Encapsulated Pathogens Research Articles

In Situ Characterization of Bioaerosols at the Single-Particle Level Using Single-Particle Mass Spectrometry: A Promising Tool for Defending Human Health against Bioaerosol Transmission.

Bioaerosol transmission is one of the important transmission pathways of COVID-19 and other infectious respiratory diseases caused by viral infection. The ability to detect bioaerosols and characterize encapsulated pathogens both in situ and in real time is crucial for early warning and monitoring of the progress of an epidemic or pandemic. The lack of a powerful analytical tool for distinguishing between bioaerosols and nonbioaerosols as well as for identification of pathogen species contained in the bioaerosols is the bottleneck in related fields. Herein, a promising solution for in situ and real-time accurate and sensitive detection of bioaeorosols is proposed by integrating single-particle aerosol mass spectrometry, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and fluorescence spectroscopy. The proposed mass spectrometry aims at detecting bioaerosols in a range of 0.5-10 μm with adequate sensitivity and specificity. This single-particle bioaerosol mass spectrometry would not only be a powerful tool that can be useful for the authorities and public health monitoring but also would be an example of advances in mass spectrometry.

Conceptual model to inform Legionella-amoebae control, including the roles of extracellular vesicles in engineered water system infections.

Extracellular vesicles (EVs or exosomes) are well described for bacterial pathogens associated with our gastrointestinal system, and more recently as a novel mechanism for environmental persistence, dissemination and infection for human enteric viruses. However, the roles played by EVs in the ancient arms race that continues between amoebae and one of their prey, Legionella pneumophila, is poorly understood. At best we know of intracellular vesicles of amoebae containing a mix of bacterial prey species, which also provides an enhanced niche for bacteriophage infection/spread. Free-living amoeba-associated pathogens have recently been recognized to have enhanced resistance to disinfection and environmental stressors, adding to previously understood (but for relatively few species of) bacteria sequestered within amoebal cysts. However, the focus of the current work is to review the likely impacts of large numbers of respiratory-sized EVs containing numerous L.pneumophila cells studied in pure and biofilm systems with mixed prey species. These encapsulated pathogens are orders of magnitude more resistant to disinfection than free cells, and our engineered systems with residual disinfectants could promote evolution of resistance (including AMR), enhanced virulence and EV release. All these are key features for evolution within a dead-end human pathogen post lung infection. Traditional single-hit pathogen infection models used to estimate the probability of infection/disease and critical environmental concentrations via quantitative microbial risk assessments may also need to change. In short, recognizing that EV-packaged cells are highly virulent units for transmission of legionellae, which may also modulate/avoid human host immune responses. Key data gaps are raised and a previous conceptual model expanded upon to clarify where biofilm EVs could play a role promoting risk as well as inform a more wholistic management program to proactively control legionellosis.

Analysis of trough levels of total IgG, IgG subclasses, measles neutralizing antibodies and IgG antibodies to encapsulated pathogens after infusion of a 5% or 10% intravenous immunoglobulin

