Metagenomics of sputum
Ontology highlight
ABSTRACT:
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| ERR710436.fastq.gz | Fastqsanger.gz | |||
| ERR710437.fastq.gz | Fastqsanger.gz | |||
| ERR710438.fastq.gz | Fastqsanger.gz | |||
| ERR710439.fastq.gz | Fastqsanger.gz | |||
| ERR710440.fastq.gz | Fastqsanger.gz |
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Project description:Sputum airway metagenomics of people with non-CF bronchiectasis
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2010-11-01 | E-GEOD-24034 | biostudies-arrayexpress
Project description:Pseudomonas aeruginosa airway infection is the leading cause of morbidity and mortality in cystic fibrosis (CF) patients. In vitro models that closely mimic CF sputum are needed to improve understanding of the pathobiology of P. aeruginosa in the CF airway. We developed an artificial sputum medium (ASMDM) that more closely resembles the composition of CF sputum than current media. In order to validate the utility of ASMDM, we used GeneChip microarrays to compare expression data of P. aeruginosa UCBPP-PA14 (PA14) in ASMDM with published data for this strain grown under the same conditions in an artificial medium containing 10% (v/v) CF sputum. Thirty-seven of 39 nutrition-related genes were differentially expressed in the same manner in both media. However, 24 quorum-sensing (QS) genes, 23 Type III secretion system and several anaerobic respiration genes were more highly expressed in ASMDM than in sputum-containing medium. When grown to stationary phase in ASMDM, PA14 differentially expressed about 50 biologically significant genes compared to stationary phase growth in Luria Broth; genes involved in iron acquisition (pfeA, fepC) and in assimilatory nitrate reduction (nasC, nirD) were upregulated, while 24 QS genes, including the regulator rhlR, lasA, rsaL, aprADEI and phenazine genes phzC2DD2EG2 were downregulated. Downregulation of QS-regulated virulence genes has been noted in chronic P. aeruginosa infection. ASMDM thus appears highly suitable for studies on gene expression of (i) P. aeruginosa strains from acutely and chronically infected CF patients and (ii) established biofilms that are a hallmark of advanced CF lung disease.
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Project description:Sputum cells collected before (visit 2) and after (visit 4) allergen challenge in asthma patients were isolated and RNA purified for analysis on gene expression arrays. Human subject recruitment part of NIH sponsored protocol as part of the Eosinophil Program Project Grant (PI: Dr. Nizar Jarjour) Sputum cell RNA collected from induced sputum cells before and 48 hours after whole-lung allergen challenge.
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Project description:Cystic fibrosis (CF) is an inherited, multi-system disease caused by dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR) protein, a ubiquitous ion channel important for epithelial hydration. A direct consequence of this dysfunction is impaired mucociliary clearance, chronic airway infection and a persistent neutrophilic inflammatory response that results in progressive loss of lung function, development of respiratory failure and premature death. Partial restoration of CFTR function is now possible for most CF patients through mutation specific CFTR modulators. Ivacaftor monotherapy produces significant clinical improvement in CF patients with the G511D mutation. Dual therapy, combining ivacaftor with lumacaftor or tezacaftor, results in modest clinical improvements in patients homozygous for F508del. More recently, triple therapy with elexacaftor/tezacaftor/ivacaftor (ETI) has led to dramatic improvements in lung function and quality of life in patients homozygous and heterozygous for F508del. Sputum proteomics is a powerful research technique capable of identifying important airway disease mechanisms by interrogating the proteome, an entire set of proteins within biological samples. It has confirmed the central role of neutrophilic immune dysregulation in CF and non-CF bronchiectasis, particularly involving the release of antimicrobial proteins and neutrophil-extracellular traps (NETs), and through impaired anti-inflammatory mechanisms. These processes produce distinct molecular signatures within the sputum proteome that become increasingly abnormal with chronic airway infection and progressive lung disease severity. In CF patients, airway and systemic inflammatory cytokines potentially related to these signatures reduce with the various forms of CFTR modulation. To date, no studies of ETI therapy in CF lung disease have assessed large-scale change in protein expression using untargeted proteomics. We hypothesised that ETI therapy would shift the sputum proteome toward health, potentially normalising airway biology in people with CF. The objectives of this study were to investigate changes in the CF sputum proteome with the introduction of ETI, correlate these with changes in clinical markers of disease severity, and make comparisons with the sputum proteome in healthy controls and in repeat samples from CF patients not suitable for ETI therapy. We also explored which molecular pathways associated with CF lung disease did not change with ETI.
