Persistence of birth mode-dependent effects on gut microbiome composition, immune system stimulation and antimicrobial resistance during the first year of life.

Caesarean section delivery (CSD) disrupts mother-to-neonate transmission of specific microbial strains and functional repertoires as well as linked immune system priming. Here we investigate whether differences in microbiome composition and impacts on host physiology persist at 1 year of age. We perform high-resolution, quantitative metagenomic analyses of the gut microbiomes of infants born by vaginal delivery (VD) or by CSD, from immediately after birth through to 1 year of life. Several microbial populations show distinct enrichments in CSD-born infants at 1 year of age including strains of Bacteroides caccae, Bifidobacterium bifidum and Ruminococcus gnavus, whereas others are present at higher levels in the VD group including Faecalibacterium prausnitizii, Bifidobacterium breve and Bifidobacterium kashiwanohense. The stimulation of healthy donor-derived primary human immune cells with LPS isolated from neonatal stool samples results in higher levels of tumour necrosis factor alpha (TNF-α) in the case of CSD extracts over time, compared to extracts from VD infants for which no such changes were observed during the first year of life. Functional analyses of the VD metagenomes at 1 year of age demonstrate a significant increase in the biosynthesis of the natural antibiotics, carbapenem and phenazine. Concurrently, we find antimicrobial resistance (AMR) genes against several classes of antibiotics in both VD and CSD. The abundance of AMR genes against synthetic (including semi-synthetic) agents such as phenicol, pleuromutilin and diaminopyrimidine are increased in CSD children at day 5 after birth. In addition, we find that mobile genetic elements, including phages, encode AMR genes such as glycopeptide, diaminopyrimidine and multidrug resistance genes. Our results demonstrate persistent effects at 1 year of life resulting from birth mode-dependent differences in earliest gut microbiome colonisation.

Birth mode is associated with earliest strain-conferred gut microbiome functions and immunostimulatory potential.

The rate of caesarean section delivery (CSD) is increasing worldwide. It remains unclear whether disruption of mother-to-neonate transmission of microbiota through CSD occurs and whether it affects human physiology. Here we perform metagenomic analysis of earliest gut microbial community structures and functions. We identify differences in encoded functions between microbiomes of vaginally delivered (VD) and CSD neonates. Several functional pathways are over-represented in VD neonates, including lipopolysaccharide (LPS) biosynthesis. We link these enriched functions to individual-specific strains, which are transmitted from mothers to neonates in case of VD. The stimulation of primary human immune cells with LPS isolated from early stool samples of VD neonates results in higher levels of tumour necrosis factor (TNF-α) and interleukin 18 (IL-18). Accordingly, the observed levels of TNF-α and IL-18 in neonatal blood plasma are higher after VD. Taken together, our results support that CSD disrupts mother-to-neonate transmission of specific microbial strains, linked functional repertoires and immune-stimulatory potential during a critical window for neonatal immune system priming.

Sequential Isolation of DNA, RNA, Protein, and Metabolite Fractions from Murine Organs and Intestinal Contents for Integrated Omics of Host-Microbiota Interactions.

The gastrointestinal microbiome plays a central role in health and disease. Imbalances in the microbiome, also referred to as dysbiosis, have recently been associated with a number of human idiopathic diseases ranging from metabolic to neurodegenerative. However, to causally link specific microorganisms or dysbiotic communities with tissue-specific and/or systemic disease-associated phenotypes, systematic in vivo studies are fundamental. Gnotobiotic mouse models have proven to be particularly useful for the elucidation of microbiota-associated characteristics as they provide a means to conduct targeted perturbations followed by analyses of induced localized and systemic effects. Here, we describe a methodology in the framework of systems biology which allows the comprehensive isolation of high quality biomolecular fractions (DNA, RNA, proteins and metabolites) from limited and/or heterogeneous sample material derived from murine brain, liver, and colon tissues, as well as from intestinal contents (fecal pellets and fecal masses). The obtained biomolecular fractions are compatible with current high-throughput genomic, transcriptomic, proteomic, and metabolomic analyses. The resulting data fulfills the premise of systematic measurements and allows the detailed study of tissue-specific and/or systemic effects of host-microbiota interactions in relation to health and disease.

Integrated meta-omic analyses of the gastrointestinal tract microbiome in patients undergoing allogeneic hematopoietic stem cell transplantation.

In patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT), treatment-induced changes to the gastrointestinal tract (GIT) microbiome have been linked to adverse outcomes, most notably graft-versus-host disease (GvHD). However, it is presently unknown whether this relationship is causal or consequential. Here, we performed an integrated meta-omic analysis to probe deeper into the GIT microbiome changes during allo-HSCT and its accompanying treatments. We used 16S and 18S rRNA gene amplicon sequencing to resolve archaea, bacteria, and eukaryotes within the GIT microbiomes of 16 patients undergoing allo-HSCT for the treatment of hematologic malignancies. These results revealed a major shift in the GIT microbiome after allo-HSCT including a marked reduction in bacterial diversity, accompanied by only limited changes in eukaryotes and archaea. An integrated analysis of metagenomic and metatranscriptomic data was performed on samples collected from a patient before and after allo-HSCT for acute myeloid leukemia. This patient developed severe GvHD, leading to death 9 months after allo-HSCT. In addition to drastically decreased bacterial diversity, the post-treatment microbiome showed a higher overall number and higher expression levels of antibiotic resistance genes (ARGs). One specific Escherichia coli strain causing a paravertebral abscess was linked to GIT dysbiosis, suggesting loss of intestinal barrier integrity. The apparent selection for bacteria expressing ARGs suggests that prophylactic antibiotic administration may adversely affect the overall treatment outcome. We therefore assert that such analyses including information about the selection of pathogenic bacteria expressing ARGs may assist clinicians in "personalizing" regimens for individual patients to improve overall outcomes.

