Immunogenicity study of a Streptococcus suis autogenous vaccine in preparturient sows and evaluation of passive maternal immunity in piglets.

BACKGROUND: Streptococcus suis is an important pathogen that causes severe diseases mostly in weaned piglets. Only available vaccines in the field are those composed of killed bacteria (bacterins) but data about their effectiveness are missing. We report here a field study on the immunological response induced by an autogenous vaccine applied in pre-parturient sows. Using a farm with recurrent S. suis serotype 7 problems, the study was divided in three experiments: (I) Sows received the vaccine at 7 and 3 weeks pre-farrowing. (II) Replacement gilts introduced to the herd received the vaccine at 4 and 7 weeks after their entry in quarantine and a boost 3 weeks pre-farrowing. (III) Gilts from experiment II received another boost 3 weeks pre-farrowing at their 3rd/4th parity. Levels, isotype profile and opsonophagocytosis capacity of the serum antibodies induced by vaccination were evaluated in sows and maternal immunity in piglets. RESULTS: In sows (I), the vaccine induced a slight, albeit significant, increase in anti-S. suis total antibodies after 2 doses when compare to basal levels already present in the animals. These antibodies showed a high opsonic capacity in vitro, highlighting their potential protective capacity. A gilt vaccination program of 3 doses (II) resulted in a significant increase in anti-S. suis total antibodies. Levels of maternal immunity transferred to piglets were high at 7 days of age, but rapidly decreased by 18 days of age. A gilt vaccination program ensued a higher transfer of maternal immunity in piglets compared to control animals; nevertheless duration was not improved at 18 day-old piglets. The vaccine response in both gilts and sows was mainly composed of IgG1 subclass, which was also the main Ig transferred to piglets. IgG2 subclass was also found in piglets, but its level was not increased by vaccination. Finally, a recall IgG1 response was induced by another boost vaccination at 3rd/4th parity (III), indicating that the vaccine induced the establishment of a lasting memory response in the herd. CONCLUSIONS: Overall, an optimal gilt/sow vaccination program might result in increased antibody responses; nevertheless duration of maternal immunity would not last long enough to protect post-weaned piglets.

Lectin-Mediated Bacterial Modulation by the Intestinal Nematode Ascaris suum.

Ascariasis is a global health problem for humans and animals. Adult Ascaris nematodes are long-lived in the host intestine where they interact with host cells as well as members of the microbiota resulting in chronic infections. Nematode interactions with host cells and the microbial environment are prominently mediated by parasite-secreted proteins and peptides possessing immunomodulatory and antimicrobial activities. Previously, we discovered the C-type lectin protein AsCTL-42 in the secreted products of adult Ascaris worms. Here we tested recombinant AsCTL-42 for its ability to interact with bacterial and host cells. We found that AsCTL-42 lacks bactericidal activity but neutralized bacterial cells without killing them. Treatment of bacterial cells with AsCTL-42 reduced invasion of intestinal epithelial cells by Salmonella. Furthermore, AsCTL-42 interacted with host myeloid C-type lectin receptors. Thus, AsCTL-42 is a parasite protein involved in the triad relationship between Ascaris, host cells, and the microbiota.

Comparative Study of Immunogenic Properties of Purified Capsular Polysaccharides from Streptococcus suis Serotypes 3, 7, 8, and 9: the Serotype 3 Polysaccharide Induces an Opsonizing IgG Response.

