Four-component Meningococcal Serogroup B Vaccine Induces Antibodies With Bactericidal Activity Against Diverse Outbreak Strains in Adolescents.

BACKGROUND: Neisseria meningitidis serogroup B (MenB) causes most meningitis outbreaks worldwide. We evaluated the ability of the 4-component MenB vaccine (4CMenB) to induce bactericidal activity against outbreak strains in adolescents. METHODS: Individual sera from 20 United States and 23 Chilean adolescents who received 2 doses of 4CMenB 2 months apart were assayed at prevaccination and 1 month after second dose using a human complement serum bactericidal antibody assay (hSBA) against a full or subset strain panel consisting of 14 MenB outbreak strains and 1 MenW hyperendemic strain collected between 2001 and 2017 in the United States, United Kingdom, and France. Bactericidal activity was determined as the percentage of adolescents with hSBA titer ≥1:4 or ≥1:8. RESULTS: One month after the second 4CMenB dose, antibodies from 65% to 100% of the US adolescents were able to kill 12 of 15 strains at 1:4 dilution. The remaining 3 strains were killed by 45%, 25%, and 15% of US adolescent sera. Similar percentages exhibited hSBA titers of ≥1:8. Across a subset of 4 strains, point estimates for the percentages of Chilean and US adolescents with hSBA titers of ≥1:4 after the second 4CMenB dose were similar (100% for strain M27703, 74% vs. 80% for M26312, 52% vs. 45% for M08 0240745), except for strain M39090 (91% vs. 65%). CONCLUSIONS: This study was the first to evaluate bactericidal activity elicited by a MenB vaccine against 15 outbreak strains. Two doses of 4CMenB elicited bactericidal activity against MenB outbreak strains and a hyperendemic MenW strain.

Antibodies Damage the Resilience of Fimbriae, Causing Them To Be Stiff and Tangled.

As adhesion fimbriae are a major virulence factor for many pathogenic Gram-negative bacteria, they are also potential targets for antibodies. Fimbriae are commonly required for initiating the colonization that leads to disease, and their success as adhesion organelles lies in their ability to both initiate and sustain bacterial attachment to epithelial cells. The ability of fimbriae to unwind and rewind their helical filaments presumably reduces their detachment from tissue surfaces with the shear forces that accompany significant fluid flow. Therefore, the disruption of functional fimbriae by inhibiting this resilience should have high potential for use as a vaccine to prevent disease. In this study, we show that two characteristic biomechanical features of fimbrial resilience, namely, the extension force and the extension length, are significantly altered by the binding of antibodies to fimbriae. The fimbriae that were studied are normally expressed on enterotoxigenic Escherichia coli, which are a major cause of diarrheal disease. This alteration in biomechanical properties was observed with bivalent polyclonal antifimbrial antibodies that recognize major pilin subunits but not with the Fab fragments of these antibodies. Thus, we propose that the mechanism by which bound antibodies disrupt the uncoiling of natural fimbria under force is by clamping together layers of the helical filament, thereby increasing their stiffness and reducing their resilience during fluid flow. In addition, we propose that antibodies tangle fimbriae via bivalent binding, i.e., by binding to two individual fimbriae and linking them together. Use of antibodies to disrupt physical properties of fimbriae may be generally applicable to the large number of Gram-negative bacteria that rely on these surface-adhesion molecules as an essential virulence factor. IMPORTANCE: Our study shows that the resiliency of colonization factor antigen I (CFA/I) and coli surface antigen 2 (CS2) fimbriae, which are current targets for vaccine development, can be compromised significantly in the presence of antifimbrial antibodies. It is unclear how the humoral immune system specifically interrupts infection after the attachment of enterotoxigenic Escherichia coli (ETEC) to the epithelial surface. Our study indicates that immunoglobulins, in addition to their well-documented role in adaptive immunity, can mechanically damage the resilience of fimbriae of surface-attached ETEC, thereby revealing a new mode of action. Our data suggest a mechanism whereby antibodies coat adherent and free-floating bacteria to impede fimbrial resilience. Further elucidation of this possible mechanism is likely to inform the development and refinement of preventive vaccines against ETEC diarrhea.

