Applications of single-cell Raman microspectroscopy in bacteria metabolism and physiology

Thesis

Bacteria play important roles in almost all ecosystems, including the natural environment and the human body, and they are extremely abundant and versatile. New generations of sequencing technologies can provide phylogenetic diversity and metagenomic profiling of microbial communities, but metabolic functions of individual microbes and their molecular mechanisms are often elusive, owing to the limited culturability and the complexity of microbial communities.

Single-cell Raman microspectroscopy (SCRM) is label-free vibrational spectroscopy that measures a collection of molecular vibrations coming from all biomolecules in a single bacterial cell. This thesis emphasises the capabilities of SCRM in a wide spectrum of applications for investigating bacterial metabolism and physiology. SCRM is applied to study bacterial phenotypes in a symbiotic setting with a plant host. SCRM is combined with deuterium-isotope-probing (DIP) and applied to study carbon metabolism in pure cultures, as well as in vivo metabolic adaptations by human-invasive pathogens. Especially, the power of single-cell analysis in studies of heterogeneous bacterial populations is explored and highlighted.

This thesis also outlines an analytical pipeline to standardise and automate the process of spectral analysis in SCRM studies, from data tidying and data preprocessing to a variety of analytical approaches for either exploratory or diagnostic purposes. By following the established pipeline and executing the developed programming script, single-cell Raman analysis can be performed in a simple and rapid manner.

To conclude, by applying SCRM to provide intrinsic biochemical fingerprints of individual bacterial cells and connecting SCRM with the power of computational analysis, this thesis provides the possibility of probing bacterial metabolism and physiological states in ever-changing and multifaceted ecological settings.

Authors: Xu, Jiabao

• DOI: 10.5287/ora-ngebqjndr
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: Single cell; Bacteria metabolism; Raman spectroscopy
• Subjects: Spectroscopy; Microbiology