Mycoplasma pneumoniae carriage in children with recurrent respiratory tract infections is associated with a less diverse and altered microbiota.

BACKGROUND: Mycoplasma pneumoniae is a common cause of community-acquired pneumonia in school-aged children and can be preceded by asymptomatic carriage. However, its role in recurrent respiratory tract infections is unclear. We studied the prevalence of M.pneumoniae carriage in children with recurrent respiratory infections and identified associated factors. METHODS: We tested M.pneumoniae carriage by qPCR in children with recurrent infections and their healthy family members in a cross-sectional study. Serum and mucosal total and M.pneumoniae-specific antibody levels were measured by ELISA and nasopharyngeal microbiota composition was characterized by 16S-rRNA sequencing. FINDINGS: Prevalence of M.pneumoniae carriage was higher in children with recurrent infections (68%) than their family members without infections (47% in siblings and 27% in parents). M.pneumoniae carriage among family members appeared to be associated with transmission within the household, likely originating from the affected child. In logistic regression corrected for age and multiple comparisons, IgA (OR 0.16 [0.06-0.37]) and total IgG deficiency (OR 0.15 [0.02-0.74]) were less prevalent in M.pneumoniae carriers (n = 78) compared to non-carriers (n = 36). In multivariable analysis, the nasopharyngeal microbiota of M.pneumoniae carriers had lower alpha diversity (OR 0.27 [0.09-0.67]) and a higher abundance of Haemophilus influenzae (OR 45.01 [2.74-1608.11]) compared to non-carriers. INTERPRETATION: M.pneumoniae carriage is highly prevalent in children with recurrent infections and carriers have a less diverse microbiota with an overrepresentation of disease-associated microbiota members compared to non-carriers. Given the high prevalence of M.pneumoniae carriage and the strong association with H. influenzae, we recommend appropriate antibiotic coverage of M.pneumoniae and H. influenzae in case of suspected pneumonia in children with recurrent respiratory tract infections or their family members. FUNDING: Wilhelmina Children's Hospital Research Fund, 'Christine Bader Stichting Irene KinderZiekenhuis', Sophia Scientific Research Foundation, ESPID Fellowship funded by Seqirus, Hypatia Fellowship funded by Radboudumc and The Netherlands Organisation for Health Research and Development (ZonMW VENI grant to LM Verhagen).

Mother-to-infant microbiota transmission and infant microbiota development across multiple body sites.

Early-life microbiota seeding and subsequent development is crucial to future health. Cesarean-section (CS) birth, as opposed to vaginal delivery, affects early mother-to-infant transmission of microbes. Here, we assess mother-to-infant microbiota seeding and early-life microbiota development across six maternal and four infant niches over the first 30 days of life in 120 mother-infant pairs. Across all infants, we estimate that on average 58.5% of the infant microbiota composition can be attributed to any of the maternal source communities. All maternal source communities seed multiple infant niches. We identify shared and niche-specific host/environmental factors shaping the infant microbiota. In CS-born infants, we report reduced seeding of infant fecal microbiota by maternal fecal microbes, whereas colonization with breastmilk microbiota is increased when compared with vaginally born infants. Therefore, our data suggest auxiliary routes of mother-to-infant microbial seeding, which may compensate for one another, ensuring that essential microbes/microbial functions are transferred irrespective of disrupted transmission routes.

Asymptomatic Viral Presence in Early Life Precedes Recurrence of Respiratory Tract Infections.

BACKGROUND: Respiratory tract infections (RTIs) in infants are often caused by viruses. Although respiratory syncytial virus (RSV), influenza virus and human metapneumovirus (hMPV) can be considered the most pathogenic viruses in children, rhinovirus (RV) is often found in asymptomatic infants as well. Little is known about the health consequences of viral presence, especially early in life. We aimed to examine the dynamics of (a)symptomatic viral presence and relate early viral detection to susceptibility to RTIs in infants. METHODS: In a prospective birth cohort of 117 infants, we tested 1304 nasopharyngeal samples obtained from 11 consecutive regular sampling moments, and during acute RTIs across the first year of life for 17 respiratory viruses by quantitative PCR. Associations between viral presence, viral (sub)type, viral load, viral co-detection and symptoms were tested by generalized estimating equation (GEE) models. RESULTS: RV was the most detected virus. RV was negatively associated [GEE: adjusted odds ratio (aOR) 0.41 (95% CI 0.18-0.92)], and hMPV, RSV, parainfluenza 2 and 4 and human coronavirus HKU1 were positively associated with an acute RTI. Asymptomatic RV in early life was, however, associated with increased susceptibility to and recurrence of RTIs later in the first year of life (Kaplan-Meier survival analysis: P = 0.022). CONCLUSIONS: Respiratory viruses, including the seasonal human coronaviruses, are often detected in infants, and are often asymptomatic. Early life RV presence is, though negatively associated with an acute RTI, associated with future susceptibility to and recurrence of RTIs. Further studies on potential ecologic or immunologic mechanisms are needed to understand these observations.

