Project description:Development of the gut microbiota is greatly impacted in preterm infants. Despite increasing knowledge about microbiota composition in preterm infants, knowledge about the functional signatures of the intestinal microbiota remains limited. The aim was to study transitions in microbiota activity during the first six postnatal weeks in ten preterm infants. A total of 64 stool samples were measured by LC-MS/MS.
Project description:Development of the gut microbiota is greatly impacted in preterm infants. Despite increasing knowledge about microbiota composition in preterm infants, knowledge about the functional signatures of the intestinal microbiota remains limited. The aim was to study transitions in microbiota activity during the first six postnatal weeks in ten preterm infants. A total of 64 stool samples were measured by LC-MS/MS.
Project description:91 preterm infant gut metaproteomes measured in technical duplicate using an eleven salt pulse 2D-LC-MS/MS method. Samples represent 17 preterm infants over the first several weeks of life, of which 6 preterm infants eventually developed necrotizing enterocolitis.
Project description:Infants are vulnerable to disseminated forms of tuberculosis and suffer disproportionately high morbidity and mortality, but the reasons for this are unknown. We hypothesized that since alveolar macrophages (AMs) are critical in the uptake and containment of Mycobacterium tuberculosis (Mtb) in the lung, their function may be impaired in early life. We developed a method of obtaining AMs during rigid bronchoscopy of healthy infants with suspected airway abnormality. RNAseq analysis of Mtb-stimulated AMs from 4 infants and 4 adults was performed.
Project description:A metaproteomics analysis was conducted on the infant fecal microbiome to characterize global protein expression in 8 samples obtained from infants with a range of early-life experiences. Samples included breast-, formula- or mixed-fed, mode of delivery, and antibiotic treatment and one set of monozygotic twins. Although label-free mass spectrometry-based proteomics is routinely used for the identification and quantification of thousands of proteins in complex samples, the metaproteomic analysis of the gut microbiome presents particular technical challenges. Among them: the extreme complexity and dynamic range of member taxa/species, the need for matched, well-annotated metagenomics databases, and the high inter-protein sequence redundancy/similarity between related members. In this study, a metaproteomic approach was developed for assessment of the biological phenotype and functioning, as a complement to 16S rRNA sequencing analysis to identify constituent taxa. A sample preparation method was developed for recovery and lysis of bacterial cells, followed by trypsin digestion, and pre-fractionation using Strong Cation Exchange chromatography. Samples were then subjected to high performance LC-MS/MS. Data was searched against the Human Microbiome Project database, and a homology-based meta-clustering strategy was used to combine peptides from multiple species into representative proteins. Bacterial taxonomies were also identified, based on species-specific protein sequences, and protein metaclusters were assigned to pathways and functional groups. The results obtained demonstrate the applicability of this approach for performing qualitative comparisons of human fecal microbiome composition, physiology and metabolism, and also provided a more detailed assessment of microbial composition in comparison to 16S rRNA.
Project description:Introduction: Thymic Stromal Lymphopoietin (TSLP) is a primarily epithelial-derived cytokine that drives type 2 allergic immune responses. Early life viral respiratory infections elicit high TSLP production, which leads to the development of type 2 inflammation and airway hyperreactivity. The goal of this study was to examine in vivo and in vitro the human airway epithelial responses leading to high TSLP production during viral respiratory infections in early infancy. Methods: A total of 129 infants (<1m – 24m, median age 10m) with severe viral respiratory infections were enrolled for in vivo (n=113), and in vitro studies (n=16). Infants were classified as “high TSLP” or “low TSLP” for values above or below the 50th percentile. High vs. low TSLP groups were compared in terms of type I-III IFN responses and production of chemokines promoting antiviral (CXCL10), neutrophilic (CXCL1, CXCL5, CXCL8), and type 2 responses (CCL11, CCL17, CCL22). Human infant airway epithelial cell (AEC) cultures were used to define the transcriptomic (RNAseq) profile leading to high vs. low TSLP responses in vitro. Results: Infants in the high TSLP group had greater in vivo type III IFN airway production (median type III IFN in high TSLP 183.2 pg/ml vs. 63.4 pg/ml in low TSLP group, p= 0.007) and increased in vitro type I-III IFN AEC responses after stimulation with a viral mimic (poly I:C). Our RNAseq data showed that infants in the high TSLP group had significant baseline upregulation of IFN signature genes (e.g., IFIT2, IFI6, MX1) and pro-inflammatory chemokine genes prior to viral mimic stimulation. Infants in the high TSLP group also showed a baseline AEC pro-inflammatory state characterized by increased production of all the chemokines assayed (e.g., CXCL10, CXCL8) in the absence of viral stimuli. Conclusion: High TSLP responses in the human infant airways are associated with pre-activated airway epithelial IFN antiviral immunity and increased baseline AEC production of pro-inflammatory chemokines. These findings present a new paradigm underlying the high production of TSLP in the human infant airway epithelium and shed light on the pathogenesis of viral respiratory illnesses during early infancy and beyond childhood.