Background: Chronic rhinitis symptoms cause significant health burden among children and can have a heterogeneous presentation. Defining phenotypes of childhood chronic rhinitis and associated pathobiology may lead to prevention or improved treatments. Objectives: We sought to identify longitudinal patterns of rhinitis symptoms in childhood and determine their associations with early life risk factors, allergic comorbidities, and nasal epithelial cell gene expression. Methods: Chronic rhinitis symptoms were evaluated from ages 1 through 11 years in 485 urban children at high risk for allergic disease in the URECA (Urban Environment and Childhood Asthma) birth cohort. We identified longitudinal rhinitis phenotypes and their relationships to early life exposures, atopic comorbidities, and patterns of nasal epithelial gene expression at age 11 years. Results: Chronic rhinitis symptoms started early in many children and were a risk factor for developing aeroallergen sensitization. We identified 4 longitudinal rhinitis phenotypes: low/minimal, persistent, persistent decreasing, and late increasing. Persistent rhinitis was most closely linked to allergic sensitization and asthma. Risk factors for persistent rhinitis included frequent colds (P < .001), antibiotic use (P < .001), and reduced exposure to common indoor aeroallergens (P 5 .003). Compared to low/minimal rhinitis phenotype, the other rhinitis phenotypes were associated with increased expression of canonical type 2 genes and decreased expression of immune response genes. Conclusions: In urban children, rhinitis symptoms often precede aeroallergen sensitization. Rhinitis phenotypes based on symptoms had distinct risk factors and nasal transcriptome. These results suggest that focusing on early life risk factors and distinct immune mechanisms may be a target to preventing chronic rhinitis in childhood.

BACKGROUND:Five distinct respiratory phenotypes based on latent classes of longitudinal patterns of wheezing, allergic sensitization. and pulmonary function measured in urban children from ages from 0 to 7 years have previously been described. OBJECTIVE:Our aim was to determine whether distinct respiratory phenotypes are associated with early-life upper respiratory microbiota development and environmental microbial exposures. METHODS:Microbiota profiling was performed using 16S ribosomal RNA-based sequencing of nasal samples collected at age 12 months (n = 120) or age 36 months (n = 142) and paired house dust samples collected at 3 months (12-month, n = 73; 36-month, n = 90) from all 4 centers in the Urban Environment and Childhood Asthma (URECA) cohort. RESULTS:In these high-risk urban children, nasal microbiota increased in diversity between ages 12 and 36 months (ß = 2.04; P = .006). Age-related changes in microbiota evenness differed significantly by respiratory phenotypes (interaction P = .0007), increasing most in the transient wheeze group. At age 12 months, respiratory illness (R2 = 0.055; P = .0001) and dominant bacterial genus (R2 = 0.59; P = .0001) explained variance in nasal microbiota composition, and enrichment of Moraxella and Haemophilus members was associated with both transient and high-wheeze respiratory phenotypes. By age 36 months, nasal microbiota was significantly associated with respiratory phenotypes (R2 = 0.019; P = .0376), and Moraxella-dominated microbiota was associated specifically with atopy-associated phenotypes. Analysis of paired house dust and nasal samples indicated that 12 month olds with low wheeze and atopy incidence exhibited the largest number of shared bacterial taxa with their environment. CONCLUSION:Nasal microbiota development over the course of early childhood and composition at age 3 years are associated with longitudinal respiratory phenotypes. These data provide evidence supporting an early-life window of airway microbiota development that is influenced by environmental microbial exposures in infancy and associates with wheeze- and atopy-associated respiratory phenotypes through age 7 years.

Early life gut microbiome composition has been correlated with childhood obesity, though microbial functional contributions to disease origins remain unclear. Here, using an infant birth cohort (n = 349) we identify a distinct fecal microbiota composition in 1-month-old infants with the lowest rate of exclusive breastfeeding, that relates with higher relative risk for obesity and overweight phenotypes at two years. Higher-risk infant fecal microbiomes exhibited accelerated taxonomic and functional maturation and broad-ranging metabolic reprogramming, including reduced concentrations of neuro-endocrine signals. In vitro, exposure of enterocytes to fecal extracts from higher-risk infants led to upregulation of genes associated with obesity and with expansion of nutrient sensing enteroendocrine progenitor cells. Fecal extracts from higher-risk infants also promoted enterocyte barrier dysfunction. These data implicate dysregulation of infant microbiome functional development, and more specifically promotion of enteroendocrine signaling and epithelial barrier impairment in the early-life developmental origins of childhood obesity.

