RESUMO
In this study we have prepared glycoconjugates with core oligosaccharides (OS) from the lipopolysaccharide (LPS) of Neisseria meningitidis, thus avoiding the neo-epitopes of the deacylated lipid A region of the derived LPS molecule identified in our previous studies. A comprehensive investigation was performed with glycoconjugates prepared from the most extended to the most truncated core OS still maintaining the conserved inner core epitope. As previously, we have established reproducible bactericidal killing of the homologous antigen elaborating strain, but a failure to kill wild-type strains. In these studies it was evident that the linker molecules used in the conjugation methodologies were dominating the immune response. However, when galE core OS based conjugates were prepared without utilizing linkers, via direct reductive amination, we failed to generate an immune response to even the homologous antigen. We also identified that immunisation with the galE antigen via linker methodologies provoked an immune response that was dependent upon key residues of the conserved inner core OS structure, whereas the immune responses to lgtB and lgtA antigens did not involve the inner core OS. This comprehensive study has, despite our best efforts, cast significant doubt as to the utility of the conserved inner core region of the meningococcal LPS as a potential vaccine antigen.
Assuntos
Vacinas Bacterianas/imunologia , Lipopolissacarídeos/imunologia , Infecções Meningocócicas/imunologia , Neisseria meningitidis/imunologia , Animais , Vacinas Bacterianas/química , Epitopos/química , Epitopos/imunologia , Lipopolissacarídeos/química , Infecções Meningocócicas/prevenção & controle , Oligossacarídeos/química , Oligossacarídeos/imunologia , Coelhos , Vacinas Conjugadas/química , Vacinas Conjugadas/imunologiaRESUMO
PURPOSE: To report a case of corneal graft rejection precipitated by severe uveitis secondary to alendronate therapy and to review the literature of relevance to this case. METHODS: A 77-year-old woman with a hypopyon and corneal graft rejection was studied for possible precipitants, including herpes viral and bacterial infection. Results were negative. She was treated unsuccessfully with systemic and topical steroids, systemic antivirals, and intraocular antibiotic therapy. RESULTS: Withdrawal of alendronate resulted in rapid resolution of intraocular inflammation and corneal edema. CONCLUSION: We recommend vigilance in corneal transplant patients on simultaneous bisphosphonate therapy. Caution is advised in the extension to human trials of animal studies investigating the use of bisphosphonates in corneal transplantation.
Assuntos
Alendronato/efeitos adversos , Conservadores da Densidade Óssea/efeitos adversos , Rejeição de Enxerto/etiologia , Ceratoplastia Penetrante , Uveíte/induzido quimicamente , Idoso , Feminino , Humanos , Osteoporose Pós-Menopausa/tratamento farmacológicoRESUMO
Monkey kidney cells productively infected with Yaba tumor poxvirus clearly exhibit plasma membrane alterations when treated with both fluorescein-labeled and unlabeled concanavalin A. The convanavalin A-mediated cytoagglutination reaction for Yaba-infected Jinet and CV-1 cells increased linearly from 12 to 16 h post-infection, reaching a maximum by 24-28 h. Treatment of either Yaba-infected CVC-1 or Jinet cells with methyl-D-glucopyranoside before or after addition of concanavalin A completely blocked or reversed the cytoaglutination response. Trypsin treatment of uninfected CV-1 or Jinet cells enhanced concanavalin A-mediated cytoagglutination properties. Conversely, trypsin treatment of Yaba-infected Jinet cells resulted in a reduced cytoagglutination response. Increasing temperature and lectin concentration enhance concanavalin A-mediated cytoagglutination for uninfected, trypsin-treated and Yaba-infected CV-1 cells. Cytosine arabinoside has little or no effect on the Yaba-induced cell cytoagglutination reaction while cycloheximide blocks the cytoagglutinatin response if added prior to 12 h post-infection. Fluorescein-labeled concanavalin A binding studies have revealed that at 4 degrees C, Yaba-infected CV-1 cells display a predominantly 'patchy' pattern of topological fluorescence, while trypsin-treated and uninfected CV-1 cells at 4 degrees C display a uniform pattern of fluorescence binding. Patchy fluorescence, indicative of concanavalin A-suspeptible, receptor-site clustering on the surface membrane, was reduced 50% if Yaba-infected CV-1 cells were treated with glutaraldehyde (2.5%) before addition of fluorescein-labeled concanavalin A at 4 degrees C. Similar pre-fixatin of trypsin-treated CV-1 cells resulted in uniform, fluorescent labelling patterns at all assay temperatures.
