[Examination of selected biological properties for swarm cells in Proteus mirabilis]. / Badanie wybranych wlasciwosci biologicznych komórek dlugich (swarm cells) Proteus mirabilis.

Certain biological properties, which generally are thought to play a role in the bacterial pathogenicity, were compared in short rods, swarm cells and penicillin-induced filaments of Proteus mirabilis S1959. Swarm cells of P. mirabilis S1959 weakly adhere to human uroepithelial cells and also do not penetrate L929 line of mouse fibroblasts. They do not show any cytotoxic activity, are poorly phagocytized by macrophages and granulocytes and start to divide by the end of the phagocytosis process. Their well-marked cell-bound haemolytic activity is correlated with fast division to the short rods. In in vivo experiments we have demonstrated the presence of long filamentous multinucleate nonseptate cells in the bladder and in the kidneys of infected animals. However there was no correlation between the number of swarm cells seen under microscopic examination and the intensity of infection.

Characteristics of spontaneously agglutinating Proteus mirabilis strains from bacteriuric patients.

Out of 210 Proteus mirabilis isolates from bacteriuric patients a total of eight spontaneously agglutinating strains were found. SDS-PAGE analysis of their lipopolysaccharides (LPS), the separation of polysaccharide fractions (PS) by gel filtration, and chemical characterization of PSs were performed. Out of the eight strains one S form and one mutant classified as Rc were detected. The remaining six strains were recognized as Ra and intermediate forms. When tested in a hematogenous infection model in mice, the P. mirabilis Rc mutant survived in kidneys for at least two weeks, while the Re mutant used as control was eliminated within 20 h after the challenge. These data indicated that strains of P. mirabilis may be pathogenic even if they express very incomplete LPS.

Galacturonic acid as the terminal constituent in the R core polysaccharide of proteus R110 (Ra) mutant.

Terminal non-reducing position of GalUA residue in the R core region of P. mirabilis R110 (Ra)mutant was established by GLC/MS analysis of methylated degraded polysaccharide. This position of GalUA is exceptional as compared with the terminal constituents present in the known structures of R core polysaccharides in Enterobacteriaceae. As it was shown in serological study, GalUA does not play a role of immunodominant in the examined Proteus R core region.

Some biological features of Proteus bacilli. 1. Comparison of Proteus mirabilis strains provided from various sources.

Some properties which may contribute to the pathogenicity of Proteus mirabilis were compared in urinary isolates and in strains provided from soil and from culture collection. Clinical isolates revealed the higher expression of all the features examined in this report: swarming growth, haemagglutination, adherence to human uroepithelial cells, urease activity and haemolytic activity. Noteworthy is the higher mean value of adherence to the uroepithelial cells in clinical strains. Three P. mirabilis urinary isolates were detected which produce an as yet unreported filterable haemolysin. However, the loss of this ability within a few months seems to suggest the temporary presence of a plasmid rapidly eliminated by the Proteus strains.

Heterogeneity of the lipopolysaccharide from Proteus mirabilis O27.

Two lipopolysaccharides (LPS) were obtained by phenol-water procedure from Proteus mirabilis O27 strain -- LPS I as sediment and LPS II from the supernatant after ultracentrifugation. There was shown a distinct predominance of the O-specific material in LPSII, whereas in LPS I the core material prevailed. Heterogeneity of the P. mirabilis.O27 lipopolysaccharide was confirmed by the sodium dodecyl sulphate-polyacrylamide gel electrophoresis, in crossed immunoelectrophoresis and by the column chromatography.

Immunochemical studies on the O-specific side chains of the heterogeneous lipopolysaccharide from Proteus mirabilis O27.

Chemically and serologically identical O-specific fractions were isolated from two different lipopolysaccharide (LPS) preparations of Proteus mirabilis O27. The release of a labile constituent in mild hydrolysis deprived those fractions of their serological activity. The compound was identified as N-acetyl-D-glucosamine and its terminal position in the molecule was established by methylation analysis. The non-carbohydrate constituent of the specific fractions: lysine and alanine are linked via their amino group. Terminal residues of N-acetylglucosamine, have to be considered as immunodeterminants of Proteus mirabilis O27. The role of other carbohydrate and non-carbohydrate constituents for the serological specificity of P. mirabilis O27 is discussed.

