Improving the management of sepsis patients through rapid metagenomic pathogen and antibiotic resistance detection in blood

Moragues Solanas, Lluis (2023) Improving the management of sepsis patients through rapid metagenomic pathogen and antibiotic resistance detection in blood. Doctoral thesis, University of East Anglia.

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Metagenomics is a promising approach to improve the diagnosis of bloodstream infections (BSI) by offering precise and sensitive genomic information on the microbial presence in blood. Long-read sequencing technologies have successfully supported the use of clinical metagenomics for the diagnosis of conditions such as respiratory illness. However, their use in BSI diagnosis has proven more challenging because of the high levels of host DNA and the low number of BSI-causing microorganisms in blood samples. In this study, several methodological steps were developed and optimised to achieve the generation of a metagenomics pipeline with optimal sensitivity. These included the optimisation of a saponin-based host DNA depletion step and a whole genome amplification method, and the creation of a bioinformatics pipeline to analyse the nanopore sequencing data. The implementation of these steps resulted in a sensitive metagenomics pipeline that could detect, in proof-of-concept experiments, E. coli and S. aureus spiked at clinically relevant concentrations in under 9 hours. However, when the same pipeline was tested on clinical blood samples, its sensitivity was lower compared to results obtained with standard culture-based results. Consequently, an improved metagenomics pipeline was developed through the implementation of a rapid bacterial enrichment step, resulting in a 12-hour protocol with increased sensitivity. In proof-of-concept experiments, the improved metagenomics pipeline allowed the identification of the four most prevalent BSI causative species (and their antimicrobial resistance determinants) when these were spiked at concentrations as low as 1 CFU/mL in blood. The testing of this pipeline on clinical blood samples from BSI patients will confirm its suitability for diagnosis.
Overall, this study has generated a clinical metagenomics pipeline that provides high levels of information in reduced times compared to standard blood cultures, having the potential to be used in the clinics and bring important advances in the BSI diagnosis field.

Item Type: Thesis (Doctoral)
Faculty \ School: Faculty of Medicine and Health Sciences > Norwich Medical School
Depositing User: Nicola Veasy
Date Deposited: 29 Jan 2024 14:46
Last Modified: 29 Jan 2024 14:46

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