Background Immunoglobulin E-mediated food allergy (IgE-FA) has emerged as a global public health concern. Immune dysregulation is an underlying mechanism for IgE-FA, caused by "dysbiosis" of the early intestinal microbiota. We investigated the association between infant gut bacterial composition and food-related atopy at age 3-5 years using a well-characterized birth cohort. Methods The study definition of IgE-FA to egg, milk, or peanut was based on physician panel retrospective review of clinical and questionnaire data collected from birth through age 3-5 years. Using 16S rRNA sequencing, we profiled the bacterial gut microbiota present in stool specimens collected at 1 and 6 months of age. Results Of 447 infants with data for analysis, 44 (9.8%) met physician panel review criteria for IgE-FA to >= 1 of the three allergens. Among children classified as IgE-FA at 3-5 years, infant stool samples showed significantly less diversity of the gut microbiota compared with the samples of children classified as no IgE-FA at age 3-5 years, especially for milk and peanut (all covariate-adjusted p's for alpha metrics <.007). Testing of individual operational taxonomic units (OTUs) revealed 6-month deficiencies in 31 OTUs for IgE-FA compared with no IgE-FA, mostly in the orders Lactobacillales, Bacteroidales, and Clostridiales. Conclusions Variations in gut microbial composition in infant stool were associated with a study definition of IgE-FA at 3-5 years of age. This included evidence of a lack of bacterial diversity, deficiencies in specific OTUs, and delayed microbial maturation. Results support dysbiosis in IgE-FA pathogenesis.

Maternal asthma status, prenatal exposures, and infant gut microbiota perturbation are associated with heightened risk of atopy and asthma risk in childhood, observations hypothetically linked by intergenerational microbial transmission. Using maternal vaginal (n = 184) and paired infant stool (n = 172) samples, we identify four compositionally and functionally distinct Lactobacillus-dominated vaginal microbiota clusters (VCs) that relate to prenatal maternal health and exposures and infant serum immunoglobulin E (IgE) status at 1 year. Variance in bacteria shared between mother and infant pairs relate to VCs, maternal allergy/asthma status, and infant IgE levels. Heritable bacterial gene pathways associated with infant IgE include fatty acid synthesis and histamine and tryptophan degradation. In vitro, vertically transmitted Lactobacillus jensenii strains induce immunosuppressive phenotypes on human antigen-presenting cells. Murine supplementation with L. jensenii reduces lung eosinophils, neutrophilic expansion, and the proportion of interleukin-4 (IL-4)+ CD4+ T cells. Thus, bacterial and atopy heritability are intimately linked, suggesting a microbial component of intergenerational disease transmission.

The path to childhood asthma is thought to initiate in utero and be further promoted by postnatal exposures. However, the underlying mechanisms remain underexplored. We hypothesized that prenatal maternal immune dysfunction associated with increased childhood asthma risk (revealed by low IFN‐γ:IL‐13 secretion during the third trimester of pregnancy) alters neonatal immune training through epigenetic mechanisms and promotes early‐life airway colonization by asthmagenic microbiota.

Author(s): Rackaityte, E; Halkias, J; Fukui, EM; Mendoza, VF; Hayzelden, C; Crawford, ED; Fujimura, KE; Burt, TD; Lynch, SV

Abstract Objective To identify features of the gut microbiome associated with multiple sclerosis activity over time. Methods We used 16S ribosomal RNA sequencing from stool of 55 recently diagnosed pediatric‐onset multiple sclerosis patients. Microbiome features included the abundance of individual microbes and networks identified from weighted genetic correlation network analyses. Prentice‐Williams‐Peterson Cox proportional hazards models estimated the associations between features and three disease activity outcomes: clinical relapses and both new/enlarging T2 lesions and new gadolinium‐enhancing lesions on brain MRI. Analyses were adjusted for age, sex, and disease‐modifying therapies. Results Participants were followed, on average, 2.1 years. Five microbes were nominally associated with all three disease activity outcomes after multiple testing correction. These included butyrate producers Odoribacter (relapse hazard ratio = 0.46, 95% confidence interval: 0.24, 0.88) and Butyricicoccus (relapse hazard ratio = 0.49, 95% confidence interval: 0.28, 0.88). Two networks of co‐occurring gut microbes were significantly associated with a higher hazard of both MRI outcomes (gadolinium‐enhancing lesion hazard ratios (95% confidence intervals) for Modules 32 and 33 were 1.29 (1.08, 1.54) and 1.42 (1.18, 1.71), respectively; T2 lesion hazard ratios (95% confidence intervals) for Modules 32 and 33 were 1.34 (1.15, 1.56) and 1.41 (1.21, 1.64), respectively). Metagenomic predictions of these networks demonstrated enrichment for amino acid biosynthesis pathways. Interpretation Both individual and networks of gut microbes were associated with longitudinal multiple sclerosis activity. Known functions and metagenomic predictions of these microbes suggest the important role of butyrate and amino acid biosynthesis pathways. This provides strong support for future development of personalized microbiome interventions to modify multiple sclerosis disease activity.

