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Rationale: Children with preschool wheezing or school-age asthma are reported to have airway microbial imbalances. Objectives: To identify clusters in children with asthma or wheezing using oropharyngeal microbiota profiles. Methods: Oropharyngeal swabs from the U-BIOPRED (Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes) pediatric asthma or wheezing cohort were characterized using 16S ribosomal RNA gene sequencing, and unsupervised hierarchical clustering was performed on the Bray-Curtis β-diversity. Enrichment scores of the Molecular Signatures Database hallmark gene sets were computed from the blood transcriptome using gene set variation analysis. Children with severe asthma or severe wheezing were followed up for 12-18 months, with assessment of the frequency of exacerbations. Measurements and Main Results: Oropharyngeal samples from 241 children (age range, 1-17 years; 40% female) revealed four taxa-driven clusters dominated by Streptococcus , Veillonella , Rothia , and Haemophilus . The clusters showed significant differences in atopic dermatitis, grass pollen sensitization, FEV 1 % predicted after salbutamol, and annual asthma exacerbation frequency during follow-up. The Veillonella cluster was the most allergic and included the highest percentage of children with two or more exacerbations per year during follow-up. The oropharyngeal clusters were different in the enrichment scores of TGF-β (transforming growth factor-β) (highest in the Veillonella cluster) and Wnt/β-catenin signaling (highest in the Haemophilus cluster) transcriptomic pathways in blood (all q values <0.05). Conclusions: Analysis of the oropharyngeal microbiota of children with asthma or wheezing identified four clusters with distinct clinical characteristics (phenotypes) that associate with risk for exacerbation and transcriptomic pathways involved in airway remodeling. This suggests that further exploration of the oropharyngeal microbiota may lead to novel pathophysiologic insights and potentially new treatment approaches.

Rationale: There is a major unmet need for improving the care of children and adolescents with severe asthma and wheeze. Identifying factors contributing to disease severity may lead to improved diagnostics, biomarkers, or therapies. The airway microbiota may be such a key factor. Objectives: To compare the oropharyngeal airway microbiota of children and adolescents with severe and mild/moderate asthma/wheeze. Methods: Oropharyngeal swab samples from school-age and preschool children in the European U-BIOPRED (Unbiased BIOmarkers in the PREDiction of respiratory disease outcomes) multicenter study of severe asthma, all receiving severity-appropriate treatment, were examined using 16S ribosomal RNA gene sequencing. Bacterial taxa were defined as amplicon sequence variants. Results: We analyzed 241 samples from four cohorts: A) 86 school-age children with severe asthma; B) 39 school-age children with mild/moderate asthma; C) 65 preschool children with severe wheeze; and D) 51 preschool children with mild/moderate wheeze. The most common bacteria were Streptococcus (mean relative abundance, 33.5%), Veillonella (10.3%), Haemophilus (7.0%), Prevotella (5.9%), and Rothia (5.5%). Age group (school-age vs. preschool) was associated with the microbiota in β-diversity analysis ( F  = 3.32, P  = 0.011) and in a differential abundance analysis (28 significant amplicon sequence variants). Among all children, we found no significant difference in the microbiota between children with severe and mild/moderate asthma/wheeze in univariable β-diversity analysis ( F  = 1.99, P  = 0.08, N  = 241), but a significant difference in a multivariable model ( F  = 2.66, P  = 0.035), including the number of exacerbations in the previous year. Age was also significant when expressed as a microbial maturity score (Spearman Rho, 0.39; P  = 4.6 × 10 -10 ); however, this score was not associated with asthma/wheeze severity. Conclusions: There was a modest difference in the oropharyngeal airway microbiota between children with severe and mild/moderate asthma/wheeze across all children but not in individual age groups, and a strong association between the microbiota and age. This suggests the oropharyngeal airway microbiota as an interesting entity in studying asthma severity, but probably without the strength to serve as a biomarker for targeted intervention.

