The glucose transporter 2 regulates CD8+ T cell function via environment sensing.

T cell activation is associated with a profound and rapid metabolic response to meet increased energy demands for cell division, differentiation and development of effector function. Glucose uptake and engagement of the glycolytic pathway are major checkpoints for this event. Here we show that the low-affinity, concentration-dependent glucose transporter 2 (Glut2) regulates the development of CD8+ T cell effector responses in mice by promoting glucose uptake, glycolysis and glucose storage. Expression of Glut2 is modulated by environmental factors including glucose and oxygen availability and extracellular acidification. Glut2 is highly expressed by circulating, recently primed T cells, allowing efficient glucose uptake and storage. In glucose-deprived inflammatory environments, Glut2 becomes downregulated, thus preventing passive loss of intracellular glucose. Mechanistically, Glut2 expression is regulated by a combination of molecular interactions involving hypoxia-inducible factor-1 alpha, galectin-9 and stomatin. Finally, we show that human T cells also rely on this glucose transporter, thus providing a potential target for therapeutic immunomodulation.

Loss of mTORC2-induced metabolic reprogramming in monocytes uncouples migration and maturation from production of proinflammatory mediators.

Monocyte migration to the sites of inflammation and maturation into macrophages are key steps for their immune effector function. Here, we show that mechanistic target of rapamycin complex 2 (mTORC2)-dependent Akt activation is instrumental for metabolic reprogramming at the early stages of macrophage-mediated immunity. Despite an increased production of proinflammatory mediators, monocytes lacking expression of the mTORC2 component Rictor fail to efficiently migrate to inflammatory sites and fully mature into macrophages, resulting in reduced inflammatory responses in vivo. The mTORC2-dependent phosphorylation of Akt is instrumental for the enhancement of glycolysis and mitochondrial respiration, required to sustain monocyte maturation and motility. These observations are discussed in the context of therapeutic strategies aimed at selective inhibition of mTORC2 activity.

Comparative epigenetic analysis of tumour initiating cells and syngeneic EPSC-derived neural stem cells in glioblastoma.

Epigenetic mechanisms which play an essential role in normal developmental processes, such as self-renewal and fate specification of neural stem cells (NSC) are also responsible for some of the changes in the glioblastoma (GBM) genome. Here we develop a strategy to compare the epigenetic and transcriptional make-up of primary GBM cells (GIC) with patient-matched expanded potential stem cell (EPSC)-derived NSC (iNSC). Using a comparative analysis of the transcriptome of syngeneic GIC/iNSC pairs, we identify a glycosaminoglycan (GAG)-mediated mechanism of recruitment of regulatory T cells (Tregs) in GBM. Integrated analysis of the transcriptome and DNA methylome of GBM cells identifies druggable target genes and patient-specific prediction of drug response in primary GIC cultures, which is validated in 3D and in vivo models. Taken together, we provide a proof of principle that this experimental pipeline has the potential to identify patient-specific disease mechanisms and druggable targets in GBM.

Lung Marginated and Splenic Murine Resident Neutrophils Constitute Pioneers in Tissue-Defense During Systemic E. coli Challenge.

The rapid response of neutrophils throughout the body to a systemic challenge is a critical first step in resolution of bacterial infection such as Escherichia coli (E. coli). Here we delineated the dynamics of this response, revealing novel insights into the molecular mechanisms using lung and spleen intravital microscopy and 3D ex vivo culture of living precision cut splenic slices in combination with fluorescent labelling of endogenous leukocytes. Within seconds after challenge, intravascular marginated neutrophils and lung endothelial cells (ECs) work cooperatively to capture pathogens. Neutrophils retained on lung ECs slow their velocity and aggregate in clusters that enlarge as circulating neutrophils carrying E. coli stop within the microvasculature. The absolute number of splenic neutrophils does not change following challenge; however, neutrophils increase their velocity, migrate to the marginal zone (MZ) and form clusters. Irrespective of their location all neutrophils capturing heat-inactivated E. coli take on an activated phenotype showing increasing surface CD11b. At a molecular level we show that neutralization of ICAM-1 results in splenic neutrophil redistribution to the MZ under homeostasis. Following challenge, splenic levels of CXCL12 and ICAM-1 are reduced allowing neutrophils to migrate to the MZ in a CD29-integrin dependent manner, where the enlargement of splenic neutrophil clusters is CXCR2-CXCL2 dependent. We show directly molecular mechanisms that allow tissue resident neutrophils to provide the first lines of antimicrobial defense by capturing circulating E. coli and forming clusters both in the microvessels of the lung and in the parenchyma of the spleen.