Mode of delivery modulates the intestinal microbiota and impacts the response to vaccination.

The gut microbiota in early life, when critical immune maturation takes place, may influence the immunogenicity of childhood vaccinations. Here we assess the association between mode of delivery, gut microbiota development in the first year of life, and mucosal antigen-specific antibody responses against pneumococcal vaccination in 101 infants at age 12 months and against meningococcal vaccination in 66 infants at age 18 months. Birth by vaginal delivery is associated with higher antibody responses against both vaccines. Relative abundances of vaginal birth-associated Bifidobacterium and Escherichia coli in the first weeks of life are positively associated with anti-pneumococcal antibody responses, and relative abundance of E. coli in the same period is also positively associated with anti-meningococcal antibody responses. In this study, we show that mode of delivery-induced microbiota profiles of the gut are associated with subsequent antibody responses to routine childhood vaccines.

Higher off-target amplicon detection rate in MiSeq v3 compared to v2 reagent kits in the context of 16S-rRNA-sequencing.

One of the most widely used techniques in microbiota research is 16S-rRNA-sequencing. Several laboratory processes have been shown to impact sequencing results, especially in low biomass samples. Low biomass samples are prone to off-target amplification, where instead of bacterial DNA, host DNA is erroneously amplified. Knowledge on the laboratory processes influencing off-target amplification and detection is however scarce. We here expand on previous findings by demonstrating that off-target amplification is not limited to invasive biopsy samples, but is also an issue in low bacterial biomass respiratory (mucosal) samples, especially when below 0.3 pg/µL. We show that off-target amplification can partly be mitigated by using gel-based library purification methods. Importantly, we report a higher off-target amplicon detection rate when using MiSeq reagent kit v3 compared to v2 (mean 13.3% vs 0.1% off-target reads/sample, respectively), possibly as a result of differences in reagents or sequencing recipes. However, since after bioinformatic removal of off-target reads, MiSeq reagent kit v3 still results in a twofold higher number of reads when compared to v2, v3 is still preferred over v2. Together, these results add to the growing knowledge base on off-target amplification and detection, allowing researchers to anticipate this problem in 16S-rRNA-based microbiome studies involving low biomass samples.

Effects of early-life antibiotics on the developing infant gut microbiome and resistome: a randomized trial.

Broad-spectrum antibiotics for suspected early-onset neonatal sepsis (sEONS) may have pronounced effects on gut microbiome development and selection of antimicrobial resistance when administered in the first week of life, during the assembly phase of the neonatal microbiome. Here, 147 infants born at ≥36 weeks of gestational age, requiring broad-spectrum antibiotics for treatment of sEONS in their first week of life were randomized 1:1:1 to receive three commonly prescribed intravenous antibiotic combinations, namely penicillin + gentamicin, co-amoxiclav + gentamicin or amoxicillin + cefotaxime (ZEBRA study, Trial Register NL4882). Average antibiotic treatment duration was 48 hours. A subset of 80 non-antibiotic treated infants from a healthy birth cohort served as controls (MUIS study, Trial Register NL3821). Rectal swabs and/or faeces were collected before and immediately after treatment, and at 1, 4 and 12 months of life. Microbiota were characterized by 16S rRNA-based sequencing and a panel of 31 antimicrobial resistance genes was tested using targeted qPCR. Confirmatory shotgun metagenomic sequencing was executed on a subset of samples. The overall gut microbial community composition and antimicrobial resistance gene profile majorly shift directly following treatment (R2 = 9.5%, adjusted p-value = 0.001 and R2 = 7.5%, adjusted p-value = 0.001, respectively) and normalize over 12 months (R2 = 1.1%, adjusted p-value = 0.03 and R2 = 0.6%, adjusted p-value = 0.23, respectively). We find a decreased abundance of Bifidobacterium spp. and increased abundance of Klebsiella and Enterococcus spp. in the antibiotic treated infants compared to controls. Amoxicillin + cefotaxime shows the largest effects on both microbial community composition and antimicrobial resistance gene profile, whereas penicillin + gentamicin exhibits the least effects. These data suggest that the choice of empirical antibiotics is relevant for adverse ecological side-effects.