Despite advances in sequencing, lack of standardization makes comparisons across studies challenging and hampers insights into the structure and function of microbial communities across multiple habitats on a planetary scale. Here we present a multi-omics analysis of a diverse set of 880 microbial community samples collected for the Earth Microbiome Project. We include amplicon (16S, 18S, ITS) and shotgun metagenomic sequence data, and untargeted metabolomics data (liquid chromatography-tandem mass spectrometry and gas chromatography mass spectrometry). We used standardized protocols and analytical methods to characterize microbial communities, focusing on relationships and co-occurrences of microbially related metabolites and microbial taxa across environments, thus allowing us to explore diversity at extraordinary scale. In addition to a reference database for metagenomic and metabolomic data, we provide a framework for incorporating additional studies, enabling the expansion of existing knowledge in the form of an evolving community resource. We demonstrate the utility of this database by testing the hypothesis that every microbe and metabolite is everywhere but the environment selects. Our results show that metabolite diversity exhibits turnover and nestedness related to both microbial communities and the environment, whereas the relative abundances of microbially related metabolites vary and co-occur with specific microbial consortia in a habitat-specific manner. We additionally show the power of certain chemistry, in particular terpenoids, in distinguishing Earth's environments (for example, terrestrial plant surfaces and soils, freshwater and marine animal stool), as well as that of certain microbes including Conexibacter woesei (terrestrial soils), Haloquadratum walsbyi (marine deposits) and Pantoea dispersa (terrestrial plant detritus). This Resource provides insight into the taxa and metabolites within microbial communities from diverse habitats across Earth, informing both microbial and chemical ecology, and provides a foundation and methods for multi-omics microbiome studies of hosts and the environment., (© 2022. The Author(s).)

Antibiotic-resistant pathogens constitute an escalating public health concern. Hence a better understanding of the underlying processes responsible for this expansion is urgently needed. Co-selection of heavy metal/biocide and antibiotic resistance genes (ARGs) has been suggested as one potential mechanism promoting the proliferation of antimicrobial resistance (AMR). This paper aims to elucidate this interplay and exploit differences in antibiotic usage to infer patterns of co-selection by the non-antibiotic factors metals and biocides in the context of pig farming. We examined 278 gut metagenomes from pigs with continuous antibiotic exposure, only at weaning and at no exposure. Metals as growth promoters and biocides as disinfectants are currently used with little restrictions in stock farming. The pigs under continuous antibiotic exposure displayed the highest co-occurrence of ARGs and other genetic elements while the pigs under limited use of antibiotics still showed abundant co-occurrences. Pathogens belonging to Enterobacteriaceae displayed increased co-occurrence phenomena, suggesting that this maintenance is not a random selection process from a mobilized pool but pertains to specific phylogenetic clades. These results suggest that metals and biocides displayed strong selective pressures on ARGs exerted by intensive farming, regardless of the current use of antibiotics., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Type 2 diabetes (T2D) management is based on combined pharmacological and lifestyle intervention approaches. While their clinical benefits are well studied, less is known about their effects on the gut microbiota. We aimed to investigate if an intensive lifestyle intervention combined with conventional standard care leads to a different gut microbiota composition compared to standard care alone treatment in individuals with T2D, and if gut microbiota is associated with the clinical benefits of the treatments. Ninety-eight individuals with T2D were randomized to either an intensive lifestyle intervention combined with standard care group (N = 64), or standard care alone group (N = 34) for 12 months. All individuals received standardized, blinded, target-driven medical therapy, and individual counseling. The lifestyle intervention group moreover received intensified physical training and dietary plans. Clinical characteristics and fecal samples were collected at baseline, 3-, 6-, 9-, and 12-month follow-up. The gut microbiota was profiled with 16S rRNA gene amplicon sequencing. There were no statistical differences in the change of gut microbiota composition between treatments after 12 months, except minor and transient differences at month 3. The shift in gut microbiota alpha diversity at all time windows did not correlate with the change in clinical characteristics, and the gut microbiota did not mediate the treatment effect on clinical characteristics. The clinical benefits of intensive lifestyle and/or pharmacological interventions in T2D are unlikely to be explained by, or causally related to, changes in the gut microbiota composition.