Metabolic regulation of T lymphocyte motility and migration.

In order to fulfill their effector and patrolling functions, lymphocytes traffic through the body and need to adapt to different tissue microenvironments. First, mature lymphocytes egress the bone marrow and the thymus into the vascular system. Circulating lymphocytes can exit the vasculature and penetrate into the tissues, either for patrolling in search for pathogens or to eliminate infection and activate the adaptive immune response. The cytoskeletal reorganization necessary to sustain migration require high levels of energy thus presenting a substantial bioenergetic challenge to migrating cells. The metabolic regulation of lymphocyte motility and trafficking has only recently begun to be investigated. In this review we will summarize current knowledge of the crosstalk between cell metabolism and the cytoskeleton in T lymphocytes, and discuss the concept that lymphocyte metabolism may reprogram in response to migratory stimuli and adapt to the different environmental cues received during recirculation in tissues.

TLR4 Polymorphism, Nasopharyngeal Bacterial Colonization, and the Development of Childhood Asthma: A Prospective Birth-Cohort Study in Finnish Children.

We aimed to explore the role of TLR4 (rs4986790) polymorphism in the nasopharyngeal (NP) bacterial colonization and its consequent impact on the development of childhood asthma. A semi-quantitative culture of NP swabs was performed on 473 children at 2 months of age and on 213 children at 13 months of age. TLR4 polymorphism was analyzed for 396 children. Children were followed from birth to the age of 7.5 years and the final outcome was physician-diagnosed asthma. The associations between TLR4 genotype, bacterial colonization, and asthma were analyzed. Children with TLR4 AG or GG genotype were more often colonized with Moraxella catarrhalis at 2 months of age (p = 0.009) and Haemophilus influenzae at 13 months of age (p = 0.018). Children who were colonized with H. influenzae at 13 months of age had a significantly higher risk of later development of asthma (p = 0.004). M. catarrhalis or H. Influenzae colonization at 2 months of age or TLR4 genotype Asp299Gly were not associated with the development of childhood asthma. TLR4 Asp299Gly polymorphism was associated with an increased risk of colonization of M. catarrhalis and H. influenzae in children. The colonization with H. influenzae at 13 months of age was associated with a higher risk of later development of childhood asthma.

Preservation of microvascular barrier function requires CD31 receptor-induced metabolic reprogramming.

Endothelial barrier (EB) breaching is a frequent event during inflammation, and it is followed by the rapid recovery of microvascular integrity. The molecular mechanisms of EB recovery are poorly understood. Triggering of MHC molecules by migrating T-cells is a minimal signal capable of inducing endothelial contraction and transient microvascular leakage. Using this model, we show that EB recovery requires a CD31 receptor-induced, robust glycolytic response sustaining junction re-annealing. Mechanistically, this response involves src-homology phosphatase activation leading to Akt-mediated nuclear exclusion of FoxO1 and concomitant ß-catenin translocation to the nucleus, collectively leading to cMyc transcription. CD31 signals also sustain mitochondrial respiration, however this pathway does not contribute to junction remodeling. We further show that pathologic microvascular leakage in CD31-deficient mice can be corrected by enhancing the glycolytic flux via pharmacological Akt or AMPK activation, thus providing a molecular platform for the therapeutic control of EB response.

Neutrophils restrain allergic airway inflammation by limiting ILC2 function and monocyte-dendritic cell antigen presentation.

Neutrophil mobilization, recruitment, and clearance must be tightly regulated as overexuberant neutrophilic inflammation is implicated in the pathology of chronic diseases, including asthma. Efforts to target neutrophils therapeutically have failed to consider their pleiotropic functions and the implications of disrupting fundamental regulatory pathways that govern their turnover during homeostasis and inflammation. Using the house dust mite (HDM) model of allergic airway disease, we demonstrate that neutrophil depletion unexpectedly resulted in exacerbated T helper 2 (TH2) inflammation, epithelial remodeling, and airway resistance. Mechanistically, this was attributable to a marked increase in systemic granulocyte colony-stimulating factor (G-CSF) concentrations, which are ordinarily negatively regulated in the periphery by transmigrated lung neutrophils. Intriguingly, we found that increased G-CSF augmented allergic sensitization in HDM-exposed animals by directly acting on airway type 2 innate lymphoid cells (ILC2s) to elicit cytokine production. Moreover, increased systemic G-CSF promoted expansion of bone marrow monocyte progenitor populations, which resulted in enhanced antigen presentation by an augmented peripheral monocyte-derived dendritic cell pool. By modeling the effects of neutrophil depletion, our studies have uncovered previously unappreciated roles for G-CSF in modulating ILC2 function and antigen presentation. More broadly, they highlight an unexpected regulatory role for neutrophils in limiting TH2 allergic airway inflammation.

