducted, and the family had no telephone connection. Outcome groups were subsequent wheeze, asthma, and healthy. The first outcome was defined as ever wheezed between the age of 2 years and 6 years. Week-by-week correlations of the determining factors with cumulative symptom scores (CSS) were calculated from weeks 2 to 52 (BILD) and weeks 8 to 52 (PASTURE). The complex dynamic interaction between the determining factors and the CSS was assessed via dynamic host-environment correlation network, quantified by a simple descriptor: trajectory function G(t). Wheeze outcomes at age 2-6 years were compared in 335 infants from BILD and 437 infants from PASTURE, and asthma outcomes were analysed at age 6 years in a merged cohort of 783 infants. FINDINGS: CSS was significantly different for wheeze and asthma outcomes and became increasingly important during infancy in direct comparison with all determining factors. Weekly symptoms were tracked for groups of infants, showing a non-linear increase with time. Using logistic regression classification, G(t) distinguished between the healthy group and wheeze or asthma groups (area under the curve>0·97, p<0·0001; sensitivity analysis confirmed significant CSS association with wheeze [BILD p=0·0002 and PASTURE p=0·068]) and G(t) was also able to distinguish between the farming and non-farming exposure groups (p<0·0001). INTERPRETATION: Similarly to other risk factors, CSS had weak sensitivity and specificity to identify risks at the individual level. At group level however, the dynamic host-environment correlation network properties (G(t)) showed excellent discriminative ability for identifying groups of infants with subsequent wheeze and asthma. Results from this study are consistent with the 2018 Lancet Commission on asthma, which emphasised the importance of dynamic interactions between risk factors during development and not the risk factors per se. FUNDING: The Swiss National Science Foundation, the Kühne Foundation, the EFRAIM study EU research grant, the FORALLVENT study EU research grant, and the Leibniz Prize.

Urinary eosinophil-derived neurotoxin is associated with reduced lung function in pediatric asthma.

INTRODUCTION: Eosinophil-derived neurotoxin (EDN) is a biomarker for eosinophilic activation. Urinary (u) EDN may allow non-invasive monitoring of asthma, but clinical recommendations are lacking. We assessed the potential of uEDN as a marker of disease activity in pediatric asthma. METHODS: We assessed urine samples of 371 children from the German ALLIANCE study cohort, from which we had: 169 preschool wheezers (<6 years), 80 asthmatics (≥6 years), and 122 healthy controls using the ImmunoCAP™ EDN Assay. Creatinine (Cr)-adjusted uEDN values were analyzed using correlations, association tests, (non) parametric statistics, multiple linear, and multivariable regression. RESULTS: uEDN/uCr values were higher in atopic versus non-atopic preschool-aged subjects (p = .035) and associated with the sum of allergen-specific IgE in younger (r = 0.24, p = .003), and older subjects (r = 0.23, p = .043). uEDN/uCr was marginally a good determinant for atopy (p = .078, for subjects aged <6 years, and p = .058 for subjects ≥6 years). Children with the T2-high phenotype had higher uEDN/uCr (p < .001) versus T2-low-irrespective of using uEDN/uCr or blood eosinophils in combination to allergen sIgE for disease phenotyping. uEDN/uCr significantly correlated with reduced lung function among asthmatics (FEV1 z-scores: r = -0.30, p = .007, and FEV1/FVC z-scores: r = -0.24, p = .038). Using multivariable modeling, uEDN/uCr was an independent determinant of FEV1 (p = .038), and to a lesser extent, FEV1/FVC (p = .080). CONCLUSIONS: uEDN/uCr may serve as a non-invasive biomarker for clinical features such as lung function in pediatric asthma. We highlight the utility of uEDN/uCr as a biomarker that can be easily assessed using widely available robust diagnostic immunoassays.

An integrated molecular risk score early in life for subsequent childhood asthma risk.