Rationale: Childhood asthma is often preceded by recurrent episodes of asthma-like symptoms, which can be triggered by both viral and bacterial agents. Recent randomized controlled trials have shown that azithromycin treatment reduces episode duration and severity through yet undefined mechanisms. Objectives: To study the influence of the airway microbiota on the effect of azithromycin treatment during acute episodes of asthma-like symptoms. Methods: Children from the COPSAC 2010 (Copenhagen Prospective Studies on Asthma in Childhood 2010) cohort with recurrent asthma-like symptoms aged 12-36 months were randomized during acute episodes to azithromycin or placebo as previously reported. Before randomization, hypopharyngeal aspirates were collected and examined by 16S ribosomal RNA gene amplicon sequencing. Measurements and Main Results: In 139 airway samples from 68 children, episode duration after randomization was associated with microbiota richness (7.5% increased duration per 10 additional operational taxonomic units [OTUs]; 95% confidence interval, 1-14%; P  = 0.025), with 15 individual OTUs (including several Neisseria and Veillonella ), and with microbial pneumotypes defined from weighted UniFrac distances (longest durations in a Neisseria -dominated pneumotype). Microbiota richness before treatment increased the effect of azithromycin by 10% per 10 additional OTUs, and more OTUs were positively versus negatively associated with an increased azithromycin effect (82 vs. 58; P  = 0.0032). Furthermore, effect modification of azithromycin was found for five individual OTUs (three OTUs increased and two OTUs decreased the effect; q  < 0.05). Conclusions: The airway microbiota in acute episodes of asthma-like symptoms is associated with episode duration and modifies the effect of azithromycin treatment of the episodes in preschool children with recurrent asthma-like symptoms. Clinical trial registered with www.clinicaltrials.gov (NCT01233297).

Environmental selection of antibiotic resistance genes (ARGs) is considered to be caused by antibiotic or metal residues, frequently used in livestock. In this study we examined three commercial poultry farms to correlate the co-occurrence patterns of antibiotic and metal residues to the presence of ARGs. We quantified 283 ARGs, 12 mobile genetic elements (MGEs), 49 targeted antibiotics, 7 heavy metals and sequenced 16S rRNA genes. The abundance and type of ARG were significantly enriched in manure while soil harbored the most diverse bacterial community. Procrustes analysis displayed significant correlations between ARGs/MGEs and the microbiome. Cadmium (Cd), arsenic (As), zinc (Zn), copper (Cu) and lead (Pb) were responsible for a majority of positive correlations to ARGs when compared to antibiotics. Integrons and transposons co-occurred with ARGs corresponding to 9 classes of antibiotics, especially Class1 integrase intI-1LC. Redundancy analysis (RDA) and Variance partitioning analysis (VPA) showed that antibiotics, metals, MGEs and bacteria explain solely 0.7%, 5.7%, 12.4%, and 21.9% of variances of ARGs in the microbial community, respectively. These results suggested that bacterial composition and horizontal gene transfer were the major factors shaping the composition of ARGs; Metals had a bigger effect on ARG profile than detected antibiotics in this study., Competing Interests: Declaration of competing interest We declare that no conflict of interest exits in this manuscript, and manuscript is approved by all authors for publication. This manuscript is original and has not been published in whole or in part previously., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Untargeted mass spectrometry is employed to detect small molecules in complex biospecimens, generating data that are difficult to interpret. We developed Qemistree, a data exploration strategy based on the hierarchical organization of molecular fingerprints predicted from fragmentation spectra. Qemistree allows mass spectrometry data to be represented in the context of sample metadata and chemical ontologies. By expressing molecular relationships as a tree, we can apply ecological tools that are designed to analyze and visualize the relatedness of DNA sequences to metabolomics data. Here we demonstrate the use of tree-guided data exploration tools to compare metabolomics samples across different experimental conditions such as chromatographic shifts. Additionally, we leverage a tree representation to visualize chemical diversity in a heterogeneous collection of samples. The Qemistree software pipeline is freely available to the microbiome and metabolomics communities in the form of a QIIME2 plugin, and a global natural products social molecular networking workflow.

Early-life microbiota has been linked to the development of chronic inflammatory diseases. It has been hypothesized that maternal vaginal microbiota is an important initial seeding source and therefore might have lifelong effects on disease risk. To understand maternal vaginal microbiota's role in seeding the child's microbiota and the extent of delivery mode-dependent transmission, we studied 665 mother-child dyads from the COPSAC 2010 cohort. The maternal vaginal microbiota was evaluated twice in the third trimester and compared with the children's fecal (at 1 week, 1 month, and 1 year of age) and airway microbiota (at 1 week, 1 month, and 3 months). Based on the concept of weighted transfer ratios (WTRs), we have identified bacterial orders for which the WTR displays patterns indicate persistent or transient transfer from the maternal vaginal microbiome, as well as orders that are shared at later time points independent of delivery mode, indicating a common reservoir., Competing Interests: MM, MR, JS, AB, CB, JT, KK, HB, SS No competing interests declared, (© 2021, Mortensen et al.)