IntroductionPatients with primary immunodeficiency (PI) are at risk of infection due to low IgG levels. We report trough levels of total IgG, IgG subclasses, measles neutralizing antibody (MVab) titers, Haemophilus influenzae type B and seven Streptococcus pneumoniae serotypes in PI patients receiving a 5% and 10% intravenous immunoglobulin (IVIG) formulation. MethodsGMX07 was a bioequivalence trial comparing Gammaplex 10% IVIG with Gammaplex 5% IVIG in patients with PI. Adult patients received the 5% for ≥ five infusions, then the 10% for ≥ five infusions, or vice versa. Pediatric patients received the 10% for 5 infusions. Doses were 300–800 mg/kg/infusion. Trough IgG levels were measured before each infusion. Trough levels for MVab, H. influenzae and S. pneumoniae titers were measured after the last infusion. Results33 adults received the 5%, 32 received the 10%. 15 pediatric patients received the 10%. In adults, median IgG trough levels = 952.5 mg/dL for the 10% and 937.0 mg/dL for the 5%. In pediatric patients, median trough level = 879.5 mg/dL. Trough levels of IgG subclasses were comparable across formulations and age groups. Adult patients had a mean MVab titer of 2908 mIU/mL with the 5% vs 3213 mIU/mL for the 10%. In pediatric patients, mean MVab titer was 2122 mIU/mL. In adults, median trough levels of IgG antibodies to H. influenzae were 2.80 µg/mL for the 5% and 2.76 µg/mL for the 10%. Trough levels of IgG antibodies to all S. pneumoniae serotypes were consistent across formulations and age groups. No clinically meaningful differences were noted between formulations, 21-day vs 28-day regimens, or age groups. DiscussionMaintaining IgG trough levels and mean MVab trough titers above protective thresholds is critical for patients with PI. No trough levels or titers fell below the protective thresholds. While thresholds for protective levels for H. influenzae or S. pneumoniae are not clearly defined, delivery of these antibodies to PI patients was demonstrated. ConclusionOur results indicate protective trough levels of IgG and antibody titers are delivered with the 5% and 10% formulations of this IVIG product, with similar results for adult and pediatric PI patients.

Isopropoxy Benzene Guanidine Ameliorates Streptococcus suis Infection In Vivo and In Vitro.

Streptococcus suis, an encapsulated zoonotic pathogen, has been reported to cause a variety of infectious diseases, such as meningitis and streptococcal-toxic-shock-like syndrome. Increasing antimicrobial resistance has triggered the need for new treatments. In the present study, we found that isopropoxy benzene guanidine (IBG) significantly attenuated the effects caused by S. suis infection, in vivo and in vitro, by killing S. suis and reducing S. suis pathogenicity. Further studies showed that IBG disrupted the integrity of S. suis cell membranes and increased the permeability of S. suis cell membranes, leading to an imbalance in proton motive force and the accumulation of intracellular ATP. Meanwhile, IBG antagonized the hemolysis activity of suilysin and decreased the expression of Sly gene. In vivo, IBG improved the viability of S. suis SS3-infected mice by reducing tissue bacterial load. In conclusion, IBG is a promising compound for the treatment of S. suis infections, given its antibacterial and anti-hemolysis activity.

Molecular mechanism of Streptococcus pneumoniae-targeting xenophagy recognition and evasion: Reinterpretation of pneumococci as intracellular bacteria.

Streptococcus pneumoniae is a major, encapsulated Gram-positive pathogen that causes diseases including community-acquired pneumonia, meningitis, and sepsis. This pathogen colonizes the nasopharyngeal epithelia asymptomatically but can often migrate to sterile tissues and cause life-threatening invasive infections (invasive pneumococcal disease). Although multivalent pneumococcal polysaccharides and conjugate vaccines are available and effective, they also have major shortcomings with respect to the emergence of vaccine-resistant serotypes. Therefore, alternative therapeutic approaches are needed, and the molecular analysis of host-pathogen interactions and their applications to pharmaceutical development and clinical practice has recently received increased attention. In this review, we introduce pneumococcal surface virulence factors involved in pathogenicity and highlight recent advances in our understanding of host autophagy recognition mechanisms against intracellular S. pneumoniae and pneumococcal evasion from autophagy.

Group B Streptococcus cpsE Is Required for Serotype V Capsule Production and Aids in Biofilm Formation and Ascending Infection of the Reproductive Tract during Pregnancy.