Streptococcus suis is an encapsulated bacterium and one of the most important swine pathogens and a zoonotic agent for which no effective vaccine exists. Bacterial capsular polysaccharides (CPSs) are poorly immunogenic, but anti-CPS antibodies are essential to the host defense against encapsulated bacteria. In addition to the previously known serotypes 2 and 14, which are nonimmunogenic, we have recently purified and described the CPS structures for serotypes 1, 1/2, 3, 7, 8, and 9. Here, we aimed to elucidate how these new structurally diverse CPSs interact with the immune system to generate anti-CPS antibody responses. CPS-stimulated dendritic cells produced significant levels of C-C motif chemokine ligand 3 (CCL3), partially via Toll-like receptor 2 (TLR2)- and myeloid differentiation factor 88-dependent pathways, and CCL2, via TLR-independent mechanisms. Mice immunized with purified serotype 3 CPS adjuvanted with TiterMax Gold produced an opsonizing IgG response, whereas other CPSs or adjuvants were negative. Mice hyperimmunized with heat-killed S. suis serotypes 3 and 9 both produced anti-CPS type 1 IgGs, whereas serotypes 7 and 8 remained negative. Also, mice infected with sublethal doses of S. suis serotype 3 produced primary anti-CPS IgM and IgG responses, of which only IgM were boosted after a secondary infection. In contrast, mice sublethally infected with S. suis serotype 9 produced weak anti-CPS IgM and IgG responses following a secondary infection. This study provides important information on the divergent evolution of CPS serotypes with highly different structural and/or biochemical properties within S. suis and their interaction with the immune system.

Explaining the Serological Characteristics of Streptococcus suis Serotypes 1 and 1/2 from Their Capsular Polysaccharide Structure and Biosynthesis.

The capsular polysaccharide (CPS) is a major virulence factor in many encapsulated pathogens, as it is the case for Streptococcus suis, an important swine pathogen and emerging zoonotic agent. Moreover, the CPS is the antigen at the origin of S. suis classification into serotypes. Hence, analyses of the CPS structure are an essential step to dissect its role in virulence and the serological relations between important serotypes. Here, the CPSs of serotypes 1 and 1/2 were purified and characterized for the first time. Chemical and spectroscopic data gave the following repeating unit sequences: [6)[Neu5Ac(α2-6)GalNAc(ß1-4)GlcNAc(ß1-3)]Gal(ß1-3)Gal(ß1-4)Glc(ß1-]n (serotype 1) and [4)[Neu5Ac(α2-6)GalNAc(ß1-4)GlcNAc(ß1-3)]Gal(ß1-4)[Gal(α1-3)]Rha(ß1-4)Glc(ß1-]n (serotype 1/2). The Sambucus nigra lectin, which recognizes the Neu5Ac(α2-6)Gal/GalNAc sequence, showed binding to both CPSs. Compared with previously characterized serotype 14 and 2 CPSs, N-acetylgalactosamine replaces galactose as the sugar bearing the sialic acid residue in the side chain. Serological analyses of the cross-reaction of serotype 1/2 with serotypes 1 and 2 and that between serotypes 1 and 14 suggested that the side chain, and more particularly the terminal sialic acid, constitutes one important epitope for serotypes 1/2 and 2. The side chain is also an important serological determinant for serotype 1, yet sialic acid seems to play a limited role. In contrast, the side chain does not seem to be part of a major epitope for serotype 14. These results contribute to the understanding of the relationship between S. suis serotypes and provide the basis for improving diagnostic tools.

Protection against Streptococcus suis Serotype 2 Infection Using a Capsular Polysaccharide Glycoconjugate Vaccine.

Streptococcus suis serotype 2 is an encapsulated bacterium and one of the most important bacterial pathogens in the porcine industry. Despite decades of research for an efficient vaccine, none is currently available. Based on the success achieved with other encapsulated pathogens, a glycoconjugate vaccine strategy was selected to elicit opsonizing anti-capsular polysaccharide (anti-CPS) IgG antibodies. In this work, glycoconjugate prototypes were prepared by coupling S. suis type 2 CPS to tetanus toxoid, and the immunological features of the postconjugation preparations were evaluated in vivo In mice, experiments evaluating three different adjuvants showed that CpG oligodeoxyribonucleotide (ODN) induces very low levels of anti-CPS IgM antibodies, while the emulsifying adjuvants Stimune and TiterMax Gold both induced high levels of IgGs and IgM. Dose-response trials comparing free CPS with the conjugate vaccine showed that free CPS is nonimmunogenic independently of the dose used, while 25 µg of the conjugate preparation was optimal in inducing high levels of anti-CPS IgGs postboost. With an opsonophagocytosis assay using murine whole blood, sera from immunized mice showed functional activity. Finally, the conjugate vaccine showed immunogenicity and induced protection in a swine challenge model. When conjugated and administered with emulsifying adjuvants, S. suis type 2 CPS is able to induce potent IgM and isotype-switched IgGs in mice and pigs, yielding functional activity in vitro and protection against a lethal challenge in vivo, all features of a T cell-dependent response. This study represents a proof of concept for the potential of glycoconjugate vaccines in veterinary medicine applications against invasive bacterial infections.