The mother's vaginal microbiota represents the first microbes to which a child is exposed when delivered vaginally. However, little is known about the composition and development of the vaginal microbiota during pregnancy and birth. Here, we analyzed the vaginal microbiota of 57 women in pregnancy week 24, 36 and at birth after rupture of membranes but before delivery, and further compared the composition with that of the gut and airways of the 1-week-old child. The vaginal community structure had dramatic changes in bacterial diversity and taxonomic distribution, yet carried an individual-specific signature. The relative abundance of most bacterial taxa increased stepwise from week 24 of pregnancy until birth, with a gradual decline of Lactobacillus. Mother-to-child vertical transfer, as suggested by sharing, was modest, with the strongest transfer being for Clostridiales followed by Lactobacillales and Enterobacteriales. In conclusion, late gestation is associated with an increase in maternal vaginal microbiota diversity, and vaginal bacteria at birth only modestly predict the composition of the neonatal microbiota.

Background: The hamadryas baboon (Papio hamadryas) is a highly social primate that lives in complex multilevel societies exhibiting a wide range of group behaviors akin to humans. In contrast to the widely studied human microbiome, there is a paucity of information on the host-associated microbiomes of nonhuman primates (NHPs). Here, our goal was to understand the microbial composition throughout different body sites of cohabiting baboons., Results: We analyzed 170 oral, oropharyngeal, cervical, uterine, vaginal, nasal and rectal samples from 16 hamadryas baboons via 16S rRNA gene sequencing. Additionally, raw Miseq sequencing data from 1041 comparable publicly available samples from the human oral cavity, gut and vagina were reanalyzed using the same pipeline. We compared the baboon and human microbiome of the oral cavity, gut and vagina, showing that the baboon microbiome is distinct from the human. Baboon cohabitants share similar microbial profiles in their cervix, uterus, vagina, and gut. The oral cavity, gut and vagina shared more bacterial amplicon sequence variants (ASVs) in group living baboons than in humans. The shared ASVs had significantly positive correlations between most body sites, suggesting a potential bacterial exchange throughout the body. No significant differences in gut microbiome composition were detected within the maternity line and between maternity lines, suggesting that the offspring gut microbiota is shaped primarily through bacterial exchange among cohabitants. Finally, Lactobacillus was not so predominant in baboon vagina as in the human vagina but was the most abundant genus in the baboon gut., Conclusions: This study is the first to provide comprehensive analyses of the baboon microbiota across different body sites. We contrast this to human body sites and find substantially different microbiomes. This group of cohabitating baboons generally showed higher microbial diversity and remarkable similarities between body sites than were observed in humans. These data and findings from one group of baboons can form the basis of future microbiome studies in baboons and be used as a reference in research where the microbiome is expected to impact human modeling with baboons.

Although organic matter may accumulate sometimes (e.g. lignocellulose in peat bog), most natural biodegradation processes are completed until full mineralization. Such transformations are often achieved by the concerted action of communities of interacting microbes, involving different species each performing specific tasks. These interactions can give rise to novel "community-intrinsic" properties, through e.g. activation of so-called "silent genetic pathways" or synergistic interplay between microbial activities and functions. Here we studied the microbial community-based degradation of keratin, a recalcitrant biological material, by four soil isolates, which have previously been shown to display synergistic interactions during biofilm formation; Stenotrophomonas rhizophila, Xanthomonas retroflexus, Microbacterium oxydans and Paenibacillus amylolyticus. We observed enhanced keratin weight loss in cultures with X. retroflexus, both in dual and four-species co-cultures, as compared to expected keratin degradation by X. retroflexus alone. Additional community intrinsic properties included accelerated keratin degradation rates and increased biofilm formation on keratin particles. Comparison of secretome profiles of X. retroflexus mono-cultures to co-cultures revealed that certain proteases (e.g. serine protease S08) were significantly more abundant in mono-cultures, whereas co-cultures had an increased abundance of proteins related to maintaining the redox environment, e.g. glutathione peroxidase. Hence, one of the mechanisms related to the community intrinsic properties, leading to enhanced degradation from co-cultures, might be related to a switch from sulfitolytic to proteolytic functions between mono- and co-cultures, respectively., Competing Interests: The authors have declared that no competing interests exist.

