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Background: Antibody levels wane rapidly after Meningococcal serogroup C conjugate (MenCC) vaccination in young children, rendering the need for an adolescent booster dose. It is not clear whether circulating memory B cells are associated with persistence of MenC-specific antibody levels., Methods: Measurement of MenC-specific IgG and IgA memory B cells and levels of serum and salivary MenC-specific IgG and IgA in healthy 10-, 12- and 15-year-olds prior to and one month and one year after a MenCC booster vaccination. All participants had received a primary MenCC vaccination nine years earlier., Results: The number of circulating MenC-specific IgG memory B cells prior to booster was low and not predictive for MenC-specific IgG responses in serum or saliva post-booster, whereas the number of MenC-specific IgA memory B cells pre-booster positively correlated with MenC-specific IgA levels in saliva post-booster (R = 0.5, P<0.05). The booster induced a clear increase in the number of MenC-specific IgG and IgA memory B cells. The number of MenC-PS-specific IgG memory B cells at 1 month post-booster was highest in the 12-year-olds. The number of MenC-specific memory B cells at one month post-booster showed no correlation with the rate of MenC-specific antibody decay throughout the first year post-booster., Conclusions: Circulating MenC-specific IgA memory B cells correlate with IgA responses in saliva, whereas circulating MenC-specific IgG memory B cells are not predictive for MenC-specific IgG responses in serum or saliva. Our results are suggestive for age-dependent differences in pre-existing memory against MenC.

Incidence of pneumococcal disease is disproportionally high in infants and elderly. Nasopharyngeal colonisation by Streptococcus pneumoniae is considered a prerequisite for disease but unlike in children, carriage in elderly is rarely detected. Here, we tested for S. pneumoniae in nasopharyngeal and saliva samples collected from community-dwelling elderly with influenza-like-illness (ILI). Trans-nasal nasopharyngeal, trans-oral nasopharyngeal and saliva samples (n = 270 per sample type) were collected during winter/spring 2011/2012 from 135 persons aged 60-89 at onset of ILI and 7-9 weeks later following recovery. After samples were tested for pneumococci by conventional culture, all plate growth was collected. DNA extracted from plate harvests was tested by quantitative-PCRs (qPCR) specific for S. pneumoniae and serotypes included in the 13-valent pneumococcal conjugated vaccine (PCV13). Pneumococci were cultured from 14 of 135 (10%) elderly with none of the sampled niches showing superiority in carriage detection. With 76/270 (28%) saliva, 31/270 (11%) trans-oral and 13/270 (5%) trans-nasal samples positive by qPCR, saliva was superior to nasopharyngeal swabs (p<0.001) in qPCR-based carriage detection. Overall, from all methods used in the study, 65 of 135 (48%) elderly carried pneumococci at least once and 26 (19%) at both study time points. The difference between carriage prevalence at ILI (n = 49 or 36%) versus recovery (n = 42 or 31%) was not significant (p = 0.38). At least 23 of 91 (25%) carriage events in 19 of 65 (29%) carriers were associated with PCV13-serotypes. We detected a large reservoir of pneumococci in saliva of elderly, with PCV13-serotype distribution closely resembling the contemporary carriage of serotypes reported in the Netherlands for PCV-vaccinated infants.

While nasopharyngeal sampling is the gold standard for the detection of Streptococcus pneumoniae carriage, historically seen, saliva sampling also seems highly sensitive for pneumococcal detection. We investigated S. pneumoniae carriage in saliva from fifty schoolchildren by conventional and molecular methods. Saliva was first culture-enriched for pneumococci, after which, DNA was extracted from all bacterial growth and tested by quantitative-PCR (qPCR) for pneumococcus-specific genes lytA and piaA. Next, serotype composition of the samples was determined by serotype-specific qPCRs, conventional-PCRs (cPCR) and sequencing of cPCR amplicons. Although only 2 (4%) of 50 samples were positive by conventional diagnostic culture, 44 (88%) were positive for pneumococci by qPCR. In total, we detected the presence of at least 81 pneumococcal strains representing 20 serotypes in samples from 44 carriers with 23 carriers (52%) positive for multiple (up to 6) serotypes. The number of serotypes detected per sample correlated with pneumococcal abundance. This study shows that saliva could be used as a tool for future pneumococcal surveillance studies. Furthermore, high rates of pneumococcal carriage and co-carriage of multiple pneumococcal strains together with a large number of serotypes in circulation suggests a ubiquitous presence of S. pneumoniae in saliva of school-aged children. Our results also suggest that factors promoting pneumococcal carriage within individual hosts may weaken competitive interactions between S. pneumoniae strains.