Immune responses induced by Clostridium difficile.

The spectrum of Clostridium difficile infections is highly variable, ranging from asymptomatic carriage to fatal colitis depending on the strain virulence and on the host, its gut microbiota and its immune response. After disruption of the gut microbiota, C. difficile pathogenesis can be divided into three steps: 1) contamination by spores and their germination; 2) multiplication of vegetative cells and intestinal colonization using colonization factors; 3) production of the toxins TcdA and TcdB, and for some strains, the binary toxin, which are responsible for the clinical signs. Three lines of defense counteract C. difficile. The first line is the epithelial barrier, which is breached by the toxins. Then, a rapid innate immune response follows, which forms the second line of defense. It provides very quick defense reactions against C. difficile but is non-specific and does not confer memory. C. difficile and its virulence factors, the toxins and colonization factors, induce a highly pro-inflammatory response, which can be either beneficial or harmful, but triggers the adaptive immunity as the third line of defense required to control the infectious process. Adaptive immunity provides a highly specific immune response against C. difficile with memory and long lasting immunity. The innate and adaptive immune responses against the toxins and surface components are analyzed as well as their role in disease susceptibility, severity and recurrences.

Identification of Poly-N-acetylglucosamine as a Major Polysaccharide Component of the Bacillus subtilis Biofilm Matrix.

Bacillus subtilis is intensively studied as a model organism for the development of bacterial biofilms or pellicles. A key component is currently undefined exopolysaccharides produced from proteins encoded by genes within the eps locus. Within this locus are four genes, epsHIJK, known to be essential for pellicle formation. We show they encode proteins synthesizing the broadly expressed microbial carbohydrate poly-N-acetylglucosamine (PNAG). PNAG was present in both pellicle and planktonic wild-type B. subtilis cells and in strains with deletions in the epsA-G and -L-O genes but not in strains deleted for epsH-K. Cloning of the B. subtilis epsH-K genes into Escherichia coli with in-frame deletions in the PNAG biosynthetic genes pgaA-D, respectively, restored PNAG production in E. coli. Cloning the entire B. subtilis epsHIJK locus into pga-deleted E. coli, Klebsiella pneumoniae, or alginate-negative Pseudomonas aeruginosa restored or conferred PNAG production. Bioinformatic and structural predictions of the EpsHIJK proteins suggest EpsH and EpsJ are glycosyltransferases (GT) with a GT-A fold; EpsI is a GT with a GT-B fold, and EpsK is an α-helical membrane transporter. B. subtilis, E. coli, and pga-deleted E. coli carrying the epsHIJK genes on a plasmid were all susceptible to opsonic killing by antibodies to PNAG. The immunochemical and genetic data identify the genes and proteins used by B. subtilis to produce PNAG as a significant carbohydrate factor essential for pellicle formation.

Protection Against Chlamydia trachomatis Infection and Upper Genital Tract Pathological Changes by Vaccine-Promoted Neutralizing Antibodies Directed to the VD4 of the Major Outer Membrane Protein.

The VD4 region from the Chlamydia trachomatis major outer membrane protein contains important neutralizing B-cell epitopes of relevance for antibody-mediated protection against genital tract infection. We developed a multivalent vaccine construct based on VD4s and their surrounding constant segments from serovars D, E, and F. Adjuvanted with cationic liposomes, this construct promoted strong immune responses to serovar-specific epitopes, the conserved LNPTIAG epitope and neutralized serovars D, E, and F. Vaccinated mice were protected against challenge, with protection defined as reduced bacterial numbers in vagina and prevention of pathological changes in the upper genital tract. Adoptive transfer of serum and T-cell depletion experiments demonstrated a dominant role for antibodies and CD4(+) T cells in the protective immune response. Integrating a multivalent VD4 construct into the sequence of the major outer membrane protein resulted in a protective and broadly neutralizing vaccine. Our findings emphasize the important role of antibodies in protection against Chlamydia trachomatis.

