Pertussis seroprevalence among adults of reproductive age (20-39 years) in fourteen European countries.

The reported incidence of pertussis in European countries varies considerably. We aimed to study specific Bordetella pertussis seroprevalence in Europe by measuring serum IgG antibody levels to pertussis toxin (anti-PT IgG). Fourteen national laboratories participated in this study including Belgium, Denmark, Finland, Greece, Hungary, Italy, Lithuania, Malta, Norway, Poland, Portugal, Romania, Spain, and Sweden. Each country collected approximately 250 samples (N = 7903) from the age groups 20-29 years (N = 3976) and 30-39 years (N = 3927) during 2010-2013. Samples were anonymous residual sera from diagnostic laboratories and were analyzed at the national laboratories by a Swedish reference method, a commercial ELISA kit, or were sent to Sweden for analysis. The median anti-PT IgG concentrations ranged from 4 to 13.6 IU/mL. The proportion of samples with anti-PT IgG ≥100 IU/mL, indicating a recent infection ranged from 0.2% (Hungary) to 5.7% (Portugal). The highest proportion of sera with anti-PT IgG levels between 50 and <100 IU/mL, indicating an infection within the last few years, was found in Portugal (12.3%) and Italy (13.9%). This study shows that the circulation of B. pertussis is quite extensive in adults, aged 20-39 years, despite well-established vaccination programs in Europe.

Alfalfa Plants (Medicago sativa L.) Expressing the 85B (MAP1609c) Antigen of Mycobacterium avium subsp. paratuberculosis Elicit Long-Lasting Immunity in Mice.

Mycobacterium avium subsp. paratuberculosis (MAP) is the etiological agent of Paratuberculosis, a contagious, untreatable, and chronic granulomatous enteritis that results in diarrhea, emaciation, and death in farmed ruminants (i.e., cattle, sheep, and goats). In this study, the Ag85B antigen from MAP was expressed in transgenic alfalfa as an attractive vaccine candidate. Agrobacterium-mediated transformation allowed the rescue of 56 putative transformed plants and transgenesis was confirmed in 19 lines by detection of the Ag85B gene (MAP1609c) by PCR. Line number 20 showed the highest Ag85B expression [840 ng Ag85B per gram of dry weight leaf tissue, 0.062% Total Soluble Protein (TSP)]. Antigenicity of the plant-made Ag85B was evidenced by its reactivity with a panel of sera from naturally MAP-infected animals, whereas immunogenicity was assessed in mice immunized by either oral or subcutaneous routes. The plant-made Ag85B antigen elicited humoral responses by the oral route when co-administered with cholera toxin as adjuvant; significant levels of anti-85B antibodies were induced in serum (IgG) and feces (IgA). Long-lasting immunity was evidenced at day 180 days post-first oral immunization. The obtained alfalfa lines expressing Ag85B constitute the first model of a plant-based vaccine targeting MAP. The initial immunogenicity assessment conducted in this study opens the path for a detailed characterization of the properties of this vaccine candidate.

IFN-γ and IL-5 whole blood response directed against mycolactone polyketide synthase domains in patients with Mycobacterium ulcerans infection.