Background: Meningococcal serogroup C (MenC) specific antibody levels decline rapidly after a single primary MenC conjugate (MenCC) vaccination in preschool children. A second MenCC vaccination during (pre)adolescence might attain longer lasting individual and herd protection. We aimed to establish an appropriate age for a (pre)adolescent MenCC booster vaccination., Methods: A phase-IV trial with healthy 10-year-olds (n = 91), 12-year-olds (n = 91) and 15-year-olds (n = 86) who were primed with a MenCC vaccine nine years earlier. All participants received a booster vaccination with the same vaccine. Serum bactericidal antibody assay titers (SBA, using baby rabbit complement), MenC-polysaccharide (MenC-PS) specific IgG, IgG subclass and avidity and tetanus-specific IgG levels were measured prior to (T0) and 1 month (T1) and 1 year (T2) after the booster. An SBA titer ≥8 was the correlate of protection., Results: 258 (96.3%) participants completed all three study visits. At T0, 19% of the 10-year-olds still had an SBA titer ≥8, compared to 34% of the 12-year-olds (P = 0.057) and 45% of the 15-year-olds (P<0.001). All participants developed high SBA titers (GMTs>30,000 in all age groups) and MenC-PS specific IgG levels at T1. IgG levels mainly consisted of IgG1, but the contribution of IgG2 increased with age. At T2, 100% of participants still had an SBA titer ≥8, but the 15-year-olds showed the highest protective antibody levels and the lowest decay., Conclusion: Nine years after primary MenCC vaccination adolescents develop high protective antibody levels in response to a booster and are still sufficiently protected one year later. Our results suggest that persistence of individual--and herd--protection increases with the age at which an adolescent booster is administered., Trial Registration: EU Clinical Trials Database 2011-000375-13 Dutch Trial Register NTR3521.

The human nasopharynx is the main reservoir for Streptococcus pneumoniae. We applied conventional and molecular methods to determine the prevalence of S. pneumoniae nasopharyngeal colonization in adults. Paired trans-orally and trans-nasally obtained nasopharyngeal samples from 268 parents of 24-month-old children were assessed for pneumococcal presence. Parents were classified as colonized when live pneumococci were recovered from either sample cultured on medium selective for S. pneumoniae. Of the 52 (19%) colonized parents 49 (18%) were culture-positive in trans-nasal and 10 (4%) in trans-oral samples. Bacterial growth was harvested from these cultures, DNA isolated and tested by quantitative-PCR (qPCR) targeting lytA and piaA genes specific for S. pneumoniae. A sample was considered positive if signals for both genes were detected. Altogether 105 (39%) individuals were classified as positive for pneumococcus by qPCR including 50 (19%) in trans-nasal and 94 (35%) in trans-oral settings. Although significantly more trans-nasal compared to trans-oral samples were culture-positive for S. pneumoniae at the primary diagnostic step (p<0.001) the opposite was observed in qPCR results (p<0.001). To confirm the presence of live pneumococcus in samples positive by qPCR but negative at the initial diagnostic step, we serially-diluted cell harvests, re-cultured and carefully examined for S. pneumoniae presence. Live pneumococci were recovered from an additional 43 parents including 42 positive in trans-oral and 4 in trans-nasal samples increasing the number of individuals culture- and qPCR-positive to 93 (35%) and positive by either of two methods to 107 (40%). There were significantly more trans-oral than trans-nasal samples positive for pneumococcus by both culture and qPCR (n = 71; 27%; vs. n = 50; 19%; p<0.05). Our data suggest that pneumococcal colonization is more common in adults than previously estimated and point towards the superiority of a trans-oral over a trans-nasal approach when testing adults for colonization with S. pneumoniae.

Respiratory infectious diseases are mainly caused by viruses or bacteria that often interact with one another. Although their presence is a prerequisite for subsequent infections, viruses and bacteria may be present in the nasopharynx without causing any respiratory symptoms. The upper respiratory tract hosts a vast range of commensals and potential pathogenic bacteria, which form a complex microbial community. This community is assumed to be constantly subject to synergistic and competitive interspecies interactions. Disturbances in the equilibrium, for instance due to the acquisition of new bacteria or viruses, may lead to overgrowth and invasion. A better understanding of the dynamics between commensals and pathogens in the upper respiratory tract may provide better insight into the pathogenesis of respiratory diseases. Here we review the current knowledge regarding specific bacterial-bacterial and viral-bacterial interactions that occur in the upper respiratory niche, and discuss mechanisms by which these interactions might be mediated. Finally, we propose a theoretical model to summarize and illustrate these mechanisms.

