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PubMed1998

Avidity of IgG for Streptococcus pneumoniae type 6B and 23F polysaccharides in infants primed with pneumococcal conjugates and boosted with polysaccharide or conjugate vaccines.

Relative avidity of IgG to Streptococcus pneumoniae type 6B and 23F polysaccharides (PSs) was measured in sera of children immunized with pneumococcal vaccines, using an EIA and the chaotropic agent thiocyanate. Infants were immunized at 2, 4, and 6 months of age with pneumococcal PS-diphtheria toxoid conjugate vaccine, and boosted at 14 months with the homologous conjugate or a pneumococcal PS vaccine. The concentrations of antibodies to 6B and 23F PSs increased significantly after the booster with both vaccines. A significant increase in the avidity of anti-6B and anti-23F antibodies was seen after the booster with conjugate but not with PS vaccine. Avidity also increased in another group of infants primed and boosted with pneumococcal PS-meningococcal protein conjugate but not in a group boosted with PS vaccine after priming with pneumococcal PS-tetanus toxoid conjugate. In the latter group, however, the avidity of anti-6B was high before boosting.

Antibodies, Bacterial↗
PubMed1985

Induction of increased antibody responses to pneumococcal type 19F polysaccharide by Klebsiella polysaccharide.

The level of antibody to pneumococcal 19F polysaccharide (PS) was studied in the serum of mice inoculated with klebsiella K2 PS or pneumococcal 19F PS or both. The serum antibody levels and the IgM plaque-forming-cell (PFC) response after immunization were higher in mice given both K2 PS and 19F PS than in mice of a control group given 19F PS alone. This immune response was also higher than the sum of PFC production in mice given 19F PS or K2 PS separately. Thus, K2 PS produced the cross-reactive antibody to 19F PS and also enhanced the magnitude of the antibody response to 19F PS. This response appeared to be specific for the 19F PS. The resulting high levels of antibody to pneumococcal 19F persisted for eight weeks after the administration of K2 PS and 19F PS. Animals immunized with both type 3 PS and K2 PS did not show an increased antibody response to pneumococcal type 3 PS. Klebsiella K-O3 antigen also stimulated the immune response to 19F PS. Higher immunologic responses to 19F PS were also induced in nude mice injected with the cross-reacting K2 PS or K47 PS and 19F PS. The ability to stimulate an antibody response to 19F PS by administering both K2 PS and 19F PS could not be transferred with spleen cells obtained from mice given both K2 PS and 19F PS.

Animals↗
PubMed1983

Immunological properties of monoclonal antibodies specific for meningococcal polysaccharides: the protective capacity of IgM antibodies specific for polysaccharide group B.

Two IgM monoclonal antibodies, MB32 and MB34 specific for meningococcal polysaccharide group B have been raised. Both were detectable by radioimmunoassay and agglutination, but only MB34 was effective in counter immunoelectrophoresis and complement fixation. MB34 was also far more potent than MB32 when tested for passive protection of mice infected with either Neisseria meningitidis group B or Escherichia coli K1. These data demonstrate that group B-specific antibodies do play a protective role in mice infected with these bacteria.

Animals↗
PubMed1993

Structural analysis of O4-reactive polysaccharides from recombinant Escherichia coli. Changes in the O-specific polysaccharide induced by cloning of the rfb genes.

In previous studies it had been shown that lipopolysaccharide from O4-specific recombinant Escherichia coli, had serological reactivities and a chemical composition that differed from wildtype O4 LPS [Haraguchi, G.E., Zähringer, U., Jann, B., Jann, K., Hull, R.A. & Hull, S.I. (1991) Microb. Pathog. 10, 351-361]. Here we present the structural elucidation of the O-specific moieties from lipopolysaccharides of some of the recombinant strains obtained in previous studies. Compositional analysis, methylation, chemical reactions and NMR spectroscopy showed that, during genetic manipulations (recombination, cosmid cloning, plasmid subcloning), a gradual structural change in the O-specific polysaccharides was observed in the recombinant strains. These changes comprised of an alteration in the position of glucose (side chain) substitution, a change in the anomeric configuration of the main-chain N-acetylglucosamine and an exchange of alpha-L-rhamnopyranose for beta-D-galactofuranose. The relevance of these results for lipopolysaccharide cloning and lipopolysaccharide biosynthesis are discussed.