Core region in Proteus mirabilis lipopolysaccharide.

Four R mutants of P. mirabilis were isolated. The composition of their degraded polysaccharides (PS) obtained from the respective lipopolysaccharides (LPS) as well as the composition and properties of the PS-fractions separated by column chromatography were examined. The results were compared with those obtained with PS of the wild type. One of the mutants could be classified as an Ra-type mutant, presenting a complete LPS core. This polysaccharide core contains: galacturonic acid, glucosamine, glucose, D-glycero-D-mannoheptose, L-glycero-D-mannoheptose in a molar ratio of 1 : 1 : 1 : 1 : 2 and 2-keto-3-deoxyoctonate. Taking into consideration the common sugars described previously in the LPS chemotypes of P. hauseri, the composition of the complete core region mentioned above represents the LPS core part of all the chemotypes, containing two different heptoses.

The linkage of lysine in the O-specific chains of Proteus mirabilis 1959.

Lipopolysaccharides of qualitatively identical but quantitatively different sugar composition were extracted from Proteus mirabilis strain 1959. The lipopolysaccharide with the higher percentage of typical O-specific constituents was subjected to partial acid hydrolysis. An oligosaccharide B22 was separated by paper chromatography and electrophoresis. It was found to be composed of equimolar amounts of D-galacturonic acid, D-galactosamine and L-lysine. Dinitrophenylation of the oligosaccharide as well as of the genuine lipopolysaccharide afforded xi-dinitrophenyl-L-lysine after acid hydrolysis, showing that lysine was linked to the disaccharide via its alpha-amino group. Further studies including the Morgan-Elson and Elson-Morgan reactions, NaBH4-reduction, hydrazinolysis and periodate oxidation revealed the structure of oligosaccharide B22 as D-galacturonyl-(1 leads to 4)-D-galactosamine with lysine attached to the carboxylic group of galacturonic acid via its alpha-amino group. Judged from its high inhibition capacity this oligosaccharide has to be considered as an essential part of the serological determinant of Proteus mirabilis 1959. The frequent occurrence of lysine and galacturonic acid in Proteus mirabilis O-serogroups and their possible significance for the respective serological specificities are discussed.

Reactivity of lipopolysaccharides from various salmonella SR and R chemotypes Ra-Re mutants with concanavalin A.

Lipopolysaccharides from different R mutants of Salmonella minnesota and Salmonella typhimurium belonging to chemotypes Ra to Re, as well as from three SR mutants of Salmonella typhimurium were selected for a study of their precipitability with Concanavalin A. Predictions as to the outcome of the reaction could be made since both the chemical structure of the Salmonella R lipopolysaccharides and structural requirements for a positive reaction with Concanavalin A are well established. Precipitation studies in the immuno-electrophoretic assay and in the microcapillary test were carried out with alkali-treated lipopolysaccharides as untreated lipopolysaccharide is too highly aggregated to allow a sufficient migration in agarose layers. Lipopolysaccharides of all mutants--except the SR mutants--were obtained by the phenol/chloroform/petroleum ether method in order to avoid contaminations by glucans or glycogen which are known to occur in phenol/water extracted lipopolysaccharides and which would lead to erroneous results. Additional precipitation studies were carried out with two other lectins of different polysaccharide specificity: Wheat Germ Agglutinin and Soybean Agglutinin. As expected, lipopolysaccharides of chemotypes Ra, Rb1, and RcP- mutants reacted strongly with Concanavalin A, whereas no reaction was demonstrable with lipopolysaccharides of chemotypes Rb2, Rb3, Rd and Re mutants. The lipopolysaccharide of an RcP+ mutant unexpectedly failed to precipitate unless it was dephosphorylated with HF. This artificially prepared RcP-lipopolysaccharide showed a strong reaction, thus demonstrating that negative charges in the direct neighborhood of reactive sugar units as in RcP+ LPS may prevent precipitation with Concanavalin A. No reactivity demonstrable by precipitation could be obtained using either Wheat Germ Agglutinin or Soybean Agglutinin with alkali-treated lipopolysaccharide even of those chemotypes which had the supposedly reactive sugar in a terminal position, such as N-acetyl-D-glucosamine in Ra mutants (Wheat Germ Agglutinin) or D-galactose in Rb2 or Rb3 mutants (Soybean Agglutinin).