Development of the immune system can be influenced by diverse extrinsic and intrinsic factors that influence the risk of disease. Severe early life respiratory syncytial virus (RSV) infection is associated with persistent immune alterations. Previously, our group had shown that adult mice orally supplemented with Lactobacillus johnsonii exhibited decreased airway immunopathology following RSV infection. Here, we demonstrate that offspring of mice supplemented with L. johnsonii exhibit reduced airway mucus and Th2 cell-mediated response to RSV infection. Maternal supplementation resulted in a consistent gut microbiome in mothers and their offspring. Importantly, supplemented maternal plasma and breastmilk, and offspring plasma, exhibited decreased inflammatory metabolites. Cross-fostering studies showed that prenatal Lactobacillus exposure led to decreased Th2 cytokines and lung inflammation following RSV infection, while postnatal Lactobacillus exposure diminished goblet cell hypertrophy and mucus production in the lung in response to airway infection. These studies demonstrate that Lactobacillus modulation of the maternal microbiome and associated metabolic reprogramming enhance airway protection against RSV in neonates.

Background Mounting evidence suggests both vitamin D and the early life gut microbiome influence childhood health outcomes. However, little is known about how these two important exposures are related. We aimed to examine associations between plasma 25-hydroxyvitamin D (25[OH]D) levels during pregnancy or at delivery (cord blood) and infant gut microbiota. Methods Maternal and cord blood 25[OH]D levels were assessed in a sample of pregnant women. Compositional analyses adjusted for race were run on the gut microbiota of their offspring at 1 and 6 months of age. Results Mean prenatal 25(OH)D level was 25.04 ± 11.62 ng/mL and mean cord blood 25(OH)D level was 10.88 ± 6.77 ng/mL. Increasing prenatal 25(OH)D level was significantly associated with decreased richness ( p = 0.028) and diversity ( p = 0.012) of the gut microbiota at 1 month of age. Both prenatal and cord 25(OH)D were significantly associated with 1 month microbiota composition. A total of 6 operational taxonomic units (OTUs) were significantly associated with prenatal 25(OH)D level (four positively and two negatively) while 11 OTUs were significantly associated with cord 25(OH)D (10 positively and one negatively). Of these, OTU 93 ( Acinetobacter ) and OTU 210 ( Corynebacterium ) , were consistently positively associated with maternal and cord 25(OH)D; OTU 64 ( Ruminococcus gnavus ) was positively associated with prenatal 25(OH)D but negatively associated with cord 25(OH)D. Conclusions Prenatal maternal and cord blood 25(OH)D levels are associated with the early life gut microbiota. Future studies are needed to understand how vitamin D and the microbiome may interact to influence child health.

Mucosal immunity develops in the human fetal intestine by 11–14 weeks gestation, yet whether viable microbes exist in utero and interact with intestinal immunity is unknown. Structures consistent with coccoid bacterial morphology, embedded in fetal meconium were evident before mid-gestation by high-resolution scanning electron microscopy (n=4). Molecular methods indicated extremely low bacterial burden and simple profiles in fetal meconium (n=40 of 50) compared to controls (n=87). A subset of Micrococcaceae-dominated (n=9) meconium associated with proportions of lamina propria PLZF+ CD161+ CD4+ T cells and divergent intestinal epithelial transcriptomes. Fetal Micrococcus luteus was isolated only in the presence of a monocyte feeder cell line. This strain grew on placental hormones, remained viable within fetal antigen presenting cells, exhibited species-specific immunomodulatory capacity and genomic features indicating fetal adaptation. Thus, viable bacteria are highly limited and inconsistently detectable in human fetal meconium at mid-gestation.