Early-life viral infections are associated with disadvantageous immune and microbiota profiles and recurrent respiratory infections.

The respiratory tract is populated by a specialized microbial ecosystem, which is seeded during and directly following birth. Perturbed development of the respiratory microbial community in early-life has been associated with higher susceptibility to respiratory tract infections (RTIs). Given a consistent gap in time between first signs of aberrant microbial maturation and the observation of the first RTIs, we hypothesized that early-life host-microbe cross-talk plays a role in this process. We therefore investigated viral presence, gene expression profiles and nasopharyngeal microbiota from birth until 12 months of age in 114 healthy infants. We show that the strongest dynamics in gene expression profiles occurred within the first days of life, mostly involving Toll-like receptor (TLR) and inflammasome signalling. These gene expression dynamics coincided with rapid microbial niche differentiation. Early asymptomatic viral infection co-occurred with stronger interferon activity, which was related to specific microbiota dynamics following, including early enrichment of Moraxella and Haemophilus spp. These microbial trajectories were in turn related to a higher number of subsequent (viral) RTIs over the first year of life. Using a multi-omic approach, we found evidence for species-specific host-microbe interactions related to consecutive susceptibility to RTIs. Although further work will be needed to confirm causality of our findings, together these data indicate that early-life viral encounters could impact subsequent host-microbe cross-talk, which is linked to later-life infections.

Microbial community networks across body sites are associated with susceptibility to respiratory infections in infants.

Respiratory tract infections are a major cause of morbidity and mortality worldwide in young children. Concepts such as the gut-lung axis have highlighted the impact of microbial communities at distal sites in mediating disease locally. However, little is known about the extent to which microbial communities from multiple body sites are linked, and how this relates to disease susceptibility. Here, we combine 16S-based rRNA sequencing data from 112 healthy, term born infants, spanning three body sites (oral cavity, nasopharynx, gut) and the first six months of life. Using a cross-niche microbial network approach, we show that, already from the first week of life on, there is a strong association between both network structure and species essential to these structures (hub species), and consecutive susceptibility to respiratory tract infections in this cohort. Our findings underline the crucial role of cross-niche microbial connections in respiratory health.

Benchmarking laboratory processes to characterise low-biomass respiratory microbiota.

The low biomass of respiratory samples makes it difficult to accurately characterise the microbial community composition. PCR conditions and contaminating microbial DNA can alter the biological profile. The objective of this study was to benchmark the currently available laboratory protocols to accurately analyse the microbial community of low biomass samples. To study the effect of PCR conditions on the microbial community profile, we amplified the 16S rRNA gene of respiratory samples using various bacterial loads and different number of PCR cycles. Libraries were purified by gel electrophoresis or AMPure XP and sequenced by V2 or V3 MiSeq reagent kits by Illumina sequencing. The positive control was diluted in different solvents. PCR conditions had no significant influence on the microbial community profile of low biomass samples. Purification methods and MiSeq reagent kits provided nearly similar microbiota profiles (paired Bray-Curtis dissimilarity median: 0.03 and 0.05, respectively). While profiles of positive controls were significantly influenced by the type of dilution solvent, the theoretical profile of the Zymo mock was most accurately analysed when the Zymo mock was diluted in elution buffer (difference compared to the theoretical Zymo mock: 21.6% for elution buffer, 29.2% for Milli-Q, and 79.6% for DNA/RNA shield). Microbiota profiles of DNA blanks formed a distinct cluster compared to low biomass samples, demonstrating that low biomass samples can accurately be distinguished from DNA blanks. In summary, to accurately characterise the microbial community composition we recommend 1. amplification of the obtained microbial DNA with 30 PCR cycles, 2. purifying amplicon pools by two consecutive AMPure XP steps and 3. sequence the pooled amplicons by V3 MiSeq reagent kit. The benchmarked standardized laboratory workflow presented here ensures comparability of results within and between low biomass microbiome studies.