Group B Streptococcus (GBS) is an encapsulated Gram-positive pathogen that causes ascending infections of the reproductive tract during pregnancy. The capsule of this organism is a critical virulence factor that has been implicated in a variety of cellular processes to promote pathogenesis. Primarily comprised of carbohydrates, the GBS capsule and its synthesis is driven by the capsule polysaccharide synthesis (cps) operon. The cpsE gene within this operon encodes a putative glycosyltransferase that is responsible for the transfer of a Glc-1-P from UDP-Glc to an undecaprenyl lipid molecule. We hypothesized that the cpsE gene product is important for GBS virulence and ascending infection during pregnancy. Our work demonstrates that a GBS cpsE mutant secretes fewer carbohydrates, has a reduced capsule, and forms less biofilm than the wild-type parental strain. We show that, compared to the parental strain, the ΔcpsE deletion mutant is more readily taken up by human placental macrophages and has a significantly attenuated ability to invade and proliferate in the mouse reproductive tract. Taken together, these results demonstrate that the cpsE gene product is an important virulence factor that aids in GBS colonization and invasion of the gravid reproductive tract.

A half-century of meningococcal vaccines

The first safe and effective vaccine for the prevention of invasive meningococcal disease was created fifty years ago. The vaccine employed a novel platform, polysaccharide capsular antigen, based on the discovery that anticapsular antibody conferred protective immunity in humans. As with most new paradigms in vaccinology, it derived from important basic research from other scientific disciplines over the preceding years. The success of the first monovalent polysaccharide vaccine in nearly eliminating invasive meningococcal disease in military settings led to accelerated advances in polysaccharide vaccine development against other serogroups of meningococcus and other encapsulated pathogens. As gaps in vaccine efficacy arose over the past half-century, new vaccine technologies and approaches were developed to address the challenges. Several of these, including conjugate vaccines and “reverse vaccinology” led to other novel, successful vaccines that have had a significant, favorable global impact on invasive meningococcal disease. The history of meningococcal vaccine discovery may provide insights into the future of vaccine efforts against other infectious threats.

Simultaneous Imaging of Dynamics and Biochemical Activities of Single Phagosomes

Phagocytosis is a major defense mechanism used by our immune system to eliminate pathogens. It proceeds in multiple steps: ligand-receptor recognition between pathogens and cell membranes in the beginning, internalization of pathogens into intracellular membrane organelles known as phagosomes, transport of phagosomes from cell edge to areas surrounding the cell nucleus, and digestion of the encapsulated pathogens through series of chemical reactions. All those steps must occur in the right order at the right time and in the right subcellular location. Abnormalities in any of those steps can lead to progression of infections and other serious human diseases, such as cystic fibrosis and chronic granulomatous disease. Thus, a fundamental understanding of phagocytosis is critical for developing methods to diagnose and treat those diseases. Unfortunately, that has been a grand challenging due to the complexity of the mechanisms involved in phagocytosis. To address the critical need, we developed nanoparticle-based sensors to probe the chemical and dynamical activities during phagocytosis. Simultaneous measurements of phagosome transport, acidification, and fusion reveal a striking correlation between the intracellular transport of phagosomes and their biochemical activities during maturation. Our results reveal a regulatory mechanism underlying the orchestration of dynamical and biochemical signaling processes during phagosome maturation.

Vitamin D and Streptococci: The Interface of Nutrition, Host Immune Response, and Antimicrobial Activity in Response to Infection.

Streptococcus species are common causes of human infection. These Gram-positive, encapsulated bacterial pathogens infect diverse anatomic spaces, leading to infections including skin and soft tissue infection, endocarditis, pneumonia, meningitis, sinusitis, otitis media, chorioamnionitis, sepsis, and even death. Risk for streptococcal infection is highest in low- and middle-income countries where micronutrient deficiency is common. Epidemiological data reveal that vitamin D deficiency is associated with enhanced risk of streptococcal infection and cognate disease outcomes. Additionally, vitamin D improves antibacterial defenses by stimulating innate immune processes such as phagocytosis and enhancing production of reactive oxygen species (oxidative burst) and antimicrobial peptides (including cathelicidin and lactoferrin), which are important for efficient killing of bacteria. This review presents the most recent published work that studies interactions between the micronutrient vitamin D, the host immune system, and pathogenic streptococci as well as comparisons with other relevant infection models.

