Isolation and biological characterization of two lipopolysaccharides and a capsular-enriched polysaccharide preparation from Haemophilus pleuropneumoniae.
A smooth-type lipopolysaccharide (HpS-LPS), a rough-type lipopolysaccharide (HpR-LPS), and a capsular-enriched polysaccharide preparation (HpC-PS) were extracted and purified from Haemophilus pleuropneumoniae serotype 5, strain J45, by the use of phenol-water (HpS-LPS) and phenol-chloroform-petroleum-ether (HpR-LPS) techniques. Chemical analysis of the HpS-LPS and HpR-LPS indicated that they contained 0.7% and 4.4% (w/w) 2-keto-3-deoxyoctonate, 11.8% and 10.4% phosphate, 0.8% and 0.8% nucleic acid, and 0.8% and 1.1% protein, respectively. The HpC-PS contained 0.3% 2-keto-3-deoxyoctonate, 1.4% phosphate, 0.2% nucleic acid, and 0.8% protein. With sodium dodecyl sulfate-polyacrylamide-gel electrophoresis, the HpS-LPS banded as a smooth-type LPS and the HpR-LPS banded as a rough-type LPS. Electrophoresis of HpC-PS indicated the presence of a broad high molecular weight band. Gelation of Limulus amoebocyte lysate developed at a minimum concentration of 8 ng/ml of HpS-LPS, 0.3 ng/ml of HpR-LPS, and 35 ng/ml of HpC-PS. The lipopolysaccharide preparations provoked a positive dermal Shwartzman reaction in rabbits and swine, a biphasic febrile response in rabbits, and a monophasic response in swine. Responses were typical of endotoxic activity with swine having greater sensitivity than rabbits. The chick embryo 50% lethal dose was calculated to be 7.3 ng for HpS-LPS, 1.6 ng for HpR-LPS, 5.1 ng for the lipopolysaccharide of Escherichia coli 0111:B4; and the HpC-PS was not toxic. In all assays, HpR-LPS was significantly more toxic than was HpS-LPS. The HpC-PS preparation was not toxic, even at high concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D002473:Cell Wall / Q000276:immunology; D002645:Chickens; D006190:Haemophilus / Q000276:immunology* / Q000302:isolation & purification; D008033:Limulus Test; D008070:Lipopolysaccharides / Q000302:isolation & purification* / Q000633:toxicity; D008168:Lung / Q000187:drug effects / Q000473:pathology; D046911:Macromolecular Substances; D008297:Male; D011001:Pleuropneumonia / Q000382:microbiology / Q000662:veterinary; D011135:Polysaccharides, Bacterial / Q000302:isolation & purification* / Q000633:toxicity; D011749:Pyrogens; D011817:Rabbits; D012882:Skin Tests; D013552:Swine; D013553:Swine Diseases / Q000382:microbiology
D008070:Lipopolysaccharides; D046911:Macromolecular Substances; D011135:Polysaccharides, Bacterial; D011749:Pyrogens