BACKGROUND: Buruli ulcer is a disease of the skin and soft tissues caused by infection with a slow growing pathogen, Mycobacterium ulcerans. A vaccine for this disease is not available but M. ulcerans possesses a giant plasmid pMUM001 that harbours the polyketide synthase (PKS) genes encoding a multi-enzyme complex needed for the production of its unique lipid toxin called mycolactone, which is central to the pathogenesis of Buruli ulcer. We have studied the immunogenicity of enzymatic domains in humans with M. ulcerans disease, their contacts, as well as non-endemic areas controls. METHODS: Between March 2013 and August 2015, heparinized whole blood was obtained from patients confirmed with Buruli ulcer. The blood samples were diluted 1 in 10 in Roswell Park Memorial Institute (RPMI) medium and incubated for 5 days with recombinant mycolactone PKS domains and mycolyltransferase antigen 85A (Ag85A). Blood samples were obtained before and at completion of antibiotic treatment for 8 weeks and again 8 weeks after completion of treatment. Supernatants were assayed for interferon-γ (IFN-γ) and interleukin-5 (IL-5) by enzyme-linked immunosorbent assay. Responses were compared with those of contacts and non-endemic controls. RESULTS: More than 80% of patients and contacts from endemic areas produced IFN-γ in response to all the antigens except acyl carrier protein type 3 (ACP3) to which only 47% of active Buruli ulcer cases and 71% of contacts responded. The highest proportion of responders in cases and contacts was to load module ketosynthase domain (Ksalt) (100%) and enoylreductase (100%). Lower IL-5 responses were induced in a smaller proportion of patients ranging from 54% after ketoreductase type B stimulation to only 21% with ketosynthase type C (KS C). Among endemic area contacts, the, highest proportion was 73% responding to KS C and the lowest was 40% responding to acyltransferase with acetate specificity type 2. Contacts of Buruli ulcer patients produced significantly higher IFN-γ and IL-5 responses compared with those of patients to PKS domain antigens and to mycolyltransferase Ag85A of M. ulcerans. There was low or no response to all the antigens in non-endemic areas controls. IFN-γ and IL-5 responses of patients improved after treatment when compared to baseline results. DISCUSSION: The major response to PKS antigen stimulation was IFN-γ and the strongest responses were observed in healthy contacts of patients living in areas endemic for Buruli ulcer. Patients elicited lower responses than healthy contacts, possibly due to the immunosuppressive effect of mycolactone, but the responses were enhanced after antibiotic treatment. A vaccine made up of the most immunogenic PKS domains combined with the mycolyltransferase Ag85A warrants further investigation.

Relationship between mold exposure, specific IgE sensitization, and clinical asthma: A case-control study.

BACKGROUND: Most of the findings related to the noxious effect of mold sensitization on asthma come from investigations based on Alternaria alternata, Cladosporium herbarum, and Aspergillus fumigatus. However, species such as Penicillium spp, Cladosporium sphaerospermum, Cladosporium cladosporioides, or Aspergillus versicolor display a more pronounced indoor tropism, and their potential harmful respiratory effects cannot be neglected. OBJECTIVE: The goal of this work was to relate mold sensitizations with asthma severity and with the level of indoor mold contamination among mold-sensitized patients with asthma and nonsensitized patients with asthma. METHODS: A case-control study was conducted and several asthma severity markers were compared between patients with asthma with and without mold sensitization. Indoor contamination of patients' dwellings was also investigated. RESULTS: Our findings confirmed the association between sensitization to A fumigatus and severity for patients with asthma in contrast with sensitization to other species. Indoor mold contamination was detected in approximately 90% of dwellings. Overall mold exposure was not associated with asthma severity. However, regardless of the sensitization, exposure to A fumigatus and Penicillium spp in dust was linked to an increased risk of severe asthma. CONCLUSION: The harmful nature of mold sensitization and mold exposure for patients with asthma was not confirmed in this study. However, sensitization to A fumigatus was associated with an increased risk for severe asthma. A better investigation of the properties of Penicillium spp is recommended because its exposure was found to be associated with a more pronounced impairment of lung function.

Evaluation of mycobacteria-specific gamma interferon and antibody responses before and after a single intradermal skin test in cattle naturally exposed to M. avium subsp. paratuberculosis and experimentally infected with M. bovis.

