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1.
Nature ; 577(7791): 543-548, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31915378

RESUMO

Although maternal antibodies protect newborn babies from infection1,2, little is known about how protective antibodies are induced without prior pathogen exposure. Here we show that neonatal mice that lack the capacity to produce IgG are protected from infection with the enteric pathogen enterotoxigenic Escherichia coli by maternal natural IgG antibodies against the maternal microbiota when antibodies are delivered either across the placenta or through breast milk. By challenging pups that were fostered by either maternal antibody-sufficient or antibody-deficient dams, we found that IgG derived from breast milk was crucial for protection against mucosal disease induced by enterotoxigenic E. coli. IgG also provides protection against systemic infection by E. coli. Pups used the neonatal Fc receptor to transfer IgG from milk into serum. The maternal commensal microbiota can induce antibodies that recognize antigens expressed by enterotoxigenic E. coli and other Enterobacteriaceae species. Induction of maternal antibodies against a commensal Pantoea species confers protection against enterotoxigenic E. coli in pups. This role of the microbiota in eliciting protective antibodies to a specific neonatal pathogen represents an important host defence mechanism against infection in neonates.


Assuntos
Anticorpos/imunologia , Escherichia coli Enterotoxigênica/imunologia , Infecções por Escherichia coli/imunologia , Infecções por Escherichia coli/prevenção & controle , Imunidade Materno-Adquirida/imunologia , Recém-Nascido/imunologia , Microbiota/imunologia , Leite Humano/imunologia , Animais , Anticorpos/sangue , Anticorpos/metabolismo , Aleitamento Materno , Reações Cruzadas/imunologia , Infecções por Escherichia coli/microbiologia , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina G/metabolismo , Masculino , Camundongos , Mães , Pantoea/imunologia , Receptores Fc/imunologia , Receptores Fc/metabolismo , Simbiose/imunologia
2.
Biochemistry ; 61(24): 2849-2855, 2022 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-35993915

RESUMO

Vaccination shows high variability in the elicited immune responses among individuals and populations for reasons still poorly understood. An increasing number of studies is supporting the evidence that gut microbiota, along with other interplaying variables, is able to modulate both humoral and cellular responses to infection and vaccination. Importantly, vaccine immunogenicity is often suboptimal at the extremes of age and also in low- and middle-income countries (LMICs), where the microbiota is believed to have an important role on immune responses. Still, contrasting findings and lack of causal evidence are calling for sophisticated methodologies to be able to overcome scientific and technical challenges to better decipher the immunomodulatory role of microbiota. In this perspective, we briefly review the status of the vaccine field in relation to the microbiome and offer possible scientific approaches to better understand the impact of the host microbiome on vaccine responsiveness.


Assuntos
Microbioma Gastrointestinal , Microbiota , Vacinas , Humanos , Vacinação
3.
Proc Natl Acad Sci U S A ; 116(1): 193-198, 2019 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-30510007

RESUMO

Glycoconjugate vaccines are among the most effective interventions for preventing several serious infectious diseases. Covalent linkage of the bacterial capsular polysaccharide to a carrier protein provides CD4+ T cells with epitopes that facilitate a memory response to the polysaccharide. Classically, the mechanism responsible for antigen processing was thought to be similar to what was known for hapten-carrier conjugates: protease digestion of the carrier protein in the endosome and presentation of a resulting peptide to the T cell receptor on classical peptide-recognizing CD4+ T cells. Recently, an alternative mechanism has been shown to be responsible for the memory response to some glycoconjugates. Processing of both the protein and the polysaccharide creates glycopeptides in the endosome of antigen-presenting cells. For presentation, the peptide portion of the glycopeptide is bound to MHCII, allowing the covalently linked glycan to activate carbohydrate-specific helper CD4+ T cells (Tcarbs). Herein, we assessed whether this same mechanism applies to conjugates prepared from other capsular polysaccharides. All of the glycoconjugates tested induced Tcarb-dependent responses except that made with group C Neisseria meningitidis; in the latter case, only peptides generated from the carrier protein were critical for helper T cell recognition. Digestion of this acid-sensitive polysaccharide, a linear homopolymer of α(2 → 9)-linked sialic acid, to the size of the monomeric unit resulted in a dominant CD4+ T cell response to peptides in the context of MHCII. Our results show that different mechanisms of presentation, based on the structure of the carbohydrate, are operative in response to different glycoconjugate vaccines.


