Chemical and immunological characterization of lipopolysaccharides from phase I and phase II Coxiella burnetii.

Lipopolysaccharides (LPSs) isolated from phase I and phase II Coxiella burnetii (LPS I and LPS II, respectively) were analyzed for chemical compositions, molecular heterogeneity by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and immunological properties. The yields of crude phenol-water extracts from phase I cells were roughly three to six times higher than those from phase II cells. Purification of LPSs by ultracentrifugation gave similar yields for both LPS I and LPS II. Purified LPS I and LPS II contained roughly 0.8 and 0.6% protein, respectively. The fatty acid constituents of the LPSs were different in composition and content, with branched-chain fatty acids representing about 15% of the total. beta-Hydroxymyristic acid was not detected in either LPS I or LPS II. A thiobarbituric acid-periodate-positive compound was evident in the LPSs; however, this component was not identified as 3-deoxy-D-mannooctulosonic acid by gas and paper chromatographies. LPS II contained D-mannose, D-glucose, D-glyceromannoheptose, glucosamine, ethanolamine, 3-deoxy-D-mannooctulosonic acid-like material, phosphate, and fatty acids. LPS I contained the unique disaccharide galactosaminuronyl glucosamine and nine unidentified components in addition to the components of LPS II. The hydrophobic, putative lipid A fraction of LPS I and LPS II contained the above constituents, but the hydrophilic fraction was devoid of ethanolamine. The LPS I disaccharide galactosaminuronyl glucosamine was found in both fractions of the acetic acid hydrolysates. Analysis of LPSs by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by silver staining indicated that LPS II was composed of only one band, whereas LPS I consisted of six or more bands with irregular spacing. Ouchterlony immunodiffusion tests demonstrated that LPS I reacted with phase I but not with phase II whole-cell hyperimmune antibody, and LPS II reacted neither with phase I nor phase II hyperimmune antibody. From these results, it was concluded that the chemical structures of LPSs from C. burnetii were different from those of the LPSs of gram-negative bacteria; however, the LPS structural variation in C. burnetii may be similar to the smooth-to-rough mutational variation of saccharide chain length in gram-negative bacteria.