The role of metagenomics as novel sequencing-based tools in analysing the virome of patients with severe community-acquired pneumonia
File(s)
Author(s)
Ainley, Adam Denis Jeremy
Type
Thesis
Abstract
Abstract
The role of metagenomics as novel sequencing-based tools in obtaining a virological diagnosis and analysing the underlying virome of patients with severe community-acquired pneumonia.
Aims:
To develop a metagenomic sequencing (MGS) workflow with the ability to detect viral pathogens in bronchoalveolar lavage samples from patients with severe community-acquired pneumonia (SCAP) and improve understanding of the respiratory virome.
Methods:
In parallel to developing a new MGS workflow, a protocol for a multi-centre study was devised whilst establishing a network of clinical centres to undertake clinical research. Ethical approval was sought and once approved led to the undertaking of The SCAP study, a multicentre clinical trial established to recruit patients with SCAP. In addition to recruiting patients from one of the 5 commissioned ECMO centres who formed part of the established network, patients were also recruited from intensive care units and a dedicated research bronchoscopy service used to sample healthy volunteers. During the recruitment stage, an evaluation and refinement of a metagenomic protocol was undertaken using a novel reference reagent. The sequencing data generated was used to establish and evaluate a bioinformatic pipeline to compliment the laboratory processes developed and a pilot study undertaken to assess the workflows detection ability.
Results:
101 patients were successfully recruited over 3 seasons and an invaluable biobank of upper and lower respiratory tract samples was curated alongside a wealth of clinical data and additional complimentary clinical samples. When applied to the BAL samples collected, MGS was able to detect a virus in 30% of subjects and showed comparable specificity and concordance to conventional diagnostics and an extended suite of molecular assays although a limit of detection was noted.
Conclusion:
The SCAP study was one of the first studies to explore the use MGS in patients with SCAP from a virological perspective and generated to date, one of the largest collections of BAL samples for MGS analysis and undertook one of few existing comparisons to a healthy cohort.
The role of metagenomics as novel sequencing-based tools in obtaining a virological diagnosis and analysing the underlying virome of patients with severe community-acquired pneumonia.
Aims:
To develop a metagenomic sequencing (MGS) workflow with the ability to detect viral pathogens in bronchoalveolar lavage samples from patients with severe community-acquired pneumonia (SCAP) and improve understanding of the respiratory virome.
Methods:
In parallel to developing a new MGS workflow, a protocol for a multi-centre study was devised whilst establishing a network of clinical centres to undertake clinical research. Ethical approval was sought and once approved led to the undertaking of The SCAP study, a multicentre clinical trial established to recruit patients with SCAP. In addition to recruiting patients from one of the 5 commissioned ECMO centres who formed part of the established network, patients were also recruited from intensive care units and a dedicated research bronchoscopy service used to sample healthy volunteers. During the recruitment stage, an evaluation and refinement of a metagenomic protocol was undertaken using a novel reference reagent. The sequencing data generated was used to establish and evaluate a bioinformatic pipeline to compliment the laboratory processes developed and a pilot study undertaken to assess the workflows detection ability.
Results:
101 patients were successfully recruited over 3 seasons and an invaluable biobank of upper and lower respiratory tract samples was curated alongside a wealth of clinical data and additional complimentary clinical samples. When applied to the BAL samples collected, MGS was able to detect a virus in 30% of subjects and showed comparable specificity and concordance to conventional diagnostics and an extended suite of molecular assays although a limit of detection was noted.
Conclusion:
The SCAP study was one of the first studies to explore the use MGS in patients with SCAP from a virological perspective and generated to date, one of the largest collections of BAL samples for MGS analysis and undertook one of few existing comparisons to a healthy cohort.
Version
Open Access
Date Issued
2022-06
Date Awarded
2023-08
Copyright Statement
Creative Commons Attribution NonCommercial NoDerivatives Licence
Advisor
Zambon, Maria
Kon, Onn Min
Openshaw, Peter
Sponsor
National Institute for Health Research (Great Britain)
Publisher Department
National Heart & Lung Institute
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)