The pathogenic yeast Candida albicans has the ability to synthesize unique sequences of beta-1,2-oligomannosides that act as adhesins, induce cytokine production, and generate protective antibodies. Depending on the growth conditions, beta-1,2-oligomannosides are associated with different carrier molecules in the cell wall. Structural evidence has been obtained for the presence of these residues in the polysaccharide moiety of the glycolipid, phospholipomannan (PLM). In this study, the refinement of purification techniques led to large quantities of PLM being extracted from Candida albicans cells. A combination of methanolysis, gas chromatography, mass spectrometry, and nuclear magnetic resonance analyses allowed the complete structure of PLM to be deduced. The lipid moiety was shown to consist of a phytoceramide associating a C(18)/C(20) phytosphingosine and C(25), C(26), or mainly C(24) hydroxy fatty acids. The spacer linking the glycan part was identified as a unique structure: -Man-P-Man-Ins-P-. Therefore, in contrast to the major class of membranous glycosphingolipids represented by mannose diinositol phosphoceramide, which is derived from mannose inositol phosphoceramide by the addition of inositol phosphate, PLM seems to be derived from mannose inositol phosphoceramide by the addition of mannose phosphate. In relation to a previous study of the glycan part of the molecule, the assignment of the second phosphorus position leads to the definition of PLM beta-1,2-oligomannosides as unbranched linear structures that may reach up to 19 residues in length. Therefore, PLM appears to be a new type of glycosphingolipid, which is glycosylated extensively through a unique spacer. The conferred hydrophilic properties allow PLM to diffuse into the cell wall in which together with mannan it presents C. albicans beta-1,2-oligomannosides to host cells.