We used direct DNA amplification from soil extracts to analyze microbial communities from an elevational transect in the German Alps by parallel metabarcoding of bacteria (16S rRNA), fungi (ITS2), and myxomycetes (18S rRNA). For the three microbial groups, 5710, 6133, and 261 operational taxonomic units (OTU) were found. For the latter group, we can relate OTUs to barcodes from fruit bodies sampled over a 4-year period. The alpha diversity of myxomycetes was positively correlated with that of bacteria. Vegetation type was found to be the main explanatory parameter for the community composition of all three groups and a substantial species turnover with elevation was observed. Bacteria and fungi display similar community responses, driven by symbiont species and plant substrate quality. Myxamoebae show a more patchy distribution, though still clearly stratified between taxa, which seems to be a response to both structural properties of the habitat and interaction with specific bacterial and fungal taxa. Finally, we report a high number of myxomycete OTUs not represented in a reference database from fructifications, which might represent novel species.

Background: Growing evidence supports the role of gut microbiota in obesity and its related disorders including type 2 diabetes. Ob/ob mice, which are hyperphagic due to leptin deficiency, are commonly used models of obesity and were instrumental in suggesting links between gut microbiota and obesity. Specific changes in their gut microbiota such as decreased microbial diversity and increased Firmicutes to Bacteroidetes ratio have been suggested to contribute to obesity via increased microbiota capacity to harvest energy. However, the differential development of ob/ob mouse gut microbiota compared to wild type microbiota and the role of hyperphagia in their metabolic impairment have not been investigated thoroughly., Results: We performed a 10-week long study in ob/ob (n = 12) and wild type control (n = 12) mice fed ad libitum. To differentiate effects of leptin deficiency from hyperphagia, we pair-fed an additional group of ob/ob mice (n = 11) based on the food consumption of control mice. Compared to control mice, ob/ob mice fed ad libitum exhibited compromised glucose metabolism and increased body fat percentage. Pair-fed ob/ob mice exhibited even more compromised glucose metabolism and maintained strikingly similar high body fat percentage at the cost of lean body mass. Acclimatization of the microbiota to our facility took up to 5 weeks. Leptin deficiency impacted gut microbial composition, explaining 18.3% of the variance. Pair-feeding also altered several taxa, although the overall community composition at the end of the study was not significantly different. We found 24 microbial taxa associations with leptin deficiency, notably enrichment of members of Lactobacillus and depletion of Akkermansia muciniphila. Microbial metabolic functions related to energy harvest, including glycan degradation, phosphotransferase systems and ABC transporters, were enriched in the ob/ob mice. Taxa previously reported as relevant for obesity were associated with body weight, including Oscillibacter and Alistipes (both negatively correlated) and Prevotella (positively correlated)., Conclusions: Leptin deficiency caused major changes in the mouse gut microbiota composition. Several microbial taxa were associated with body composition. Pair-fed mice maintained a pre-set high proportion of body fat despite reduced calorie intake, and exhibited more compromised glucose metabolism, with major implications for treatment options for genetically obese individuals.

Background: Exposure to arsenic and cadmium is common. Epidemiological and animal studies have suggested that exposure to these two heavy metals can cause metabolic health problems, including type 2 diabetes (T2DM). It has been hypothesized that T2DM could be mediated through the gut microbiome and the metabolites it produces. Although many studies have investigated the association between the gut microbiome and T2DM, few have focused on the connection to arsenic and cadmium., Results: We applied 16S rRNA gene amplicon sequencing and untargeted LC-MS/MS metabolomics to examine the changes in the gut microbiome and metabolite profiles of exposed mice to relevant levels of cadmium and arsenic in the drinking water over two weeks. Cadmium chloride (Cd) exposure significantly changed the mice gut microbiome and resulted in a significantly lower microbial diversity whereas sodium arsenite (As) caused a non-significant decrease in microbial diversity. For Cd and As treatment respectively, we identified 5 and 2 phyla with significant changes and 42 and 24 genera. Bacterial genera that were observed to decline upon both treatments, included several butyrate-producers. Both As and Cd treatment perturbed the metabolome significantly, with 50 ppm Cd compound exposure having the greatest effect when compared to 50 ppm As compound exposure. Two unidentified features were differentially abundant in the As group, while 33 features changed in the Cd group. Differential abundance analysis of all bile acid associated molecular components showed differences under both treatments. Finally, integrative network analysis via bipartite correlation networks suggested that several genera, including the metabolically important Blautia, Eisenbergiella, Clostridium&#95;XlVa, etc. declined in numbers of metabolite interactions., Conclusions: These results demonstrated that As and Cd exposure caused significant changes to the gut microbiome and metabolome by affecting bile acids, amino acids and taxa associated with metabolic health., (Copyright © 2019. Published by Elsevier Ltd.)

