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PubMed | Imperial College London, Laboratory of Pediatric Infectious Diseases and Radboud University Nijmegen
Type: Journal Article | Journal: The Journal of infectious diseases | Year: 2016

Streptococcus pneumoniae is a major cause of life-threatening infections. Complement activation plays a vital role in opsonophagocytic killing of pneumococci in blood. Initial complement activation via the classical and lectin pathways is amplified through the alternative pathway amplification loop. Alternative pathway activity is inhibited by complement factor H (FH). Our study demonstrates the functional consequences of the variability in human serum FH levels on host defense. Using an in vivo mouse model combined with human in vitro assays, we show that the level of serum FH correlates with the efficacy of opsonophagocytic killing of pneumococci. In summary, we found that FH levels determine a delicate balance of alternative pathway activity, thus affecting the resistance to invasive pneumococcal disease. Our results suggest that variation in FH expression levels, naturally occurring in the human population, plays a thus far unrecognized role in the resistance to invasive pneumococcal disease.


PubMed | University of Groningen, Laboratory of Pediatric Infectious Diseases, Laboratory of Pediatric Oncology and Radboud University Nijmegen
Type: Journal Article | Journal: The Journal of infectious diseases | Year: 2016

For many bacterial respiratory infections, development of (severe) disease is preceded by asymptomatic colonization of the upper airways. For Streptococcus pneumoniae, the transition to severe lower respiratory tract infection is associated with an increase in nasopharyngeal colonization density. Insight into how the mucosal immune system restricts colonization may provide new strategies to prevent clinical symptoms. Several studies have provided indirect evidence that the mucosal adjuvant cholera toxin subunit B (CTB) may confer nonspecific protection against respiratory infections. Here, we show that CTB reduces the pneumococcal load in the nasopharynx, which required activation of the caspase-1/11 inflammasome, mucosal T cells, and macrophages. Our findings suggest that CTB-dependent activation of the local innate response synergizes with noncognate T cells to restrict bacterial load. Our study not only provides insight into the immunological components required for containment and clearance of pneumococcal carriage, but also highlights an important yet often understudied aspect of adjuvants.


PubMed | NIZO food research B.V., Emory University, The Netherlands ; NIZO food research B.V., Laboratory of Pediatric Infectious Diseases and 2 more.
Type: | Journal: Microbiome | Year: 2015

Several cohort studies have indicated associations between S. pneumoniae and other microbes in the nasopharynx. To study causal relationships between the nasopharyngeal microbiome and pneumococcal carriage, we employed an experimental human pneumococcal carriage model. Healthy adult volunteers were assessed for pneumococcal carriage by culture of nasal wash samples (NWS). Those without natural pneumococcal carriage received an intranasal pneumococcal inoculation with serotype 6B or 23F. The composition of the nasopharyngeal microbiome was longitudinally studied by 16S rDNA pyrosequencing on NWS collected before and after challenge.Among 40 selected volunteers, 10 were natural carriers and 30 were experimentally challenged. At baseline, five distinct nasopharyngeal microbiome profiles were identified. The phylogenetic distance between microbiomes of natural pneumococcal carriers was particularly large compared to non-carriers. A more diverse microbiome prior to inoculation was associated with the establishment of pneumococcal carriage. Perturbation of microbiome diversity upon pneumococcal challenge was strain specific. Shifts in microbiome profile occurred after pneumococcal exposure, and those volunteers who acquired carriage more often diverted from their original profile. S. pneumoniae was little prominent in the microbiome of pneumococcal carriers.Pneumococcal acquisition in healthy adults is more likely to occur in a diverse microbiome and appears to promote microbial heterogeneity.


Zoll J.,Radboudumc | Rahamat-Langendoen J.,Radboudumc | Ahout I.,Laboratory of Pediatric Infectious Diseases | Jonge M.I.,Laboratory of Pediatric Infectious Diseases | And 5 more authors.
Journal of Clinical Virology | Year: 2015

Background: Acute respiratory tract infections (RTI) cause substantial morbidity during childhood, and are responsible for the majority of pediatric infectious diseases. Although most acute RTI are thought to be of viral origin, viral etiology is still unknown in a significant number of cases. Objectives: Multiplexed whole genome sequencing (WGS) was used for virome determination directly on clinical samples as proof of principle for the use of deep sequencing techniques in clinical diagnosis of viral infections. Study design: WGS was performed with nucleic acids from sputum and nasopharyngeal aspirates from four pediatric patients with known respiratory tract infections (two patients with human rhinovirus, one patient with human metapneumovirus and one patient with respiratory syncytial virus), and from four pediatric patients with PCR-negative RTI, and two control samples. Results: Viral infections detected by routine molecular diagnostic methods were confirmed by WGS; in addition, typing information of the different viruses was generated. In three out of four samples from pediatric patients with PCR-negative respiratory tract infections and the two control samples, no causative viral pathogens could be detected. In one sample from a patient with PCR-negative RTI, rhinovirus type-C was detected. Almost complete viral genomes could be assembled and in all cases virus species could be determined. Conclusions: Our study shows that, in a single run, viral pathogens can be detected and characterized, providing information for clinical assessment and epidemiological studies. We conclude that WGS is a powerful tool in clinical virology that delivers comprehensive information on the viral content of clinical samples. © 2015 Elsevier B.V.