This study reports on the diagnostic potential of IFN-γ release assays and serology for Mycobacterium bovis in six naturally M. avium subsp. paratuberculosis (Map) exposed bulls of which four were intratracheally infected with a Belgian field strain of M. bovis. Heparinized blood, serum and fecal samples were collected at regular time intervals for mycobacteria-specific IFN-γ release assays, antibody analysis and for Map culture respectively. Single intradermal skin test (SIT) with bovine tuberculin (PPD-B) was performed on day 115 and animals were sacrificed on day 133 after M. bovis infection. Organs were collected and stored for histopathological examination, modified Ziehl-Neelsen staining and bacteriological analysis of M. bovis and Map by culture and RT-PCR. Prior to infection five animals showed positive IFN-γ responses to avian PPD (PPD-A) and four were positive in Map PCR (IS900) on faeces. Three M. bovis infected animals reacted as early as day 14 with sustained higher PPD-B than PPD-A specific IFN-γ responses, whereas the fourth animal (with the strongest PPD-A response prior to infection) showed sustained higher PPD-B specific IFN-γ levels only a day 56 after infection. Two of the infected animals had a sustained positive IFN-γ response to the ESAT-6/CFP-10/TB7.7 (QuantiFERON®-TB Gold) peptide cocktail as early as day 14, among which the animal with the initial high PPD-A response. Later during infection, positive responses were found to ESAT-6 peptides in three infected bulls and to CFP-10 peptides in all four infected bulls. One of the control animals reacted intermittently to the ESAT-6/CFP10/TB7.7 cocktail. Prior to SIT, weak but positive MPB83/MBP70 specific antibody responses were detected in two of the infected bulls. All four M. bovis infected bulls reacted with a positive skin test and showed, as reported by others, increased mycobacteria specific IFN-γ production and increased positive responses in MPB83/MBP70 specific serology after SIT. At autopsy, M. bovis lesions were detected in all four experimentally infected bulls. Our results indicate that in Map exposed cattle, M. bovis diagnosis using IFN-γ assays needs a combination of PPD-B/A and ESAT-6/CFP10 for early and optimal sensitivity and that sensitivity of MPB83/MBP70 serodiagnosis is dramatically increased by prior skin testing. Map exposure did not interfere with the development of SIT in M. bovis infected animals, but resulted in a false positive M. bovis specific IFN-γ and antibody response after SIT in one of the two control animals (which remained negative in skin-test).

Field performance of six Mycobacterium avium subsp. paratuberculosis antigens in a 20h interferon gamma release assay in Belgium.

Paratuberculosis, caused by Mycobacterium avium subsp. paratuberculosis (Map), is a chronic granulomatous enteritis which primarily affects domestic and wild ruminants, resulting in serious economic losses for dairy and beef industry around the world. There is no satisfactory cure or vaccine, and actual diagnostic tests need improvement, particularly for the initial stages of the disease. Map specific cell-mediated immune responses may allow early detection of the infection at subclinical stages. In this study, over a period of 39 months, we collected 548 blood samples in two culture-confirmed Map-infected herds, 95 blood samples in five dairy herds that scored negative during 3 consecutive years of Map serology testing and 79 samples in three culture-confirmed M. bovis infected herds. Based on criteria of bacteriology, serology and ratio of IFN-γ induced with bovine and avian purified protein derivative of tuberculin (PPD-B/PPD-A), we classified the samples in four groups: 415 samples as Map-exposed/infected (MAP), 58 samples as aspecific reactors (AR), 179 samples as non-responders (NI) and 70 samples as M. bovis infected (TB). Age of the animals influenced the IFN-γ response in the MAP group, with PPD specific IFN-γ levels (but not PPD-B/PPD-A IFN-γ ratio) being significantly higher in animals <18 months of age. Map specific antibodies were detected by IDEXX ELISA in 13/415 (3%) sera of the MAP group, whereas fecal culture was positive for only 7/405 (1.7%) samples. Animals in the MAP group could therefore be considered being at the very early stage of Map infection. Six purified, recombinant Map antigens (Ag5, Ag6, MAP1637c, MAP0388, MAP3547c and MAP0586c), previously identified using combined advanced proteomic or reverse genomic approaches, were tested for their diagnostic potential in a 20h IFN-γ release assay. In the age group >18 months old, Ag5 and MAP0388 were recognized by only 10.1% and 7.7% of the animals in the MAP group, whereas a total of 38.6.%, 29.4%, 25.6% and 39.0% of the animals in the MAP group reacted to Ag6, MAP1637c, MAP3547c and MAP0586c respectively. None of the animals in the TB group reacted to Ag6, MAP1637c or MAP586c. Except for MAP0388, the % of reactors in the MAP group was significantly higher in animals <18 months old: 28.0%, 24.0%, 45.5%, 47.1%, 49.8% and 47.4% respectively. Further studies of these candidates and their combination are needed to confirm their diagnostic potential for the detection of early Map infection.

Survival of Mycobacterium tuberculosis and Mycobacterium bovis BCG in lysosomes in vivo.

