Identification of clinical and environmental isolates of Legionella pneumophila by analysis of outer-membrane proteins, ubiquinones and fatty acids

Abstract

47 strains of L. pneumophila, 9 reference strains of eight serogroups, 15 clinical isolates and 23 water-derived strains from different cities were analysed with respect to three chemical constituents of the cell envelope: outer- membrane proteins (OMPs), ubiquinones and fatty acids. The OMPs were obtained by Sarkosyl-extraction of a total membrane preparation and analysed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PA-GE). The ubiquinone content was quantitated by high-performance liquid chromatography (HPLC) and the fatty acid composition was established by gas-liquid chromatography coupled with mass spectrometry (GLC-MS). In all strains the OMP yielded the known characteristic pattern with the major component of 29 kilodalton. The ubiquinone content was very consistent within different strains revealing Q-12 with 65% as the predominant feature. Q-11 and Q-13 were present in amounts of 11% and 23%, respectively. Q-14 was found in trace amounts of about 0.65%. Only a very few ubiquinones with side chains shorter than 11 residues were present. GLC-MS allowed an unequivocal identification of each fatty acid. The branched chain fatty acid i-16:0 was the predominant feature with 33% followed by the unsaturated straight-chain acid 16:1. In this respect earlier reports by other authors were confirmed but minor compounds not detected so far could be revealed as 15:1, the doublet 18:1 and i-18:0, 19:1 and 20:1. OMP profiles are easy to establish and to interpret. They are well suited for identification of the species L. pneumophila. Ubiquinone analysis permits rapid but preliminary species identification, whereas fatty acid analysis by GLC-MS is an intricate technique providing a valuable chemical marker for L. pneumophila. The biochemical analyses presented here can supplement immunological studies, clarify immunological findings and help to identify new serogroups of L. pneumophila.