Assuntos
Imunidade Adaptativa/imunologia , Glicoconjugados/imunologia , Polissacarídeos Bacterianos/imunologia , Vacinas Conjugadas/imunologia , Imunidade Adaptativa/efeitos dos fármacos , Animais , Formação de Anticorpos/imunologia , Antígenos/imunologia , Epitopos/imunologia , Feminino , Citometria de Fluxo , Glicoconjugados/farmacologia , Vacinas Meningocócicas/imunologia , Vacinas Meningocócicas/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Ácido N-Acetilneuramínico , Neisseria meningitidis/imunologia , Polissacarídeos Bacterianos/farmacologia , Linfócitos T Auxiliares-Indutores/imunologia , Vacinas Conjugadas/farmacologia
4.
Proc Natl Acad Sci U S A ; 116(14): 7062-7070, 2019 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-30872471

RESUMO

Francisella tularensis is the causative agent of tularemia, a category A bioterrorism agent. The lipopolysaccharide (LPS) O antigen (OAg) of F. tularensis has been considered for use in a glycoconjugate vaccine, but conjugate vaccines tested so far have failed to confer protection necessary against aerosolized pulmonary bacterial challenge. When F. tularensis OAg was purified under standard conditions, the antigen had a small molecular size [25 kDa, low molecular weight (LMW)]. Using milder extraction conditions, we found the native OAg had a larger molecular size [80 kDa, high molecular weight (HMW)], and in a mouse model of tularemia, a glycoconjugate vaccine made with the HMW polysaccharide coupled to tetanus toxoid (HMW-TT) conferred better protection against intranasal challenge than a conjugate made with the LMW polysaccharide (LMW-TT). To further investigate the role of OAg size in protection, we created an F. tularensis live vaccine strain (LVS) mutant with a significantly increased OAg size [220 kDa, very high molecular weight (VHMW)] by expressing in F. tularensis a heterologous chain-length regulator gene (wzz) from the related species Francisella novicida Immunization with VHMW-TT provided markedly increased protection over that obtained with TT glycoconjugates made using smaller OAgs. We found that protective antibodies recognize a length-dependent epitope better expressed on HMW and VHMW antigens, which bind with higher affinity to the organism.


Assuntos
Vacinas Bacterianas/imunologia , Francisella tularensis/imunologia , Glicoconjugados/imunologia , Antígenos O/imunologia , Tularemia , Animais , Vacinas Bacterianas/farmacologia , Feminino , Glicoconjugados/farmacologia , Glicoconjugados/fisiologia , Camundongos , Camundongos Endogâmicos BALB C , Antígenos O/farmacologia , Tularemia/imunologia , Tularemia/patologia , Tularemia/prevenção & controle
5.
Proc Natl Acad Sci U S A ; 116(52): 26157-26166, 2019 Dec 26.
Artigo em Inglês | MEDLINE | ID: mdl-31811024

RESUMO

The mammalian immune system is tolerized to trillions of microbes residing on bodily surfaces and can discriminate between symbionts and pathogens despite their having related microbial structures. Mechanisms of innate immune activation and the subsequent signaling pathways used by symbionts to communicate with the adaptive immune system are poorly understood. Polysaccharide A (PSA) of Bacteroides fragilis is the model symbiotic immunomodulatory molecule. Here we demonstrate that PSA-dependent immunomodulation requires the Toll-like receptor (TLR) 2/1 heterodimer in cooperation with Dectin-1 to initiate signaling by the downstream phosphoinositide 3-kinase (PI3K) pathway, with consequent CREB-dependent transcription of antiinflammatory genes, including antigen presentation and cosignaling molecules. High-resolution LC-MS/MS analysis of PSA identified a previously unknown small molecular-weight, covalently attached bacterial outer membrane-associated lipid that is required for activation of antigen-presenting cells. This archetypical commensal microbial molecule initiates a complex collaborative integration of Toll-like receptor and C-type lectin-like receptor signaling mechanisms culminating in the activation of the antiinflammatory arm of the PI3K pathway that serves to educate CD4+ Tregs to produce the immunomodulatory cytokine IL-10. Immunomodulation is a key function of the microbiome and is a focal point for developing new therapeutic agents.