Expanding the Scope of Ugi Multicomponent Bioconjugation to Produce Pneumococcal Multivalent Glycoconjugates as Vaccine Candidates.

Conjugate vaccines against encapsulated pathogens like Streptococcus pneumoniae face many challenges, including the existence of multiple serotypes with a diverse global distribution that constantly requires new formulations and higher coverage. Multivalency is usually achieved by combining capsular polysaccharide-protein conjugates from invasive serotypes, and for S. pneumoniae, this has evolved from 7- up to 20-valent vaccines. These glycoconjugate formulations often contain high concentrations of carrier proteins, which may negatively affect glycoconjugate immune response. This work broadens the scope of an efficient multicomponent strategy, leading to multivalent pneumococcal glycoconjugates assembled in a single synthetic operation. The bioconjugation method, based on the Ugi four-component reaction, enables the one-pot incorporation of two different polysaccharide antigens to a tetanus toxoid carrier, thus representing the fastest approach to achieve multivalency. The reported glycoconjugates incorporate three combinations of capsular polysaccharides 1, 6B, 14, and 18C from S. pneumoniae. The glycoconjugates were able to elicit functional specific antibodies against pneumococcal strains comparable to those shown by mixtures of the two monovalent glycoconjugates.

Chemical structure and genetic organization of the E. coli O6:K15 capsular polysaccharide

Capsular polysaccharides are important virulence factors in pathogenic bacteria. Characterizing the structural components and biosynthetic pathways for these polysaccharides is key to our ability to design vaccines and other preventative therapies that target encapsulated pathogens. Many gram-negative pathogens such as Neisseria meningitidis and Escherichia coli express acidic capsules. The E. coli K15 serotype has been identified as both an enterotoxigenic and uropathogenic pathogen. Despite its relevance as a disease-causing serotype, the associated capsular polysaccharide remains poorly characterized. We describe in this report the chemical structure of the K15 polysaccharide, based on chemical analysis and nuclear magnetic resonance (NMR) data. The repeating structure of the K15 polysaccharide consists of 4)-α-GlcpNAc-(1 → 5)-α-KDOp-(2 → partially O-acetylated at 3-hydroxyl of GlcNAc. We also report, the organization of the gene cluster responsible for capsule biosynthesis. We identify genes in this cluster that potentially encode an O-acetyltransferase, an N-acetylglucosamine transferase, and a KDO transferase consistent with the structure we report.

Audit of Post-Splenectomy Prophylaxis in a Single Tertiary Center in Slovenia: Where Are We and What Should Be Done?

Background: Patients who have had splenectomies are at increased risk for severe infections and overwhelming sepsis caused by pneumococci, meningococci, Haemophilus influenzae, and other encapsulated pathogens. Prophylactic measures are important to reduce mortality in this group; among them are vaccinations, antibiotic prophylaxis, and education. We performed a retrospective audit of post-splenectomy prophylactic measures, focusing on vaccinations and antibiotic prophylaxis coverage. Patients and Methods: We included 156 adult patients who had splenectomies between January 2010 and December 2018 in a study conducted at a single tertiary medical center. Data regarding vaccinations, antibiotic prophylaxis, and severe infections in the post-splenectomy period were obtained from medical records and supplemented by a patient-devised questionnaire. Results: Overall 121 of 156 (77.5%) of patients received a basic pneumococcal vaccination, although only 27 of 70 (38.6%) received re-vaccination after five years. Lower vaccination coverage was observed for meningococcal diseases and Haemophilus influenzae type b (Hib) disease with 57.0% (89/156) and 55.7% (87/156) vaccination coverage, respectively. A difference in pneumococcal vaccination coverage between patients after elective splenectomy (81/91; 89.0%) and non-elective splenectomy (40/65; 61.5%) was observed. However, no difference was observed among meningococcal and Hib vaccination coverage. Antibiotic prophylaxis was recommended to patients more frequently after elective splenectomy (41/82; 50.0%) than non-elective (16/56; 28.6%) yet after an elective splenectomy, patients discontinued the prophylaxis more often than those after a non-elective surgery. Overall, only 30 of 146 (20.5%) of patients received antibiotic prophylaxis over two years. In the observed period, 12.3% of patients suffered a severe infection. Conclusion: Our study shows that there is adequate basic pneumococcal vaccination coverage among patients after a splenectomy particularly after an elective splenectomy, but there is a lack of and an inadequate implementation of other prophylactic measures. There is an urgent need for an organized approach involving better education of healthcare practitioners as well as patients concerning the risk of asplenia. In addition, an improved long-term follow-up is needed including establishing a central registry for the asplenic patients in Slovenia.