Identification and validation of a linear protective neutralizing epitope in the ß-pore domain of alpha toxin.

The plethora of virulence factors associated with Staphylococcus aureus make this bacterium an attractive candidate for a molecularly-designed epitope-focused vaccine. This approach, which necessitates the identification of neutralizing epitopes for incorporation into a vaccine construct, is being evaluated for pathogens where conventional approaches have failed to elicit protective humoral responses, like HIV-1 and malaria, but may also hold promise for pathogens like S. aureus, where the elicitation of humoral immunity against multiple virulence factors may be required for development of an effective vaccine. Among the virulence factors employed by S. aureus, animal model and epidemiological data suggest that alpha toxin, a multimeric ß-pore forming toxin like protective antigen from Bacillus anthracis, is particularly critical, yet no candidate neutralizing epitopes have been delineated in alpha toxin to date. We have previously shown that a linear determinant in the 2ß2-2ß3 loop of the pore forming domain of B. anthracis protective antigen is a linear neutralizing epitope. Antibody against this site is highly potent for neutralizing anthrax lethal toxin in vitro and for protection of rabbits in vivo from virulent B. anthracis. We hypothesized that sequences in the ß-pore of S. aureus alpha toxin that share structural and functional homology to ß-pore sequences in protective antigen would contain a similarly critical neutralizing epitope. Using an in vivo mapping strategy employing peptide immunogens, an optimized in vitro toxin neutralization assay, and an in vivo dermonecrosis model, we have now confirmed the presence of this epitope in alpha toxin, termed the pore neutralizing determinant. Antibody specific for this determinant neutralizes alpha toxin in vitro, and is highly effective for mitigating dermonecrosis and bacterial growth in a mouse model of S. aureus USA300 skin infection. The delineation of this linear neutralizing determinant in alpha toxin could facilitate the development of an epitope-focused vaccine against S. aureus.

Antibodies mediate formation of neutrophil extracellular traps in the middle ear and facilitate secondary pneumococcal otitis media.

Otitis media (OM) (a middle ear infection) is a common childhood illness that can leave some children with permanent hearing loss. OM can arise following infection with a variety of different pathogens, including a coinfection with influenza A virus (IAV) and Streptococcus pneumoniae (the pneumococcus). We and others have demonstrated that coinfection with IAV facilitates the replication of pneumococci in the middle ear. Specifically, we used a mouse model of OM to show that IAV facilitates the outgrowth of S. pneumoniae in the middle ear by inducing middle ear inflammation. Here, we seek to understand how the host inflammatory response facilitates bacterial outgrowth in the middle ear. Using B cell-deficient infant mice, we show that antibodies play a crucial role in facilitating pneumococcal replication. We subsequently show that this is due to antibody-dependent neutrophil extracellular trap (NET) formation in the middle ear, which, instead of clearing the infection, allows the bacteria to replicate. We further demonstrate the importance of these NETs as a potential therapeutic target through the transtympanic administration of a DNase, which effectively reduces the bacterial load in the middle ear. Taken together, these data provide novel insight into how pneumococci are able to replicate in the middle ear cavity and induce disease.

Chronic Mycoplasma conjunctivitis in house finches: host antibody response and M. gallisepticum VlhA expression.

Previous studies have shown that house finch field isolates of Mycoplasma gallisepticum (MG) vary in virulence and ability to induce an antibody response. After experimental inoculation, MG causes persistent, severe disease in a subset of individuals. In this study, we further characterized MG infection using five field isolates, with an emphasis on chronically diseased birds. After experimental inoculation of house finches, MG load was measured by quantitative PCR and anti-MG antibody responses were measured by ELISAs. Birds with chronic disease had significantly higher pathogen loads and antibody responses than did birds without chronic disease. Using a monoclonal antibody (MAb86) specific for a variant of the MG VlhA adhesin and immunodominant surface protein, we show that VlhA expression differs among MG isolates in this study, and that in vivo VlhA changes occur in house finches infected with MG. Overall, our results suggest that chronic MG disease has a strong pathogen-mediated component.