Carbohydrate Sequence↗
PubMed1992

Cloning of a locus involved in Streptococcus mutans intracellular polysaccharide accumulation and virulence testing of an intracellular polysaccharide-deficient mutant.

The streptococcal transposon Tn916 (Tcr) was used to isolate mutants of Streptococcus mutans altered in glycogen accumulation to investigate whether glycogenlike intracellular polysaccharides (IPS) play an important role in S. mutans-induced caries formation. S. mutans UA130 (serotype c) was transformed with the Escherichia coli plasmid pAM620 (Tn916), and the resultant transposon libraries were screened for glycogen content by iodine staining. A transposon mutant, designated SMS201, demonstrated a glycogen-deficient phenotype on glucose-enriched medium. Quantitative electron microscopy confirmed that IPS concentrations were significantly reduced in SMS201 relative to the wild-type progenitor strain, UA130. Importantly, reversion to wild type correlated at all times with loss of the transposon. Transposon excisants were used to facilitate cloning of the streptococcal gene(s) involved in glycogen biosynthesis and storage. A 2.1-kb chromosomal determinant (glgR) which encodes a putative regulator of S. mutans glycogen accumulation was isolated. A stable deletion mutation (delta glgR) was subsequently generated in E. coli and introduced into S. mutans by allelic exchange. The resultant glycogen synthesis-deficient mutant, SMS203, demonstrated a significantly reduced cariogenic potential (P less than 0.01) on the buccal, sulcal, and proximal surfaces of teeth in germfree rats, relative to animals challenged with the glycogen synthesis-proficient progenitor strain, UA130. These observations confirm previous reports (J. M. Tanzer, M. L. Freedman, F. N. Woodiel, R. L. Eifert, and L. A. Rinehimer, p. 597-616, in H. M. Stiles, W. J. Loesche, and T. L. O'Brien, ed., Proceedings in Microbiology. Aspects of Dental Caries. Special Supplement to Microbiology Abstracts, vol. 3, 1976) which implicate IPS as significant contributors to the S. mutans cariogenic process.

Animals↗
PubMed1993

Immunogenicity of Vibrio vulnificus capsular polysaccharides and polysaccharide-protein conjugates.

Opaque colony morphology has been correlated to Vibrio vulnificus virulence. However, the number of capsular serotypes expressed by virulent isolates is unknown. In an effort to produce anticapsule sera, capsular polysaccharide (CPS) from three opaque V. vulnificus strains was purified and characterized. Purified CPSs were acidic and contained considerable amounts of hexosamine and trace quantities of protein and nucleic acid. CPS purified from strain C7184 was poorly immunogenic for rabbits and mice, since repeated injection produced little detectable anticapsular antibody. To improve immunogenicity, CPS-protein conjugates were prepared from adipic acid hydrazide derivatives of CPS purified from each strain and carbodiimide as a coupling reagent. The immunogenicity of C7184 CPS was enhanced by conjugation to keyhole limpet hemocyanin, since injection into mice elicited production of anticapsular antibodies, the level of which was dependent on the dose and time since initial immunization. Injection of rabbits with CPS-protein conjugates also produced anticapsular antibodies. Staphylococcus aureus cells armed with each of the three anticapsular antibodies coagglutinated only the homologous opaque strain, indicating the existence of at least three capsular types. Further screening of 32 opaque and translucent V. vulnificus isolates revealed only three cross-reacting strains. These results suggest the presence of numerous V. vulnificus capsular types.

Agglutination Tests↗
PubMed1994

Epitope specificity of rabbit immunoglobulin G (IgG) elicited by pneumococcal type 23F synthetic oligosaccharide- and native polysaccharide-protein conjugate vaccines: comparison with human anti-polysaccharide 23F IgG.