BACKGROUND: Numerous children present with early wheeze symptoms, yet solely a subgroup develops childhood asthma. Early identification of children at risk is key for clinical monitoring, timely patient-tailored treatment, and preventing chronic, severe sequelae. For early prediction of childhood asthma, we aimed to define an integrated risk score combining established risk factors with genome-wide molecular markers at birth, complemented by subsequent clinical symptoms/diagnoses (wheezing, atopic dermatitis, food allergy). METHODS: Three longitudinal birth cohorts (PAULINA/PAULCHEN, n = 190 + 93 = 283, PASTURE, n = 1133) were used to predict childhood asthma (age 5-11) including epidemiological characteristics and molecular markers: genotype, DNA methylation and mRNA expression (RNASeq/NanoString). Apparent (ap) and optimism-corrected (oc) performance (AUC/R2) was assessed leveraging evidence from independent studies (Naïve-Bayes approach) combined with high-dimensional logistic regression models (LASSO). RESULTS: Asthma prediction with epidemiological characteristics at birth (maternal asthma, sex, farm environment) yielded an ocAUC = 0.65. Inclusion of molecular markers as predictors resulted in an improvement in apparent prediction performance, however, for optimism-corrected performance only a moderate increase was observed (upto ocAUC = 0.68). The greatest discriminate power was reached by adding the first symptoms/diagnosis (up to ocAUC = 0.76; increase of 0.08, p = .002). Longitudinal analysis of selected mRNA expression in PASTURE (cord blood, 1, 4.5, 6 years) showed that expression at age six had the strongest association with asthma and correlation of genes getting larger over time (r = .59, p < .001, 4.5-6 years). CONCLUSION: Applying epidemiological predictors alone showed moderate predictive abilities. Molecular markers from birth modestly improved prediction. Allergic symptoms/diagnoses enhanced the power of prediction, which is important for clinical practice and for the design of future studies with molecular markers.

Immune modulation by rural exposures and allergy protection.

BACKGROUND: Growing up on traditional farms protects children from the development of asthma and allergies. However, we have identified distinct asthma-protective factors, such as poultry exposure. This study aims to examine the biological effect of rural exposure in China. METHODS: We recruited 67 rural children (7.4 ± 0.9 years) and 79 urban children (6.8 ± 0.6 years). Depending on the personal history of exposure to domestic poultry (DP), rural children were further divided into those with DP exposure (DP+ , n = 30) and those without (DP- , n = 37). Blood samples were collected to assess differential cell counts and expression of immune-related genes. Dust samples were collected from poultry stables inside rural households. In vivo activities of nasal administration of DP dust extracts were tested in an ovalbumin-induced asthma model. RESULTS: There was a stepwise increase in the percentage of eosinophils (%) from rural DP+ children (median = 1.65, IQR = [1.28, 3.75]) to rural DP- children (3.40, [1.70, 6.50]; DP+ vs. DP- , p = .087) and to the highest of their urban counterparts (4.00, [2.00, 7.25]; urban vs. DP+ , p = .017). Similarly, rural children exhibited reduced mRNA expression of immune markers, both at baseline and following lipopolysaccharide (LPS) stimulation. Whereas LPS stimulation induced increased secretion of Th1 and proinflammatory cytokines in rural DP+ children compared to rural DP- children and urban children. Bronchoalveolar lavage of mice with intranasal instillation of dust extracts from DP household showed a significant decrease in eosinophils as compared to those of control mice (p < .05). Furthermore, DP dust strongly inhibited gene expression of Th2 signature cytokines and induced IL-17 expression in the murine asthma model. CONCLUSIONS: Immune responses of rural children were dampened compared to urban children and those exposed to DP had further downregulated immune responsiveness. DP dust extracts ameliorated Th2-driven allergic airway inflammation in mice. Determining active protective components in the rural environment may provide directions for the development of primary prevention of asthma.

The role of regulatory B cells in immune regulation and childhood allergic asthma.

BACKGROUND: As the most common chronic disease in childhood, asthma displays a major public health problem worldwide with the incidence of those affected rising. As there is currently no cure for allergic asthma, it is mandatory to get a better understanding of the underlying molecular mechanism. MAIN BODY: By producing IgE antibodies upon allergen contact, B cells play a pivotal role in allergic asthma. Besides that, IL-10-secreting B cell subsets, namely regulatory B cells (Bregs), are reported in mice and humans to play a role in allergic asthma. In humans, several Breg subsets with distinct phenotypic and functional properties are identified among B cells at different maturational and differentiation stages that exert anti-inflammatory functions by expressing several suppressor molecules. Emerging research has focused on the role of Bregs in allergic asthma as well as their role for future diagnostic and preventive strategies. CONCLUSION: Knowledge about the exact function of human Bregs in allergic asthma is still very limited. This review aims to summarize the current knowledge on Bregs. We discuss different human Breg subsets, several ways of Breg induction as well as the mechanisms through which they exert immunoregulatory functions, and their role in (childhood) allergic asthma.