Structural stability of Burkholderia cenocepacia biofilms is reliant on eDNA structure and presence of a bacterial nucleic acid binding protein.

Cystic fibrosis (CF) is the most common lethal inherited genetic disorder affection Caucasians. Even with medical advances, CF is life-shortening with patients typically surviving only to age 38. Infection of the CF lung by Burkholderia cenocepacia presents exceptional challenges to medical management of these patients as clinically this microbe is resistant to virtually all antibiotics, is highly transmissible and infection of CF patients with this microbe renders them ineligible for lung transplant, often the last lifesaving option. Here we have targeted two abundant components of the B. cenocepacia biofilm for immune intervention: extracellular DNA and DNABII proteins, the latter of which are bacterial nucleic acid binding proteins. Treatment of B. cenocepacia biofilms with antiserum directed at one of these DNABII proteins (integration host factor or IHF) resulted in significant disruption of the biofilm. Moreover, when anti-IHF mediated destabilization of a B. cenocepacia biofilm was combined with exposure to traditional antibiotics, B. cenocepacia resident within the biofilm and thereby typically highly resistant to the action of antibiotics, were now rendered susceptible to killing. Pre-incubation of B. cenocepacia with anti-IHF serum prior to exposure to murine CF macrophages, which are normally unable to effectively degrade ingested B. cenocepacia, resulted in a statistically significant increase in killing of phagocytized B. cenocepacia. Collectively, these findings support further development of strategies that target DNABII proteins as a novel approach for treatment of CF patients, particularly those whose lungs are infected with B. cenocepacia.

The capsular polysaccharide Vi from Salmonella typhi is a B1b antigen.

Vaccination with purified capsular polysaccharide Vi Ag from Salmonella typhi can protect against typhoid fever, although the mechanism for its efficacy is not clearly established. In this study, we have characterized the B cell response to this vaccine in wild-type and T cell-deficient mice. We show that immunization with typhoid Vi polysaccharide vaccine rapidly induces proliferation in B1b peritoneal cells, but not in B1a cells or marginal zone B cells. This induction of B1b proliferation is concomitant with the detection of splenic Vi-specific Ab-secreting cells and protective Ab in Rag1-deficient B1b cell chimeras generated by adoptive transfer-induced specific Ab after Vi immunization. Furthermore, Ab derived from peritoneal B cells is sufficient to confer protection against Salmonella that express Vi Ag. Expression of Vi by Salmonella during infection did not inhibit the development of early Ab responses to non-Vi Ags. Despite this, the protection conferred by immunization of mice with porin proteins from Salmonella, which induce Ab-mediated protection, was reduced postinfection with Vi-expressing Salmonella, although protection was not totally abrogated. This work therefore suggests that, in mice, B1b cells contribute to the protection induced by Vi Ag, and targeting non-Vi Ags as subunit vaccines may offer an attractive strategy to augment current Vi-based vaccine strategies.

Anti-peptidoglycan antibodies and Fcγ receptors are the key mediators of inflammation in Gram-positive sepsis.

Gram-positive bacteria are an important public health problem, but it is unclear how they cause systemic inflammation in sepsis. Our previous work showed that peptidoglycan (PGN) induced proinflammatory cytokines in human cells by binding to an unknown extracellular receptor, followed by phagocytosis leading to the generation of NOD ligands. In this study, we used flow cytometry to identify host factors that supported PGN binding to immune cells. PGN binding required plasma, and plasma from all tested healthy donors contained IgG recognizing PGN. Plasma depleted of IgG or of anti-PGN Abs did not support PGN binding or PGN-triggered cytokine production. Adding back intact but not F(ab')2 IgG restored binding and cytokine production. Transfection of HEK293 cells with FcγRIIA enabled PGN binding and phagocytosis. These data establish a key role for anti-PGN IgG and FcγRs in supporting inflammation to a major structural element of Gram-positive bacteria and suggest that anti-PGN IgG contributes to human pathology in Gram-positive sepsis.