Mycobacterium tuberculosis is one of the most successful pathogens known, having infected more than a third of the global population. An important strategy for intracellular survival of pathogenic mycobacteria relies on their capacity to resist delivery to lysosomes, instead surviving within macrophage phagosomes. Several factors of both mycobacterial and host origin have been implicated in this process. However, whether or not this strategy is employed in vivo is not clear. Here we show that in vivo, following intravenous infection, M. tuberculosis and Mycobacterium bovis BCG initially survived by resisting lysosomal transfer. However, after prolonged infection the bacteria were transferred to lysosomes yet continued to proliferate. A M. bovis BCG mutant lacking protein kinase G (PknG), that cannot avoid lysosomal transfer and is readily cleared in vitro, was found to survive and proliferate in vivo. The ability to survive and proliferate in lysosomal organelles in vivo was found to be due to an altered host environment rather than changes in the inherent ability of the bacteria to arrest phagosome maturation. Thus, within an infected host, both M. tuberculosis and M. bovis BCG adapts to infection-specific host responses. These results are important to understand the pathology of tuberculosis and may have implications for the development of effective strategies to combat tuberculosis.

Influence of maternal vaccination against diphtheria, tetanus, and pertussis on the avidity of infant antibody responses to a pertussis containing vaccine in Belgium.

Maternal antibodies induced by vaccination during pregnancy cross the placental barrier and can close the susceptibility gap to pertussis in young infants up to the start of primary immunization. As not only the quantity but also the quality of circulating antibodies is important for protection, we assessed whether maternal immunization affects the avidity of infant vaccine-induced IgG antibodies, in the frame of a prospective clinical trial on pregnancy vaccination in Belgium. Infants born from Tdap (Boostrix®) vaccinated (N = 55) and unvaccinated (N = 26) mothers were immunized with a hexavalent pertussis containing vaccine (Infanrix Hexa®) at 8, 12 and 16 weeks, followed by a fourth dose at 15 months of age. Right before and one month after this fourth vaccine dose, the avidity of IgG antibodies against diphtheria toxin (DT), tetanus toxin (TT), pertussis toxin (PT), filamentous hemagglutinin (FHA) and pertactin (Prn) was determined using 1.5 M ammonium thiocyanate as dissociating agent. In both groups, antibody avidity was moderate for TT, PT, FHA and Prn and low for DT after priming. After a fourth dose, antibody avidity increased significantly to high avidity for TT and PT, whereas it remained moderate for FHA and Prn and low for DT. The avidity correlated positively with antibody level in both study groups, yet not significantly for PT. When comparing both study groups, only PT-specific antibodies showed significantly lower avidity in infants born from vaccinated than from unvaccinated mothers after the fourth vaccine dose. The clinical significance of lower avidity of vaccine induced infant antibodies after maternal vaccination, if any, needs further investigation.

Virulence and immunogenicity of genetically defined human and porcine isolates of M. avium subsp. hominissuis in an experimental mouse infection.

Mycobacterium avium subsp. hominissuis (Mah) represents a health concern for humans and to a lesser extent for pigs, but its zoonotic potential remains elusive. Using multispacer sequence typing (MST) we previously identified 49 different genotypes of Mah among Belgian clinical and porcine isolates, with 5 MSTs shared by both hosts. Using experimental intranasal infection of BALB/c mice, we compared the virulence and immunogenicity of porcine and clinical human isolates with shared genotype or with a genotype only found in humans or pigs. Bacterial replication was monitored for 20 weeks in lungs, spleen and liver and mycobacteria specific spleen cell IFN-γ, IL-10 and IL-17 production as well as serum antibody responses were analyzed. Isolates varied in virulence, with human and porcine isolates sharing MST22 genotype showing a thousand fold higher bacterial replication in lungs and more dissemination to spleen and liver than the human and porcine MST91 isolates. Virulent MST22 type was also associated with progressive suppression of IFN-γ and IL-17 responses, and increased IL-10 production. Whole genome sequencing of the two virulent isolates with MST22 genotype and two avirulent isolates of genotype MST91 and comparison with two well-studied M. avium subsp. hominissuis reference strains i.e. Mah 104 and Mah TH135, identified in the two MST22 isolates nine specific virulence factors of the mammalian cell entry family, that were identical with Mah 104 strain. Despite the obvious limitations of the mouse model, a striking link of virulence and identity at the genome level of porcine and human isolates with the same multisequence type, for which no correlation of place of residence (humans) or farm of origin (pigs) was observed, seems to point to the existence in the environment of certain genotypes of Mah which may be more infectious both for humans and pigs than other genotypes.