Cremers A.J.H.,Laboratory of Pediatric Infectious Diseases | Hagen F.,Canisius Wilhelmina Hospital | Hermans P.W.M.,Laboratory of Pediatric Infectious Diseases | Hermans P.W.M.,Crucell | And 2 more authors.
European Journal of Clinical Microbiology and Infectious Diseases | Year: 2014

Detection of pneumococcal DNA in blood could be a fast alternative for blood culture in invasive pneumococcal disease (IPD). In this study we compared the diagnostic value of the serum pneumococcal DNA load between different clinical syndromes in adults with bacteremic pneumococcal infections, also after initiation of antibiotic treatment. Adults hospitalized with a blood culture proven pneumococcal infection between December 2008 and June 2013 were retrospectively included. Pneumococcal DNA loads in corresponding serum samples were determined by qPCR. Data on clinical diagnosis, course of disease and antibiotic treatment were extracted from medical records. For 53 IPD cases eligible stored serum samples were retrieved. The proportion of samples positive in qPCR was lower in uncomplicated pneumonia compared with other clinical syndromes (59.5 % vs. 100 %, p=0.005). The pneumococcal DNA load was higher in cases other than uncomplicated pneumonia (p=0.043) as well as in more severe disease (p-values 0.018, 0.029 and 0.003 for PSI Risk Class IV/V, ICU admission and mortality, respectively). Both detection of pneumococcal DNA and distribution of load did not significantly change over the first days of hospitalization despite treatment with appropriate antibiotics. Detection of pneumococcal DNA in serum was more sensitive in clinical syndromes other than uncomplicated pneumonia. Furthermore, the pneumococcal DNA load was associated with the type of IPD and severity of disease. Since the serum pneumococcal DNA load seemed unaffected by antibiotic treatment during the first days of IPD, it may offer an alternative for culture methods after prior antibiotic use. © 2014 Springer-Verlag.


PubMed | Laboratory of Pediatric Infectious Diseases, Radboudumc and Center for Molecular and Biomolecular Informatics
Type: | Journal: Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology | Year: 2015

Acute respiratory tract infections (RTI) cause substantial morbidity during childhood, and are responsible for the majority of pediatric infectious diseases. Although most acute RTI are thought to be of viral origin, viral etiology is still unknown in a significant number of cases.Multiplexed whole genome sequencing (WGS) was used for virome determination directly on clinical samples as proof of principle for the use of deep sequencing techniques in clinical diagnosis of viral infections.WGS was performed with nucleic acids from sputum and nasopharyngeal aspirates from four pediatric patients with known respiratory tract infections (two patients with human rhinovirus, one patient with human metapneumovirus and one patient with respiratory syncytial virus), and from four pediatric patients with PCR-negative RTI, and two control samples.Viral infections detected by routine molecular diagnostic methods were confirmed by WGS; in addition, typing information of the different viruses was generated. In three out of four samples from pediatric patients with PCR-negative respiratory tract infections and the two control samples, no causative viral pathogens could be detected. In one sample from a patient with PCR-negative RTI, rhinovirus type-C was detected. Almost complete viral genomes could be assembled and in all cases virus species could be determined.Our study shows that, in a single run, viral pathogens can be detected and characterized, providing information for clinical assessment and epidemiological studies. We conclude that WGS is a powerful tool in clinical virology that delivers comprehensive information on the viral content of clinical samples.


Shak J.R.,Emory University | Cremers A.J.H.,Laboratory of Pediatric Infectious Diseases | Gritzfeld J.F.,Respiratory Infection Group | De Jonge M.I.,Laboratory of Pediatric Infectious Diseases | And 4 more authors.
PLoS ONE | Year: 2014