Tonsils in the oral cavity are an important source of contamination during pig slaughter, but have not received as much attention as faecal contamination. In the present study, ten pigs were sampled from tonsils, faeces and three different areas on each carcass. The samples were analysed by both culturing of Escherichia coli and Yersinia enterocolitica and by 16S rRNA gene sequencing to characterize the bacterial communities. Comparing culture data from deep tonsil tissue and tonsil surface showed similar numbers of E. coli but significantly higher numbers of Y. enterocolitica in the deep tissue samples. Microbiota analysis showed similar bacterial communities in the two sample types at phylum level, while comparison at genus level showed significant differences between the relative abundance of several genera in the two sample types. The finding of a significantly higher relative abundance of Yersinia in tonsil tissue compared to tonsil surface supported the culture analysis. The microbiota analysis also investigated characteristics of the bacterial community that could discriminate bacterial transfer from tonsils and faeces to the carcass during slaughter. The microbiota analyses demonstrated that Fusobacteria and Proteobacteria are the most abundant phyla in tonsils, while Firmicutes showed the highest relative abundance in faeces. The dominating phylum on carcasses was Proteobacteria. Besides Proteobacteria, the swabbing area on the forepart of the carcass, showed a higher relative abundance of Firmicutes and Fusobacteria compared to swabbing areas on the rear part and mid-section of the carcass. Principal coordinate analysis showed clear clustering of samples based on sample source (tonsils, faeces and carcass). Carcass swab samples from the forepart tended to cluster closer to the tonsil samples compared to carcass swab samples from the rear part and mid-section. Identification of the genera Fusobacterium, Moraxella, Actinobacillus and non-E. coli genera of the family Enterobacteriaceae in carcass swabs could indicate tonsil contamination, while faecal contamination would more likely include higher prevalence of bacteria belonging to the class of Clostridia. The present study supports that it is possible to identify bacterial groups that are indicative for either tonsil or faecal carcass contamination. The level and composition of Enterobacteriaceae on the carcasses did, however, indicate that other sources of meat contamination than tonsils and faeces may be important, such as the process environment., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Scope: The impact of dietary protein types on the gut microbiome is scarcely studied. The aim of the present study is therefore to examine the effects of lean-seafood and non-seafood proteins on the gut microbiome composition and activity and elucidate potential associations to cardiovascular disease (CVD) risk factors., Methods: A crossover intervention study in which 20 healthy subjects consumed two diets that varied in protein source was conducted. 1 H NMR spectroscopy and 16S rDNA sequencing analyses were applied to characterize fecal metabolites and gut microbiota composition, respectively., Results: A twofold increase in fecal trimethylamine excretion was observed after the lean-seafood diet period. Circulating TAG and the total to high-density lipoprotein (HDL) cholesterol ratio as well as circulating TMAO levels were each associated with specific gut bacteria. Following the non-seafood diet period, a decreased relative abundance of Clostridium cluster IV and a tendency toward an increased Firmicutes/Bacteroidetes ratio were found., Conclusions: Lean-seafood and non-seafood diets differentially modulate the gut microbiome composition and activity. Furthermore, the gut microbiota composition seems to affect circulating TMAO levels and CVD risk factors., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: We performed a 12-month cohort study of the stability and resilience of the intestinal microbiota of healthy children in daycare in Denmark in relation to diarrheal events and exposure to known risk factors for gastrointestinal health such as travelling and antibiotic use. In addition, we analyzed how gut microbiota recover from such exposures., Results: We monitored 32 children in daycare aged 1-6 years. Fecal samples were submitted every second month during a one-year observational period. Information regarding exposures and diarrheal episodes was obtained through questionnaires. Bacterial communities were identified using 16S rRNA gene sequencing. The core microbiota (mean abundance > 95%) dominated the intestinal microbiota, and none of the tested exposures (diarrheal events, travel, antibiotic use) were associated with decreases in the relative abundance of the core microbiota. Samples exhibited lower intra-individual variation than inter-individual variation. Half of all the variation between samples was explained by which child a sample originated from. Age explained 7.6-9.6% of the variation, while traveling, diarrheal events, and antibiotic use explained minor parts of the beta diversity. We found an age-dependent increase of alpha diversity in children aged 1-3 years, and while diarrheal events caused a decrease in alpha diversity, a recovery time of 40-45 days was observed. Among children having had a diarrheal event, we observed a 10x higher relative abundance of Prevotella. After travelling, a higher abundance of two Bacteroides species and 40% less Lachnospiraceae were seen. Antibiotic use did not correlate with changes in the abundance of any bacteria., Conclusion: We present data showing that Danish children in daycare have stable intestinal microbiota, resilient to the exposures investigated. An early age-dependent increase in the diversity was demonstrated. Diarrheal episodes decreased alpha diversity with an estimated recovery time of 40-45 days.

