Revealing the Fate of Isomeric Monounsaturated Fatty Acids in Enterococcus faecalis Membrane Lipids and Their...

Previous work suggests that Enterococcus faecalis uses exogenous oleic acid (FA 18:1(9z)) to increase its tolerance against the membrane-targeting antimicrobial, daptomycin. However, the specific roles of OA and the endogenous positional isomer, cis-vaccenic acid (FA 18:1(11z)), in both daptomycin-susceptible (Dap-S) and daptomycin-resistant (Dap-R) strains have not been fully explored. We performed lipidomics using hydrophilic interaction liquid chromatography and reversed-phase liquid chromatography-ion mobility-mass spectrometry to identify alterations in the lipid composition of Dap-S (S613) and Dap-R (R712) strain pairs of E. faecalis following OA or CV supplementation. Lipidomics showed that total PG intensity in Dap-S was only impacted by OA, while total DGDG intensity was impacted by both OA and CV. However, total PG and DGDG intensities in Dap-R were not impacted by either FA. Both strains produced significantly more lipids containing two 18:1 acyl tails following OA supplementation, while CV had a similar effect on Dap-S only, indicating that Dap-S and Dap-R respond to OA and CV differently. Additionally, the preferences for OA vs CV, alone and in mixtures, were distinguished using ozonolysis and deuterium-labeling techniques that can resolve C═C positional isomers. Growth and survival assays show that Dap-S responds differently to daptomycin when it is cultured with OA or CV. Together, these results reveal that E. faecalis strains with genetic determinants for daptomycin resistance respond differently, in membrane lipid composition and overall growth, to biologically relevant FA 18:1 isomers compared to strains that acquire daptomycin tolerance solely through exogenous fatty acid uptake.