Abstract
The anaerobic bacteria of the Bacteroides fragilis group including Bacteroides thetaiotaomicron, B. fragilis, Bacteroides vulgatus, and Bacteroides ovatus in genus Bacteroides are among the most commonly found human gut microbiota. They are generally commensal but are also opportunistic pathogens. Both the inner and outer membranes of the Bacteroides cell envelope contain abundant lipids with diversified structures, and dissection of the lipid composition of the inner and outer membrane fractions is important for understanding the biogenesis of this multilaminate wall structure. Here, we describe mass spectrometry-based approaches to delineate in detail the lipidome of the membrane and the outer membrane vesicle of the bacteria cells. We identified 15 lipid class/subclasses (>100 molecular species), including sphingolipid families [dihydroceramide (DHC), glycylseryl (GS) DHC, DHC-phosphoinositolphosphoryl-DHC (DHC-PIP-DHC), ethanolamine phosphorylceramide, inositol phosphorylceramide (IPC), serine phosphorylceramide, ceramide-1-phosphate, and glycosyl ceramide], phospholipids [phosphatidylethanolamine, phosphatidylinositol (PI), and phosphatidylserine], peptide lipids (GS-, S-, and G-lipids) and cholesterol sulfate, of which several have not been reported previously, or have similar structures to those found in Porphyromonas gingivalis, the periodontopathic bacterium in oral microbiota. The new DHC-PIPs-DHC lipid family is found only in B. vulgatus, which, however, lacks the PI lipid family. The galactosyl ceramide family is exclusively present in B. fragilis, which nevertheless lacks IPC and PI lipids. The lipidomes as revealed in this study demonstrate the lipid diversity among the various strains and the utility of multiple-stage mass spectrometry (MS(n)) with high-resolution mass spectrometry in the structural elucidation of complex lipids.