17q21 Variants Disturb Mucosal Host Defense in Childhood Asthma.

Rationale: The strongest genetic risk factor for childhood-onset asthma, the 17q21 locus, is associated with increased viral susceptibility and disease-promoting processes.Objectives: To identify biological targets underlying the escalated viral susceptibility associated with the clinical phenotype mediated by the 17q21 locus.Methods: Genome-wide transcriptome analysis of nasal brush samples from 261 children (78 healthy, 79 with wheezing at preschool age, 104 asthmatic) within the ALLIANCE (All-Age-Asthma) cohort, with a median age of 10.0 (range, 1.0-20.0) years, was conducted to explore the impact of their 17q21 genotype (SNP rs72163891). Concurrently, nasal secretions from the same patients and visits were collected, and high-sensitivity mesoscale technology was employed to measure IFN protein levels.Measurements and Main Results: This study revealed that the 17q21 risk allele induces a genotype- and asthma/wheeze phenotype-dependent enhancement of mucosal GSDMB expression as the only relevant 17q21-encoded gene in children with preschool wheeze. Increased GSDMB expression correlated with the activation of a type-1 proinflammatory, cell-lytic immune, and natural killer signature, encompassing key genes linked to an IFN type-2-signature (IFNG, CXCL9, CXCL10, KLRC1, CD8A, GZMA). Conversely, there was a reduction in IFN type 1 and type 3 expression signatures at the mRNA and protein levels.Conclusions: This study demonstrates a novel disease-driving mechanism induced by the 17q21 risk allele. Increased mucosal GSDMB expression is associated with a cell-lytic immune response coupled with compromised airway immunocompetence. These findings suggest that GSDMB-related airway cell death and perturbations in the mucosal IFN signature account for the increased vulnerability of 17q21 risk allele carriers to respiratory viral infections during early life, opening new options for future biological interventions.The All-Age-Asthma (ALLIANCE) cohort is registered at www.clinicaltrials.gov (pediatric arm, NCT02496468).

From preschool wheezing to asthma: Immunological determinants.

Asthma represents a chronic respiratory disease affecting millions of children worldwide. The transition from preschool wheezing to school-age asthma involves a multifaceted interplay of various factors, including immunological aspects in early childhood. These factors include complex cellular interactions among different immune cell subsets, induction of pro-inflammatory mediators and the molecular impact of environmental factors like allergens or viral infections on the developing immune system. Furthermore, the activation of specific genes and signalling pathways during this early phase plays a pivotal role in the manifestation of symptoms and subsequent development of asthma. Early identification of the propensity or risk for asthma development, for example by allergen sensitisation and viral infections during this critical period, is crucial for understanding the transition from wheeze to asthma. Favourable immune regulation during a critical 'window of opportunity' in early childhood can induce persistent changes in immune cell behaviour. In this context, trained immunity, including memory function of innate immune cells, has significant implications for understanding immune responses, potentially shaping long-term immunological outcomes based on early-life environmental exposures. Exploration of these underlying immune mechanisms that drive disease progression will provide valuable insights to understand childhood asthma development. This will be instrumental to develop preventive strategies at different stages of disease development for (i) inhibiting progression from wheeze to asthma or (ii) reducing disease severity and (iii) uncovering novel therapeutic strategies and contributing to more tailored and effective treatments for childhood asthma. In the long term, this shall empower healthcare professionals to develop evidence-based interventions that reduce the burden of asthma for children, families and society overall.

Childhood asthma phenotypes and endotypes: a glance into the mosaic.

