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E Romanowska

Publications and source records attributed to E Romanowska.

At least 73 records · Page 4Linked to original sources

Citrobacter lipopolysaccharides: structure elucidation of the O-specific polysaccharide from strain PCM 1487 by mass spectrometry, one-dimensional and two-dimensional 1H-NMR spectroscopy and methylation analysis.

Structural studies on the O-specific polysaccharide of Citrobacter PCM 1487 lipopolysaccharide, using methylation analysis, Smith degradation and 1H-NMR spectroscopy, indicate that it consists of the trisaccharide repeating units (formula, see text) In this structure, 4-deoxy-D-araHex stands for 4-deoxy-D-arabino-hexose.

Carbohydrate Conformation↗

The enterobacterial common-antigen, a cyclic polysaccharide.

Structural studies of the enterobacterial common-antigen, using chemical methods and fast-atom-bombardment mass spectrometry, indicate that it is a cyclic polysaccharide, composed of four, five, and, to a smaller extent, six trisaccharide repeating-units. In the structure of the antigen, given below, D-Fuc4NAc stands for 4-acetamido-4,6-dideoxy-D-galactose.

Antigens, Bacterial↗

Characterization of anti-Citrobacter 036 specific polysaccharide monoclonal antibodies.

Monoclonal mouse antibodies specific for the 0 antigen of Citrobacter 036, a homopolymer of beta (1----2)-linked 4-deoxy-D-arabinohexose, were generated by the hybridoma technique. Balb/c mice were immunized with killed whole-cell vaccine and initial selection of active clones was based on enzyme-linked immunosorbent assay (ELISA) employing purified lipopolysaccharide (LPS). Concentrated culture supernatants from selected hybrid cultures were used to identify 10 0-antigen specific monoclonal antibodies using the multiple criteria of immunoprecipitation of 0 chains and LPS, inhibition by acid hydrolyzed 0 chains in the screening ELISA, and antibody class analysis. Four monoclonal antibodies were chosen for further study using dose-dependent 0-chain inhibition of ELISA and passive hemagglutination, passive hemolysis, and bacterial agglutination titres. When screened with Citrobacter serotypes known to contain the sugar 4-deoxy-D-arabinose, passive hemagglutination tests showed that the two monoclonal antibodies examined possessed titres which could be correlated with the reported 4-deoxy-D-arabinohexose content of the respective LPS's. This sugar is an antigenically important unit of several Citrobacter serotypes as defined by these well-characterized monoclonal antibodies.

Animals↗

Immunochemical characterization of lipopolysaccharides of Citrobacter chemotypes CC-G and CC-H.

Lipopolysaccharides of Citrobacter serotypes 04, 027 and 036 indicate strong cross-reactivity in passive hemagglutination and their O-specific chains are homopolymers of (beta 1----2)--linked 4-deoxy-D-arabinohexose. Citrobacter serotype 023 cannot be placed in chemotype CC-H, since its O-specific polysaccharide does not contain 4-deoxy-hexose. The possibility exists that 4-deoxy-D-arabinohexosyl side chains of 023-specific-polysaccharide disappeared as a result of a mutation of the parent strain. Although Citrobacter strain PCM 1487 contains 4-deoxy-D-arabinohexose and belongs to the chemotype CC-G, it cannot be included to the serotypes 04, 027 or 036. Instead, it represents a new serotype, with O-specific polysaccharide composed of 4-deoxy-D-arabinohexose, N-acetylglucosamine and N-acetylgalactosamine.

Carbohydrates↗

Enterobacterial common antigen-tetanus toxoid conjugate as immunogen.

The methods of limited periodate oxidation and reductive amination were used to obtain covalently linked enterobacterial common antigen (ECA) with tetanus toxoid. This procedure is simple and gives a good yield of the conjugate with high ECA content (molecular ratio of ECA to tetanus toxoid, 6:1). The ECA-tetanus toxoid conjugate is immunogenic in rabbits, in contrast to free ECA or a mixture of ECA with proteins. This conjugate produces high levels of ECA-specific immunoglobulin G antibodies, which can be used as a standard serum.

Animals↗

Immunological activities of purified preparations of enterobacterial common antigen.

The immunological activities of three purified preparations of enterobacterial common antigen (ECA) obtained by different procedures were studied. ECA-Ma (method of A. Marx) was from Salmonella typhimurium TV149 (Ra mutant), ECA-My (method of H. Mayer) was from S. montevideo, and ECA-Ro (method of E. Romanowska) was from Shigella sonnei phase I. These preparations, on a weight basis, neutralized similar amounts of ECA antibodies, indicating that the serological activities were comparable. Neither ECA-My nor ECA-Ro elicited specific delayed-type hypersensitivity skin reactions at 24 or 48 h in immunized guinea pigs. ECA-Ma, as well as the nonpurified preparations of the antigens used for immunization, elicited reactions at 24 h but not at 48 h. Thus, ECA-specific delayed-type hypersensitivity was not detected in immunized guinea pigs. Striking differences were noted in the immunogenicity of these antigens, ECA-Ma being highly immunogenic in the rabbit in contrast to ECA-My and ECA-Ro. ECA-Ma was a potent mitogen for guinea pig spleen cells, stimulating high levels of DNA synthesis; ECA-My was only slightly active. The three antigens were mitogenic to spleen cells from both CBA/J and C3H/HeJ mice, although not to the same degree, indicating that this effect is not due to contaminating lipopolysaccharide, since the latter strain of mice is resistant to endotoxin. Since an ECA-Ma extract made from an ECA-negative mutant proved to be mitogenic to murine spleen cells, the mitogenicity is not due to the ECA haptenic determinant. The mitogenic effect is polyclonal in nature, ECA-Ma producing a maximum response on day 3. Thus, the ECA preparations are both B-cell mitogens and polyclonal activators in murine spleen cells. From these studies it is evident that the biological and immunological activities of these purified antigens depend not only on the haptenic determinant but also on associated or bound components of the preparations.