Development of a Dot-Blot Assay for the Detection of Mould-Specific IgE in the Belgian Population.

BACKGROUND: Data on mould sensitization in the general population are scarce and mostly on Aspergillus fumigatus, Alternaria alternata and Cladosporium herbarum. OBJECTIVES: To validate a dot-blot assay for the detection of specific IgE and evaluate the prevalence of mould sensitization in a healthy population. METHODS: The dot-blot assay was validated against the CAP test. Sensitization rate to ten common indoor and outdoor mould species in 344 serum samples was calculated. For each serum with more than one reactivity, the "major sensitization" defined as the strongest response against a single mould species was calculated. RESULTS: Intra- and inter-assay variations were both below 20%, and the positivity threshold of the test was of 0.418 kU/L for A. fumigatus. Correlation with CAP results was strong. The overall prevalence of sensitization was 32.8%, and the commonest sensitizations were against A. alternaria, A. flavus and A. niger (around 15%). The most frequent "major reactivities" were against A. niger and A. alternata (20-30%). In 25.1% of the samples, "major reactivities" were directed against a group of moulds commonly found indoor (Penicillium spp., Aspergillus versicolor, Cladosporium sphaerospermum and Cladosporium cladosporioides). CONCLUSIONS: The dot-blot assay was validated for the detection of mould-specific IgE. In the general population, sensitization to indoor species was common and accounted for 25% of overall mould sensitizations.

Inflammatory Properties and Adjuvant Potential of Synthetic Glycolipids Homologous to Mycolate Esters of the Cell Wall of Mycobacterium tuberculosis.

The cell wall of mycobacteria is characterised by glycolipids composed of different classes of mycolic acids (MAs; alpha-, keto-, and methoxy-) and sugars (trehalose, glucose, and arabinose). Studies using mutant Mtb strains have shown that the structure of MAs influences the inflammatory potential of these glycolipids. As mutant Mtb strains possess a complex mixture of glycolipids, we analysed the inflammatory potential of single classes of mycolate esters of the Mtb cell wall using 38 different synthetic analogues. Our results show that synthetic trehalose dimycolate (TDM) and trehalose, glucose, and arabinose monomycolates (TMM, GMM, and AraMM) activate bone marrow-derived dendritic cells in terms of the production of pro-inflammatory cytokines (IL-6 and TNF-α) and reactive oxygen species, upregulation of costimulatory molecules, and activation of NLRP3 inflammasome by a mechanism dependent on Mincle. These findings demonstrate that Mincle receptor can also recognise pentose esters and seem to contradict the hypothesis that production of GMM is an escape mechanism used by pathogenic mycobacteria to avoid recognition by the innate immune system. Finally, our experiments indicate that TMM and GMM, as well as TDM, can promote Th1 and Th17 responses in mice in an OVA immunisation model, and that further analysis of their potential as novel adjuvants for subunit vaccines is warranted.

The Effect of Maternal Pertussis Immunization on Infant Vaccine Responses to a Booster Pertussis-Containing Vaccine in Vietnam.

BACKGROUND: Maternal vaccination with an acellular pertussis (aP)-containing vaccine is a recommended strategy in a growing number of industrialized countries, to protect young infants from disease. Little is known on the effect of this strategy in low- and middle-income countries. Following a previous report on the effect of adding a pertussis and diphtheria component to the tetanus vaccination program in pregnant women in Vietnam, we report on infant immune responses to a booster aP vaccine dose in this randomized controlled clinical trial. METHODS: Thirty infants of Tdap (tetanus, diphtheria, and acellular pertussis)-vaccinated pregnant women and 37 infants of women vaccinated with a tetanus-only vaccine received a fourth aP-containing vaccine dose in the second year of life. Blood was taken 1 month after the fourth infant dose. Immunoglobulin G (IgG) antibodies against pertussis toxin (PT), filamentous hemagglutinin (FHA), pertactin (Prn), tetanus toxoid (TT), and diphtheria toxoid (DT) were measured using commercially available enzyme-linked immunosorbent assays (ELISA). RESULTS: One month after the booster dose, significantly lower antibody titers were measured in the Tdap group for anti-TT IgG (P < .001) only. Anti-DT IgG, anti-PT IgG, anti-Prn IgG, and anti-FHA IgG antibody titers were comparable for both groups. A rise in antibody concentrations was elicited for all (except DT) antigens after boosting. CONCLUSIONS: The present results indicate that the blunting of infant pertussis responses induced by maternal immunization, measured after a primary series of aP vaccines, was resolved with the booster aP vaccine dose. These results add to the evidence for national and international decision makers on maternal immunization as a vaccination strategy for protection of young infants against infectious diseases.