BACKGROUND: Asthma is an inflammatory lung disease that constitutes the most common noncommunicable chronic disease in childhood. Childhood asthma shows large heterogeneity regarding onset of disease, symptoms, severity, prognosis, and response to therapy. MAIN BODY: Evidence suggests that this variability is due to distinct pathophysiological mechanisms, which has led to an exhaustive research effort to understand and characterize these distinct entities currently designated as "endotypes." Initially, studies focused on identifying specific groups using clinical variables yielding different "clinical phenotypes." In addition, the identification of specific patterns based on inflammatory cell counts and cytokine data has resulted in "inflammatory endotypes." More recently, an increasing number of molecular data from high-throughput technology ("omics" data) have allowed to investigate more complex "molecular endotypes." CONCLUSION: A better definition and comprehension of childhood asthma heterogeneity is key for improving diagnosis and treatment. This review aims at summarizing the current knowledge on this topic and discusses some limitations in their application as well as recommendations for future studies.

Asthma and allergic diseases: Cross talk of immune system and environmental factors.

Air pollution and immune-related diseases including allergy and asthma are constantly on the rise worldwide. Thus, a comprehensive investigation of environmentally induced immune regulation is required for a deeper understanding of disease pathogenesis, progression as well as prevention. Here, we summarize the current knowledge on environmental factors such as microbiome or geographical locations with harmful or protective effects for human health and their different routes of exposure. This review comprises a brief outline regarding the latest findings on the interaction of environmental factors with innate and adaptive regulation of the immune system, exemplarily for one protective and one harmful environmental factor, respectively.

What Have Mechanistic Studies Taught Us About Childhood Asthma?

Childhood asthma is a chronic heterogeneous syndrome consisting of different disease entities or phenotypes. The immunologic and cellular processes that occur during asthma development are still not fully understood but represent distinct endotypes. Mechanistic studies have examined the role of gene expression, protein levels, and cell types in early life development and the manifestation of asthma, many under the influence of environmental stimuli, which can be both protective and risk factors for asthma. Genetic variants can regulate gene expression, controlled partly by different epigenetic mechanisms. In addition, environmental factors, such as living space, nutrition, and smoking, can contribute to these mechanisms. All of these factors produce modifications in gene expression that can alter the development and function of immune and epithelial cells and subsequently different trajectories of childhood asthma. These early changes in a partially immature immune system can have dramatic effects (e.g., causing dysregulation), which in turn contribute to different disease endotypes and may help to explain differential responsiveness to asthma treatment. In this review, we summarize published studies that have aimed to uncover distinct mechanisms in childhood asthma, considering genetics, epigenetics, and environment. Moreover, a discussion of new, powerful tools for single-cell immunologic assays for phenotypic and functional analysis is included, which promise new mechanistic insights into childhood asthma development and therapeutic and preventive strategies.

A compilation answering 50 questions on monkeypox virus and the current monkeypox outbreak.

The current monkeypox disease (MPX) outbreak constitutes a new threat and challenge for our society. With more than 55,000 confirmed cases in 103 countries, World Health Organization declared the ongoing MPX outbreak a Public Health Emergency of International Concern (PHEIC) on July 23, 2022. The current MPX outbreak is the largest, most widespread, and most serious since the diagnosis of the first case of MPX in 1970 in the Democratic Republic of the Congo (DRC), a country where MPX is an endemic disease. Throughout history, there have only been sporadic and self-limiting outbreaks of MPX outside Africa, with a total of 58 cases described from 2003 to 2021. This figure contrasts with the current outbreak of 2022, in which more than 55,000 cases have been confirmed in just 4 months. MPX is, in most cases, self-limiting; however, severe clinical manifestations and complications have been reported. Complications are usually related to the extent of virus exposure and patient health status, generally affecting children, pregnant women, and immunocompromised patients. The expansive nature of the current outbreak leaves many questions that the scientific community should investigate and answer in order to understand this phenomenon better and prevent new threats in the future. In this review, 50 questions regarding monkeypox virus (MPXV) and the current MPX outbreak were answered in order to provide the most updated scientific information and to explore the potential causes and consequences of this new health threat.

Trajectories of cough without a cold in early childhood and associations with atopic diseases.