Animals↗

Effect of proteins on the immunogenicity of enterobacterial common antigen.

Enterobacterial common antigen isolated by two independent extraction procedures was found to precipitate with a number of basic or hydrophobic proteins. Complexes of enterobacterial common antigen with protamine sulfate, with methylated bovine serum albumin or with a fraction of outer membrane proteins of two different Shigella wild types proved to be highly immunogenic in rabbits upon intravenous immunization, in contrast to the enterobacterial common antigen preparations by themselves. This explains why crude isolates of enterobacterial common antigen usually are good immunogens in contrast to the isolated antigen, which was described to be either not or only very poorly immunogenic.

Antibodies, Bacterial↗

Serological analysis of core oligosaccharides of Shigella flexneri serotype 6 and its R mutants lipopolysaccharides.

The core oligosaccharides originating from Shigella flexneri 6 S strains and R mutants were examined by the complement fixation inhibition and passive hemolysis inhibition tests using antisera against the complete and partially degraded core regions. Strong cross reactivity between Shigella flexneri 6, Shigella sonnei phase II and E. coli C core fractions which all are of R1 type was observed. Terminal beta-D-glucosyl groups and alpha-D-galactosyl-1,2-alpha-D-galactosyl sequences are essential elements of R1 type immunodeterminants. Heptose region of the R1 cores seems not be involved in their serological specificity.

Antigens, Bacterial↗

Heterogeneity of Shigella sonnei phase I O-antigen.

Shigella sonnei phase I and phase II lipopolysaccharides were examined using passive hemagglutination, immunodiffusion and immunoelectrophoresis. Moreover, O-antigen preparations of phase I obtained by three different methods were separated in SDS polyacrylamide gel electrophoresis into two fractions: L1 and L2, whereas phase II lipopolysaccharide showed one fraction (LI) only. The results of the serological investigations of three mentioned above preparations indicate that the heterogeneity of phase I O-antigen is natural, and not caused by extraction procedures. Shigella sonnei phase I cells produce two types of lipopolysaccharide molecules: I) with core regions substituted by phase I-specific polysaccharide chains, and 2) with core regions unsubstituted, having phase II specificity.

Antigens, Bacterial↗

Studies on virulence of Shigella flexneri. Protective effect of outer membrane proteins.

Groups of mice were immunized intraperitoneally with proteins isolated from outer membrane of virulent strain of Shigella flexneri 3a and its avirulent variant. All the animals were found to survive challenge with LD100 of virulent Sh. flexneri 3a. Similar protective effect against challenge with Sh. flexneri 3a gave also immunization of mice with outer membrane proteins isolated from Sh. flexneri of other serotypes (2a, 4a, 6, and X) and from Escherichia coli, Hafnia alvei and Shigella sonnei.

Animals↗

Structural studies on Shigella flexneri serotype 6 core region.

The core structure of Shigella flexneri serotype 6 has been investigated. The methods used in these studies involved methylation analysis, periodate oxidation, Smith degradation and enzymic modifications of core oligosaccharides isolated from two smooth strains and four rough mutants. As a result the complete core region was established (Formula: see text) where DGalp = D-galactopyranose, DGlcp = D-glucopyranose, L alpha DHepp = L-glycero-D-mannoheptapyranose, dOclA = 3-deoxy-D-manno-octulosonic, DGlcNp = 2-amino-2-deoxy-D-glucopyranose and P--PEtN = pyrophosphorylethanolamine (partially substituted). In the R mutants examined two types of mutation have been observed: (1) defect in heptose region (lack of D-glucosamine) or (2) defects in hexose region of the core.

Carbohydrate Sequence↗

Protection against keratoconjunctivitis shigellosa induced by immunization with outer membrane proteins of Shigella spp.

Active immunization of guinea pigs and rabbits with outer membrane proteins (OMP) isolated from Shigella flexneri 3a and Shigella sonnei phase I protected the animals against keratoconjunctivitis shigellosa induced with the homologous or heterologous strain. Protection was also achieved in rabbits after passive immunization with anti-OMP immune serum. Active immunization with lipopolysaccharide of S. flexneri 3a did not protect rabbits against keratoconjunctivitis shigellosa.

Animals↗

Outer membrane proteins of Shigella sonnei. I. Characterization of phase I, phase II and R-form Shigella sonnei.

The cell envelope of Shigella sonnei phase I, phase II and R-form was fractionated inot outer and cytoplasmic membrane components by means of sucrose density gradient centrifugation. The protein composition of the outer membrane has been analyzed by SDS-polyacrylamide gel electrophoresis. The outer membrane preparations contained 15-17 proteins. The major proteins of the outer membrane were of apparent molecular weights: 27,000, 28,000 and 31,000. Their amount varied depending on the structural defects of lipopolysaccharide.

Bacterial Proteins↗

Outer membrane proteins of Shigella sonnei. II. Comparative studies on virulent and avirulent strains of phase I.

The proteins of the outer membrane of virulent and avirulent Shigella sonnei phase I strains were characterized by SDS-polyarcylamide gel electrophoresis and their molecular weights determined. Significant quantitative differences were found in composition of the outer membrane proteins between virulent and avirulent strains. These differences were confirmed in electrofocusing experiments and disc-crossed immunoelectrophoresis.

Bacterial Proteins↗