Assuntos
Transformação Celular Viral , Receptores de Concanavalina A/metabolismo , Vírus do Tumor do Macaco de Yaba/imunologia , Testes de Aglutinação , Animais , Linhagem Celular , Transformação Celular Viral/efeitos dos fármacos , Concanavalina A , Cicloeximida/farmacologia , Citarabina/farmacologia , Haplorrinos , Rim , Tripsina/farmacologia , Vírus do Tumor do Macaco de Yaba/efeitos dos fármacosRESUMO
This chapter presents the application of capillary electrophoresis coupled to electrospray mass spectrometry (CE-ES-MS) for the analysis of complex bacterial lipopolysaccharides (LPS) from pathogenic strains of Haemophilus influenzae and Neisseria meningitidis. A discussion is included of the development of electrophoretic conditions conducive to trace-level enrichment and separation of closely related glycoforms and isoforms, which provided sensitive detection of glycolipids from as little as five bacterial colonies. The chapter also describes the use of mixed MS scanning functions to aid the identification of specific functionalities and immunodeterminants of LPS, such as pyrophosphoethanolamine, phosphocholine, and N-acetyl neuraminic acid (Neu5Ac), which represent less than 2% of the overall LPS population. The combination of high-resolution capillary electrophoresis with sensitive tandem mass spectrometry (MS/MS) provides a unique analytical tool to probe the subtle structural changes resulting from oligosaccharide branching and location of substituted LPS isoforms. The ability to detect a diverse LPS population over a wide dynamic range of expression using CE-MS enables the correlation of structural changes between bacterial strains and isogenic mutants to assign functional gene relationship.
Assuntos
Eletroforese Capilar/métodos , Glicolipídeos/química , Lipopolissacarídeos/química , Espectrometria de Massas por Ionização por Electrospray/métodos , Acilação , Eletroforese , Haemophilus influenzae/metabolismo , Íons , Espectrometria de Massas , Modelos Químicos , Mutação , Neisseria meningitidis/metabolismo , Oligossacarídeos/química , Isoformas de Proteínas , Sensibilidade e Especificidade , Fatores de TempoRESUMO
Gene probes are used in virtually all disciplines of the life sciences. Probes will probably have their greatest impact in health care when used as diagnostic tools to detect and identify micro-organisms responsible for infectious disease. This review will use development of gene probes for the clinical laboratory as a theme and review recent patents and publications which have nurtured probe development.
Assuntos
Sondas de DNA , Sondas RNA , Animais , Amplificação de Genes , Humanos , Hibridização de Ácido NucleicoRESUMO
Structural studies of Bordetella endotoxins (LPSs) have revealed remarkable differences: (i) between their LPSs and those of other bacterial pathogens; (ii) among the LPSs of the seven identified Bordetella species; and (iii) among the LPSs of some Bordetella strains. The lipid As have the "classical" bisphosphorylated diglucosamine backbone but tend to have fewer and species-specific fatty acid components compared to those of other genera. Nevertheless, three strains of B. bronchiseptica have at least three different fatty acid distributions; however, the recently identified B. hinzii and B. trematum LPSs had identical lipid A structures. The B. pertussis core is a dodecasaccharide multi-branched structure bearing amino and carboxylic groups. Another unusual feature is the presence of free amino sugars in the central core region and a complex distal trisaccharide unit containing five amino groups of which four are acetylated and one is methylated. The B. pertussis LPS does not have O-chains and that of B. trematum had only a single O-unit, unlike the LPSs of all the other species of the smooth-type. The O-chain-free cores of non-B. pertussis LPSs were always built on the B. pertussis core model but most were species-specifically incomplete. The LPS structures of three B. bronchiseptica strains were found to be different from each other. The O-chains of B. bronchiseptica and B. parapertussis were almost identical and had some features in common with B. hinzii O-chain. Serological analyses are consistent with the determined LPS structures.
