Prokaryotic lipidomics: development of a method to study the phospholipids in E. coli and B. subtilis

Abstract

The way in which prokaryotes, particularly Gram-negative bacteria, regulate their lipid compositions in response to changes in their growth media can provide insights into how bacteria adapt to life in oligotrophic environments. In this thesis, it is reported how electrospray ionisation mass spectrometry (ESI-MS) techniques can be adapted to map the impact of growth conditions on the regulation of the phospholipid biosynthetic pathways in E. coli and B. subtilis. Reliable counting of bacterial number densities is one of the main barriers to achieving reproducible quantitative lipid composition profiles. Consequently an improved method has been developed, based on an automated cell counter, to characterize the bacterial number densities and cell size for each of the samples used in the ESI-MS experiments. This approach has enabled the observation of unexpected changes in both the relative and absolute amounts of the phospholipid species isolated from bacteria grown under different conditions. These studies have investigated the adaptation of E. coli and B. subtilis to a glucose deplete growth medium. In E. coli the effect of growth in a nutrient rich growth medium, phosphate depletion and CTAB addition was investigated. In B. subtilis initial studies were performed into nitrogen depletion, myristic acid addition and DOPC addition. The changes in phospholipid classes, as well as in the homologues within each class, that we have observed may be related to changes in the fluxes within the lipid biosynthetic pathway of the organism. The expectation is that studying the changes in growth conditions using E. coli and B. subtilis as a model organism could help with the understanding of biofilm formation and help towards a greater understanding of bacterial adaptation to stressful environments.

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ORCID for George S. Attard: orcid.org/0000-0001-8304-0742

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