6.
Molecules ; 27(19)2022 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-36234967

RESUMO

Glycoconjugate vaccines have contributed enormously to reducing and controlling encapsulated bacterial infections for over thirty years. Glycoconjugate vaccines are based on a carbohydrate antigen that is covalently linked to a carrier protein; this is necessary to cause T cell responses for optimal immunogenicity, and to protect young children. Many interdependent parameters affect the immunogenicity of glycoconjugate vaccines, including the size of the saccharide antigen. Here, we examine and discuss the impact of glycan chain length on the efficacy of glycoconjugate vaccines and report the methods employed to size polysaccharide antigens, while highlighting the underlying reaction mechanisms. A better understanding of the impact of key parameters on the immunogenicity of glycoconjugates is critical to developing a new generation of highly effective vaccines.


Assuntos
Glicoconjugados , Açúcares , Antígenos , Carboidratos , Proteínas de Transporte , Criança , Pré-Escolar , Humanos , Polissacarídeos , Vacinas Conjugadas
7.
Bioconjug Chem ; 26(12): 2507-13, 2015 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-26549104

RESUMO

A solid-phase conjugation method was developed and applied to the synthesis of an O-antigen based glycoconjugate vaccine against Salmonella Typhimurium, with CRM197 as the carrier protein. Copper-free click chemistry was used as the conjugation chemistry, after derivatizing the sugar and the protein components with alkyne and azido linkers, respectively. This chemistry has the advantage of not deactivating functional groups during the conjugation step, thereby allowing the recycling of unreacted components. The activated carrier protein was adsorbed to an anion exchange matrix and quantitatively conjugated to the O-antigen. The resulting conjugate was eluted from the resin free of unconjugated sugar which was previously removed by simple washing steps. Unreacted O-antigen was recycled by addition to a new batch of resin-CRM197 resulting in further quantitative protein conjugation. This process has advantages in relation to reduction of costs for production of conjugate vaccines, allowing unreacted sugar recovery and simplifying the purification of the glycoconjugate.


Assuntos
Química Click/métodos , Glicoconjugados/química , Antígenos O/química , Salmonella typhimurium/química , Técnicas de Síntese em Fase Sólida/métodos , Vacinas Conjugadas/química , Proteínas de Bactérias/química , Humanos , Modelos Moleculares , Infecções por Salmonella/microbiologia , Infecções por Salmonella/prevenção & controle
8.
Angew Chem Int Ed Engl ; 54(45): 13198-203, 2015 Nov 02.
Artigo em Inglês | MEDLINE | ID: mdl-26350581

RESUMO

A series of glycoconjugates with defined connectivity were synthesized to investigate the impact of coupling Salmonella typhimurium O-antigen to different amino acids of CRM197 protein carrier. In particular, two novel methods for site-selective glycan conjugation were developed to obtain conjugates with single attachment site on the protein, based on chemical modification of a disulfide bond and pH-controlled transglutaminase-catalyzed modification of lysine, respectively. Importantly, conjugation at the C186-201 bond resulted in significantly higher anti O-antigen bactericidal antibody titers than coupling to K37/39, and in comparable titers to conjugates bearing a larger number of saccharides. This study demonstrates that the conjugation site plays a role in determining the immunogenicity in mice and one single attachment point may be sufficient to induce high levels of bactericidal antibodies.


Assuntos
Glicoconjugados/química , Glicoconjugados/imunologia , Antígenos O/química , Antígenos O/imunologia , Vacinas contra Salmonella/química , Vacinas contra Salmonella/imunologia , Salmonella typhimurium/imunologia , Animais , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Moleculares , Conformação Molecular , Salmonella typhimurium/química
9.
Front Mol Biosci ; 10: 1201693, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37261327

RESUMO

Vaccines are cost-effective tools for reducing morbidity and mortality caused by infectious diseases. The rapid evolution of pneumococcal conjugate vaccines, the introduction of tetravalent meningococcal conjugate vaccines, mass vaccination campaigns in Africa with a meningococcal A conjugate vaccine, and the recent licensure and introduction of glycoconjugates against S. Typhi underlie the continued importance of research on glycoconjugate vaccines. More innovative ways to produce carbohydrate-based vaccines have been developed over the years, including bioconjugation, Outer Membrane Vesicles (OMV) and the Multiple antigen-presenting system (MAPS). Several variables in the design of these vaccines can affect the induced immune responses. We review immunogenicity studies comparing conjugate vaccines that differ in design variables, such as saccharide chain length and conjugation chemistry, as well as carrier protein and saccharide to protein ratio. We evaluate how a better understanding of the effects of these different parameters is key to designing improved glycoconjugate vaccines.