Reduced phagocytosis and killing of Cryptococcus neoformans biofilm-derived cells by J774.16 macrophages is associated with fungal capsular production and surface modification

Cryptococcus neoformans is an opportunistic encapsulated pathogen that causes life-threatening meningoencephalitis in individuals with immunosuppression. We compared the interactions of C. neoformans planktonic and biofilm-derived cells with J774.16 macrophage-like cells. Planktonic cells are more phagocytized and killed by J774.16 cells than biofilm-derived fungal cells. Biofilm-derived cryptococci possess larger capsule size and release significantly more capsular polysaccharide than planktonic cells in culture. Biofilm-derived fungi exhibited upregulation of genes involved in capsular production. Capsular-specific monoclonal antibody 18B7 demonstrated differential binding to the surface of planktonic and biofilm-derived cryptococci providing a plausible strategy for fungal evasion of macrophages and persistence. Future studies are necessary to elucidate how C. neoformans biofilm-derived cells regulate their virulence factors when interacting with cells of the immune system.

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(β1-3)D-FucNAc4N(α1-]n for serotype 7 and [2)L-Rha(α1-P-4)D-ManNAc(β1-4)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.

The Buoyancy of Cryptococcus neoformans Is Affected by Capsule Size.

Cryptococcus neoformans is an environmental pathogenic fungus with a worldwide geographical distribution that is responsible for hundreds of thousands of human cryptococcosis cases each year. During infection, the yeast undergoes a morphological transformation involving capsular enlargement that increases microbial volume. To understand the factors that play a role in environmental dispersal of C. neoformans and C. gattii, we evaluated the cell density of Cryptococcus using Percoll isopycnic gradients. We found differences in the cell densities of strains belonging to C. neoformans and C. gattii species complexes. The buoyancy of C. neoformans strains varied depending on growth medium. In minimal medium, the cryptococcal capsule made a major contribution to the cell density such that cells with larger capsules had lower density than those with smaller capsules. Removing the capsule, by chemical or mechanical methods, increased the C. neoformans cell density and reduced buoyancy. Melanization of the C. neoformans cell wall, which also contributes to virulence, produced a small but consistent increase in cell density. Encapsulated C. neoformans sedimented much more slowly in seawater as its density approached the density of water. Our results suggest a new function for the capsule whereby it can function as a flotation device to facilitate transport and dispersion in aqueous fluids.IMPORTANCE The buoyancy of a microbial cell is an important physical characteristic that may affect its transportability in fluids and interactions with tissues during infection. The polysaccharide capsule surrounding C. neoformans is required for infection and dissemination in the host. Our results indicate that the capsule has a significant effect on reducing cryptococcal cell density, altering its sedimentation in seawater. Modulation of microbial cell density via encapsulation may facilitate dispersal for other important encapsulated pathogens.

A Streptococcus suis Live Vaccine Suppresses Streptococcal Toxic Shock-Like Syndrome and Provides Sequence Type-Independent Protection.