Group B Streptococcus and Streptococcus suis capsular polysaccharides induce chemokine production by dendritic cells via Toll-like receptor 2- and MyD88-dependent and -independent pathways.

Streptococcus agalactiae (also known as group B Streptococcus [GBS]) and Streptococcus suis are encapsulated streptococci causing severe septicemia and meningitis. Bacterial capsular polysaccharides (CPSs) are poorly immunogenic, but anti-CPS antibodies are essential to the host defense against encapsulated bacteria. The mechanisms underlying anti-CPS antibody responses are not fully elucidated, but the biochemistry of CPSs, particularly the presence of sialic acid, may have an immunosuppressive effect. We investigated the ability of highly purified S. suis and GBS native (sialylated) CPSs to activate dendritic cells (DCs), which are crucial actors in the initiation of humoral immunity. The influence of CPS biochemistry was studied using CPSs extracted from different serotypes within these two streptococcal species, as well as desialylated CPSs. No interleukin-1ß (IL-1ß), IL-6, IL-12p70, tumor necrosis factor alpha (TNF-α), or IL-10 production was observed in S. suis or GBS CPS-stimulated DCs. Moreover, these CPSs exerted immunosuppressive effects on DC activation, as a diminution of gamma interferon (IFN-γ)-induced B cell-activating factor of the tumor necrosis factor family (BAFF) expression was observed in CPS-pretreated cells. However, S. suis and GBS CPSs induced significant production of CCL3, via partially Toll-like receptor 2 (TLR2)- and myeloid differentiation factor 88 (MyD88)-dependent pathways, and CCL2, via TLR-independent mechanisms. No major influence of CPS biochemistry was observed on the capacity to induce chemokine production by DCs, indicating that DCs respond to these CPSs in a patterned way rather than a structure-dedicated manner.

Structure determination of Streptococcus suis serotype 14 capsular polysaccharide.

The capsular polysaccharide (CPS) of Streptococcus suis serotype 14 was purified, chemically modified, and characterized. Sugar and absolute configuration analyses gave the following CPS composition: D-Gal, 3; D-Glc, 1; D-GlcNAc, 1; D-Neu5Ac, 1. The Sambucus nigra lectin, which recognizes the Neu5Ac(α2-6)Gal/GalNAc sequence, showed binding to the native CPS. Sialic acid was found to be terminal, and the CPS was quantitatively desialylated by mild acid hydrolysis. It was also submitted to periodate oxidation followed by borohydride reduction and Smith degradation. Sugar and methylation analyses, (1)H and (13)C nuclear magnetic resonance, and mass spectrometry of the native CPS or of its specifically modified products allowed to determine the repeating unit sequence: [6)[Neu5Ac(α2-6)Gal(ß1-4)GlcNAc(ß1-3)]Gal(ß1-3)Gal(ß1-4)Glc(ß1-](n). S. suis serotype 14 CPS has an identical sialic acid-containing side chain as serotype 2 CPS, but differs by the absence of rhamnose in its composition. The same side chain is also present in group B Streptococcus type Ia CPS, except that in the latter sialic acid is 2,3- rather than 2,6-linked to the following galactose. A correlation between the S. suis CPS sequence and genes of the serotype 14 cps locus encoding putative glycosyltransferases and polymerase responsible for the biosynthesis of the repeating unit is proposed.

Impact of Protein Glycosylation on the Design of Viral Vaccines.