Colonization and Succession within the Human Gut Microbiome by Archaea, Bacteria, and Microeukaryotes during the First Year of Life.

Perturbations to the colonization process of the human gastrointestinal tract have been suggested to result in adverse health effects later in life. Although much research has been performed on bacterial colonization and succession, much less is known about the other two domains of life, archaea, and eukaryotes. Here we describe colonization and succession by bacteria, archaea and microeukaryotes during the first year of life (samples collected around days 1, 3, 5, 28, 150, and 365) within the gastrointestinal tract of infants delivered either vaginally or by cesarean section and using a combination of quantitative real-time PCR as well as 16S and 18S rRNA gene amplicon sequencing. Sequences from organisms belonging to all three domains of life were detectable in all of the collected meconium samples. The microeukaryotic community composition fluctuated strongly over time and early diversification was delayed in infants receiving formula milk. Cesarean section-delivered (CSD) infants experienced a delay in colonization and succession, which was observed for all three domains of life. Shifts in prokaryotic succession in CSD infants compared to vaginally delivered (VD) infants were apparent as early as days 3 and 5, which were characterized by increased relative abundances of the genera Streptococcus and Staphylococcus, and a decrease in relative abundance for the genera Bifidobacterium and Bacteroides. Generally, a depletion in Bacteroidetes was detected as early as day 5 postpartum in CSD infants, causing a significantly increased Firmicutes/Bacteroidetes ratio between days 5 and 150 when compared to VD infants. Although the delivery mode appeared to have the strongest influence on differences between the infants, other factors such as a younger gestational age or maternal antibiotics intake likely contributed to the observed patterns as well. Our findings complement previous observations of a delay in colonization and succession of CSD infants, which affects not only bacteria but also archaea and microeukaryotes. This further highlights the need for resolving bacterial, archaeal, and microeukaryotic dynamics in future longitudinal studies of microbial colonization and succession within the neonatal gastrointestinal tract.

Integrated multi-omics of the human gut microbiome in a case study of familial type 1 diabetes.

The gastrointestinal microbiome is a complex ecosystem with functions that shape human health. Studying the relationship between taxonomic alterations and functional repercussions linked to disease remains challenging. Here, we present an integrative approach to resolve the taxonomic and functional attributes of gastrointestinal microbiota at the metagenomic, metatranscriptomic and metaproteomic levels. We apply our methods to samples from four families with multiple cases of type 1 diabetes mellitus (T1DM). Analysis of intra- and inter-individual variation demonstrates that family membership has a pronounced effect on the structural and functional composition of the gastrointestinal microbiome. In the context of T1DM, consistent taxonomic differences were absent across families, but certain human exocrine pancreatic proteins were found at lower levels. The associated microbial functional signatures were linked to metabolic traits in distinct taxa. The methodologies and results provide a foundation for future large-scale integrated multi-omic analyses of the gastrointestinal microbiome in the context of host-microbe interactions in human health and disease.

Sources and Functions of Extracellular Small RNAs in Human Circulation.

Various biotypes of endogenous small RNAs (sRNAs) have been detected in human circulation, including microRNAs, transfer RNAs, ribosomal RNA, and yRNA fragments. These extracellular sRNAs (ex-sRNAs) are packaged and secreted by many different cell types. Ex-sRNAs exhibit differences in abundance in several disease states and have, therefore, been proposed for use as effective biomarkers. Furthermore, exosome-borne ex-sRNAs have been reported to elicit physiological responses in acceptor cells. Exogenous ex-sRNAs derived from diet (most prominently from plants) and microorganisms have also been reported in human blood. Essential issues that remain to be conclusively addressed concern the (a) presence and sources of exogenous ex-sRNAs in human bodily fluids, (b) detection and measurement of ex-sRNAs in human circulation, (c) selectivity of ex-sRNA export and import, (d) sensitivity and specificity of ex-sRNA delivery to cellular targets, and (e) cell-, tissue-, organ-, and organism-wide impacts of ex-sRNA-mediated cell-to-cell communication. We survey the present state of knowledge of most of these issues in this review.

Community-integrated omics links dominance of a microbial generalist to fine-tuned resource usage.

Microbial communities are complex and dynamic systems that are primarily structured according to their members' ecological niches. To investigate how niche breadth (generalist versus specialist lifestyle strategies) relates to ecological success, we develop and apply an integrative workflow for the multi-omic analysis of oleaginous mixed microbial communities from a biological wastewater treatment plant. Time- and space-resolved coupled metabolomic and taxonomic analyses demonstrate that the community-wide lipid accumulation phenotype is associated with the dominance of the generalist bacterium Candidatus Microthrix spp. By integrating population-level genomic reconstructions (reflecting fundamental niches) with transcriptomic and proteomic data (realised niches), we identify finely tuned gene expression governing resource usage by Candidatus Microthrix parvicella over time. Moreover, our results indicate that the fluctuating environmental conditions constrain the accumulation of genetic variation in Candidatus Microthrix parvicella likely due to fitness trade-offs. Based on our observations, niche breadth has to be considered as an important factor for understanding the evolutionary processes governing (microbial) population sizes and structures in situ.