Microbial and clinical factors are related to recurrence of symptoms after childhood lower respiratory tract infection.

Childhood lower respiratory tract infections (LRTI) are associated with dysbiosis of the nasopharyngeal microbiota, and persistent dysbiosis following the LRTI may in turn be related to recurrent or chronic respiratory problems. Therefore, we aimed to investigate microbial and clinical predictors of early recurrence of respiratory symptoms as well as recovery of the microbial community following hospital admission for LRTI in children. To this end, we collected clinical data and characterised the nasopharyngeal microbiota of 154 children (4 weeks-5 years old) hospitalised for a LRTI (bronchiolitis, pneumonia, wheezing illness or mixed infection) at admission and 4-8 weeks later. Data were compared to 307 age-, sex- and time-matched healthy controls. During follow-up, 66% of cases experienced recurrence of (mild) respiratory symptoms. In cases with recurrence of symptoms during follow-up, we found distinct nasopharyngeal microbiota at hospital admission, with higher levels of Haemophilus influenzae/haemolyticus, Prevotella oris and other gram-negatives and lower levels of Corynebacterium pseudodiphtheriticum/propinquum and Dolosigranulum pigrum compared with healthy controls. Furthermore, in cases with recurrence of respiratory symptoms, recovery of the microbiota was also diminished. Especially in cases with wheezing illness, we observed a high rate of recurrence of respiratory symptoms, as well as diminished microbiota recovery at follow-up. Together, our results suggest a link between the nasopharyngeal microbiota composition during LRTI and early recurrence of respiratory symptoms, as well as diminished microbiota recovery after 4-8 weeks. Future studies should investigate whether (speed of) ecological recovery following childhood LRTI is associated with long-term respiratory problems.

Individual and Group Response of Treatment with Ivacaftor on Airway and Gut Microbiota in People with CF and a S1251N Mutation.

Ivacaftor has been shown to restore the functionality of the S1251N (also known as c.3752G>A) mutated CFTR, which may cause alterations in both airway and gut physiology and micro-environment, resulting in a change of microbiota in these organs. The aim of the present study was to analyze the effects of ivacaftor on the microbial community composition of both airway and gut in subjects with CF carrying one S1251N mutation, using a 16S rRNA gene-based sequencing approach. In 16 subjects with CF, repetitive samples from airways and gut were collected just before, and 2 months after, and, for 8 patients, also 9 and 12 months after, start of ivacaftor. 16S rRNA based sequencing identified 344 operational taxonomical units (OTUs) in a total of 139 samples (35 nasopharyngeal, 39 oropharyngeal, 29 sputum, and 36 fecal samples). Ivacaftor significantly enhanced bacterial diversity and overall microbiota composition in the gut (p < 0.01). There were no significant changes in the overall microbial composition and alpha diversity in upper and lower airways of these patients after ivacaftor treatment. Treatment with ivacaftor induces changes in gut microbiota whereas airway microbiota do not change significantly over time.

Nasopharyngeal Microbiota Profiles in Rural Venezuelan Children Are Associated With Respiratory and Gastrointestinal Infections.

BACKGROUND: Recent research suggests that the microbiota affects susceptibility to both respiratory tract infections (RTIs) and gastrointestinal infections (GIIs). In order to optimize global treatment options, it is important to characterize microbiota profiles across different niches and geographic/socioeconomic areas where RTI and GII prevalences are high. METHODS: We performed 16S sequencing of nasopharyngeal swabs from 209 Venezuelan Amerindian children aged 6 weeks-59 months who were participating in a 13-valent pneumococcal conjugate vaccine (PCV13) study. Using random forest models, differential abundance testing, and regression analysis, we determined whether specific bacteria were associated with RTIs or GIIs and variation in PCV13 response. RESULTS: Microbiota compositions differed between children with or without RTIs (P = .018) or GIIs (P = .001). Several species were associated with the absence of infections. Some of these health-associated bacteria are also observed in developed regions, such as Corynebacterium (log2(fold change [FC]) = 3.30 for RTIs and log2(FC) = 1.71 for GIIs), while others are not commonly observed in developed regions, such as Acinetobacter (log2(FC) = 2.82 and log2(FC) = 5.06, respectively). Klebsiella spp. presence was associated with both RTIs (log2(FC) = 5.48) and GIIs (log2(FC) = 7.20). CONCLUSIONS: The nasopharyngeal microbiota of rural Venezuelan children included several bacteria that thrive in tropical humid climates. Interestingly, nasopharyngeal microbiota composition not only differed in children with an RTI but also in those with a GII, which suggests a reciprocal interplay between the 2 environments. Knowledge of region-specific microbiota patterns enables tailoring of preventive and therapeutic approaches.