Antibodies generated against conserved antigens expressed by bacteria and allergen-bearing fungi suppress airway disease.

There has been a sharp rise in allergic asthma and asthma-related deaths in the developed world, in contrast to many childhood illnesses that have been reduced or eliminated. The hygiene hypothesis proposes that excessively sanitary conditions early in life result in autoimmune and allergic phenomena because of a failure of the immune system to receive proper microbial stimulation during development. We demonstrate that Abs generated against conserved bacterial polysaccharides are reactive with and dampen the immune response against chitin and Aspergillus fumigatus. A reduction in Ag uptake, cell influx, cell activation, and cytokine production occurred in the presence of anti-polysaccharide Abs, resulting in a striking decrease in the severity of allergic airway disease in mice. Overall, our results suggest that Ag exposure--derived from environmental sources, self-antigens, or vaccination--during the neonatal period has dramatic effects on the adult Ab response and modifies the development of allergic airway disease.

IgY stability in eggs stored at room temperature or at +4°C.

1. The aim of this study was to investigate the stability of immunoglobulin-Y (IgY) in stored eggs from immunised hens. 2. Eggs from individual hens were randomised and stored for up to one month at room temperature, or for up to 6 months at +4°C. IgY was extracted from the egg yolks and the antibody activities were tested by ELISA. 3. There was no significant reduction in antibody titres with egg storage under these conditions. 4. Egg yolks of immunised chickens provide an inexpensive source of large amounts of polyclonal antibodies for use in immunotherapy and immunoassays. By collecting eggs from different immunised hens and pooling their yolks, it should be possible to reduce batch-to-batch variation.

Antibodies anti-Shiga toxin 2 B subunit from chicken egg yolk: isolation, purification and neutralization efficacy.

Shiga toxins (Stx1 and Stx2) are the main virulence factors of enterohemorrhagic Escherichia coli (EHEC), a foodborne pathogen associated with diarrhea, hemorrhagic colitis and hemolytic uremic syndrome. The aim of this study was to evaluate the antibodies against Stx2 obtained from egg yolks of laying hens immunized with a recombinant Stx2B subunit. A high specific response in serum was observed 25 days after the first immunization and IgY antibodies were extracted from day 47th and purified from egg yolk. A concentration of 0.84 mg of total IgY/ml of egg yolk was obtained, of which 8% were antigen specific. The ability of anti-Stx2B IgY to recognize Stx2B and Stx2 either in solid-phase or in solution were evaluated and compared with anti-Stx2B rabbit antibodies by Western blotting and ELISA. The protective efficacy of IgY against Stx2 was determined by in vitro and in vivo experiments. The results show that IgY was able to recognize Stx2B and Stx2 in denatured conditions, attached to a solid-phase and free in solution. The anti-Stx2B IgY could effectively block the biological activity of Stx2 on Vero cells and protect mice from Stx2 challenge. The data suggest that immunization of hens with Stx2B could be a strategy to obtain at low cost a relatively high concentration of anti-Stx2 egg yolk IgY, able to neutralize Stx2 lethal activity. IgY technology could be an useful tool for research, diagnosis and therapy of EHEC infection.

Salmonella Typhimurium oral challenge model in mature broilers: bacteriological, immunological, and growth performance aspects.