Streptococcus pneumoniae type 23F capsular polysaccharide (PS23F) consitss of a repeating glycerol-phosphorylated branched tetrasaccharide. The immunogenicities of the following related antigens were investigated: (i) a synthetic trisaccharide comprising the backbone of one repeating unit, (ii) a synthetic tetrasaccharide comprising the complete repeating unit, and (iii) native PS23F (all three conjugated to keyhole limpet hemocyanin [KLH]) and (iv) formalin-killed S. pneumoniae 23F. All antigens except the trisaccharide-KLH conjugate induced relatively high anti-PS23F antibody levels in rabbits. The epitope specificity of such antibodies was then studied by means of an inhibition immunoassay. The alpha(1-->2)-linked L-rhamnose branch was shown to be immunodominant for immunoglobulin G (IgG) induced by tetrasaccharide-KLH, PS23F-KLH, and killed S. pneumoniae 23F: in most sera L-rhamnose totally inhibited the binding of IgG to PS23F. Thus, there appears to be no major difference in epitope specificity between IgG induced by tetrasaccharide-KLH and that induced by antigens containing the polymeric form of PS23F. Human anti-PS23F IgG (either vaccine induced or naturally acquired) had a different epitope specificity: none of the inhibitors used, including L-rhamnose and tetrasaccharide-KLH, exhibited substantial inhibition. These observations suggest that the epitope recognized by human IgG on PS23F is larger than the epitope recognized by rabbit IgG. Both human and rabbit antisera efficiently opsonized type 23F pneumococci, as measured in a phagocytosis assay using human polymorphonuclear leukocytes.

Animals↗
PubMed1994

Polysaccharides in Fungi. XXXIV. A polysaccharide from the fruiting bodies of Amanita muscaria and the antitumor activity of its carboxymethylated product.

A water-insoluble, alkali-soluble, glucan (AM-APP), [alpha]D +160 degrees in 0.4 M NaOH, was isolated from the alkaline extract of the fruiting bodies of Amanita muscaria. The results of chemical and spectroscopic investigations indicate that AM-APP is a linear (1 --> 3)-alpha-D-glucan with a molecular weigh estimated by gel chromatography of about 42000. Its carboxymethylated product (AM-APP-CM) showed potent antitumor activity against sarcoma 180 in mice, although the native polysaccharide (AM-APP) had little effect. The distribution of carboxymethyl groups in the molecule was analyzed by gas chromatography and mass spectrometry. The degree of substitution of carboxymethyl groups was 0.95 and the substituents were located at O-2, at O-4, at O-6, at O-2 and O-6, and at O-4 and O-6 on glucose.

Acetates↗
PubMed1995

Polysaccharides in fungi. XXXV. Anti diabetic activity of an acidic polysaccharide from the fruiting bodies of Tremella aurantia.

An acidic polysaccharide (TAP) was isolated from a hot-water extract of the fruiting bodies of Tremella aurantia. It showed remarkable hypoglycemic activity in normal mice and two diabetic mouse models, streptozotocin-induced diabetes and genetic diabetes, following intraperitoneal administration. Continuous oral administration of TAP solution (0.5g/l) for a long period was found to be also effective in hyperglycemia in glucose-loaded mice and no harmful physical effects were found. TAP had an [alpha]D -7 degrees in water, and its molecular weight was estimated to be about 1500000. TAP is composed of mannose, xylose, glucuronic acid and glucose (molar ratio, 4:2:1:0.3), and it contains 2.2% of O-acetyl groups.

Animals↗
PubMed1996

Polysaccharides in fungi. XXXVI. Hypoglycemic activity of a polysaccharide (CS-F30) from the cultural mycelium of Cordyceps sinensis and its effect on glucose metabolism in mouse liver.

A polysaccharide (CS-F30) obtained from the cultural mycelium of Cordyceps sinensis showed potent hypoglycemic activity in genetic diabetic mice after intraperitoneal administration, and the plasma glucose level was quickly reduced in normal and streptozotocin-induced diabetic mice after intravenous administration. Administration of CS-F-30 to normal mice significantly increased the activities of hepatic glucokinase, hexokinase and glucose-6-phosphate dehydrogenase, although the glycogen content in the liver was reduced. Furthermore, CS-F30 lowered the plasma triglyceride level and cholesterol level in mice.

Animals↗
PubMed1992

Polysaccharides in fungi. XXX. Antitumor and immunomodulating activities of two polysaccharides from the fruiting bodies of Armillariella tabescens.

The effects of two polysaccharides, AT-HW and AT-AL obtained from the fruiting bodies of Armillariella tabescens on murine sarcoma 180 tumor and peritoneal macrophages were examined at intraperitoneal administration. AT-HW from the hot-water extract and AT-AL from the alkaline extract significantly inhibited the tumor, and the results of different administration schedule and phagocytic system blockade suggested that the mechanism of AT-AL differed from that of AT-HW and branched (1----3)-beta-D-glucans. AT-HW and AT-AL showed reticuloendothelial system-potentiating activity, increased the number of peritoneal exudate cells, activated on macrophages (acid phosphatase activity, glucose consumption, superoxide anion production), and enhanced mitogenic reaction, although AT-HW did not produce superoxide anion in vitro.