We, and others, have shown that the microbial community composition of early infancy stool is associated with the incidence of asthma in pre-school age and young children. Our objective was to determine if there is an association between early life gut microbiota and specifically current atopic asthma at age 10 years. We studied a general-risk ethnically diverse birth cohort (WHEALS) from metropolitan Detroit (Michigan, USA). Stool samples were collected at approximately ages 1 and 6 months and evaluated using bacterial (16S) sequencing. During a clinical follow-up evaluation at age 10 years, a board-certified allergist reviewed the subject’s history of any breathing difficulties, performed an examination, evaluated skin tests and spirometry and classified the child as having current asthma (yes/no). Asthmatic children were defined as having atopic asthma if they had concurrent positive results to ≥2 of 11 tested allergen-specific IgEs. Generalized estimating equations (GEE) were used to test whether differences in alpha diversity metrics over time were related to current atopic asthma at age 10. Three hundred sixty-three (363) children are included in analyses: 51 with allergic asthma at age 10, and 312 without. One hundred thirty-one (131) children had a 1-month sample only, 91 had a 6-month sample only, and 141 had both (i.e., 504 total bacterial samples: 272 1-month and 232 6-month). Although at both 1 and 6 months separately there were no significant differences in alpha diversity by allergic asthma at age 10 (p=0.22, p=0.39, respectively), the slope of the diversity trajectory was lower in children with allergic asthma at age 10 versus children without (β=0.44 versus 0.65, respectively, with an interaction p value = 0.08). This pattern was found only in females. Children with current allergic asthma at age 10 appeared to accumulate bacterial diversity in early life at a slower rate than children without allergic asthma at age 10; however, this effect was found in females only. These results emphasize the importance of appropriate microbial development in early life, and suggests that both accelerated microbial development at 1-month as well as delayed microbial development at 6-months may increase the risk of allergic asthma at age 10 years.

The impact of the maternal prenatal immune profile on the neonatal epigenome and the trajectory to asthma during childhood remains uncharacterized. Epigenome-wide DNA methylation at 850,000 CpG sites was profiled in 182 children from the Infant Immune Study with information for childhood asthma (2-9 years) and maternal immune profile (IFNg/IL-13 ratio produced by mitogen-stimulated maternal peripheral blood immune cells isolated during the third trimester of pregnancy), which is associated with childhood asthma. Differentially methylated CpG sites (DMCs) associated with differences in maternal immune profile [log10-transformed values or lowest vs. highest ratio quartiles] were identified by linear regression (FDR<0.05). Differences in the maternal prenatal IFNγ/IL-13 ratio were significantly associated with differential methylation at 191 neonatal DMCs, whereas 986 DMCs were identified comparing the lowest and highest quartiles of maternal prenatal IFNγ/IL-13 ratio. Among these, 181 were also differentially methylated in children who did or did not develop asthma by age 9 years. Neonatal methylation at ZFYVE9, a strong maternal IFNγ/IL-13 ratio-associated DMC in a TGFB pathway gene, was also associated with the child's immune profile (IFNγ/IL-13 ratio) at age 3-months. Our data provide evidence for an epigenetic trajectory to asthma in which the neonatal methylome is influenced by the maternal prenatal immune profile and in turn influences the inception of childhood asthma. Epigenetic modifications of the TGFB pathway at birth may provide a functional link between the maternal immune profile during pregnancy and the development of the child's immune response in early life.

As microbial therapeutics are increasingly being tested in diverse patient populations, it is essential to understand the host and environmental factors influencing the microbiome. Through analysis of 1,359 gut microbiome samples from 946 healthy donors of the Milieu Intérieur cohort, we detail how microbiome composition is associated with host factors, lifestyle parameters, and disease states. Using a genome-based taxonomy, we found biological sex was the strongest driver of community composition. Additionally, bacterial populations shift across decades of life (age 20–69), with Bacteroidota species consistently increased with age while Actinobacteriota species, including Bifidobacterium, decreased. Longitudinal sampling revealed that short-term stability exceeds interindividual differences. By accounting for these factors, we defined global shifts in the microbiomes of patients with non-gastrointestinal tumors compared with healthy donors. Together, these results demonstrated that the microbiome displays predictable variations as a function of sex, age, and disease state. These variations must be considered when designing microbiome-targeted therapies or interpreting differences thought to be linked to pathophysiology or therapeutic response.