Assuntos
Bordetella/imunologia , Endotoxinas/química , Lipídeo A/química , Bordetella/genética , Sequência de Carboidratos , Cromatografia em Camada Fina , Eletroforese em Gel de Poliacrilamida , Endotoxinas/genética , Ácidos Graxos/análise , Espectrometria de Massas/métodos , Dados de Sequência Molecular , Estrutura MolecularRESUMO
We have developed a solid-phase ELISA to study the human immune response to inner core lipopolysaccharide (LPS) of Neisseria meningitidis (Nm) using structurally defined glycolipids from a genetically defined mutant (galE) of a serogroup B Nm strain. Previous studies had demonstrated that a galE (inner core) LPS epitope is conserved in approximately 70% Nm strains and was accessible to antibody in fully encapsulated wild-type Nm strains. A murine monoclonal antibody, MAb B5, raised to a galE mutant of serogroup B Nm strain, immunotype L3 (B.15.P1.7,16) was used to determine the specificity of the inner core LPS ELISA by inhibition studies using purified galE LPS and human sera. The intra-assay coefficient of variation (CV) was 5-6% and inter-assay CV was 19-22%. Using this ELISA, significant differences in the geometric mean titres (GMTs) of naturally occurring serum antibodies (specific to inner core LPS) between healthy adults (18-65 years, N=54) and healthy infants (3-4 months, N=144) of both IgG and IgM classes were found (P<0.0001). GMTs were expressed in galE arbitrary units (AU/ml) (95% confidence intervals): IgG antibodies in adults 5.7 (5. 0,6.9) and in infants 1.1 (1.0,1.3); IgM antibodies in adults 7.7 (5. 7,10.4), and in infants 0.85 (0.7,1.1). In age-matched children aged 26-113 months a difference (P=0.04) in specific IgG was found in healthy infants and infants in the acute phase of invasive Nm disease (GMT (95%CI) in AU/ml: in healthy infants 7.7 (5.3,11.0), in acute phase infants 4.2 (2.5,7.2). However, there was no difference in specific IgM (P=0.98) between these groups healthy infants 4.7 (3. 1,7.0), acute phase 4.6 (2.9, 7.4). In eleven children (5-181 months) there were differences in the GMTs of specific IgG and IgM (P=0.02, P=0.008 respectively) between paired acute and convalescent sera (GMT) (95%CI) in AU/ml: IgG acute 1.95 (0.98, 3.8), convalescent 5.2 (2.2,12.4); IgM acute 1.78 (1.05,3.0), convalescent 4.38 (2.6,7.3). We conclude that ELISA is a specific, sensitive and reproducible method for the detection of antibodies to inner core LPS of Nm and that an epitope defined by MAb B5 can be immunogenic in infants and adults. These findings are relevant to the potential candidacy of inner core LPS as a vaccine.