Introduction: Accurate analyses of microbiota composition of low-density communities (10(3)-10(4) bacteria/sample) can be challenging. Background DNA from chemicals and consumables, extraction biases as well as differences in PCR efficiency can significantly interfere with microbiota assessment. This study was aiming to establish protocols for accurate microbiota analysis at low microbial density., Methods: To examine possible effects of bacterial density on microbiota analyses we compared microbiota profiles of serial diluted saliva and low (nares, nasopharynx) and high-density (oropharynx) upper airway communities in four healthy individuals. DNA was extracted with four different extraction methods (Epicentre Masterpure, Qiagen DNeasy, Mobio Powersoil and a phenol bead-beating protocol combined with Agowa-Mag-mini). Bacterial DNA recovery was analysed by 16S qPCR and microbiota profiles through GS-FLX-Titanium-Sequencing of 16S rRNA gene amplicons spanning the V5-V7 regions., Results: Lower template concentrations significantly impacted microbiota profiling results. With higher dilutions, low abundant species were overrepresented. In samples of <10(5) bacteria per ml, e.g. DNA <1 pg/µl, microbiota profiling deviated from the original sample and other dilutions showing a significant increase in the taxa Proteobacteria and decrease in Bacteroidetes. In similar low density samples, DNA extraction method determined if DNA levels were below or above 1 pg/µl and, together with lysis preferences per method, had profound impact on microbiota analyses in both relative abundance as well as representation of species., Conclusion: This study aimed to interpret microbiota analyses of low-density communities. Bacterial density seemed to interfere with microbiota analyses at < than 10(6) bacteria per ml or DNA <1 pg/µl. We therefore recommend this threshold for working with low density materials. This study underlines that bias reduction is crucial for adequate profiling of especially low-density bacterial communities.

Background: Meningococci produce a penta-acylated instead of hexa-acylated lipid A when their lpxL1 gene is inactivated. Meningococcal strains with such lipid A endotoxin variants have been found previously in adult meningitis patients, where they caused less blood coagulopathy because of decreased TLR4 activation., Methods: A cohort of 448 isolates from patients with invasive meningococcal disease in the Netherlands were screened for the ability to induce IL-6 in monocytic cell Mono Mac 6 cells. The lpxL1 gene was sequenced of isolates, which show poor capacity to induce IL-6.. Clinical characteristics of patients were retrieved from hospital records., Results: Of 448 patients, 29 (6.5%) were infected with meningococci expressing a lipid A variant strain. Lipid A variation was not associated with a specific serogroup or genotype. Infections with lipid A variants were associated with older age (19.3 vs. 5.9 (median) years, p = 0.007) and higher prevalence of underlying comorbidities (39% vs. 17%; p = 0.004) compared to wild-type strains. Patients infected with lipid A variant strains had less severe infections like meningitis or shock (OR 0.23; 95%CI 0.09-0.58) and were less often admitted to intensive care (OR 0.21; 95%CI 0.07-0.60) compared to wild-type strains, independent of age, underlying comorbidities or strain characteristics., Conclusions: In adults with meningococcal disease lipid A variation is rather common. Infection with penta-acylated lipid A variant meningococci is associated with a less severe disease course.

To understand the role of human microbiota in health and disease, we need to study effects of environmental and other epidemiological variables on the composition of microbial communities. The composition of a microbial community may depend on multiple factors simultaneously. Therefore we need multivariate methods for detecting, analyzing and visualizing the interactions between environmental variables and microbial communities. We provide two different approaches for multivariate analysis of these complex combined datasets: (i) We select variables that correlate with overall microbiota composition and microbiota members that correlate with the metadata using canonical correlation analysis, determine independency of the observed correlations in a multivariate regression analysis, and visualize the effect size and direction of the observed correlations using heatmaps; (ii) We select variables and microbiota members using univariate or bivariate regression analysis, followed by multivariate regression analysis, and visualize the effect size and direction of the observed correlations using heatmaps. We illustrate the results of both approaches using a dataset containing respiratory microbiota composition and accompanying metadata. The two different approaches provide slightly different results; with approach (i) using canonical correlation analysis to select determinants and microbiota members detecting fewer and stronger correlations only and approach (ii) using univariate or bivariate analyses to select determinants and microbiota members detecting a similar but broader pattern of correlations. The proposed approaches both detect and visualize independent correlations between multiple environmental variables and members of the microbial community. Depending on the size of the datasets and the hypothesis tested one can select the method of preference.