10.
Viruses ; 15(5)2023 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-37243247

RESUMO

The humoral response after vaccination was evaluated in 1248 individuals who received different COVID-19 vaccine schedules. The study compared subjects primed with adenoviral ChAdOx1-S (ChAd) and boosted with BNT162b2 (BNT) mRNA vaccines (ChAd/BNT) to homologous dosing with BNT/BNT or ChAd/ChAd vaccines. Serum samples were collected at two, four and six months after vaccination, and anti-Spike IgG responses were determined. The heterologous vaccination induced a more robust immune response than the two homologous vaccinations. ChAd/BNT induced a stronger immune response than ChAd/ChAd at all time points, whereas the differences between ChAd/BNT and BNT/BNT decreased over time and were not significant at six months. Furthermore, the kinetic parameters associated with IgG decay were estimated by applying a first-order kinetics equation. ChAd/BNT vaccination was associated with the longest time of anti-S IgG negativization and with a slow decay of the titer over time. Finally, analyzing factors influencing the immune response by ANCOVA analysis, it was found that the vaccine schedule had a significant impact on both the IgG titer and kinetic parameters, and having a Body Mass Index (BMI) above the overweight threshold was associated with an impaired immune response. Overall, the heterologous ChAd/BNT vaccination may offer longer-lasting protection against SARS-CoV-2 than homologous vaccination strategies.


Assuntos
Vacinas contra COVID-19 , COVID-19 , Humanos , Estudos Longitudinais , Vacina BNT162 , COVID-19/prevenção & controle , SARS-CoV-2 , Vacinação , ChAdOx1 nCoV-19 , Imunoglobulina G , Anticorpos Antivirais , Anticorpos Neutralizantes
11.
Front Cell Infect Microbiol ; 11: 808005, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-35118012

RESUMO

Carbohydrates are ubiquitous molecules expressed on the surface of nearly all living cells, and their interaction with carbohydrate-binding proteins is critical to many immunobiological processes. Carbohydrates are utilized as antigens in many licensed vaccines against bacterial pathogens. More recently, they have also been considered as adjuvants. Interestingly, unlike other types of vaccines, adjuvants have improved immune response to carbohydrate-based vaccine in humans only in a few cases. Furthermore, despite the discovery of many new adjuvants in the last years, aluminum salts, when needed, remain the only authorized adjuvant for carbohydrate-based vaccines. In this review, we highlight historical and recent advances on the use of glycans either as vaccine antigens or adjuvants, and we review the use of currently available adjuvants to improve the efficacy of carbohydrate-based vaccines. A better understanding of the mechanism of carbohydrate interaction with innate and adaptive immune cells will benefit the design of a new generation of glycan-based vaccines and of immunomodulators to fight both longstanding and emerging diseases.


Assuntos
Doenças Transmissíveis , Vacinas , Adjuvantes Imunológicos , Antígenos , Carboidratos , Humanos
12.
Pathogens ; 10(6)2021 Jun 09.
Artigo em Inglês | MEDLINE | ID: mdl-34207575

RESUMO

GMMA, outer membrane vesicles resulting from hyperblebbing mutated bacterial strains, are a versatile vaccine platform for displaying both homologous and heterologous antigens. Periplasmic expression is a popular technique for protein expression in the lumen of the blebs. However, the ability of internalized antigens to induce antibody responses has not been extensively investigated. Herein, the Neisseria meningitidis factor H binding protein (fHbp) was heterologously expressed in the lumen of O-antigen positive (OAg+) and O-antigen negative (OAg-) Salmonella Typhimurium GMMA. Only the OAg- GMMA induced an anti-fHbp IgG response in mice if formulated on Alum, although it was weak and much lower compared to the recombinant fHbp. The OAg- GMMA on Alum showed partial instability, with possible exposure of fHbp to the immune system. When we chemically conjugated fHbp to the surface of both OAg+ and OAg- GMMA, these constructs induced a stronger functional response compared to the fHbp immunization alone. Moreover, the OAg+ GMMA construct elicited a strong response against both the target antigens (fHbp and OAg), with no immune interference observed. This result suggests that antigen localization on GMMA surface can play a critical role in the induction of an effective immune response and can encourage the development of GMMA based vaccines delivering key protective antigens on their surface.