Adjuvants, Immunologic↗
PubMed2000

Capsular polysaccharide typing of domestic mastitis-causing Staphylococcus aureus strains and its potential exploration of bovine mastitis vaccine development. I. Capsular polysaccharide typing, isolation and purification of the strains.

One hundred seven isolates of Staphylococcus aureus from bovine mastitis were investigated for colony morphology in serum-soft agar (SSA), autoagglutination in salt, and capsular serotype. Capsular polysaccharide (CP) was purified and quantified from the extracts of clinical isolates. Overall, 89 isolates (83.2%) were diffuse in the SSA, without any difference in the proportion of diffuse colony between type 5 and type 8 strains. Some strains exhibited compact colonies in the SSA and expressed CP as determined by an enzyme-linked immunosorbent assay, indicating that compact morphology does not exclude encapsulation. The majority of the strains (11/12) showed autoagglutination in the salt aggregation test. The serotype 336 accounted for 46.7% of the isolates followed by serotype 5 (12.1%) and serotype 8 (12.1%). Particularly, twenty-six (24.3%) isolates reacted with two serotypes; 7 for type 8/336 and 19 for type 5/336. Five isolates (4.7%) were nontypeable with monoclonal antibodies specific for CP serotype 5, 8, or 336. The CP concentration in culture supernatants varied with the serotypes, and the total amount of CP produced by cells grown in a liquid medium was much less than that produced by cells grown on a solid medium. The Western blotting indicated that the CP bands of S. aureus serotype 5 and 8 were ranged in the molecular mass of 58-84 kilodalton (kDa), with additional bands in the region of approximately >/= 48 or </= 84 kDa.

Agglutination Tests↗
PubMed1989

Chromotropic character of bacterial acidic polysaccharides: Part II--Induction of metachromasy in cationic dye pinacyanol chloride by Klebsiella K10 capsular polysaccharide.

The acidic capsular polysaccharide isolated from Klebsiella K10 exhibited chromotropic character with respect to induction of metachromasy in the cationic dye pinacyanol chloride (1-ethyl-2-[3-(1-ethyl-2(1H)-quinolylidene)propenyl]quinolinium chloride). Klebsiella K10 polymer consists of hexasaccharide repeating units containing one residue of glucuronic acid along with other neutral sugars in each repeating unit. It induces a metachromatic blue shift in the visible absorption spectrum of the dye from 600 nm to 500 nm. The spectral changes have been studied during interaction of the dye cations with the polyanions at different polymer/dye molar ratios. The polyanion-dye compounds are formed with polymer/dye stoichiometry of 1:1, indicating formation of stacking conformation. The complete reversal of polymer-induced metachromasy has also been observed by the addition of ethanol and urea.

Carbocyanines↗
PubMed1977

Multiple polysaccharide antigens of group B streptococcus, type Ia: emphasis on a sialic acid type-specific polysaccharide.

Group B streptococci, type Ia (strain 090/14/4), were subjected to sequential extraction procedures and the various extracts were fractionated by a combination of DEAE-cellulose chromatography and gel-filtration. Three major antigens were isolated: the conventional group-specific and type-specific carbohydrates and an acidic polysaccharide consisting of galactose, glucose, glucosamine, and sialic acid. Immunochemical data suggest that the acidic antigen, with the exception of the immunodominant sialic acid, is structurally similar to the conventional pH 2.0 extracted type-specific antigen. Removal of the terminal sialic acid residues from the acidic antigen by mild acid hydrolysis resulted in a residual polymer which was chemically and immunologically analogous to the type-specific carbohydrate. Precipitin analysis focused attention to the cross-reactivity between thia acidic antigen and anti-types Ib, Ic, and III sera.

Animals↗
PubMed1995

Functional differences in idiotypically defined IgG1 anti-polysaccharide antibodies elicited by vaccination with Haemophilus influenzae type B polysaccharide-protein conjugates.