BACKGROUND: Although children can frequently experience a cough that affects their quality of life, few epidemiological studies have explored cough without a cold during childhood. OBJECTIVES: The objective of the study was to describe the latent class trajectories of cough from one to 10 years old and analyse their association with wheezing, atopy and allergic diseases. METHODS: Questions about cough, wheeze and allergic diseases were asked at 1, 1.5, 2, 3, 4, 5, 6 and 10 years of age in the European prospective cohort of Protection against Allergy: STUdy in Rural Environment (PASTURE). Specific IgE assays were performed at 10 years of age. Questions regarding a cough without a cold were used to build a latent class model of cough over time. RESULTS: Among the 961 children included in the study, apart from the never/infrequent trajectory (59.9%), eight trajectories of cough without a cold were identified: five grouped acute transient classes (24.1%), moderate transient (6.8%), late persistent (4.8%) and early persistent (4.4%). Compared with the never/infrequent trajectory, the other trajectories were significantly associated with wheezing, asthma and allergic rhinitis. For asthma, the strongest association was with the early persistent trajectory (ORa  = 31.00 [14.03-68.51]), which was inversely associated with farm environment (ORa  = 0.39 [0.19-0.77]) and had a high prevalence of cough triggers and unremitting wheeze. Late and early persistent trajectories were also associated with food allergy. Atopic sensitization was only associated with the late persistent trajectory. CONCLUSION: Late and early persistent coughs without a cold are positively associated with atopic respiratory diseases and food allergy. Children having recurrent cough without a cold with night cough and triggers would benefit from an asthma and allergy assessment. Growing up on a farm is associated with reduced early persistent cough.

Parental occupational exposures prior to conception and offspring wheeze and eczema during first year of life.

PURPOSE: Parental exposures prior to conception might influence asthma and allergy risk in offspring. As occupational exposures are established risk factors for asthma and allergies, we investigated if parental occupational exposures prior to conception cause wheeze and eczema in offspring during the first year of life. METHODS: We analysed data of 436 families from an offspring cohort based on a follow-up study of German participants of the International Study of Asthma and Allergies in Childhood (ISAAC). Offspring cohort data was collected between 2009 and 2019. Occupational exposures were based on participants' work histories and measured by a Job-Exposure-Matrix. We used Bayesian logistic regression models for analysis. Inference and confounder selection were based on directed acyclic graphs. RESULTS: In mothers, for both allergic and irritative occupational exposures prior to conception suggestive effects on offspring eczema during the first year of life were found (allergens: odds ratio (OR) 1.22, 95% compatibility interval (CI) 0.92-1.57; irritants: OR 1.36, 95% CI 0.99-1.77), while no relation with wheeze was suggested. CONCLUSIONS: Our results suggest that reduction of asthma-related occupational exposures might not only reduce the burden of disease for occupationally induced or aggravated asthma and allergies in employees but also in their children.

Continuous Rather Than Solely Early Farm Exposure Protects From Hay Fever Development.

BACKGROUND: An important window of opportunity for early-life exposures has been proposed for the development of atopic eczema and asthma. OBJECTIVE: However, it is unknown whether hay fever with a peak incidence around late school age to adolescence is similarly determined very early in life. METHODS: In the Protection against Allergy-Study in Rural Environments (PASTURE) birth cohort potentially relevant exposures such as farm milk consumption and exposure to animal sheds were assessed at multiple time points from infancy to age 10.5 years and classified by repeated measure latent class analyses (n = 769). Fecal samples at ages 2 and 12 months were sequenced by 16S rRNA. Hay fever was defined by parent-reported symptoms and/or physician's diagnosis of hay fever in the last 12 months using questionnaires at 10.5 years. RESULTS: Farm children had half the risk of hay fever at 10.5 years (adjusted odds ratio [aOR] 0.50; 95% CI 0.31-0.79) than that of nonfarm children. Whereas early life events such as gut microbiome richness at 12 months (aOR 0.66; 95% CI 0.46-0.96) and exposure to animal sheds in the first 3 years of life (aOR 0.26; 95% CI 0.06-1.15) were determinants of hay fever, the continuous consumption of farm milk from infancy up to school age was necessary to exert the protective effect (aOR 0.35; 95% CI 0.17-0.72). CONCLUSIONS: While early life events determine the risk of subsequent hay fever, continuous exposure is necessary to achieve protection. These findings argue against the notion that only early life exposures set long-lasting trajectories.

Timing of Blood Sample Processing Affects the Transcriptomic and Epigenomic Profiles in CD4+ T-cells of Atopic Subjects.