In this study, Salmonella enterica serovar Typhimurium challenge models were tested to identify the best conditions under which to perform the experimental infection of 3-wk-old broilers. Such a model would be useful to study the efficiency of therapeutic treatments that could take place at the end of the grow-out period. Salmonella-free chicks were obtained from a breeder flock vaccinated with Salmonella. Intestinal maternal immunity was monitored by ELISA analyses at 2, 9, and 16 d of age. Data indicated that protection of maternal origin was not maintained over time and was drastically reduced at 9 d of age (P < 0.01). At 21 d of age, chickens were orally inoculated with Salmonella Typhimurium. The effects of the oral challenge dose (0, 3 × 10(3), 3 × 10(6), and 3 × 10(9) cfu/bird) and vancomycin pretreatment (no administration or 25 mg/bird) on intestinal immune responses, growth performance, and Salmonella colonization of chickens were investigated. After infection, the mucosal immune response was rapid, with increased (P < 0.01) anti-Salmonella Typhimurium IgA titers measured at 8 d postinfection in intestinal contents. A linear relationship (P < 0.05) existed between specific IgA levels in intestinal and cecal contents and the challenge dose inoculated. None of the challenge protocols caused mortality or clinical symptoms after infection. Nevertheless, the experimental infection induced a significant deterioration of growth performance. The pretreatment with 25 mg of vancomycin at 3 h before Salmonella inoculation was able to establish stable infection rates among the population of 3-wk-old infected chickens. Nevertheless, Salmonella shedding was not stable over the rearing period, and the bacteria seemed to be naturally eliminated from most birds at 22 d postinfection. This natural clearance of the gut, which was related, at least in part, to the intestinal immune response, should limit the usability of the created mature challenge model within 1 to 2 wk after inoculation.

Neisseria lactamica selectively induces mitogenic proliferation of the naive B cell pool via cell surface Ig.

Neisseria lactamica is a commensal bacteria that colonizes the human upper respiratory tract mucosa during early childhood. In contrast to the closely related opportunistic pathogen Neisseria meningitidis, there is an absence of adaptive cell-mediated immunity to N. lactamica during the peak age of carriage. Instead, outer membrane vesicles derived from N. lactamica mediate a B cell-dependent proliferative response in mucosal mononuclear cells that is associated with the production of polyclonal IgM. We demonstrate in this study that this is a mitogenic human B cell response that occurs independently of T cell help and any other accessory cell population. The ability to drive B cell proliferation is a highly conserved property and is present in N. lactamica strains derived from diverse clonal complexes. CFSE staining of purified human tonsillar B cells demonstrated that naive IgD(+) and CD27(-) B cells are selectively induced to proliferate by outer membrane vesicles, including the innate CD5(+) subset. Neither purified lipooligosaccharide nor PorB from N. lactamica is likely to be responsible for this activity. Prior treatment of B cells with pronase to remove cell-surface Ig or treatment with BCR-specific Abs abrogated the proliferative response to N. lactamica outer membrane vesicles, suggesting that this mitogenic response is dependent upon the BCR.

Secretory-IgA antibodies play an important role in the immunity to Streptococcus pneumoniae.

This study was designed to investigate whether secretory-IgA (S-IgA) Abs induced by a pneumococcal surface protein A (PspA)-based nasal vaccine are necessary for prevention of streptococcal colonization. Mice nasally immunized with PspA plus a plasmid expressing Flt3 ligand (pFL) cDNA as a mucosal adjuvant showed significantly higher levels of PspA-specific S-IgA and IgG Ab responses in both plasma and nasal washes when compared with naive mice. Although IgA(-/-) mice given nasal PspA plus pFL had significantly high levels of PspA-specific IgG Abs, high numbers of CFUs were detected in nasal washes and nasal passages. In contrast, vaccinated wild-type mice showed essentially no bacteria in the nasal cavity. Further, a nasal vaccine consisting of PspA plus pFL effectively reduced pre-existing Streptococcus pneumoniae in the nasal cavity. These results show that PspA-based vaccine-induced specific S-IgA Abs play a necessary role in the regulation of S. pneumoniae colonization in the nasal cavity.

Contribution of cell surface protein antigen c of Streptococcus mutans to platelet aggregation.

