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1.
J Biol Chem ; 290(32): 19512-26, 2015 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-26109072

RESUMO

The incidence of multidrug-resistant Enterococcus faecium hospital infections has been steadily increasing. With the goal of discovering new vaccine antigens, we systematically fractionated and purified four distinct surface carbohydrates from E. faecium endocarditis isolate Tx16, shown previously to be resistant to phagocytosis in the presence of human serum. The two most abundant polysaccharides consist of novel branched heteroglycan repeating units that include signature sugars altruronic acid and legionaminic acid, respectively. A minor high molecular weight polysaccharide component was recognized as the fructose homopolymer levan, and a glucosylated lipoteichoic acid (LTA) was identified in a micellar fraction. The polysaccharides were conjugated to the CRM197 carrier protein, and the resulting glycoconjugates were used to immunize rabbits. Rabbit immune sera were evaluated for their ability to kill Tx16 in opsonophagocytic assays and in a mouse passive protection infection model. Although antibodies raised against levan failed to mediate opsonophagocytic killing, the other glycoconjugates induced effective opsonic antibodies, with the altruronic acid-containing polysaccharide antisera showing the greatest opsonophagocytic assay activity. Antibodies directed against either novel heteroglycan or the LTA reduced bacterial load in mouse liver or kidney tissue. To assess antigen prevalence, we screened a diverse collection of blood isolates (n = 101) with antibodies to the polysaccharides. LTA was detected on the surface of 80% of the strains, and antigens recognized by antibodies to the two major heteroglycans were co-expressed on 63% of these clinical isolates. Collectively, these results represent the first steps toward identifying components of a glycoconjugate vaccine to prevent E. faecium infection.


Assuntos
Anticorpos Antibacterianos/biossíntese , Antígenos de Bactérias/química , Vacinas Bacterianas/imunologia , Enterococcus faecium/imunologia , Infecções por Bactérias Gram-Positivas/prevenção & controle , Animais , Antibacterianos/uso terapêutico , Anticorpos Antibacterianos/sangue , Antígenos de Bactérias/imunologia , Carga Bacteriana/efeitos dos fármacos , Proteínas de Bactérias/química , Proteínas de Bactérias/imunologia , Vacinas Bacterianas/administração & dosagem , Vacinas Bacterianas/biossíntese , Vacinas Bacterianas/genética , Sequência de Carboidratos , Modelos Animais de Doenças , Farmacorresistência Bacteriana Múltipla , Enterococcus faecium/química , Feminino , Frutanos/química , Frutanos/imunologia , Infecções por Bactérias Gram-Positivas/sangue , Infecções por Bactérias Gram-Positivas/imunologia , Infecções por Bactérias Gram-Positivas/microbiologia , Humanos , Soros Imunes/química , Lipopolissacarídeos/química , Lipopolissacarídeos/imunologia , Camundongos , Dados de Sequência Molecular , Proteínas Opsonizantes/química , Proteínas Opsonizantes/imunologia , Coelhos , Ácidos Siálicos/química , Ácidos Siálicos/imunologia , Ácidos Teicoicos/química , Ácidos Teicoicos/imunologia , Ácidos Urônicos/química , Ácidos Urônicos/imunologia , Vacinas Conjugadas
2.
Carbohydr Res ; 351: 98-107, 2012 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-22341920

RESUMO

The original structure of Streptococcus pneumoniae capsular polysaccharide (CPS) serotype 6C was proposed based on chemical degradation and tandem mass analysis [J. Clin. Microbiol.2007, 45, 1225-1233]. In order to confirm the repeat unit structure and assign the stereochemical structure, the CPS 6C and the known CPS 6A were fully characterized by NMR spectroscopy. Full (1)H and (13)C NMR spectra assignments of CPS 6C and CPS 6A were achieved based on DQCOSY, TOCSY, HSQC, HMBC, and NOESY analysis. These analyses confirmed the published structure of CPS 6A and established the repeat unit structure of the CPS 6C as: →2)-α-D-Glcp-(1→3)-α-D-Glcp-(1→3)-α-L-Rhap-(1→3)-D-Ribitol-(5→phosphate-.


Assuntos
Polissacarídeos Bacterianos/química , Streptococcus pneumoniae/química , Cápsulas , Sequência de Carboidratos , Espectroscopia de Ressonância Magnética , Dados de Sequência Molecular , Monossacarídeos/química , Controle de Qualidade , Estereoisomerismo
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