Assuntos
Anticorpos Antibacterianos/sangue , Ensaio de Imunoadsorção Enzimática/métodos , Lipopolissacarídeos/imunologia , Neisseria meningitidis/imunologia , Doença Aguda , Adolescente , Adulto , Idoso , Animais , Anticorpos Monoclonais , Sequência de Carboidratos , Estudos de Casos e Controles , Criança , Pré-Escolar , Ensaio de Imunoadsorção Enzimática/estatística & dados numéricos , Genes Bacterianos , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Lactente , Lipopolissacarídeos/química , Infecções Meningocócicas/imunologia , Camundongos , Pessoa de Meia-Idade , Dados de Sequência Molecular , Mutação , Neisseria meningitidis/classificação , Neisseria meningitidis/genética , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , SorotipagemRESUMO
Experiments were carried out to compare the sensitivity of Ancylostoma ceylanicum and Necator americanus to ivermectin (IVM) and pyrantel in vitro. Loss of motility and inhibition of ingestion by IVM were compared and A. ceylanicum was found to be approximately 40-50 times more sensitive to IVM than N. americanus. Both species showed a similar sensitivity to pyrantel. Uptake of [3H]IVM across the cuticle was compared and shown to be unlikely to account for the differences in sensitivity to IVM between the two species.
Assuntos
Ancylostoma/efeitos dos fármacos , Antinematódeos/farmacologia , Ivermectina/farmacologia , Necator/efeitos dos fármacos , Animais , Antinematódeos/farmacocinética , Cricetinae , Interações Medicamentosas , Feminino , Inulina/farmacocinética , Ivermectina/farmacocinética , Masculino , Pirantel/farmacologiaRESUMO
Representative strains of Bordetella bronchiseptica and B. parapertussis were found to produce smooth lipopolysaccharides (LPS) having identical antigenic O-polysaccharide components composed of linear unbranched polymers of 1,4-linked 2,3-diacetamido-2,3-dideoxy-alpha-L-galacto-pyranosyluronic acid residues. These LPSs differed from the LPS of B. pertussis which produces only rough-type LPS, devoid of O-polysaccharide. While B. bronchiseptica and B. parapertussis had chemically and immunologically identical O-polysaccharide structures, their core oligosaccharide components differed. The core oligosaccharide of B. parapertussis was chemically distinct from the core of B. bronchiseptica which appeared to be structurally and immunologically similar to a core oligosaccharide of B. pertussis.
Assuntos
Antígenos de Bactérias/química , Bordetella bronchiseptica/imunologia , Bordetella/imunologia , Lipopolissacarídeos/química , Eletroforese em Gel de Poliacrilamida , Espectroscopia de Ressonância Magnética , Especificidade da EspécieRESUMO
By deletion mutagenesis in the entire meningococcal chromosome, we have previously identified the icsA gene, which encodes the glycosyltransferase required for adding GlcNAc to Hep-II in the inner core of meningococcal LPS. This gene has homology to several LPS glycosyltransferases, notably to rfaK from Salmonella typhimurium and bplH from Bordetella pertussis, both of which encode GlcNAc transferases. Directly upstream of icsA is an ORF showing significant homology to the hypothetical protein HI0653 from the Haemophilus influenzae genome sequence, and to a lesser degree to putative glycosyltransferases from Streptococcus thermophilus and Yersinia enterocolitica. Insertional inactivation of this ORF resulted in a meningococcal strain with truncated LPS. We have named this new LPS-involved gene icsB. Differences in binding of monoclonal antibodies and in mobility on Tricine-SDS-PAGE showed that LPS from icsA and icsB mutants is similar but not identical. On the basis of these results, we postulated that the new gene encodes the glycosyltransferase required for adding Glc to Hep-I. Structural analysis of purified mutant LPS by electrospray mass spectrometry was used to verify this hypothesis. The composition determined for icsA and icsB is lipidA-(KDO)2-(Hep)2.PEA and lipidA-(KDO)2-(Hep)2.PEA-GlcNAc, respectively. The icsA and icsB genes thus form an operon encoding the glycosyltransferases required for chain elongation from the lipidA-(KDO)2-(Hep)2 basal structure, with IcsA first adding GlcNAc to Hep-II and IcsB subsequently adding Glc to Hep-I. Only then is completion of the lacto-N-neotetraose structure possible through the action of the IgtA-E genes.