Streptococcus suis is an encapsulated zoonotic pathogen. Increasing antimicrobial resistance invokes the need for effective vaccines. Despite many attempts to develop an effective vaccine, none is currently available. A capsular polysaccharide (CPS)-expressing attenuated mutant 2015033 was constructed by deleting 5 virulence-associated factors (sly, scpA, ssnA, fhb, and ssads) in an infective S. suis strain SC19. The safety and immune effect of 2015033 were determined both in vitro and in vivo. Deletion of 5 genes did not impact the growth ability and CPS generation of 2015033, and the mutant exhibited no cytotoxicity in different cell models. 2015033 was more easily eliminated by innate immunity both in vitro and in vivo. In addition, 2015033 showed a diminished invasive ability in different mouse organs (brain, lung, and liver) and avirulent properties in mice associated with weak inflammation-inducing ability. Immunization with 2015033 triggered T cell-dependent immunity, suppressed streptococcal toxic shock-like syndrome, and conferred sequence type-independent protection to mice during infection. This study presents the feasibility of the strategy of multigene deletion for the development of promising live vaccines against invasive encapsulated pathogens.

Enzymatic Hydrolysis of Pneumococcal Capsular Polysaccharide Renders the Bacterium Vulnerable to Host Defense.

Despite a century of investigation, Streptococcus pneumoniae remains a major human pathogen, causing a number of diseases, such as pneumonia, meningitis, and otitis media. Like many encapsulated pathogens, the capsular polysaccharide (CPS) of S. pneumoniae is a critical component for colonization and virulence in mammalian hosts. This study aimed to evaluate the protective role of a glycoside hydrolase, Pn3Pase, targeting the CPS of type 3 S. pneumoniae, which is one of the most virulent serotypes. We have assessed the ability of Pn3Pase to degrade the capsule on a live type 3 strain. Through in vitro assays, we observed that Pn3Pase treatment increases the bacterium's susceptibility to phagocytosis by macrophages and complement-mediated killing by neutrophils. We have demonstrated that in vivo Pn3Pase treatment reduces nasopharyngeal colonization and protects mice from sepsis caused by type 3 S. pneumoniae Due to the increasing shifts in serotype distribution, the rise in drug-resistant strains, and poor immune responses to vaccine-included serotypes, it is necessary to investigate approaches to combat pneumococcal infections. This study evaluates the interaction of pneumococcal CPS with the host at molecular, cellular, and systemic levels and offers an alternative therapeutic approach for diseases caused by S. pneumoniae through enzymatic hydrolysis of the CPS.

STAT1 regulates marginal zone B cell differentiation in response to inflammation and infection with blood-borne bacteria.

Marginal zone B (MZ B) cells can rapidly produce antibody in response to infection with blood-borne encapsulated pathogens. Although TLR-mediated activation of MZ B is known to trigger humoral immune response, the signal cascade directing this response remains undefined. Here, we demonstrate that STAT1 plays an essential role in TLR-mediated antibody response of MZ B cells. Further, the TLR-induced IgM response is impaired in a type I and type II IFN-independent manner. Although activation, proliferation, and apoptosis are not affected, both differentiation into plasma cells and IgM production are impaired in Stat1-/- MZ B cells. Interestingly, STAT1 directly regulates the expression of Prdm1 (encodes BLIMP-1) by binding to its promoter, and Prdm1 expression is reduced in Stat1-/- MZ B cells. Restoration of BLIMP-1 to cells rescues TLR-induced IgM response. Moreover, Stat1-/- mice are more susceptible to S. pneumoniae infection, which can be rescued by the serum of bacteria-primed WT mice. The increased susceptibility to S. pneumoniae infection in Stat1-/- mice is also intrinsic to STAT1 requirement in MZ B cells. Collectively, these results define a differential regulation of TLR-mediated activation and differentiation of MZ B cells by STAT1 and reveal a STAT1-dependent, but IFN-independent, antibody response during infection and inflammation.

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.

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(β1-4)GlcNAc(β1-3)]Gal(β1-3)Gal(β1-4)Glc(β1-]n (serotype 1) and [4)[Neu5Ac(α2-6)GalNAc(β1-4)GlcNAc(β1-3)]Gal(β1-4)[Gal(α1-3)]Rha(β1-4)Glc(β1-]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.