Glycans play crucial roles in various biological processes such as cell proliferation, cell-cell interactions, and immune responses. Since viruses co-opt cellular biosynthetic pathways, viral glycosylation mainly depends on the host cell glycosylation machinery. Consequently, several viruses exploit the cellular glycosylation pathway to their advantage. It was shown that viral glycosylation is strongly dependent on the host system selected for virus propagation and/or protein expression. Therefore, the use of different expression systems results in various glycoforms of viral glycoproteins that may differ in functional properties. These differences clearly illustrate that the choice of the expression system can be important, as the resulting glycosylation may influence immunological properties. In this review, we will first detail protein N- and O-glycosylation pathways and the resulting glycosylation patterns; we will then discuss different aspects of viral glycosylation in pathogenesis and in vaccine development; and finally, we will elaborate on how to harness viral glycosylation in order to optimize the design of viral vaccines. To this end, we will highlight specific examples to demonstrate how glycoengineering approaches and exploitation of different expression systems could pave the way towards better self-adjuvanted glycan-based viral vaccines.

S-Layer From Lactobacillus brevis Modulates Antigen-Presenting Cell Functions via the Mincle-Syk-Card9 Axis.

C-type lectin receptors (CLRs) are pattern recognition receptors that are crucial in the innate immune response. The gastrointestinal tract contributes significantly to the maintenance of immune homeostasis; it is the shelter for billions of microorganisms including many genera of Lactobacillus sp. Previously, it was shown that host-CLR interactions with gut microbiota play a crucial role in this context. The Macrophage-inducible C-type lectin (Mincle) is a Syk-coupled CLR that contributes to sensing of mucosa-associated commensals. In this study, we identified Mincle as a receptor for the Surface (S)-layer of the probiotic bacteria Lactobacillus brevis modulating GM-CSF bone marrow-derived cells (BMDCs) functions. We found that the S-layer/Mincle interaction led to a balanced cytokine response in BMDCs by triggering the release of both pro- and anti-inflammatory cytokines. In contrast, BMDCs derived from Mincle-/-, CARD9-/- or conditional Syk-/- mice failed to maintain this balance, thus leading to an increased production of the pro-inflammatory cytokines TNF and IL-6, whereas the levels of the anti-inflammatory cytokines IL-10 and TGF-ß were markedly decreased. Importantly, this was accompanied by an altered CD4+ T cell priming capacity of Mincle-/- BMDCs resulting in an increased CD4+ T cell IFN-γ production upon stimulation with L. brevis S-layer. Our results contribute to the understanding of how commensal bacteria regulate antigen-presenting cell (APC) functions and highlight the importance of the Mincle/Syk/Card9 axis in APCs as a key factor in host-microbiota interactions.

Capsular polysaccharide switching in Streptococcus suis modulates host cell interactions and virulence.

The capsular polysaccharide (CPS) of Streptococcus suis defines various serotypes based on its composition and structure. Though serotype switching has been suggested to occur between S. suis strains, its impact on pathogenicity and virulence remains unknown. Herein, we experimentally generated S. suis serotype-switched mutants from a serotype 2 strain that express the serotype 3, 4, 7, 8, 9, or 14 CPS. The effects of serotype switching were then investigated with regards to classical properties conferred by presence of the serotype 2 CPS, including adhesion to/invasion of epithelial cells, resistance to phagocytosis by macrophages, killing by whole blood, dendritic cell-derived pro-inflammatory mediator production and virulence using mouse and porcine infection models. Results demonstrated that these properties on host cell interactions were differentially modulated depending on the switched serotypes, although some different mutations other than loci of CPS-related genes were found in each the serotype-switched mutant. Among the serotype-switched mutants, the mutant expressing the serotype 8 CPS was hyper-virulent, whereas mutants expressing the serotype 3 or 4 CPSs had reduced virulence. By contrast, switching to serotype 7, 9, or 14 CPSs had little to no effect. These findings suggest that serotype switching can drastically alter S. suis virulence and host cell interactions.

Characterization and Protective Activity of Monoclonal Antibodies Directed against Streptococcus suis Serotype 2 Capsular Polysaccharide Obtained Using a Glycoconjugate.

