Topographic heterogeneity of lung microbiota in end-stage idiopathic pulmonary fibrosis: the Microbiome in Lung Explants-2 (MiLEs-2) study.

BACKGROUND: Lung microbiota profiles in patients with early idiopathic pulmonary fibrosis (IPF) have been associated with disease progression; however, the topographic heterogeneity of lung microbiota and their roles in advanced IPF are unknown. METHODS: We performed a retrospective, case-control study of explanted lung tissue obtained at the time of lung transplantation or rapid autopsy from patients with IPF and other chronic lung diseases (connective tissue disease-associated interstitial lung disease (CTD-ILD), cystic fibrosis (CF), COPD and donor lungs unsuitable for transplant from Center for Organ Recovery and Education (CORE)). We sampled subpleural tissue and airway-based specimens (bronchial washings and airway tissue) and quantified bacterial load and profiled communities by amplification and sequencing of the 16S rRNA gene. FINDINGS: Explants from 62 patients with IPF, 15 patients with CTD-ILD, 20 patients with CF, 20 patients with COPD and 20 CORE patients were included. Airway-based samples had higher bacterial load compared with distal parenchymal tissue. IPF basilar tissue had much lower bacterial load compared with CF and CORE lungs (p<0.001). No microbial community differences were found between parenchymal tissue samples from different IPF lobes. Dirichlet multinomial models revealed an IPF cluster (29%) with distinct composition, high bacterial load and low alpha diversity, exhibiting higher odds for acute exacerbation or death. INTERPRETATION: IPF explants had low biomass in the distal parenchyma of all three lobes with higher bacterial load in the airways. The discovery of a distinct subgroup of patients with IPF with higher bacterial load and worse clinical outcomes supports investigation of personalised medicine approaches for microbiome-targeted interventions.

The Dynamics of Interacting Bacterial and Fungal Communities of the Mouse Colon Following Antibiotics.

We tested two hypotheses concerning the dynamics of intestinal microbial communities of young mice following antibiotic-induced disturbance. The first is that disturbance of the bacterial community causes disturbance of the fungal community. Our results were consistent with that hypothesis. Antibiotics significantly altered bacterial community structure. Antibiotics also altered fungal community structure, significantly increasing the relative abundance of Candida lusitaniae, a known pathogen, while simultaneously significantly decreasing the relative abundances of several other common fungal species. The result was a temporary decrease in fungal diversity. Moreover, bacterial load was negatively correlated with the relative abundances of Candida lusitaniae and Candida parapsilosis, while it was positively correlated with the relative abundances of many other fungal species. Our second hypothesis is that control mice serve as a source of probiotics capable of invading intestines of mice with disturbed microbial communities and restoring pre-antibiotic bacterial and fungal communities. However, we found that control mice did not restore disturbed microbial communities. Instead, mice with disturbed microbial communities induced disturbance in control mice, consistent with the hypothesis that antibiotic-induced disturbance represents an alternate stable state that is easier to achieve than to correct. Our results indicate the occurrence of significant interactions among intestinal bacteria and fungi and suggest that the stimulation of certain bacterial groups may potentially be useful in countering the dominance of fungal pathogens such as Candida spp. However, the stability of disturbed microbial communities could complicate recovery.

Metagenomic identification of severe pneumonia pathogens in mechanically-ventilated patients: a feasibility and clinical validity study.

BACKGROUND: Metagenomic sequencing of respiratory microbial communities for pathogen identification in pneumonia may help overcome the limitations of culture-based methods. We examined the feasibility and clinical validity of rapid-turnaround metagenomics with Nanopore™ sequencing of clinical respiratory specimens. METHODS: We conducted a case-control study of mechanically-ventilated patients with pneumonia (nine culture-positive and five culture-negative) and without pneumonia (eight controls). We collected endotracheal aspirates and applied a microbial DNA enrichment method prior to metagenomic sequencing with the Oxford Nanopore MinION device. For reference, we compared Nanopore results against clinical microbiologic cultures and bacterial 16S rRNA gene sequencing. RESULTS: Human DNA depletion enabled in depth sequencing of microbial communities. In culture-positive cases, Nanopore revealed communities with high abundance of the bacterial or fungal species isolated by cultures. In four cases with resistant clinical isolates, Nanopore detected antibiotic resistance genes corresponding to the phenotypic resistance in antibiograms. In culture-negative pneumonia, Nanopore revealed probable bacterial pathogens in 1/5 cases and Candida colonization in 3/5 cases. In controls, Nanopore showed high abundance of oral bacteria in 5/8 subjects, and identified colonizing respiratory pathogens in other subjects. Nanopore and 16S sequencing showed excellent concordance for the most abundant bacterial taxa. CONCLUSIONS: We demonstrated technical feasibility and proof-of-concept clinical validity of Nanopore metagenomics for severe pneumonia diagnosis, with striking concordance with positive microbiologic cultures, and clinically actionable information obtained from sequencing in culture-negative samples. Prospective studies with real-time metagenomics are warranted to examine the impact on antimicrobial decision-making and clinical outcomes.