14.
PLoS One ; 10(10): e0139847, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26445460

RESUMO

Nontyphoidal Salmonellae, principally S. Typhimurium and S. Enteritidis, are a major cause of invasive bloodstream infections in sub-Saharan Africa with no vaccine currently available. Conjugation of lipopolysaccharide O-antigen to a carrier protein constitutes a promising vaccination strategy. Here we describe a rational process to select the most appropriate isolates of Salmonella as source of O-antigen for developing a bivalent glycoconjugate vaccine. We screened a library of 30 S. Typhimurium and 21 S. Enteritidis in order to identify the most suitable strains for large scale O-antigen production and generation of conjugate vaccines. Initial screening was based on growth characteristics, safety profile of the isolates, O-antigen production, and O-antigen characteristics in terms of molecular size, O-acetylation and glucosylation level and position, as determined by phenol sulfuric assay, NMR, HPLC-SEC and HPAEC-PAD. Three animal isolates for each serovar were identified and used to synthesize candidate glycoconjugate vaccines, using CRM197 as carrier protein. The immunogenicity of these conjugates and the functional activity of the induced antibodies was investigated by ELISA, serum bactericidal assay and flow cytometry. S. Typhimurium O-antigen showed high structural diversity, including O-acetylation of rhamnose in a Malawian invasive strain generating a specific immunodominant epitope. S. Typhimurium conjugates provoked an anti-O-antigen response primarily against the O:5 determinant. O-antigen from S. Enteritidis was structurally more homogeneous than from S. Typhimurium, and no idiosyncratic antibody responses were detected for the S. Enteritidis conjugates. Of the three initially selected isolates, two S. Typhimurium (1418 and 2189) and two S. Enteritidis (502 and 618) strains generated glycoconjugates able to induce high specific antibody levels with high breadth of serovar-specific strain coverage, and were selected for use in vaccine production. The strain selection approach described is potentially applicable to the development of glycoconjugate vaccines against other bacterial pathogens.


Assuntos
Glicoconjugados/química , Antígenos O/química , Infecções por Salmonella/prevenção & controle , Vacinas contra Salmonella/imunologia , Vacinas Conjugadas/imunologia , Acetilação , Anticorpos Antibacterianos/análise , Anticorpos Antibacterianos/imunologia , Proteínas de Bactérias/química , Proteínas de Bactérias/imunologia , Proteínas de Bactérias/metabolismo , Cromatografia em Gel , Cromatografia Líquida de Alta Pressão , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Espectroscopia de Ressonância Magnética , Salmonella enteritidis/imunologia , Salmonella enteritidis/isolamento & purificação , Salmonella enteritidis/metabolismo , Salmonella typhimurium/imunologia , Salmonella typhimurium/isolamento & purificação , Salmonella typhimurium/metabolismo
15.
Carbohydr Res ; 385: 1-8, 2014 Feb 19.
Artigo em Inglês | MEDLINE | ID: mdl-24384528

RESUMO

Salmonella Typhimurium is the major cause of invasive nontyphoidal Salmonella disease in Africa, with high mortality among children and HIV-infected individuals. Currently, no vaccine is available for use in humans. Antibodies directed against the O-polysaccharide of the lipopolysaccharide molecule of Salmonella mediate bacterial killing and are protective, and conjugation of the O-polysaccharide to a carrier protein represents a possible strategy for vaccine development. Here we have purified the O-polysaccharide from six different strains of S. Typhimurium and fully characterized them using analytical methods including HPLC-SEC, HPAEC-PAD, GC, GC-MS, 1D and 2D NMR spectroscopy. All the O-polysaccharide samples showed a similar bimodal molecular mass distribution, but differed with respect to the amount and position of O-acetylation and glucosylation. For some strains, O-acetyl groups were found not only on C-2 of abequose (factor 5 specificity), but also on C-2 and C-3 of rhamnose; glucose was found to be linked 1→4 or 1→6 to galactose in different amounts according to the strain of origin. This structural variability could have an impact on the immunogenicity of corresponding glycoconjugate vaccines and different strains need to be evaluated in order to identify the appropriate source of O-polysaccharide to use for the development of a candidate conjugate vaccine with broad coverage against S. Typhimurium.


Assuntos
Antígenos O/química , Salmonella typhimurium/imunologia , Acetilação , Sequência de Carboidratos , Cromatografia Gasosa-Espectrometria de Massas , Conformação Molecular , Ressonância Magnética Nuclear Biomolecular
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