Streptococcus suis serotype 2 is an encapsulated bacterium and an important swine pathogen. Opsonizing antibody responses targeting capsular polysaccharides (CPSs) are protective against extracellular pathogens. To elucidate the protective activity of monoclonal antibodies (mAbs) directed against S. suis serotype 2 CPS, mice were immunized with a serotype 2 CPS-glycoconjugate and three hybridomas were isolated; of which, two were murine IgMs and the other a murine IgG1. Whereas the IgMs (mAbs 9E7 and 13C8) showed different reactivity levels with S. suis serotypes 1, 1/2, 2 and 14, the IgG1 (mAb 16H11) was shown to be serotype 2-specific. All mAbs targeted the sialylated chain of the CPSs. Using an opsonophagocytosis assay, the IgMs were opsonizing towards the S. suis serotypes to which they cross-react, while the IgG1 failed to induce bacterial elimination. In a model of mouse passive immunization followed by a lethal challenge with S. suis serotype 2, the IgG1 and IgM cross-reacting only with serotype 14 (mAb 13C8) failed to protect, while the IgM cross-reacting with serotypes 1, 1/2, and 14 (mAb 9E7) was shown to be protective by limiting bacteremia. These new mAbs show promise as new S. suis diagnostic tools, as well as potential for therapeutic applications.

Structure determination of Streptococcus suis serotypes 7 and 8 capsular polysaccharides and assignment of functions of the cps locus genes involved in their biosynthesis.

Streptococcus suis serotypes 7 and 8 are counted among the top six S. suis serotypes causing clinical disease in pigs. Yet, limited information is available on these serotypes. Since S. suis serotyping system is based upon capsular polysaccharide (CPS) antigenicity and the CPS is considered a major virulence factor for encapsulated pathogens, here we determined for the first time the chemical compositions and structures of serotypes 7 and 8 CPSs. Chemical and spectroscopic data gave the following repeating unit sequences: [3)L-Rha(α1-P-2)D-Gal(α1-4)D-GlcA(ß1-3)D-FucNAc4N(α1-]n for serotype 7 and [2)L-Rha(α1-P-4)D-ManNAc(ß1-4)D-Glc(α1-]n for serotype 8. As serotype 8 CPS is identical to Streptococcus pneumoniae type 19F CPS, dot-blot analyses showed a strong reaction of the 19F polysaccharide with reference anti-S. suis serotype 8 rabbit serum. A correlation between S. suis serotypes 7 and 8 sequences and genes of those serotypes' loci encoding putative glycosyltransferases and polymerases responsible for the biosynthesis of the repeating units was tentatively established. Knowledge of CPS structure and composition will contribute to better dissect the role of this bacterial component in the pathogenesis of the disease caused by S. suis serotypes 7 and 8.

A platform for glycoengineering a polyvalent pneumococcal bioconjugate vaccine using E. coli as a host.

Chemical synthesis of conjugate vaccines, consisting of a polysaccharide linked to a protein, can be technically challenging, and in vivo bacterial conjugations (bioconjugations) have emerged as manufacturing alternatives. Bioconjugation relies upon an oligosaccharyltransferase to attach polysaccharides to proteins, but currently employed enzymes are not suitable for the generation of conjugate vaccines when the polysaccharides contain glucose at the reducing end, which is the case for ~75% of Streptococcus pneumoniae capsules. Here, we use an O-linking oligosaccharyltransferase to generate a polyvalent pneumococcal bioconjugate vaccine with polysaccharides containing glucose at their reducing end. In addition, we show that different vaccine carrier proteins can be glycosylated using this system. Pneumococcal bioconjugates are immunogenic, protective and rapidly produced within E. coli using recombinant techniques. These proof-of-principle experiments establish a platform to overcome limitations of other conjugating enzymes enabling the development of bioconjugate vaccines for many important human and animal pathogens.

Streptococcus suis serotype 3 and serotype 18 capsular polysaccharides contain di-N-acetyl-bacillosamine.