Author Correction: Impact of delivery mode-associated gut microbiota dynamics on health in the first year of life.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Impact of delivery mode-associated gut microbiota dynamics on health in the first year of life.

The early-life microbiome appears to be affected by mode of delivery, but this effect may depend on intrapartum antibiotic exposure. Here, we assess the effect of delivery mode on gut microbiota, independent of intrapartum antibiotics, by postponing routine antibiotic administration to mothers until after cord clamping in 74 vaginally delivered and 46 caesarean section born infants. The microbiota differs between caesarean section born and vaginally delivered infants over the first year of life, showing enrichment of Bifidobacterium spp., and reduction of Enterococcus and Klebsiella spp. in vaginally delivered infants. The microbiota composition at one week of life is associated with the number of respiratory infections over the first year. The taxa driving this association are more abundant in caesarean section born children, providing a possible link between mode of delivery and susceptibility to infectious outcomes.

Interaction between the nasal microbiota and S. pneumoniae in the context of live-attenuated influenza vaccine.

Streptococcus pneumoniae is the main bacterial pathogen involved in pneumonia. Pneumococcal acquisition and colonization density is probably affected by viral co-infections, the local microbiome composition and mucosal immunity. Here, we report the interactions between live-attenuated influenza vaccine (LAIV), successive pneumococcal challenge, and the healthy adult nasal microbiota and mucosal immunity using an experimental human challenge model. Nasal microbiota profiles at baseline are associated with consecutive pneumococcal carriage outcome (non-carrier, low-dense and high-dense pneumococcal carriage), independent of LAIV co-administration. Corynebacterium/Dolosigranulum-dominated profiles are associated with low-density colonization. Lowest rates of natural viral co-infection at baseline and post-LAIV influenza replication are detected in the low-density carriers. Also, we detected the fewest microbiota perturbations and mucosal cytokine responses in the low-density carriers compared to non-carriers or high-density carriers. These results indicate that the complete respiratory ecosystem affects pneumococcal behaviour following challenge, with low-density carriage representing the most stable ecological state.

Bacterial and viral respiratory tract microbiota and host characteristics in children with lower respiratory tract infections: a matched case-control study.

BACKGROUND: Lower respiratory tract infections (LRTIs) are a leading cause of childhood morbidity and mortality. Potentially pathogenic organisms are present in the respiratory tract in both symptomatic and asymptomatic children, but their presence does not necessarily indicate disease. We aimed to assess the concordance between upper and lower respiratory tract microbiota during LRTIs and the use of nasopharyngeal microbiota to discriminate LRTIs from health. METHODS: First, we did a prospective study of children aged between 4 weeks and 5 years who were admitted to the paediatric intensive care unit (PICU) at Wilhelmina Children's Hospital (Utrecht, Netherlands) for a WHO-defined LRTI requiring mechanical ventilation. We obtained paired nasopharyngeal swabs and deep endotracheal aspirates from these participants (the so-called PICU cohort) between Sept 10, 2013, and Sept 4, 2016. We also did a matched case-control study (1:2) with the same inclusion criteria in children with LRTIs at three Dutch teaching hospitals and in age-matched, sex-matched, and time-matched healthy children recruited from the community. Nasopharyngeal samples were obtained at admission for cases and during home visits for controls. Data for child characteristics were obtained by questionnaires and from pharmacy printouts and medical charts. We used quantitative PCR and 16S rRNA-based sequencing to establish viral and bacterial microbiota profiles, respectively. We did sparse random forest classifier analyses on the bacterial data, viral data, metadata, and the combination of all three datasets to distinguish cases from controls. FINDINGS: 29 patients were enrolled in the PICU cohort. Intra-individual concordance in terms of viral microbiota profiles (96% agreement [95% CI 93-99]) and bacterial microbiota profiles (58 taxa with a median Pearson's r 0·93 [IQR 0·62-0·99]; p<0·05 for all 58 taxa) was high between nasopharyngeal and endotracheal aspirate samples, supporting the use of nasopharyngeal samples as proxy for lung microbiota during LRTIs. 154 cases and 307 matched controls were prospectively recruited to our case-control cohort. Individually, bacterial microbiota (area under the curve 0·77), viral microbiota (0·70), and child characteristics (0·80) poorly distinguished health from disease. However, a classification model based on combined bacterial and viral microbiota plus child characteristics distinguished children with LRTIs from their matched controls with a high degree of accuracy (area under the curve 0·92). INTERPRETATION: Our data suggest that the nasopharyngeal microbiota can serve as a valid proxy for lower respiratory tract microbiota in childhood LRTIs, that clinical LRTIs in children result from the interplay between microbiota and host characteristics, rather than a single microorganism, and that microbiota-based diagnostics could improve future diagnostic and treatment protocols. FUNDING: Spaarne Gasthuis, University Medical Center Utrecht, and the Netherlands Organization for Scientific Research.