Assuntos
Lipopolissacarídeos/metabolismo , Neisseria meningitidis/genética , Acetilglucosamina/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Southern Blotting , Glicosiltransferases/genética , Glicosiltransferases/metabolismo , Heptoses/metabolismo , Dados de Sequência Molecular , Mutagênese/fisiologia , Neisseria meningitidis/classificação , Neisseria meningitidis/enzimologia , Proteínas Recombinantes/genética , SorotipagemRESUMO
Research into the relationship between the Type A behavior pattern and coronary heart disease suggests that the anger-hostility-aggression (AHA!) syndrome is directly related to total serum cholesterol and low-density lipoproteins. The present study involved an investigation of the specific components of the AHA! syndrome related to blood lipid levels in 98 healthy men. The disposition to experience and express anger when frustrated, criticized, or treated unfairly (angry reaction, a component of trait anger) was related to total serum cholesterol and to low-density lipoprotein levels. Age and diet also predicted levels of these lipids, but each was unrelated to angry reaction. These results suggest that in healthy men, the experience of strong angry affect in reaction to perceived rejection, criticism, or unfair treatment may be health-toxic because of its relationship to elevated unfavorable serum lipids.
Assuntos
Ira , Hostilidade , Lipídeos/sangue , Personalidade Tipo A , Adulto , Agressão , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
Recent information-processing studies have suggested that a selective attention deficit may be involved in the symptomatology of obsessive-compulsive disorder (OCD). In this study, individuals diagnosed with OCD were distinguished from those with panic disorder and from control participants by their relatively poorer performance on a series of psychometric tasks of selective attention. These results are interpreted as supporting the hypothesis of a diminished ability of people with OCD to selectively ignore competing external (sensory) and internal (cognitive) stimuli, especially intrusive thoughts.
Assuntos
Transtorno do Deficit de Atenção com Hiperatividade/etiologia , Transtorno Obsessivo-Compulsivo/complicações , Transtorno de Pânico/complicações , Adulto , Análise de Variância , Transtornos de Ansiedade/diagnóstico , Transtornos de Ansiedade/etiologia , Atenção/fisiologia , Transtorno do Deficit de Atenção com Hiperatividade/diagnóstico , Transtornos Cognitivos/diagnóstico , Transtornos Cognitivos/etiologia , Depressão/diagnóstico , Depressão/etiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Testes Psicológicos , Desempenho Psicomotor , Tempo de ReaçãoRESUMO
The present investigation describes the use of on-line chromatographic preconcentration coupled to capillary zone electrophoresis-electrospray mass spectrometry (cPC-CZE-ES-MS) for trace level analysis of negatively charged lipopolysaccharides (LPS) obtained from pathogenic strains of Haemophilus influenzae. The analytical performance of two different types of adsorption media [i.e., C18 irregular particles and poly(styrene-divinylbenzene) membrane] for anionic analytes was first evaluated using a mixture of peptide standards to determine the overall sensitivity of this approach. These chromatographic preconcentrators provided an enhancement of sample loadings of up to 5 microliters with good linear response and low nM concentration detection limits for most peptides investigated. The application of cPC-CZE-ES-MS is further demonstrated for extracts of O-deacylated LPS obtained from H. influenzae strain Eagan. In combination with novel enzymatic releasing methods using proteinase K, this technique provides unparalleled sensitivity and enabled the identification of LPS surface antigens from as little as five bacterial colonies.