Background: The CRM197-conjugated 7-valent pneumococcal vaccine (PCV7) is protective against vaccine serotype disease and nasopharyngeal carriage. Data on PCV7-induced mucosal antibodies in relation to systemic or natural anticapsular antibodies are scarce., Methods: In a randomized controlled setting, children received PCV7 at age 2 and 4 months (2-dose group), at age 2, 4 and 11 months (2+1-dose group) or no PCV7 (control group). From 188 children paired saliva samples were collected at 12 and 24 months of age. From a subgroup of 15 immunized children also serum samples were collected. IgG and IgA antibody-levels were measured by multiplex immunoassay., Results: At 12 months, both vaccine groups showed higher serum and saliva IgG-levels against vaccine serotypes compared with controls which sustained until 24 months for most serotypes. Salivary IgG-levels were 10-20-fold lower compared to serum IgG, however, serum and saliva IgG-levels were highly correlated. Serum and salivary IgA-levels were higher in both vaccine groups at 12 months compared with controls, except for serotype 19F. Higher salivary IgA levels remained present for most serotypes in the 2+1-dose group until 24 months, but not in the 2-dose group. Salivary IgA more than IgG, increased after documented carriage of serotypes 6B, 19F and 23F In contrast to IgG, salivary IgA-levels were comparable with serum, suggesting local IgA-production., Conclusions: PCV7 vaccination results in significant increases in salivary IgG and IgA-levels, which are more pronounced for IgG when compared to controls. In contrast, salivary anticapsular IgA-levels seemed to respond more to natural boosting. Salivary IgG and IgA-levels correlate well with systemic antibodies, suggesting saliva might be useful as potential future surveillance tool.

Background: Shifts in pneumococcal serotypes following introduction of 7-valent pneumococcal conjugate vaccine (PCV-7) may alter the presence of other bacterial pathogens co-inhabiting the same nasopharyngeal niche., Methodology/principal Findings: Nasopharyngeal prevalence rates of S. pneumoniae, S. aureus, H. influenzae and M. catarrhalis were investigated before, 3 and 4.5 years after introduction of PCV-7 in the national immunisation program in children at 11 and 24 months of age, and parents of 24-month-old children (n≈330/group) using conventional culture methods. Despite a virtual disappearance of PCV-7 serotypes over time, similar overall pneumococcal rates were observed in all age groups, except for a significant reduction in the 11-month-old group (adjusted Odds Ratio after 4.5 years 0.48, 95% Confidence Interval 0.34-0.67). Before, 3 and 4.5 years after PCV-7 implementation, prevalence rates of S. aureus were 5%, 9% and 14% at 11 months of age (3.59, 1.90-6.79) and 20%, 32% and 34% in parents (1.96, 1.36-2.83), but remained similar at 24 months of age, respectively. Prevalence rates of H. influenzae were 46%, 65% and 65% at 11 months (2.22, 1.58-3.13), 52%, 73% and 76% at 24 months of age (2.68, 1.88-3.82) and 23%, 30% and 40% in parents (2.26, 1.58-3.33), respectively. No consistent changes in M. catarrhalis carriage rates were observed over time., Conclusions/significance: In addition to large shifts in pneumococcal serotypes, persistently higher nasopharyngeal prevalence rates of S. aureus and H. influenzae were observed among young children and their parents after PCV-7 implementation. These findings may have implications for disease incidence and antibiotic treatment in the post-PCV era.