Loss of Microbial Topography between Oral and Nasopharyngeal Microbiota and Development of Respiratory Infections Early in Life.

Rationale: The respiratory microbiota is increasingly being appreciated as an important mediator in the susceptibility to childhood respiratory tract infections (RTIs). Pathogens are presumed to originate from the nasopharyngeal ecosystem.Objectives: To investigate the association between early life respiratory microbiota and development of childhood RTIs.Methods: In a prospective birth cohort (Microbiome Utrecht Infant Study: MUIS), we characterized the oral microbiota longitudinally from birth until 6 months of age of 112 infants (nine regular samples/subject) and compared them with nasopharyngeal microbiota using 16S-rRNA-based sequencing. We also characterized oral and nasopharynx samples during RTI episodes in the first half year of life.Measurements and Main Results: Oral microbiota were driven mostly by feeding type, followed by age, mode of delivery, and season of sampling. In contrast to our previously published associations between nasopharyngeal microbiota development and susceptibility to RTIs, oral microbiota development was not directly associated with susceptibility to RTI development. However, we did observe an influx of oral taxa, such as Neisseria lactamica, Streptococcus, Prevotella nanceiensis, Fusobacterium, and Janthinobacterium lividum, in the nasopharyngeal microbiota before and during RTIs, which was accompanied by reduced presence and abundance of Corynebacterium, Dolosigranulum, and Moraxella spp. Moreover, this phenomenon was accompanied by reduced niche differentiation indicating loss of ecological topography preceding confirmed RTIs. This loss of ecological topography was further augmented by start of daycare, and linked to consecutive development of symptomatic infections.Conclusions: Together, our results link the loss of topography to subsequent development of RTI episodes. This may lead to new insights for prevention of RTIs and antibiotic use in childhood.

Respiratory Microbiota Predicts Clinical Disease Course of Acute Otorrhea in Children With Tympanostomy Tubes.

BACKGROUND: Acute otitis media (AOM) is one of the most common childhood infections, generally thought to be caused by ascension of bacteria from the nasopharynx (NP) to the middle ear. Using 16S ribosomal RNA-based sequencing, we evaluated the relationship between the NP and middle ear fluid (MEF) microbiota in children with AOM with tympanostomy tubes (AOMT) as a proxy for AOM and explored whether microbiota profiling predicts natural disease course. METHODS: Microbiota profiles of paired NP and MEF samples of 94 children below 5 years of age with uncomplicated AOMT were determined. RESULTS: Local diversity (P < 0.001) and overall microbiota composition (P < 0.001) of NP and MEF samples differed significantly, although paired NP and MEF samples were much more similar than unpaired samples (P < 0.001). High qualitative agreement between the presence of individual bacteria in both niches was observed. Abundances of Pseudomonas aeruginosa, Staphylococcus aureus, Streptococcus pyogenes, Turicella otitidis, Klebsiella pneumoniae and Haemophilus spp. were strongly correlated between the 2 niches. Additionally, P. aeruginosa, S. aureus, T. otitidis and Streptococcus pneumoniae abundance in NP were predictive of the presence of a range of oral types of bacteria in MEF. Interestingly, there was no association between Moraxella catarrhalis in NP and MEF samples, which was highly present in NP but virtually absent in MEF. Finally, the NP microbiota composition could predict duration of AOMT, even better than MEF microbiota. CONCLUSIONS: We observed substantial correlations between paired NP and MEF microbiota in children with AOMT. Our data also suggest that NP microbiota profiling deserves further exploration as tool for future treatment decisions.