Assuntos
Eletroforese Capilar/métodos , Lipopolissacarídeos/análise , Espectrometria de Massas/métodos , Sequência de Aminoácidos , Sequência de Carboidratos , Haemophilus influenzae/química , Lipopolissacarídeos/química , Dados de Sequência Molecular , Sensibilidade e EspecificidadeRESUMO
In the catastrophic misinterpretation model of panic Clark [Behav. Res. Ther. 24(1986)1461] proposes that panic attacks result from the misinterpretation of autonomic arousal stimuli as precursors to a physical or psychological emergency. The model has been widely examined, with many researchers suggesting that this specific cognitive bias is implicated in both the phenomenon of panic, and the aetiology and maintenance of panic disorder. Various research methodologies have provided only partial or inconclusive support for the model as being uniquely associated with panic, and as a cognitive process underpinning the experience of panic. This paper reviews the body of existing evidence and its implications for the model and proposes future research directions. The influence of implicit operational definitions of key terms in the catastrophic misinterpretation literature (e.g. 'catastrophe', 'threat', 'anxiety-related') are examined, and clarifications proposed. Inconsistencies and limitations in the measurement of catastrophic misinterpretation are highlighted, and subsequently developments to measurement instruments are proposed.
Assuntos
Nível de Alerta , Mecanismos de Defesa , Transtorno de Pânico/psicologia , Teoria Psicológica , Atenção , Humanos , IndividualidadeRESUMO
The aqueous-phase lipopolysaccharide isolated from Pasteurella haemolytica serotype T10 cells by the phenol-water extraction method was found to be S-type lipopolysaccharide which possessed O-antigenic polysaccharide chains composed only of D-galactose residues. Structural analysis of the O-polysaccharide, using a combination of 1D and 2D 1H- and 13C-n.m.r. methods, led to the identification of the disaccharide repeating-unit as [----3)-alpha-D-Galp-(1----3)-beta-D-Galf-(1----]n. The serological cross-reactivity between P. haemolytica serotypes T4 and T10 can now be related to the structural similarity of the antigenic LPS O-polysaccharides.
Assuntos
Antígenos de Bactérias , Lipopolissacarídeos , Espectroscopia de Ressonância Magnética , Pasteurella/imunologia , Antígenos de Bactérias/imunologia , Configuração de Carboidratos , Dissacarídeos , Lipopolissacarídeos/imunologia , Estrutura Molecular , Antígenos O , Pasteurella/classificação , SorotipagemRESUMO
Lipopolysaccharide was isolated from Pasteurella haemolytica biotype A, serotype 1 by using the phenol-water extraction procedure. Hydrolysis with mild acid afforded a high-molecular-weight antigenic O-chain. On the basis of 1D and 2D NMR spectral studies and microanalytical chemical methods, the O-polysaccharide was determined to be a linear polymer of a trisaccharide repeating unit having the structure -->3)-beta-D-Galp-(1-->3)-beta-D-GalpNAc-(1-->4)-beta-D-Galp-(1--> This O-polysaccharide antigen is expressed by several P. haemolytica biotype A serotypes.
Assuntos
Antígenos de Bactérias/química , Mannheimia haemolytica/química , Polissacarídeos Bacterianos/química , Antígenos de Bactérias/isolamento & purificação , Sequência de Carboidratos , Espectroscopia de Ressonância Magnética , Mannheimia haemolytica/imunologia , Dados de Sequência Molecular , Antígenos O , Polissacarídeos Bacterianos/imunologia , Polissacarídeos Bacterianos/isolamento & purificação , Análise de Sequência , SorotipagemRESUMO
The specific capsular polysaccharide produced by Rhodococcus equi serotype 2 is a high-molecular-weight acidic polymer composed of D-glucose, D-mannose, D-glucuronic acid and 3-O-[(S)-1-carboxyethyl]-L-rhamnose in equimolar proportions. Structural analysis, employing a combination of chemical and n.m.r. techniques, established that the polysaccharide is composed of linear repeating tetrasaccharide units. (formula; see text) in which the beta-D-mannose residues carry O-acetyl groups at O-2 and O-3 to the extent of 1.7 mol equivalents. Unequivocal determination of the absolute chirality of the 3-O-[(S)-1-carboxyethyl]-alpha-L-rhamnose residues was achieved by chemical correlation with an authentic synthetic sample. The 1H and 13C-n.m.r. resonances of the native and O-deacetylated serotype 2 polysaccharides were fully assigned by homo- and heteronuclear chemical-shift correlation methods.