Colonization of the nasopharynx by Streptococcus pneumoniae is a necessary precursor to pneumococcal diseases that result in morbidity and mortality worldwide. The nasopharynx is also host to other bacterial species, including the common pathogens Staphylococcus aureus, Haemophilus influenzae, and Moraxella catarrhalis. To better understand how these bacteria change in relation to pneumococcal colonization, we used species-specific quantitative PCR to examine bacterial densities in 52 subjects 7 days before, and 2, 7, and 14 days after controlled inoculation of healthy human adults with S. pneumoniae serotype 6B. Overall, 33 (63%) of subjects carried S. pneumoniae post-inoculation. The baseline presence and density of S. aureus, H. influenzae, and M. catarrhalis were not statistically associated with likelihood of successful pneumococcal colonization at this study's sample size, although a lower rate of pneumococcal colonization in the presence of S. aureus (7/14) was seen compared to that in the presence of H. influenzae (12/16). Among subjects colonized with pneumococci, the number also carrying either H. influenzae or S. aureus fell during the study and at 14 days post-inoculation, the proportion carrying S. aureus was significantly lower among those who were colonized with S. pneumoniae (p = 0.008) compared to non-colonized subjects. These data on bacterial associations are the first to be reported surrounding experimental human pneumococcal colonization and show that co-colonizing effects are likely subtle rather than absolute. © 2014 Shak et al.


Cremers A.J.H.,Laboratory of Pediatric Infectious Diseases | Cremers A.J.H.,Laboratory of Medical Immunology | Kokmeijer I.,Laboratory of Pediatric Infectious Diseases | Kokmeijer I.,Laboratory of Medical Immunology | And 6 more authors.
International Journal of Medical Microbiology | Year: 2014

Introduction: The clinical severity and course of invasive pneumococcal disease (IPD) differs substantially between patients. Streptococcus pneumoniae harbors large genetic variability. Zinc metalloproteinase C (ZmpC), a secreted pneumococcal protein involved in neutrophil extravasation, inflammation and tissue remodeling, is present in a minority of IPD isolates. We investigated whether the presence of zmpC was associated with the clinical manifestation of IPD. Material and methods: IPD patients admitted to two Dutch hospitals between 2000 and 2013 were included in the study. Detailed clinical data were collected and the serotype and presence of zmpC were determined in the corresponding blood culture isolates. Results: ZmpC was present in 21% of the 542 included IPD cases and was mainly associated with serotypes 8, 4, 33A/F and 11A/D. Infection with S. pneumoniae positive for zmpC was more frequently observed in females (p= 0.048) and patients with a history of smoking (p= 0.033). Although no relation to clinical syndrome was observed, zmpC positive cases more often presented with cough, dyspnea and sepsis (p-values 0.026, 0.001 and 0.018), and more frequently required ICU admission (p= 0.011) compared to zmpC negative cases. Conclusion: The presence of zmpC was associated with a more severe clinical manifestation of IPD. This study demonstrates that information on pneumococcal genetic background may be useful to identify vulnerable individuals, to monitor clinical presentation and to predict the course of IPD. © 2014 Elsevier GmbH.


Ahout I.,Laboratory of Pediatric Infectious Diseases | Ferwerda G.,Laboratory of Pediatric Infectious Diseases | de Groot R.,Laboratory of Pediatric Infectious Diseases
Journal of Infection | Year: 2013

Seasonal influenza infections cause a high burden of disease for the whole community every year. Effective vaccines are available and used worldwide in adults and children. Discussion is ongoing as to whether influenza vaccination for children should be implemented in the National Immunization Program (NIP). Is there enough evidence to support routine influenza vaccination in children? In this review we briefly discuss the influenza viruses and the available vaccines. Subsequently, the current data available on influenza vaccination is reviewed and weighed against the Dutch criteria for the introduction of new vaccines into the NIP. © 2013.


PubMed | Respiratory Infection Group, Laboratory of Pediatric Infectious Diseases and Emory University
Type: Journal Article | Journal: PloS one | Year: 2014

Colonization of the nasopharynx by Streptococcus pneumoniae is a necessary precursor to pneumococcal diseases that result in morbidity and mortality worldwide. The nasopharynx is also host to other bacterial species, including the common pathogens Staphylococcus aureus, Haemophilus influenzae, and Moraxella catarrhalis. To better understand how these bacteria change in relation to pneumococcal colonization, we used species-specific quantitative PCR to examine bacterial densities in 52 subjects 7 days before, and 2, 7, and 14 days after controlled inoculation of healthy human adults with S. pneumoniae serotype 6B. Overall, 33 (63%) of subjects carried S. pneumoniae post-inoculation. The baseline presence and density of S. aureus, H. influenzae, and M. catarrhalis were not statistically associated with likelihood of successful pneumococcal colonization at this studys sample size, although a lower rate of pneumococcal colonization in the presence of S. aureus (7/14) was seen compared to that in the presence of H. influenzae (12/16). Among subjects colonized with pneumococci, the number also carrying either H. influenzae or S. aureus fell during the study and at 14 days post-inoculation, the proportion carrying S. aureus was significantly lower among those who were colonized with S. pneumoniae (p = 0.008) compared to non-colonized subjects. These data on bacterial associations are the first to be reported surrounding experimental human pneumococcal colonization and show that co-colonizing effects are likely subtle rather than absolute.

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