The development of immune responses in infants is central to establishing a balanced system that reacts appropriately to infectious stimuli but does not induce altered disease states with potential long term sequelae. Studies have linked severe RSV infection during early-life with an enhanced likelihood of developing childhood asthma. We have previously shown that supplementing adult mice with a probiotic species identified in allergen-protected animals, L. johnsonii, can attenuate pathologic responses in animals with viral and allergen induced disease. In these studies, we supplemented female mice with L. johnsonii by oral gavage prior to breeding and during pregnancy. The neonatal mice were infected with RSV at day 6 of age after 8 days of infection (day 14) some were evaluated. Subsequently, some of the neonatal miss were sensitized with allergen into the airway 4 or 12 weeks later and examined after a final challenge at 3 weeks post-sensitization. Our studies show that early-life RSV infection leads to long-term effects leaving mice prone to exacerbation upon secondary allergen exposure at 4 to 12 weeks post-infection. Offspring from the L. johnsonii supplemented mice were protected from an early life RSV infection and from the subsequent allergen-induced exacerbated responses. These early life protective responses reduced Th2 responses and mucus and were associated with an altered neonatal microbiome. Using cross-fostering studies we were able to demonstrate both in utero and post natal protective effects on the offspring that were dependent upon the supplementation of the mother with L. johnsonii. Assessment of the plasma and milk metabolic profiles identified an altered metabolite phenotype in the different cross-fostered neonates that may be useful in assessing allergen susceptible and/or protected early immune environments during early life. Thus, we have identified a maternal microbiome associated metabolomic effect associated with protective in offspring that is related to an altered immune phenotype in the lung during RSV infection and subsequent allergic asthma responses. These results identify that maternal environments on offspring have both in utero and post-natal effects and may help guide future clinical research toward evaluating the maternal impact on development of the infant's immune/allergic responses.

Analysis of 1,363 deeply sequenced gut microbiome samples from 946 healthy donors of the Milieu Intérieur cohort provides new opportunities to discover how the gut microbiome is associated with host factors and lifestyle parameters. Using a genome-based taxonomy to achieve higher resolution analysis, we found an enrichment of Prevotella species in males, and that bacterial profiles are dynamic across five decades of life (20-69), with Bacteroidota species consistently increased with age while Actinobacteriota species, including Bifidobacterium , decreased. Longitudinal sampling revealed short-term stability exceeds inter-individual differences; however, the degree of stability was variable between donors and influenced by their baseline community composition. We then integrated the microbiome results with systemic immunophenotypes to show that host/microbe associations discovered in animal models, such as T regulatory cells and short chain fatty acids, could be validated in human data. These results will enable personalized medicine approaches for microbial therapeutics and biomarkers.

Mucosal immunity influences host-microbial interactions and is evident in the human fetal intestine by 11–14 weeks of gestation; the developing intestine is populated by lymphoid aggregates, memory T cells and dendritic cells capable of responding to microbial stimuli. However, whether intestinal encounters with viable microbes occur in utero and shape immune maturation has not been investigated. Here, we identified subsets of fetal meconium relatively enriched in Lactobacillus or Micrococcaceae, using culture-independent methods, which related to divergent epithelial cell layer transcriptomes and proportions of lamina propria PLZF+ CD161+ CD4+ T cells in paired intestinal samples. Mimicking conditions in the fetal intestine permitted isolation of viable Lactobacillus and Micrococcus strains from fetal meconium. In contrast with phylogenetically related reference strains, fetal isolates utilized placental hormones for growth, remained viable within antigen presenting cells, and exhibited species-specific capacity to promote immune homeostasis. Fetal Lactobacillus isolates reduced activation of antigen presenting cells, while the Micrococcus isolate induced IL-10 production in antigen presenting cells and inhibited IFNγ production by memory PLZF+T cells. Fetal isolates were identified as strains of Lactobacillus jensenii or Micrococcus luteus by whole genome sequencing, with the latter encoding strain-specific genes for intracellular survival, immune modulation, and steroid uptake. These data suggest that pre-natal immunity is influenced by bacterial exposure in the fetal intestine, identifying a previously unknown component of human mucosal immune development.

Microbial dysbiosis commonly occurs in patients with inflammatory bowel diseases (IBD). Exogenous causes of dysbiosis such as antibiotics and diet are well described, but host derived causes are understudied. A20 is a potent regulator of signals triggered by microbial pattern molecules, and A20 regulates susceptibility to intestinal inflammation in mice and in humans. We now report that mice lacking A20 expression in dendritic cells, A20FL/FL CD11c-Cre mice (or A20dDC mice), spontaneously develop colitogenic intestinal dysbiosis that is evident upon weaning and precedes the onset of colitis. Intestines from A20dDC mice express increased amounts of Reg3β and Reg3γ, but not Ang4. A20 deficient DCs promote gut microbiota perturbation in the absence of adaptive lymphocytes. Moreover, A20 deficient DCs directly induce expression of Reg3β and Reg3γ but not Ang 4 in normal intestinal epithelial cell enteroid cultures in the absence of other cell types. These findings reveal a pathophysiological pathway in which defective expression of an IBD susceptibility gene in DCs drives aberrant expression of anti-bacterial peptides and luminal dysbiosis that in turn confers host susceptibility to intestinal inflammation.