Streptococcus suis serotype 3 is counted among the S. suis serotypes causing clinical disease in pigs. Yet, limited information is available on this serotype. Here we determined for the first time the chemical composition and structure of serotype 3 capsular polysaccharide (CPS), a major bacterial virulence factor and the antigen at the origin of S. suis classification into serotypes. Chemical and spectroscopic data gave the repeating unit sequence for serotype 3: [4)D-GlcA (ß1-3)d-QuiNAc4NAc(ß1-]n. To the best of our knowledge, this is the first report of di-N-acetyl-d-bacillosamine (QuiNAc4NAc) containing polysaccharides in Streptococci and the second time this rare diamino sugar has been observed in a Gram-positive bacterial species since its initial report. This led to the identification of homologues of UDP-QuiNAc4NAc synthesis genes in S. suis serotype 18. Thus, the repeating unit sequence for serotype 18 is: [3)d-GalNAc(α1-3)[d-Glc (ß1-2)]d-GalA4OAc(ß1-3)d-GalNAc(α1-3)d-QuiNAc4NAc(α1-]n. A correlation between S. suis serotypes 3 and 18 CPS sequences and genes of these serotypes' cps loci encoding putative glycosyltransferases and polymerase responsible for the biosynthesis of the repeating unit was tentatively established. Knowledge of CPS structure and composition will contribute to better dissect the role of this bacterial component in the pathogenesis of S. suis serotypes 3 and 18.

Capsular Sialyltransferase Specificity Mediates Different Phenotypes in Streptococcus suis and Group B Streptococcus.

The capsular polysaccharide (CPS) represents a key virulence factor for most encapsulated streptococci. Streptococcus suis and Group B Streptococcus (GBS) are both well-encapsulated pathogens of clinical importance in veterinary and/or human medicine and responsible for invasive systemic diseases. S. suis and GBS are the only Gram-positive bacteria which express a sialylated CPS at their surface. An important difference between these two sialylated CPSs is the linkage between the side-chain terminal galactose and sialic acid, being α-2,6 for S. suis but α-2,3 for GBS. It is still unclear how sialic acid may affect CPS production and, consequently, the pathogenesis of the disease caused by these two bacterial pathogens. Here, we investigated the role of sialic acid and the putative effect of sialic acid linkage modification in CPS synthesis using inter-species allelic exchange mutagenesis. To this aim, a new molecular biogenetic approach to express CPS with modified sialic acid linkage was developed. We showed that sialic acid (and its α-2,6 linkage) is crucial for S. suis CPS synthesis, whereas for GBS, CPS synthesis may occur in presence of an α-2,6 sialyltransferase or in absence of sialic acid moiety. To evaluate the effect of the CPS composition/structure on sialyltransferase activity, two distinct capsular serotypes within each bacterial species were compared (S. suis serotypes 2 and 14 and GBS serotypes III and V). It was demonstrated that the observed differences in sialyltransferase activity and specificity between S. suis and GBS were serotype unrestricted. This is the first time that a study investigates the interspecies exchange of capsular sialyltransferase genes in Gram-positive bacteria. The obtained mutants represent novel tools that could be used to further investigate the immunomodulatory properties of sialylated CPSs. Finally, in spite of common CPS structural characteristics and similarities in the cps loci, sialic acid exerts differential control of CPS expression by S. suis and GBS.

Porcine Dendritic Cells as an In Vitro Model to Assess the Immunological Behaviour of Streptococcus suis Subunit Vaccine Formulations and the Polarizing Effect of Adjuvants.

An in vitro porcine bone marrow-derived dendritic cell (DC) culture was developed as a model for evaluating immune polarization induced by adjuvants when administered with immunogens that may become vaccine candidates if appropriately formulated. The swine pathogen Streptococcus suis was chosen as a prototype to evaluate proposed S. suis vaccine candidates in combination with the adjuvants Poly I:C, Quil A ®, Alhydrogel ®, TiterMax Gold ® and Stimune ®. The toll-like receptor ligand Poly I:C and the saponin Quil A ® polarized swine DC cytokines towards a type 1 phenotype, with preferential production of IL-12 and TNF-α. The water-in-oil adjuvants TiterMax Gold ® and Stimune ® favoured a type 2 profile as suggested by a marked IL-6 release. In contrast, Alhydrogel ® induced a type 1/type 2 mixed cytokine profile. The antigen type differently modified the magnitude of the adjuvant effect, but overall polarization was preserved. This is the first comparative report on swine DC immune activation by different adjuvants. Although further swine immunization studies would be required to better characterize the induced responses, the herein proposed in vitro model is a promising approach that helps assessing behaviour of the vaccine formulation rapidly at the pre-screening stage and will certainly reduce numbers of animals used while advancing vaccinology science.