Optimal pre-analytical conditions for blood sample processing and isolation of selected cell populations for subsequent transcriptomic and epigenomic studies are required to obtain robust and reproducible results. This pilot study was conducted to investigate the potential effects of timing of CD4+ T-cell processing from peripheral blood of atopic and non-atopic adults on their transcriptomic and epigenetic profiles. Two heparinized blood samples were drawn from each of three atopic and three healthy individuals. For each individual, CD4+ T-cells were isolated from the first blood sample within 2 h (immediate) or from the second blood sample after 24 h storage (delayed). RNA sequencing (RNA-Seq) and histone H3K27 acetylation chromatin immunoprecipitation sequencing (ChIP-Seq) analyses were performed. A multiplicity of genes was shown to be differentially expressed in immediately processed CD4+ T-cells from atopic versus healthy subjects. These differences disappeared when comparing delayed processed cells due to a drastic change in expression levels of atopy-related genes in delayed processed CD4+ T-cells from atopic donors. This finding was further validated on the epigenomic level by examining H3K27 acetylation profiles. In contrast, transcriptomic and epigenomic profiles of blood CD4+ T-cells of healthy donors remained rather unaffected. Taken together, for successful transcriptomics and epigenomics studies, detailed standard operation procedures developed on the basis of samples from both healthy and disease conditions are implicitly recommended.

Rural environment reduces allergic inflammation by modulating the gut microbiota.

Rural environments and microbiota are linked to a reduction in the prevalence of allergies. However, the mechanism underlying the reduced allergies modulated by rural residency is unclear. Here, we assessed gut bacterial composition and metagenomics in urban and rural children in the EuroPrevall-INCO cohort. Airborne dusts, including mattress and rural henhouse dusts, were profiled for bacterial and fungal composition by amplicon sequencing. Mice were repeatedly exposed to intranasal dust extracts and evaluated for their effects on ovalbumin (OVA)-induced allergic airway inflammation, and gut microbiota restoration was validated by fecal microbiota transplant (FMT) from dust-exposed donor mice. We found that rural children had fewer allergies and unique gut microbiota with fewer Bacteroides and more Prevotella. Indoor dusts in rural environments harbored higher endotoxin level and diversity of bacteria and fungi, whereas indoor urban dusts were enriched with Aspergillus and contained elevated pathogenic bacteria. Intranasal administration of rural dusts before OVA sensitization reduced respiratory eosinophils and blood IgE level in mice and also led to a recovery of gut bacterial diversity and Ruminiclostridium in the mouse model. FMT restored the protective effect by reducing OVA-induced lung eosinophils in recipient mice. Together, these results support a cause-effect relationship between exposure to dust microbiota and allergy susceptibility in children and mice. Specifically, rural environmental exposure modulated the gut microbiota, which was essential in reducing allergy in children from Southern China. Our findings support the notion that the modulation of gut microbiota by exposure to rural indoor dust may improve allergy prevention.

In vitro neutrophil migration is associated with inhaled corticosteroid treatment and serum cytokines in pediatric asthma.

Background: Different asthma phenotypes are driven by molecular endotypes. A Th1-high phenotype is linked to severe, therapy-refractory asthma, subclinical infections and neutrophil inflammation. Previously, we found neutrophil granulocytes (NGs) from asthmatics exhibit decreased chemotaxis towards leukotriene B4 (LTB4), a chemoattractant involved in inflammation response. We hypothesized that this pattern is driven by asthma in general and aggravated in a Th1-high phenotype. Methods: NGs from asthmatic nd healthy children were stimulated with 10 nM LTB4/100 nM N-formylmethionine-leucyl-phenylalanine and neutrophil migration was documented following our prior SiMA (simplified migration assay) workflow, capturing morphologic and dynamic parameters from single-cell tracking in the images. Demographic, clinical and serum cytokine data were determined in the ALLIANCE cohort. Results: A reduced chemotactic response towards LTB4 was confirmed in asthmatic donors regardless of inhaled corticosteroid (ICS) treatment. By contrast, only NGs from ICS-treated asthmatic children migrate similarly to controls with the exception of Th1-high donors, whose NGs presented a reduced and less directed migration towards the chemokines. ICS-treated and Th1-high asthmatic donors present an altered surface receptor profile, which partly correlates with migration. Conclusions: Neutrophil migration in vitro may be affected by ICS-therapy or a Th1-high phenotype. This may be explained by alteration of receptor expression and could be used as a tool to monitor asthma treatment.