INTRODUCTION: Streptococcus mutans is considered to be one of the pathogens that cause infective endocarditis. The purpose of the present study was to examine the properties of S. mutans with regard to platelet aggregation by focusing on its high molecular protein antigen c (PAc). METHODS: The platelet aggregation properties of six clinical strains and one isogenic mutant strain of S. mutans were analysed using an aggregometer and confocal microscopy, as well as with an inhibition assay of platelet aggregation using anti-PAc serum. RESULTS: S. mutans strains with PAc expression induced platelet aggregation, while a PAc-deficient mutant and two clinical isolates with no PAc expression did not. When platelets were pretreated with higher amounts of anti-PAc serum, the platelet aggregation rate was reduced in a dose-dependent manner, indicating that PAc binds directly to platelets. CONCLUSION: S. mutans PAc is involved in human platelet aggregation and may be one of the virulence factors in the pathogenesis of infective endocarditis.

A novel ICOS-independent, but CD28- and SAP-dependent, pathway of T cell-dependent, polysaccharide-specific humoral immunity in response to intact Streptococcus pneumoniae versus pneumococcal conjugate vaccine.

Polysaccharide (PS)- and protein-specific murine IgG responses to intact Streptococcus pneumoniae (Pn) are both dependent on CD4(+) T cell help, B7-dependent costimulation, and CD40/CD40 ligand interactions. However, the primary PS-specific, relative to protein-specific, IgG response terminates more rapidly, requires a shorter period of T cell help and B7-dependent costimulation, and fails to generate memory. In light of the critical role for ICOS/ICOS ligand interactions in sustaining T cell-dependent Ig responses and promoting germinal center reactions, we hypothesized that this interaction was nonessential for PS-specific IgG responses to Pn. We now demonstrate that ICOS(-/-), relative to wild-type, mice elicit a normal PS-specific IgG isotype response to Pn, despite marked inhibition of both the primary and secondary IgG anti-protein (i.e., PspA, PspC, and PsaA) response. A blocking anti-ICOS ligand mAb injected during primary Pn immunization inhibits both the primary anti-protein response and the generation of protein-specific memory, but has no effect when injected during secondary immunization. In contrast to Pn, both PS- and protein-specific IgG responses to a pneumococcal conjugate vaccine are inhibited in ICOS(-/-) mice. ICOS(-/-) mice immunized with intact Pn or conjugate exhibit nearly complete abrogation in germinal center formation. Finally, although mice that lack the adaptor molecule SAP (SLAM-associated protein) resemble ICOS(-/-) mice (and can exhibit decreased ICOS expression), we observe that the PS-specific, as well as protein-specific, IgG responses to both Pn and conjugate are markedly defective in SAP(-/-) mice. These data define a novel T cell-, SAP-, and B7-dependent, but ICOS-independent, extrafollicular pathway of Ig induction.

Staphylococcus aureus-induced plasmacytoid dendritic cell activation is based on an IgG-mediated memory response.

Type I IFNs represent a major antimicrobial defense mechanism due to their property of enhancing immune responses by priming both innate and adaptive immune cells. Plasmacytoid dendritic cells (pDC) are the major source of type I IFN in the human body and represent innate immune cells involved in first-line defense against invading pathogens. Although pDC activation has been extensively studied upon stimulation with synthetic TLR ligands, viruses, and intracellular bacteria, there is only scarce information on extracellular bacteria. In this study we show that the triggering of human pDC-derived IFN-alpha secretion by Staphylococcus aureus is independent of TLR2 and specific for coagulase-positive staphylococci. Specificity of the pDC response to S. aureus is independent of the bacterial virulence factors protein A and alpha-toxin but is mediated by Ag-specific IgG and CD32. S. aureus-induced pDC activation can be blocked by inhibitory DNA oligonucleotides and chloroquine, suggesting that engagement of TLR7/9 by bacterial nucleic acids after CD32-mediated uptake of these compounds may play a central role in this process. Altogether, we propose that in marked contrast to nonselective TLR2-dependent activation of most innate immune cells, pDC activation by S. aureus represents an Ag-specific memory response since it requires the presence of class-switched immunoglobulins.