Exploring Immune Development in Infants With Moderate to Severe Atopic Dermatitis.

Background: Atopic dermatitis (AD) is the most common chronic inflammatory skin disease in infancy with a complex pathology. In adults, the clinical severity of AD has been associated with increases in T helper cell type (Th) 2, Th22, and Th17 serum markers, including high levels of CC chemokine ligand (CCL) 17 and CCL22 chemokines. Objective: To explore the possible association between serum chemokine levels and AD severity in infants with moderate-to-severe AD and elevated immunoglobulin E (IgE). Subjects and methods: Serum samples (n = 41) obtained from a randomized, double-blind, and clinical dietary intervention study were used to study biomarkers in infants with AD. Baseline- and post-intervention samples (4 months) were used, six chemokines and nine ratios thereof were analyzed using Luminex and correlated to AD severity. In the initial study, the infants were randomized to receive extensively hydrolyzed whey-based formula without (control) or with short-chain galacto-oligosaccharides/long-chain fructo-oligosaccharides (9:1) and Bifidobacterium breve M-16V (active). Results: 31 Infants up to 11 months of age, with an objective-SCORAD score (oSCORAD) ≥ 20 and elevated total-IgE and/or specific-IgE levels were included. In time, the median oSCORAD decreased in both groups by -8 (control, p < 0.05; active, p < 0.01). Irrespective of dietary intervention, several changes in Th2 chemokines (CCL17 and CCL22), inflammatory chemokine (CCL20), and the Th1 chemokine, CXC chemokine ligand (CXCL) 9, were detected over time. Overall CCL17 correlated to oSCORAD (r = 0.446, p < 0.01). After 4 months of dietary intervention, CXCL9 was higher (p < 0.01) in the active group compared with control [active, 2.33 (1.99-2.89); controls, 1.95 (1.77-2.43) log 10 median (range)]. In addition, a reduction in Th2/Th1 chemokine ratios for CCL17/CXCL9, CCL22/CXCL9, CCL20/CXCL10, and CCL20/CXCL11 was detected associated with the active intervention. Conclusion: While this study is small and exploratory in nature, these data contribute to immune biomarker profiling and understanding of AD in infants.

Maturation of the Infant Respiratory Microbiota, Environmental Drivers, and Health Consequences. A Prospective Cohort Study.

RATIONALE: Perinatal and postnatal influences are presumed important drivers of the early-life respiratory microbiota composition. We hypothesized that the respiratory microbiota composition and development in infancy is affecting microbiota stability and thereby resistance against respiratory tract infections (RTIs) over time. OBJECTIVES: To investigate common environmental drivers, including birth mode, feeding type, antibiotic exposure, and crowding conditions, in relation to respiratory tract microbiota maturation and stability, and consecutive risk of RTIs over the first year of life. METHODS: In a prospectively followed cohort of 112 infants, we characterized the nasopharyngeal microbiota longitudinally from birth on (11 consecutive sample moments and the maximum three RTI samples per subject; in total, n = 1,121 samples) by 16S-rRNA gene amplicon sequencing. MEASUREMENTS AND MAIN RESULTS: Using a microbiota-based machine-learning algorithm, we found that children experiencing a higher number of RTIs in the first year of life already demonstrate an aberrant microbial developmental trajectory from the first month of life on as compared with the reference group (0-2 RTIs/yr). The altered microbiota maturation process coincided with decreased microbial community stability, prolonged reduction of Corynebacterium and Dolosigranulum, enrichment of Moraxella very early in life, followed by later enrichment of Neisseria and Prevotella spp. Independent drivers of these aberrant developmental trajectories of respiratory microbiota members were mode of delivery, infant feeding, crowding, and recent antibiotic use. CONCLUSIONS: Our results suggest that environmental drivers impact microbiota development and, consequently, resistance against development of RTIs. This supports the idea that microbiota form the mediator between early-life environmental risk factors for and susceptibility to RTIs over the first year of life.