Pulmonary environmental cues drive group 2 innate lymphoid cell dynamics in mice and humans.

Group 2 innate lymphoid cells (ILC2s) are enriched in mucosal tissues (e.g., lung) and respond to epithelial cell-derived cytokines initiating type 2 inflammation. During inflammation, ILC2 numbers are increased in the lung. However, the mechanisms controlling ILC2 trafficking and motility within inflamed lungs remain unclear and are crucial for understanding ILC2 function in pulmonary immunity. Using several approaches, including lung intravital microscopy, we demonstrate that pulmonary ILC2s are highly dynamic, exhibit amoeboid-like movement, and aggregate in the lung peribronchial and perivascular spaces. They express distinct chemokine receptors, including CCR8, and actively home to CCL8 deposits located around the airway epithelium. Within lung tissue, ILC2s were particularly motile in extracellular matrix-enriched regions. We show that collagen-I drives ILC2 to markedly change their morphology by remodeling their actin cytoskeleton to promote environmental exploration critical for regulating eosinophilic inflammation. Our study provides previously unappreciated insights into ILC2 migratory patterns during inflammation and highlights the importance of environmental guidance cues in the lung in controlling ILC2 dynamics.

Interleukin-10 gene polymorphism rs1800896 is associated with post-bronchiolitis asthma at 11-13 years of age.

AIM: Interleukin-10 (IL-10) is an anti-inflammatory cytokine that is involved with bronchiolitis and asthma. We evaluated associations between four IL-10 polymorphisms, namely rs1800871, rs1800872, rs1800890 and rs1800896, and post-bronchiolitis asthma in young adolescents. METHODS: The cohort consisted of 125 children hospitalised for bronchiolitis at Tampere University Hospital, Finland, in 2000-2004, at less than six months of age. At 11-13 years, asthma diagnoses and asthma-presumptive symptoms, allergic rhinitis and use of inhaled corticosteroids (ICS) were registered. Data on the four polymorphisms and their genotypes, haplotypes and allele frequencies were analysed in relation to asthma, allergic rhinitis and asthma medication. RESULTS: The variant IL-10 rs1800896 genotype was associated with less persistent asthma at five to seven and 11-13 years of age (4.3 versus 15.2%, p = 0.04) than the wild genotype and less ICS use during the previous 12 months (5.4 versus 18.2%, p = 0.03), as was the variant allele G. Allele A was associated with more persistent asthma and ICS use. The significant differences between the variant and wild genotypes were lost in adjusted logistic regression, but the direction of the association remained. CONCLUSION: IL-10 rs1800896 gene polymorphism was associated with post-bronchiolitis asthma at 11-13 years of age in children hospitalised for bronchiolitis at less than six months of age.

Polymorphisms in the promoter region of IL10 gene are associated with virus etiology of infant bronchiolitis.

BACKGROUND: Bronchiolitis is the most common infection leading to hospitalization in infancy. Interleukin-10 (IL-10) is an anti-inflammatory cytokine, and in our previous study, IL10 gene rs1800896 (- 1082A/G) polymorphism was associated with viral etiology of infant bronchiolitis. The objective of this study was to evaluate the associations between IL10 single nucleotide polymorphisms (SNPs) at rs1800890 (- 3575A/T), rs1800871 (- 819C/T) or rs1800872 (- 592C/A) either alone or combined with the SNP at rs1800896 (- 1082G/A), and the etiology and severity of infant bronchiolitis. METHODS: Data on four IL10 SNPs were available from 135 full-term infants, hospitalized for bronchiolitis at age less than 6 months, and from 378 to 400 controls. Viral etiology was studied, and oxygen support, feeding support and the length of stay in hospital were recorded during bronchiolitis hospitalization. RESULTS: Infants with rhinovirus bronchiolitis had the IL10 rs1800890 variant AT or TT genotype less often (18.2%) than controls (63.3%, P = 0.03), and likewise, had the IL10 rs1800896 variant AG or GG genotype less often (27.3%) than controls (65.5%, P = 0.009). Twenty-eight infants with bronchiolitis had the variant-variant Grs1800896Trs1800890 haplotype, and none of them had rhinovirus infection. The IL10 rs1800871 or rs1800872 genotypes showed no associations with viruses. No association was found between any genotypes and bronchiolitis severity measures. CONCLUSION: IL10 rs1800890 and rs1800896 polymorphisms differed between infants with rhinovirus bronchiolitis and controls, but not between infants with respiratory syncytial virus bronchiolitis and controls.