A single amino acid polymorphism in the glycosyltransferase CpsK defines four Streptococcus suis serotypes.

The capsular polysaccharide (CPS) is the major virulence factor of the emerging zoonotic pathogen Streptococcus suis. CPS differences are also the basis for serological differentiation of the species into 29 serotypes. Serotypes 2 and 1/2, which possess identical gene content in their cps loci, express CPSs that differ only by substitution of galactose (Gal) by N-acetylgalactosamine (GalNAc) in the CPS side chain. The same sugar substitution differentiates the CPS of serotypes 14 and 1, whose cps loci are also identical in gene content. Here, using mutagenesis, CPS structural analysis, and protein structure modeling, we report that a single amino acid polymorphism in the glycosyltransferase CpsK defines the enzyme substrate predilection for Gal or GalNAc and therefore determines CPS composition, structure, and strain serotype. We also show that the different CPS structures have similar antiphagocytic properties and that serotype switching has limited impact on the virulence of S. suis.

Structure determination of Streptococcus suis serotype 9 capsular polysaccharide and assignment of functions of the cps locus genes involved in its biosynthesis.

Streptococcus suis serotype 9 is the most prevalent S. suis serotype in several European countries. In spite of its pathogenicity for pigs and increasing zoonotic potential, limited information is available on this serotype. Here we determined for the first time the chemical composition and structure of serotype 9 capsular polysaccharide (CPS), a major bacterial virulence factor and the antigen at the origin of S. suis classification into serotypes. Chemical and spectroscopic data gave the repeating unit sequence: [3)Glcol-6-P-3-[D-Gal(α1-2)]D-Gal(ß1-3)D-Sug(ß1-3)L-Rha(α1-)]n. Compared to previously characterized S. suis CPSs (serotypes 1, 1/2, 2 and 14), serotype 9 CPS does not contain sialic acid but contains a labile 4-keto sugar (2-acetamido-2,6-dideoxy-ß-D-xylo-hexopyranos-4-ulose), one particular feature of this serotype. A correlation between S. suis serotype 9 CPS sequence and genes of this serotype cps locus encoding putative glycosyltransferases and polymerase responsible for the biosynthesis of the repeating unit was tentatively established. Knowledge of CPS structure and composition will contribute to better dissect the role of this bacterial component in the pathogenesis of S. suis serotype 9.

Glycoengineered Outer Membrane Vesicles: A Novel Platform for Bacterial Vaccines.

The World Health Organization has indicated that we are entering into a post-antibiotic era in which infections that were routinely and successfully treated with antibiotics can now be lethal due to the global dissemination of multidrug resistant strains. Conjugate vaccines are an effective way to create a long-lasting immune response against bacteria. However, these vaccines present many drawbacks such as slow development, high price, and batch-to-batch inconsistencies. Alternate approaches for vaccine development are urgently needed. Here we present a new vaccine consisting of glycoengineered outer membrane vesicles (geOMVs). This platform exploits the fact that the initial steps in the biosynthesis of most bacterial glycans are similar. Therefore, it is possible to easily engineer non-pathogenic Escherichia coli lab strains to produce geOMVs displaying the glycan of the pathogen of interest. In this work we demonstrate the versatility of this platform by showing the efficacy of geOMVs as vaccines against Streptococcus pneumoniae in mice, and against Campylobacter jejuni in chicken. This cost-effective platform could be employed to generate vaccines to prevent infections caused by a wide variety of microbial agents in human and animals.