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Project description:Several microRNAs (miRs) have been described as potential biomarkers in liquid biopsies and in the context of allergic asthma, while therapeutic effects on the airway expression of miRs remain elusive. In this study, we investigated epigenetic miR-associated mechanisms in the sputum of grass pollen allergic patients with and without allergen specific immunotherapy (AIT). Induced sputum samples of healthy controls (HC), AIT treated and untreated grass pollen allergic rhinitis patients with (AA) and without asthma (AR) were profiled using miR microarray and transcriptome microarray analysis of the same samples. miR targets were predicted in silico and used to identify inverse regulation. Local PGE2 levels were measured using ELISA. Two Hundred and fifty nine miRs were upregulated in the sputum of AA patients compared with HC, while only one was downregulated. The inverse picture was observed in induced sputum of AIT-treated patients: while 21 miRs were downregulated, only 4 miRs were upregulated in asthmatics upon AIT. Of these 4 miRs, miR3935 stood out, as its predicted target PTGER3, the prostaglandin EP3 receptor, was downregulated in treated AA patients compared with untreated. The levels of its ligand PGE2 in the sputum supernatants of these samples were increased in allergic patients, especially asthmatics, and downregulated after AIT. Finally, local PGE2 levels correlated with ILC2 frequencies, secreted sputum IL13 levels, inflammatory cell load, sputum eosinophils and symptom burden.While profiling the sputum of allergic patients for novel miR expression patterns, we uncovered an association between miR3935 and its predicted target gene, the prostaglandin E3 receptor, which might mediate AIT effects through suppression of the PGE2-PTGER3 axis.
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Project description:Pseudomonas aeruginosa airway infection is the leading cause of morbidity and mortality in cystic fibrosis (CF) patients. In vitro models that closely mimic CF sputum are needed to improve understanding of the pathobiology of P. aeruginosa in the CF airway. We developed an artificial sputum medium (ASMDM) that more closely resembles the composition of CF sputum than current media. In order to validate the utility of ASMDM, we used GeneChip microarrays to compare expression data of P. aeruginosa UCBPP-PA14 (PA14) in ASMDM with published data for this strain grown under the same conditions in an artificial medium containing 10% (v/v) CF sputum. Thirty-seven of 39 nutrition-related genes were differentially expressed in the same manner in both media. However, 24 quorum-sensing (QS) genes, 23 Type III secretion system and several anaerobic respiration genes were more highly expressed in ASMDM than in sputum-containing medium. When grown to stationary phase in ASMDM, PA14 differentially expressed about 50 biologically significant genes compared to stationary phase growth in Luria Broth; genes involved in iron acquisition (pfeA, fepC) and in assimilatory nitrate reduction (nasC, nirD) were upregulated, while 24 QS genes, including the regulator rhlR, lasA, rsaL, aprADEI and phenazine genes phzC2DD2EG2 were downregulated. Downregulation of QS-regulated virulence genes has been noted in chronic P. aeruginosa infection. ASMDM thus appears highly suitable for studies on gene expression of (i) P. aeruginosa strains from acutely and chronically infected CF patients and (ii) established biofilms that are a hallmark of advanced CF lung disease. PA14 was grown in four different ways: 1. Logarithmic growth for early gene expression Cells were grown in MOPS-Glucose and separately in ASMDM. The average of two biological duplicates in each case was compared to the other to determine differential gene expression. 2. Stationary phase growth for gene expression Cells were grown in Luria Broth and separately in ASMDM. The average of two biological duplicates in each case was compared to the other to determine differential gene expression.
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