Background: High rates of potentially pathogenic bacteria and respiratory viruses can be detected in the upper respiratory tract of healthy children. Investigating presence of and associations between these pathogens in healthy individuals is still a rather unexplored field of research, but may have implications for interpreting findings during disease., Methodology/principal Findings: We selected 986 nasopharyngeal samples from 433 6- to 24-month-old healthy children that had participated in a randomized controlled trial. We determined the presence of 20 common respiratory viruses using real-time PCR. Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis and Staphylococcus aureus were identified by conventional culture methods. Information on risk factors was obtained by questionnaires. We performed multivariate logistic regression analyses followed by partial correlation analysis to identify the overall pattern of associations. S. pneumoniae colonization was positively associated with the presence of H. influenzae (adjusted odds ratio 1.60, 95% confidence interval 1.18-2.16), M. catarrhalis (1.78, 1.29-2.47), human rhinoviruses (1.63, 1.19-2.22) and enteroviruses (1.97, 1.26-3.10), and negatively associated with S. aureus presence (0.59, 0.35-0.98). H. influenzae was positively associated with human rhinoviruses (1.63, 1.22-2.18) and respiratory syncytial viruses (2.78, 1.06-7.28). M. catarrhalis colonization was positively associated with coronaviruses (1.99, 1.01-3.93) and adenoviruses (3.69, 1.29-10.56), and negatively with S. aureus carriage (0.42, 0.25-0.69). We observed a strong positive association between S. aureus and influenza viruses (4.87, 1.59-14.89). In addition, human rhinoviruses and enteroviruses were positively correlated (2.40, 1.66-3.47), as were enteroviruses and human bocavirus, WU polyomavirus, parainfluenza viruses, and human parechovirus. A negative association was observed between human rhinoviruses and coronaviruses., Conclusions/significance: Our data revealed high viral and bacterial prevalence rates and distinct bacterial-bacterial, viral-bacterial and viral-viral associations in healthy children, hinting towards the complexity and potential dynamics of microbial communities in the upper respiratory tract. This warrants careful consideration when associating microbial presence with specific respiratory diseases.

Background: Whooping cough is a respiratory disease caused by Bordetella pertussis, which induces mucosal IgA antibodies that appear to be relevant in protection. Serum IgA responses are measured after pertussis infection and might provide an additional role in pertussis diagnostics. However, the possible interfering role for pertussis vaccinations in the induction of serum IgA antibodies is largely unknown., Methods/principal Findings: We compared serum IgA responses in healthy vaccinated children between 1 and 10 years of age with those in children who despite vaccinations recently were infected with Bordetella pertussis. All children have been vaccinated at 2, 3, 4 and 11 months of age with either the Dutch whole-cell pertussis (wP) vaccine or an acellular pertussis (aP) vaccine and additionally received an aP booster vaccination at 4 years of age. Serum IgA responses to pertussis toxin (PT), filamentous heamagglutinin (FHA) and pertactin (Prn) were measured with a fluorescent multiplex bead-based immuno-assay. An ELISPOT-assay was used for the detection of IgA-memory B-cells specific to these antigens. Serum IgA levels to all pertussis vaccine antigens were significantly higher in infected children compared with healthy children. High correlations between anti-PT, anti-FHA or anti-Prn IgA and IgG levels were found in infected children and to some degree in wP primed children, but not at all in aP primed children. Highest numbers of IgA-pertussis-specific memory B-cells were observed after infection and generally comparable numbers were found after wP and aP vaccination., Conclusions: This study provides new insight in the diagnostic role for serum IgA responses against PT in vaccinated children. Since aP vaccines induce high serum IgG levels that interfere with pertussis diagnostics, serum IgA-PT levels will provide an additional diagnostic role. High levels of serum IgA for PT proved specific for recent pertussis infection with reasonable sensitivity, whereas the role for IgA levels against FHA and Prn in diagnosing pertussis remains controversial.

Background: All children and adolescents between 1 and 19 years of age in The Netherlands received a single meningococcal serogroup C conjugate (MenCC) vaccine in 2002. During follow-up 4-5 years later, the persistence of MenC polysaccharide-specific IgG was found to be dependent on age of vaccination with higher IgG levels in the oldest immunized age categories., Methods and Findings: Two cross-sectional population-based serum banks, collected in 1995/1996 and in 2006/2007, were used for this study. We measured MenC polysaccharide-specific IgM, the IgG1 and IgG2 subclasses and determined the avidity of the IgG antibodies. We report that the age-related persistence of IgG after immunization with the MenCC vaccine seemed to result from an increase of IgG2 levels with age, while IgG1 levels remained stable throughout the different age-cohorts. Furthermore, an age-related increase in IgM levels was observed, correlating with the persistence of IgG antibodies with age. It is noteworthy that the increase in IgG2 correlated with a reduced IgG-avidity with age., Conclusion: These date indicate that the classical characteristics of a T-cell-dependent antibody response as elicited by protein based vaccines might not be completely applicable when conjugate vaccines are administered to older children and adolescents up to 18 years of age. The response elicited by the MenCC vaccine seemed to be more a mixture of both T cell dependent and T cell independent responses in terms of humoral immunological characteristics.