Toll-like receptor 1 and 10 gene polymorphisms are linked to postbronchiolitis asthma in adolescence.

AIM: Toll-like receptors (TLR) are innate immunity molecules and our previous studies found that TLR1 gene polymorphism was associated with postbronchiolitis asthma at one to six years of age, as was TLR10 at five to seven years of age. This study examined any associations at 11-13 years of age. METHODS: This prospective follow-up study was part of an ongoing evaluation of children admitted to Tampere University Hospital, Finland, for bronchiolitis in 2001-2004 at less than six months of age. We evaluated the association of TLR1 rs5743618 and TLR10 rs4129009 polymorphisms with asthma and asthma medication in 125 children aged 11-13 years. RESULTS: Associations were measured as adjusted odd ratios (aOR) with 95% confidence intervals (95% CI). The variant TLR1 rs5743618 (aOR 4.04, 95% CI 0.99-13.01) and TLR10 rs4129009 (aOR 7.02, 95% CI 1.56-31.53) genotypes increased the risk of needing inhaled corticosteroids (ICSs) at 11-13 years of age. The variant TLR10 genotype (aOR 7.69, 95% CI 1.35-43.95) increased the risk of persistent asthma continuing from five to seven years of age until 11-13 years of age. The results were similar when the combined genotypes were analysed. [Correction added on 3 October 2017, after online publication: The data in the variant TRL1 rs5743618 genotype were incorrect and have been corrected in this version.] CONCLUSION: Polymorphisms in both the TLR1 and TLR10 genes may increase the risk of asthma at 11-13 years after infant bronchiolitis.

Haplotype of the Interleukin 17A gene is associated with osteitis after Bacillus Calmette-Guerin vaccination.

Bacillus Calmette-Guerin (BCG) osteitis was more common in Finland than elsewhere at the time when universal BCG vaccinations were given to Finnish newborns. There is evidence that IL-17 plays a role in the defense against tuberculosis. The aim of this study was to evaluate the associations of IL17A rs4711998, IL17A rs8193036 and IL17A rs2275913 single-nucleotide polymorphisms (SNPs) with the risk of BCG osteitis after newborn vaccination. IL17A rs4711998, rs8193036 and rs2275913 SNPs were determined in 131 adults had presented with BCG osteitis after newborn BCG vaccination. We analyzed, using the HaploView and PLINK programs, whether allele or haplotype frequencies of these SNPs differ between the former BCG osteitis patients and Finnish population controls. Of the three IL17A SNPs studied, rs4711998 associated nominally with BCG osteitis; minor allele frequency was 0.215 in 130 BCG osteitis cases and 0.298 in 99 controls (p = 0.034). Frequency of the second common haplotype (GTA) differed significantly between BCG osteitis cases and controls (0.296 vs. 0.184, p = 0.040 after multi-testing correction). The GTA haplotype of the IL17A SNPs rs4711998, rs8193036 and rs2275913 was associated with osteitis after BCG vaccination.

Interleukin 17A gene polymorphism rs2275913 is associated with osteitis after the Bacillus Calmette-Guérin vaccination.

AIM: Interleukin-17 (IL-17) appears to promote the host's defence against mycobacterial infections. This study evaluated the association between IL17A gene polymorphism and the risk of Bacillus Calmette-Guérin (BCG) osteitis after newborn vaccination and between IL17A gene polymorphism and IL-17A concentrations in serum. METHODS: IL17A rs2275913 gene polymorphisms and serum IL-17A concentrations were studied in 132 adults aged 21-49 years from across Finland, who had BCG osteitis in infancy after a newborn BCG vaccination. The subjects were recruited in 2007-2008, and their whole-blood samples were sent to the National Institute for Health and Welfare, Turku, Finland. Their genotypes and minor allele frequencies were compared with 405 population-based unvaccinated controls aged two to three months from a prospective birth cohort study. RESULTS: The genotypes and allele frequencies of IL17A rs2275913 differed significantly between the former BCG osteitis patients and controls. The genotype was variant in 75.8% of cases and 64.0% of controls (p = 0.012), and the minor allele frequency was 50.0% in the cases and 41.6% of the controls (p = 0.009). Serum IL-17 concentrations did not differ significantly between the cases with wild or variant genotypes. CONCLUSION: IL17A rs2275913 gene polymorphism was associated with a risk of BCG osteitis after vaccination.

