Heterogeneity of the Lipopolysaccharide from Pseudomonas aeruginosa

Abstract

Lipopolysaccharide isolated from Pseudomonas aeruginosa PACl and its phage-resistant mutant was degraded by mild acid hydrolysis into lipid A and three major polysaccharide-containing fractions which were separated on Sephadex G-75. The low-molecular-weight fraction contained glucose, rhamnose, heptose, galactosamine, alanine and phosphate. The higher-molecular-weight fractions consisted mainly of glucose, rhamnose and glucosamine together with amino compounds. Alkaline degradation of the lipopolysaccharide produced at least four different species each of which contained a low-molecular-weight polysaccharide similar if not identical to that produced by acid hydrolysis. Under certain growth conditions an abnormal lipopolysaccharide was produced which was defective in the low-molecular-weight polysaccharide and contained mainly high-molecular-weight material. Strains of different serotype yielded lipopolysaccharides which also exhibited heterogeneity but contained a low-molecular-weight polysaccharide similar to that obtained from strain PAC 1 and PAClR. It is suggested that each strain of P. aeruginosa may produce several lipopolysaccharides each containing a polysaccharide common to all. The relative proportions of the various lipopolysaccharides may be changed by growth conditions. The lipopolysaccharide of Pseudornonas aeruginosa PAC1 can be isolated by phenol extraction [l] and the lipid A split off by hydrolysis with 1 % acetic acid at 100°C for 60 min [2]. This releases partially degraded polysaccharide which can be separated by fractionation on Sephadex G-50 into high and lowmolecular-weight fractions of different composition. A mutant defective in rhamnose metabolism lacks the high-molecular-weight fraction [2] and we suggested that this might correspond to a side-chain if the pseudomonad lipopolysaccharide had a structure comparable to that of the Enterobacteriaceae (see [3] for review). Similarly the major low-molecular-weight fraction might be derived from a core polysaccharide common to all strains of P.aeruginosa. Evidence for a common core polysaccharide consisting mainly of glucose, rhamnose, heptose, galactosamine, phosphorus and probably alanine has since been obtained in several different strains of P. aeruginosa [4-91.There is much more variation in the highmolecular-weight fractions released by acetic acid hydrolysis of the isolated lipopolysaccharide. In a preliminary survey of the chemical composition and structure of lipopolysaccharide isolated from strains of P. aeruginosa of different Habs [lo] serotype, we found the high-molecular-weight fractions were responsible for the serological specificity of the original lipopolysaccharide and varied markedly in chemical composition [4]. In some strains, particularly those of Habs type 3, several high-molecular-weight fractions were detected, while in others, for example a strain of Habs type 2A, the high-molecular-weight fractions isolated on Sephadex G-75 contained no detectable neutral sugars but retained their antigenic specificity and contain...

Heterogeneity of lipopolysaccharides from Pseudomonas aeruginosa: analysis of lipopolysaccharide chain length

“…These populations represent sets of molecules with 0 antigens of different lengths which are made in large amounts. Other investigators have also demonstrated, using gel permeation chromatography in combination with other methods, that the LPS from members of the family Enterobacteriaceae and other gram-negative bacteria could be resolved into at least two main populations of LPS, differing in the length of their O-polysaccharide chain (9,26,30,32,34). In the results presented here, we found that the LPS from strains 503 and 1715 ( Fig.…”

“…2 and 3, respectively) of the LPS molecules in the low-molecular-weight population and 30 to 50% in the high-molecular-weight fractions (36,48). It (47), antibiotic susceptibility (la, 4, 18), LPS aggregate structure (47), bacteriophage recognition (25,27), immunochemical characterization (8,9,49), virulence (11,50), protection against the bactericidal action of serum (21,41), polyclonal B cell aativation, and macrophage cytotoxicity (43). The low level of LPS on P. aeruginosa that contains a long 0 polymer, however, may be sufficient to form a uniform cover over the cell, since the surface is inaccessible to rough-core-specific monoclonal antibodies (52).…”

“…The characterization of P. aeruginosa 0-specific polysac-* Corresponding author. charides has been complicated in some cases by chemical heterogeneity of the polysaccharide chains (8,9, 30,58). In some instances, the polymeric material has been resolved into amino sugar-rich and neutral sugar-rich fractions (30, 58).…”

“…Structural microheterogeneity in several regions of LPS molecules from members of the family Enterobacteriaceae (2,14,22,37,38,48,51) and Pseudomonas aeruginosa strains (33,57) has been demonstrated. Of the several methods used to separate the subclasses of LPS from individual strains, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) (20,24,45,48) and gel filtration (9,26,30,32,34,48) are the best. Either of these two methods by themselves, however, may be insufficient to completely characterize the high-and low-molecular-weight fractions of LPS.…”

The Specificity of Agglutination Reactions of Pseudomonas Aeruginosa With O Antisera

“…Our study has confirmed that the substance responsible for polyagglutinability is heat stable and that the antibody against it is distinct from 0 antibody. This is supported by the finding that chemical analysis of the polysaccharide portion of the cell-wall lipopolysaccharide, which is associated with the specificity of the 0 antigen (Chester, Meadow and Pitt, 1973), shows no significant difference between a typable culture and a PAculture derived from it (Chester and Meadow, 1975).…”