Background: Heptavalent pneumococcal conjugate vaccine (PCV7) shifts nasopharyngeal colonisation with vaccine serotype pneumococci towards nonvaccine serotypes. Because of the reported negative association of vaccine serotype pneumococci and Staphylococcus aureus in the nasopharynx, we explored the effect of PCV7 on nasopharyngeal colonisation with S. aureus in children and parents., Methodology/principal Findings: This study was part of a randomised controlled trial on the effect of PCV7 on pneumococcal carriage, enrolling healthy newborns who were randomly assigned (1:1:1) to receive PCV7 (1) at 2 and 4 months of age (2) at 2, 4 and 11 months or (3) no PCV7 (controls). Nasopharyngeal colonisation of S. aureus was a planned secondary outcome. Nasopharyngeal swabs were obtained from all children over a 2-year period with 6-months interval and from one parent at the child's age of 12 and 24 months and cultured for Streptococcus pneumoniae and S. aureus. Between July 2005 and February 2006, 1005 children were enrolled and received either 2-doses of PCV7 (n = 336), 2+1-doses (336) or no dose (n = 333) before PCV7 implementation in the Dutch national immunization program. S. aureus colonisation had doubled in children in the 2+1-dose group at 12 months of age compared with unvaccinated controls (10.1% versus 5.0%; p = 0.019). A negative association for co-colonisation of S. pneumoniae and S. aureus was observed for both vaccine serotype (adjusted odds ratio (aOR) 0.53, 95% confidence interval (CI) 0.38-0.74) and nonvaccine serotype pneumococci (aOR 0.67, 95% CI 0.52-0.88)., Conclusions/significance: PCV7 induces a temporary increase in S. aureus colonisation in children around 12 months of age after a 2+1-dose PCV7 schedule. The potential clinical consequences are unknown and monitoring is warranted., Trial Registration: ClinicalTrials.gov NCT00189020.

Background: In 2002 a Meningococcal serogroup C (MenC) conjugate vaccine, with tetanus toxoid as carrier protein, was introduced in the Netherlands as a single-dose at 14 months of age. A catch-up campaign was performed targeting all individuals aged 14 months to 18 years. We determined the MenC-specific immunity before and after introduction of the MenC conjugate (MenCC) vaccine., Methods and Findings: Two cross-sectional population-based serum banks, collected in 1995/1996 (n = 8539) and in 2006/2007 (n = 6386), were used for this study. The main outcome measurements were the levels of MenC polysaccharide(PS)-specific IgG and serum bactericidal antibodies (SBA) after routine immunization, 4-5 years after catch-up immunization or by natural immunity. There was an increasing persistence of PS-specific IgG and SBA with age in the catch-up immunized cohorts 4-5 years after their MenCC immunization (MenC PS-specific IgG, 0.25 microg/ml (95%CI: 0.19-0.31 microg/ml) at age 6 years, gradually increasing to 2.34 microg/ml,(95%CI: 1.70-3.32 microg/ml) at age 21-22 years). A comparable pattern was found for antibodies against the carrier protein in children immunized above 9 years of age. In case of vaccination before the age of 5 years, PS-specific IgG was rapidly lost. For all age-cohorts together, SBA seroprevalence (> or =8) increased from 19.7% to 43.0% in the pre- and post-MenC introduction eras, respectively. In non-immunized adults the SBA seroprevalence was not significantly different between the pre- and post-MenC introduction periods, whereas PS-specific IgG was significantly lower in the post-MenC vaccination (GMT, age > or =25 years, 0.10 microg/ml) era compared to the pre-vaccination (GMT, age > or =25 years, 0.43 microg/ml) era., Conclusion: MenCC vaccination administered above 5 years of age induced high IgG levels compared to natural exposure, increasing with age. In children below 14 months of age and non-immunized cohorts lower IgG levels were observed compared to the pre-vaccination era, whereas functional levels remained similar in adults. Whether the lower IgG poses individuals at increased risk for MenC disease should be carefully monitored. Large-scale introduction of a MenCC vaccine has led to improved protection in adolescents, but in infants a single-dose schedule may not provide sufficient protection on the long-term and therefore a booster-dose early in adolescence should be considered.