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

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Abstract
Lipopolysaccharides (LPS) isolated from phase I and phase II C. burnetii (LPS I and LPS II, respectively) were analyzed for chemical compositions, molecular heterogeneity by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunological properties. The yields of crude phenol-water extracts from phase I cells were roughly 3-6 times higher than those from phase II cells. Purification of LPS 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 LPS were different in compositon and content, with branched-chain fatty acids representing .apprx. 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 LPS; this component was not identified as 3-deoxy-D-mannooctulosonic acid by gas and paper chromatographies. LPS II contained D-mannose, D-glucose, 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 9 unidentified components in addition to the components of LPS II. The hydrophogic, 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 LPS by SDS-PAGE followed by Ag staining indicated that LPS II was composed of only 1 band, wheres LPS I consisted .gtoreq. 6 bands with irregular spacing. Ouchterlony immunoduffusion 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. Thus, the chemical structures of LPS from C. burnetii are different from those of the LPS 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.