Introduction: Gut microbial communities are critical players in the pathogenesis of obesity. Pregnancy is associated with increased bacterial load and changes in gut bacterial diversity. Sparse data exist regarding composition of gut microbial communities in obesity combined with pregnancy., Material and Methods: Banked tissues were collected under sterile conditions during necropsy, from three non-obese (nOb) and four obese (Ob) near-term pregnant baboons. Sequences were assigned taxonomy using the Ribosomal Database Project classifier. Microbiome abundance and its difference between distinct groups were assessed by a nonparametric test., Results: Three families predominated in both the nOb and Ob colonic microbiome: Prevotellaceae (25.98% and 32.71% respectively), Ruminococcaceae (12.96% and 7.48%), and Lachnospiraceae (8.78% and 11.74%). Seven families of the colon microbiome displayed differences between Ob and nOb groups., Conclusion: Changes in gut microbiome in pregnant obese animals open the venue for dietary manipulation in pregnancy., (© 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Macrolides are commonly prescribed for respiratory infections and asthma-like episodes in children. While their clinical benefits have been proved, concerns regarding the side-effects of their therapeutic use have been raised. Here we assess the short- and long-term impacts of azithromycin on the gut microbiota of young children., Methods: We performed a randomized, double-blind, placebo-controlled trial in a group of children aged 12-36 months, diagnosed with recurrent asthma-like symptoms from the COPSAC 2010 cohort. Each acute asthma-like episode was randomized to a 3-day course of azithromycin oral solution of 10 mg/kg per day or placebo. Azithromycin reduced episode duration by half, which was the primary end-point and reported previously. The assessment of gut microbiota after treatment was the secondary end-point and reported in this study. Fecal samples were collected 14 days after randomization (N = 59, short-term) and again at age 4 years (N = 49, long-term, of whom N = 18 were placebo treated) and investigated by 16S rRNA gene amplicon sequencing., Findings: Short-term, azithromycin caused a 23% reduction in observed richness and 13% reduction in Shannon diversity. Microbiota composition was shifted primarily in the Actinobacteria phylum, especially a reduction of abundance in the genus Bifidobacterium. Long-term (13-39 months after treatment), we did not observe any differences between the azithromycin and placebo recipients in their gut microbiota composition., Interpretation: Azithromycin treatment induced a perturbation in the gut microbiota 14 days after randomization but did not have long-lasting effects on the gut microbiota composition. However, it should be noted that our analyses included a limited number of fecal samples for the placebo treated group at age 4 years. FUND: Lundbeck Foundation, Danish Ministry of Health, Danish Council for Strategic Research, Capital Region Research Foundation, China Scholarship Council., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

Unicellular, eukaryotic organisms (protists) play a key role in soil food webs as major predators of microorganisms. However, due to the polyphyletic nature of protists, no single universal barcode can be established for this group, and the structure of many protistean communities remains unresolved. Plasmodial slime moulds (Myxogastria or Myxomycetes) stand out among protists by their formation of fruit bodies, which allow for a morphological species concept. By Sanger sequencing of a large collection of morphospecies, this study presents the largest database to date of dark-spored myxomycetes and evaluate a partial 18S SSU gene marker for species annotation. We identify and discuss the use of an intraspecific sequence similarity threshold of 99.1% for species differentiation (OTU picking) in environmental PCR studies (ePCR) and estimate a hidden diversity of putative species, exceeding those of described morphospecies by 99%. When applying the identified threshold to an ePCR data set (including sequences from both NGS and cloning), we find 64 OTUs of which 21.9% had a direct match (>99.1% similarity) to the database and the remaining had on average 90.2 ± 0.8% similarity to their best match, thus thought to represent undiscovered diversity of dark-spored myxomycetes., (© 2017 John Wiley & Sons Ltd.)