Polymorphism in the gene encoding toll-like receptor 10 may be associated with asthma after bronchiolitis.

Toll-like receptors (TLRs) recognise microbes that contribute to the severity of bronchiolitis and the subsequent risk of asthma. We evaluated whether post-bronchiolitis asthma was associated with polymorphisms in the TLR3 rs3775291, TLR4 rs4986790, TLR7 rs179008, TLR8 rs2407992, TLR9 rs187084, and TLR10 rs4129009 genes. The gene polymorphisms were studied at the age of 6.4 years (mean) in 135 children hospitalised for bronchiolitis in infancy. The outcome measure was current or previous asthma. Current asthma was more common (30%) in children with the variant AG or GG genotype in the TLR10 rs4129009 gene versus those who were homozygous for the major allele A (11%) (p = 0.03). The adjusted odds ratio (aOR) was 4.30 (95% CI 1.30-14.29). Asthma ever was more common (34.6%) in girls with the TLR7 variant AT or TT genotype versus those who were homozygous for the major allele A (12.5%) (p = 0.03). The adjusted OR was 3.93 (95% CI 1.06-14.58). Corresponding associations were not seen in boys. There were no significant associations between TLR3, TLR4, TLR8, or TLR9 polymorphisms and post-bronchiolitis asthma. Polymorphism in the TLR10 gene increases and in the TLR7 gene may increase the risk of asthma in preschool-aged children after infant bronchiolitis.

Polymorphisms of Mannose-binding Lectin and Toll-like Receptors 2, 3, 4, 7 and 8 and the Risk of Respiratory Infections and Acute Otitis Media in Children.

BACKGROUND: Mannose-binding lectin (MBL) and toll-like receptors (TLRs) are important components of the innate immune system. We assessed the susceptibility of children with genetic variants in these factors to respiratory infections, rhinovirus infections and acute otitis media. METHODS: In a prospective cohort study, blood samples from 381 Finnish children were analyzed for polymorphisms in MBL2 at codons 52, 54 and 57, TLR2 Arg753Gln, TLR3 Leu412Phe, TLR4 Asp299Gly, TLR7 Gln11Leu and TLR8 Leu651Leu. Children were followed up for respiratory infections until 24 months of age with daily diaries. Polymerase chain reaction and antigen tests were used for detection of respiratory viruses from nasal swabs. RESULTS: Children with MBL variant genotype had a mean of 59 days with symptoms of respiratory infection per year, compared with 49 days in those with wild-type (P = 0.01). TLR8 polymorphisms were associated with an increased risk and TLR7 polymorphisms with a decreased risk of recurrent rhinovirus infections (P = 0.02 for both). TLR2 polymorphisms were associated with recurrent acute otitis media (P = 0.02). MBL polymorphisms were associated with an increased and TLR7 polymorphisms with a decreased risk of rhinovirus-associated acute otitis media (P = 0.03 and P = 0.006, respectively). CONCLUSIONS: Genetic polymorphisms in MBL and TLRs promote susceptibility to or protection against respiratory infections. In addition to environmental factors, genetic variations may explain why some children are more prone to respiratory infections.

IL-10 gene polymorphism is associated with preschool atopy and early-life recurrent wheezing after bronchiolitis in infancy.

