Leptospira express mainly di- N -acetylated nonulosonic acids. Nonulosonic acids were released from Leptospira isolates and fluorescently derivatized with DMB followed by HPLC as described in Materials and Methods. Selected peaks were subjected to electrospray ionization mass spectrometry. Pse and Leg refer to the di-N-acetylated nonulosonic acids pseudaminic and legionaminic acids, closely related isomers with an identical DMB-derivatized mass of 451. Kdo is a related eight-carbon backbone monosaccharide common to the core region of lipopolysaccharide. All MS data are shown from 400–500 m/z, except for representative MS data shown for peak b (Kdo), shown from 300–400 m/z. Each of these strains was analyzed in 2–3 independent experiments with similar results.