Extreme habitats are not only limited to natural environments, but also exist in manmade systems, for instance, household appliances such as dishwashers. Limiting factors, such as high temperatures, high and low pHs, high NaCl concentrations, presence of detergents, and shear force from water during washing cycles, define microbial survival in this extreme system. Fungal and bacterial diversity in biofilms isolated from rubber seals of 24 different household dishwashers was investigated using next-generation sequencing. Bacterial genera such as Pseudomonas , Escherichia , and Acinetobacter , known to include opportunistic pathogens, were represented in most samples. The most frequently encountered fungal genera in these samples belonged to Candida , Cryptococcus , and Rhodotorula , also known to include opportunistic pathogenic representatives. This study showed how specific conditions of the dishwashers impact the abundance of microbial groups and investigated the interkingdom and intrakingdom interactions that shape these biofilms. The age, usage frequency, and hardness of incoming tap water of dishwashers had significant impact on bacterial and fungal community compositions. Representatives of Candida spp. were found at the highest prevalence (100%) in all dishwashers and are assumed to be one of the first colonizers in recently purchased dishwashers. Pairwise correlations in tested microbiomes showed that certain bacterial groups cooccur, as did the fungal groups. In mixed bacterial-fungal biofilms, early adhesion, contact, and interactions were vital in the process of biofilm formation, where mixed complexes of bacteria and fungi could provide a preliminary biogenic structure for the establishment of these biofilms. IMPORTANCE Worldwide demand for household appliances, such as dishwashers and washing machines, is increasing, as is the number of immunocompromised individuals. The harsh conditions in household dishwashers should prevent the growth of most microorganisms. However, our research shows that persisting polyextremotolerant groups of microorganisms in household appliances are well established under these unfavorable conditions and supported by the biofilm mode of growth. The significance of our research is in identifying the microbial composition of biofilms formed on dishwasher rubber seals, how diverse abiotic conditions affect microbiota, and which key microbial members were represented in early colonization and contamination of dishwashers, as these appliances can present a source of domestic cross-contamination that leads to broader medical impacts., (Copyright © 2018 Raghupathi et al.)

The originally published version of this Article contained an incorrect version of Figure 3 that was introduced following peer review and inadvertently not corrected during the production process. Both versions contain the same set of abundance data, but the incorrect version has the children's asthma status erroneously disconnected from the abundance data, thereby producing the non-representative p values and graphic presentations. These errors have now been rectified, with the correct version of Figure 3 replaced in both the PDF and HTML versions of the Article.

The composition of the human gut microbiome matures within the first years of life. It has been hypothesized that microbial compositions in this period can cause immune dysregulations and potentially cause asthma. Here we show, by associating gut microbial composition from 16S rRNA gene amplicon sequencing during the first year of life with subsequent risk of asthma in 690 participants, that 1-year-old children with an immature microbial composition have an increased risk of asthma at age 5 years. This association is only apparent among children born to asthmatic mothers, suggesting that lacking microbial stimulation during the first year of life can trigger their inherited asthma risk. Conversely, adequate maturation of the gut microbiome in this period may protect these pre-disposed children.

Background: The airways of healthy humans harbor a distinct microbial community. Perturbations in the microbial community have been associated with disease, yet little is known about the formation and development of a healthy airway microbiota in early life. Our goal was to understand the establishment of the airway microbiota within the first 3 months of life. We investigated the hypopharyngeal microbiota in the unselected COPSAC 2010 cohort of 700 infants, using 16S rRNA gene sequencing of hypopharyngeal aspirates from 1 week, 1 month, and 3 months of age., Results: Our analysis shows that majority of the hypopharyngeal microbiota of healthy infants belong to each individual's core microbiota and we demonstrate five distinct community pneumotypes. Four of these pneumotypes are dominated by the genera Staphylococcus, Streptococcus, Moraxella, and Corynebacterium, respectively. Furthermore, we show temporal pneumotype changes suggesting a rapid development towards maturation of the hypopharyngeal microbiota and a significant effect from older siblings. Despite an overall common trajectory towards maturation, individual infants' microbiota are more similar to their own, than to others, over time., Conclusions: Our findings demonstrate a consolidation of the population of indigenous bacteria in healthy airways and indicate distinct trajectories in the early development of the hypopharyngeal microbiota.