Correction: Capture-Recapture Estimators in Epidemiology with Applications to Pertussis and Pneumococcal Invasive Disease Surveillance.

[This corrects the article DOI: 10.1371/journal.pone.0159832.].

Capture-Recapture Estimators in Epidemiology with Applications to Pertussis and Pneumococcal Invasive Disease Surveillance.

INTRODUCTION: Surveillance networks are often not exhaustive nor completely complementary. In such situations, capture-recapture methods can be used for incidence estimation. The choice of estimator and their robustness with respect to the homogeneity and independence assumptions are however not well documented. METHODS: We investigated the performance of five different capture-recapture estimators in a simulation study. Eight different scenarios were used to detect and combine case-information. The scenarios increasingly violated assumptions of independence of samples and homogeneity of detection probabilities. Belgian datasets on invasive pneumococcal disease (IPD) and pertussis provided motivating examples. RESULTS: No estimator was unbiased in all scenarios. Performance of the parametric estimators depended on how much of the dependency and heterogeneity were correctly modelled. Model building was limited by parameter estimability, availability of additional information (e.g. covariates) and the possibilities inherent to the method. In the most complex scenario, methods that allowed for detection probabilities conditional on previous detections estimated the total population size within a 20-30% error-range. Parametric estimators remained stable if individual data sources lost up to 50% of their data. The investigated non-parametric methods were more susceptible to data loss and their performance was linked to the dependence between samples; overestimating in scenarios with little dependence, underestimating in others. Issues with parameter estimability made it impossible to model all suggested relations between samples for the IPD and pertussis datasets. For IPD, the estimates for the Belgian incidence for cases aged 50 years and older ranged from 44 to58/100,000 in 2010. The estimates for pertussis (all ages, Belgium, 2014) ranged from 24.2 to30.8/100,000. CONCLUSION: We encourage the use of capture-recapture methods, but epidemiologists should preferably include datasets for which the underlying dependency structure is not too complex, a priori investigate this structure, compensate for it within the model and interpret the results with the remaining unmodelled heterogeneity in mind.

Mycobacterium tuberculosis-associated synthetic mycolates differentially exert immune stimulatory adjuvant activity.

Mycolic acids (MAs) are highly hydrophobic long-chain α-alkyl ß-hydroxy fatty acids present in the cell wall of Mycobacterium tuberculosis (Mtb) as a complex mixture of molecules with a common general structure but with variable functional groups in the meromycolate chain. In this study, we addressed the relationship between the MA molecular structure and their contribution to the development of T-cell immune responses. Hereto, we used the model antigen ovalbumin and single synthetic MAs, differing in oxygenation class and cis versus trans proximal cyclopropane configuration, as immune stimulatory agents. Subcutaneous delivery of liposome-formulated MAs with a proximal cis cyclopropane elicited antigen-specific Th1 and cytotoxic T-cell immune responses, whereas intratracheal immunization elicited pulmonary Th17 responses. These immune stimulatory activities depended not only on the cis versus trans proximal cyclopropane configuration but also on the MA oxygenation class. Our study thus shows that both the presence and nature of the functional groups in the meromycolate chain affect the immune stimulatory adjuvant activity of Mtb mycolates and suggests that Mtb bacilli may impact on the host protective immune response by modulating the cis versus trans stereochemistry of its mycolates as well as by altering the oxygenation class of the meromycolate functional group.

A Belgian Serosurveillance/Seroprevalence Study of Diphtheria, Tetanus and Pertussis Using a Luminex xMAP Technology-Based Pentaplex.