We investigated the relationship between the form of the Haemophilus influenzae type B (Hib) polysaccharide (PS)-protein conjugate vaccine, Id expression, and Ab quality. Two post-vaccination pools were prepared from sera of infants vaccinated with either Hib PS oligomers coupled to CRM197, a mutant diphtheria toxin (HbOC), or with higher m.w. Hib PS coupled to an outer membrane protein complex of Neisseria meningitidis group B (Hib-OMP). The mean anti-Hib PS Ab avidity of the serum pool from the infants vaccinated with HbOC was threefold higher than that of the pool from infants vaccinated with Hib-OMP. Using sequential immunoabsorption, three IgG1 idiotypically-defined anti-Hib PS fractions were isolated from each of the serum pools: Hibld-1, Hibld-2, and a Hibld-1/-2-depleted population, designated Hibld-0. Hibld-1 and Hibld-2 are idiotypic markers for anti-Hib PS Abs expressing kappa II-A2 and lambda VII V regions, respectively. Hibld-1 anti-Hib PS Abs had significantly higher avidity, 2- to 19-fold higher in vitro bactericidal activity, and were more protective against Hib bacteremia in infant rats, than the respective Hibld-2 Abs isolated from each of the pools. Comparing the two vaccines, Hibld-1 anti-Hib PS Abs elicited by HbOC had significantly higher avidity and 10-fold higher bactericidal and rat protective activity than the Hibld-1 Abs elicited by Hib-OMP. These findings demonstrate that the molecular form of the Hib PS immunogen dictates both V region usage and quality of Ab function.

Animals↗
PubMed1998

Gamma 3 gene-disrupted mice selectively deficient in the dominant IgG subclass made to bacterial polysaccharides undergo normal isotype switching after immunization with polysaccharide-protein conjugate vaccines.

Bacterial polysaccharides (PS) are T-independent type 2 Ags that elicit restricted Ab responses of IgM and IgG3 in mice and IgM and predominantly IgG2 in humans. Immunodeficiency in the dominant IgG subclass made to PS is associated with chronic sinus and pulmonary infections with PS-encapsulated bacteria. To elucidate the biologic role of the dominant IgG subclass in the immune response to PS and to make an animal model of human IgG subclass deficiency, we generated mice with a targeted disruption of the exon encoding the CH1 domain of the gamma 3 heavy-chain constant region gene. Homozygotes had no detectable serum IgG3, and their splenocytes did not produce IgG3 after LPS stimulation. IgG3(-/-) mice immunized with PS from Pseudomonas aeruginosa LPS O-side chain or Streptococcus pneumoniae type 19F capsule did not produce any IgG3 anti-PS Abs, in contrast to wild-type mice in which IgG3 was the major IgG subclass. Immunizing both wild-type and IgG3(-/-) mice with 19F PS-protein conjugate elicited IgG1 Abs. We conclude that IgG3(-/-) mice have a selective deficiency in the dominant murine IgG subclass made to T-independent type 2 Ags and may be a useful animal model of IgG subclass deficiency. In addition, we show that the anti-PS Ab class switching to IgG1 that occurs when mice are immunized with a PS-protein conjugate vaccine does not require sequential Ig expression or an intact, upstream gamma 3 heavy-chain gene.

Animals↗
PubMed2002

Ordering in aqueous polysaccharide solutions. II. Optical rotation and heat capacity of aqueous solutions of a triple-helical polysaccharide schizophyllan.

Deuterium oxide solutions of schizophyllan, a triple-helical polysaccharide, undergoing an order-disorder transition centered at 17 degrees C, were studied by optical rotation (OR) and heat capacity (C(p)) to elucidate the molecular mechanism of the transition and water structure in the solution and frozen states. The ordered structure at low temperature consisted of the side chains and water in the vicinity forming an ordered hydrogen-bonded network surrounding the helix core and was disordered at higher temperature. In the solution state appeared clearly defined transition curves in both the OR and C(p) data. The results for three samples of different molecular weights were analyzed theoretically, treating this transition as a typical linear cooperative transition from the ordered to disordered states and explained quantitatively if the molecular weight polydispersity of the sample was considered. The excess heat capacity C(EX)(p) defined as the C(p) minus the contributions from schizophyllan and D(2)O was estimated. In the frozen state it increased with raising temperature above 150 K until the mixture melted. This was compared with the dielectric increment observed in this temperature range and ascribed to unfreezable water. From the heat capacity and dielectric data, unfreezable water is mobile but more ordered than free water. In the solution state, the excess heat capacity originates from the interactions of D(2)O molecules as bound water and structured water, and so forth. Thus the schizophyllan triple helix molds water into various structures of differing orders in solution and in the solid state.

Adjuvants, Immunologic↗