Assuntos
Polissacarídeos Bacterianos/química , Rhodococcus/análise , Configuração de Carboidratos , Sequência de Carboidratos , Espectroscopia de Ressonância Magnética , Dados de Sequência Molecular , Estrutura Molecular , Rhodococcus/classificação , SorotipagemRESUMO
Structural analysis of the specific capsular polysaccharide produced by Rhodococcus equi serotype 7 indicated it to be a high-molecular-weight polymer consisting of equal molar amounts of D-galactose, D-mannose, L-rhamnose, and pyruvic acid. By employing a combination of chemical and NMR techniques, it was established that the polysaccharide is composed of the linear repeating trisaccharide units: [formula: see text] -->3)-alpha-D-Galp-(1-->3)-alpha-D-Manp-(1-->3)-alpha-L-Rhap -(1-->, in which the cyclic pyruvic acid acetal groups bridging the O-4 and O-6 positions of the alpha-D-Manp residues have the S-configuration. The 1H and 13C NMR spectra of the native and pyruvic acetal-free polysaccharides were fully assigned.
Assuntos
Cápsulas Bacterianas/química , Polissacarídeos Bacterianos/química , Rhodococcus equi/química , Sequência de Carboidratos , Isótopos de Carbono , Galactose/análise , Espectroscopia de Ressonância Magnética , Manose/análise , Dados de Sequência Molecular , Polissacarídeos Bacterianos/isolamento & purificação , Prótons , Piruvatos/análise , Ácido Pirúvico , Ramnose/análise , Rhodococcus equi/imunologia , Análise de SequênciaRESUMO
The specific capsular polysaccharide produced by Rhodococcus equi serotype 4 was found to be a high-molecular-weight acidic polymer composed of D-glucose, D-mannose, pyruvic acid and a previously unidentified 5-amino-3,5-dideoxynonulosonic (rhodaminic) acid in the proportions 2:1:1:1. Structural analysis, employing a combination of microanalytical methods, nuclear magnetic resonance spectroscopy, and mass spectrometric techniques, established that the polysaccharide consisted of linear repeating tetrasaccharide units having the sequence of residues shown below. In the native polysaccharide, the rhodaminic acid residues were present as their acetamido derivatives (RhoANAc) and carried 1-carboxyethylidene groups that bridged the O-7 and O-9 positions. Treatment of the capsular polysaccharide with dilute acetic acid and/or anhydrous hydrogen fluoride under hydrolytic/solvolytic conditions, resulted in the formation of four different oligosaccharide species. The 1H and 13C NMR resonances of these oligosaccharide fragments and of the native serotype 4 capsular polysaccharides were fully assigned by homo- and heteronuclear chemical shift correlation methods.
Assuntos
Polissacarídeos/química , Rhodococcus equi/química , Animais , Configuração de Carboidratos , Sequência de Carboidratos , Espectroscopia de Ressonância Magnética , Dados de Sequência Molecular , Polissacarídeos/isolamento & purificação , Polissacarídeos Bacterianos/química , Polissacarídeos Bacterianos/isolamento & purificação , Sorotipagem , Espectrometria de Massas de Bombardeamento Rápido de ÁtomosRESUMO
The capsular polysaccharide of A. pleuropneumoniae serotype 10 was found to be composed of a linear disaccharide repeating unit. By use of methylation analysis, partial hydrolysis, g.l.c.--and f.a.b.- m.s., and 1D and 2D n.m.r. studies, the polysaccharide was determined to be a polymer of a repeating disaccharide unit composed of D-mannose and 3-deoxy-D-manno-octulosonic acid (Kdo) having the structure: [formula; see text]