BACKGROUND: Variations in the genes that regulate innate immunity responses may be associated with susceptibility to asthma or atopy after early-life bronchiolitis. The aim of this study was to evaluate the association between four different polymorphisms of the IL-10 gene at rs1800871, rs1800872, rs1800890, and rs1800896, either alone or in combination, and post-bronchiolitis asthma or allergies at 5-7 years of age. METHODS: Data on single nucleotide polymorphisms (SNP) of IL-10 rs1800896 (-1082G/A), rs1800871 (-819C/T), rs1800872 (-592C/A), and IL-10 rs1800890 (-3575T/A) were available for 135 children. Polymorphisms and their associations with asthma and allergies were studied in 135 preschool-aged children who had been hospitalized for bronchiolitis at age 0-6 months. Their parents were interviewed to record the children's history with asthma and allergies from infancy to the present. RESULTS: At 6.4 years (mean), asthma was present in 17 children (12.6%), while recurrent wheezing during the first 7 years of life was present in 39 (28.9%) children. Fifty-three (39.3%) study participants had current atopy (atopic eczema or allergic rhinitis). Eight (72%) of 11 children with the IL-10 rs1800896, IL-10 rs1800871, and IL-10 rs1800872 combination AA + CT + CA had current atopy (P = 0.02 vs. 38% in other genotype combinations). Twenty-three (56%) children with the IL-10 rs1800871C/T or IL-10 rs1800872C/A genotype had present atopy versus 34 (38%) with other IL-10 genotypes (P = 0.03). Between 2 years and 3 years of age, 27% of ATA haplotype carriers had asthma versus 13.7% of other haplotype carriers (P = 0.02). CONCLUSIONS: IL-10 polymorphisms at rs1800871, rs1800872, rs1800890, and rs1800896 seem to be associated with elevated allergies and/or recurrent wheezing risk in later childhood, after early-life bronchiolitis. Pediatr Pulmonol. 2017;52:14-20. © 2016 Wiley Periodicals, Inc.

Post-bronchiolitis wheezing is associated with toll-like receptor 9 rs187084 gene polymorphism.

Innate immunity receptors play a critical role in host defence, as well as in allergy and asthma. The aim of this exploratory study was to evaluate whether there are associations between TLR7 rs179008, TLR8 rs2407992, TLR9 rs187084 or TLR10 rs4129009 polymorphisms and viral findings, clinical characteristics or subsequent wheezing in infants with bronchiolitis. In all, 135 full-term infants were hospitalized for bronchiolitis at age less than 6 months: 129 of them were followed-up until the age of 1.5 years. The outcome measures were repeated wheezing, use of inhaled corticosteroids, atopic dermatitis during the first 1.5 years of life and total serum immunoglobulin E (IgE). There were no significant associations between the genotypes or allele frequencies of TLR7 rs179008, TLR8 rs2407992, TLR9 rs187084 or TLR10 rs4129009 polymorphisms and clinical characteristics or the severity of bronchiolitis during hospitalization. During follow-up, repeated wheezing was more common in children with TLR9 rs187084 variant genotype CC (30.5%) than in children with TLR9 wild-type genotype TT (12.2%) (p = 0.02, aOR 2.73, 95% CI 1.02-7.29). The TLR10 rs4129009 minor allele G was associated with elevated total serum IgE. TLR9 rs187084 gene polymorphism may be associated with post-bronchiolitis wheezing, and TLR10 rs4129009 gene polymorphism may be associated with atopy.

Burden of Recurrent Respiratory Tract Infections in Children: A Prospective Cohort Study.

BACKGROUND: The burden of recurrent respiratory infections is unclear. We identified young children with recurrent respiratory infections in order to characterize the clinical manifestations, risk factors and short-term consequences. METHODS: In this prospective cohort study, 1089 children were followed from birth to 2 years of age for respiratory infections by a daily symptom diary. Nasal swabs taken during respiratory infections were analyzed for viruses from 714 children. Nasopharyngeal swabs collected at 2 months of age were cultured for bacteria. The 10% of children with the highest number of annual respiratory illness days were defined to have recurrent respiratory tract infections. RESULTS: The 90th percentile in the number of annual respiratory illness days was 98. Children above this limit (n = 109) had a median of 9.6 acute respiratory infections per year. Rhinovirus was detected in 58% of their infections. Of the children with recurrent infections, 60% were diagnosed with at least 3 episodes of acute otitis media, 73% received at least 3 antibiotic treatments and 21% were hospitalized for an acute respiratory infection. Tympanostomy was performed for 35% and adenoidectomy for 13% of the children. Asthma was diagnosed in 12% by 24 months of age. Older siblings increased the risk of recurrent respiratory infections. Early nasopharyngeal colonization with Streptococcus pneumoniae was common in children who later developed recurrent infections. CONCLUSIONS: Children with recurrent respiratory infections frequently use health care services and antibiotics, undergo surgical procedures and are at risk for asthma in early life. Having older siblings increases the risk of recurrent infections.