Serosurveillance and seroprevalence studies are an essential tool to monitor vaccine-preventable diseases. We have developed a magnetic bead-based pentaplex immunoassay (MIA) for the simultaneous detection of IgG antibodies against diphtheria toxin (DT), tetanus toxin (TT), pertussis toxin (PT), filamentous hemagglutinin (FHA) and pertactin (Prn). The in-house pentaplex MIA showed a good correlation with commercial ELISAs with correlation coefficients between 0.89 for PT and 0.98 for TT. Intra- and inter-assay variability was <10%. A total of 670 anonymized serum samples collected in 2012 in Belgian adults (ages 20-29.9 years) were analyzed. Geometric mean concentrations (GMC) were 0.2 (0.13-0.29) IU/mL for DT, 0.63 (0.45-0.82) IU/mL for TT, 3.9 (2.6-5.8) IU/mL for PT, 16.3 (11.7-22.7) IU/mL for FHA and 15.4 (10.1-23.6) IU/mL for Prn. Antibody concentrations were below the protective level of 0.1 IU/mL in 26.4% of the sera for DT and in 8.6% of the sera for TT. Anti-PT IgG concentrations indicative of recent pertussis infection (>125 IU/mL) were detected in 1.2% of the subjects. High anti-PT antibodies were not correlated with high antibodies against any of the four other vaccine antigens. This pentaplex MIA will be used for a new large-scale Belgian serosurveillance/seroprevalence study of diphtheria, tetanus and pertussis.

Innate signaling by mycobacterial cell wall components and relevance for development of adjuvants for subunit vaccines.

INTRODUCTION: Pathogen recognition receptors (PRRs) recognize pathogen-associated molecular patterns, triggering the induction of inflammatory innate responses and contributing to the development of specific adaptive immune responses. Novel adjuvants have been developed based on agonists of PRRs. Areas covered: Lipid pathogen-associated molecular patterns (PAMPs) present in the cell wall of mycobacteria are revised, with emphasis on agonists of C-type lectin receptors, signaling pathways, and preclinical data supporting their use as novel adjuvants inducing cell-mediated immune responses. Their potential use as lipid antigens in novel tuberculosis subunit vaccines is also discussed. Expert commentary: Few adjuvants are licensed for human use and mainly favour antibody-mediated protective immunity. Use of lipid PAMPs that trigger cell-mediated immune responses could lead to the development of adjuvants for vaccines against intracellular pathogens and cancer.

Pertussis vaccination during pregnancy in Belgium: Follow-up of infants until 1 month after the fourth infant pertussis vaccination at 15 months of age.

Vaccination of pregnant women with a pertussis containing vaccine is a recommended strategy in some industrialized countries, to protect young infants from severe disease. One of the effects of the presence of high titers of passively acquired maternal antibodies in young infants is blunting of immune responses to infant vaccination. We present infant immune responses to a fourth pertussis containing vaccine dose at 15 months of age, as a follow-up of previously presented data. In a prospective cohort study, women were either vaccinated with an acellular pertussis vaccine (Boostrix(®)) during pregnancy (vaccine group) or received no vaccine (control group). All infants were vaccinated with Infanrix Hexa(®) according to the standard Belgian vaccination schedule (8/12/16 weeks, 15 months). We report results from blood samples collected before and 1 month after the fourth vaccine dose. Immunoglobulin G (IgG) antibodies against pertussis toxin (PT), filamentous hemagglutinin (FHA), pertactin (Prn), tetanus toxoid (TT) and diphtheria toxoid (DT) were measured using commercially available ELISA tests. Antibody levels were expressed in International Units per milliliter. Demographic characteristics were similar in the vaccine and control group. Before the fourth vaccine dose, significantly lower antibody titers were measured in the vaccine group compared to the control group for anti-Prn IgG (p=0.003) and anti-DT IgG (p=0.023), with a steep decay of antibody titers since post-primary vaccination. One month after the fourth dose, antibody titers were only significantly lower in the vaccine group for anti-PT IgG (p=0.006). For all antigens, there was a rise in antibody titer after the fourth vaccine dose. The present results indicate still a minor blunting effect 1 month after a fourth vaccine dose for anti-PT antibodies. However, a good humoral immune response on all measured antigens was elicited in both groups of children. The clinical significance of such blunting effect is yet unknown. Clinicaltrials.gov identifier: NCT01698346.