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1.
Ann Intern Med ; 174(5): 585-594, 2021 05.
Article in English | MEDLINE | ID: mdl-33587687

ABSTRACT

BACKGROUND: Zika virus (ZIKV) may cause severe congenital disease after maternal-fetal transmission. No vaccine is currently available. OBJECTIVE: To assess the safety and immunogenicity of Ad26.ZIKV.001, a prophylactic ZIKV vaccine candidate. DESIGN: Phase 1 randomized, double-blind, placebo-controlled clinical study. (ClinicalTrials.gov: NCT03356561). SETTING: United States. PARTICIPANTS: 100 healthy adult volunteers. INTERVENTION: Ad26.ZIKV.001, an adenovirus serotype 26 vector encoding ZIKV M-Env, administered in 1- or 2-dose regimens of 5 × 1010 or 1 × 1011 viral particles (vp), or placebo. MEASUREMENTS: Local and systemic adverse events; neutralization titers by microneutralization assay (MN50) and T-cell responses by interferon-γ enzyme-linked immunospot and intracellular cytokine staining; and protectivity of vaccine-induced antibodies in a subset of participants through transfer in an exploratory mouse ZIKV challenge model. RESULTS: All regimens were well tolerated, with no safety concerns identified. In both 2-dose regimens, ZIKV neutralizing titers peaked 14 days after the second vaccination, with geometric mean MN50 titers (GMTs) of 1065.6 (95% CI, 494.9 to 2294.5) for 5 × 1010 vp and 956.6 (595.8 to 1535.8) for 1 × 1011 vp. Titers persisted for at least 1 year at a GMT of 68.7 (CI, 26.4-178.9) for 5 × 1010 vp and 87.0 (CI, 29.3 to 258.6) for 1 × 1011 vp. A 1-dose regimen of 1 × 1011 vp Ad26.ZIKV.001 induced seroconversion in all participants 56 days after the first vaccination (GMT, 103.4 [CI, 52.7 to 202.9]), with titers persisting for at least 1 year (GMT, 90.2 [CI, 38.4 to 212.2]). Env-specific cellular responses were induced. Protection against ZIKV challenge was observed after antibody transfer from participants into mice, and MN50 titers correlated with protection in this model. LIMITATION: The study was conducted in a nonendemic area, so it did not assess safety and immunogenicity in a flavivirus-exposed population. CONCLUSION: The safety and immunogenicity profile makes Ad26.ZIKV.001 a promising candidate for further development if the need reemerges. PRIMARY FUNDING SOURCE: Janssen Vaccines and Infectious Diseases.


Subject(s)
Viral Vaccines/immunology , Zika Virus Infection/prevention & control , Adenoviridae/immunology , Adult , Animals , Double-Blind Method , Female , Humans , Male , Mice , United States , Zika Virus/immunology , Zika Virus Infection/immunology
2.
PLoS One ; 12(3): e0170884, 2017.
Article in English | MEDLINE | ID: mdl-28288168

ABSTRACT

The respiratory pathogen Streptococcus pneumoniae is a major cause of diseases such as otitis media, pneumonia, sepsis and meningitis. The first step towards infection is colonization of the nasopharynx. Recently, it was shown that agglutinating antibodies play an important role in the prevention of mucosal colonization with S. pneumoniae. Here, we present a novel method to quantify antibody-dependent pneumococcal agglutination in a high-throughput manner using flow cytometry. We found that the concentration of agglutinating antibodies against pneumococcal capsule are directly correlated with changes in the size and complexity of bacterial aggregates, as measured by flow cytometry and confirmed by light microscopy. Using the increase in size, we determined the agglutination index. The cutoff value was set by measuring a series of non-agglutinating antibodies. With this method, we show that not only anti-polysaccharide capsule antibodies are able to induce agglutination but that also anti-PspA protein antibodies have agglutinating capabilities. In conclusion, we have described and validated a novel method to quantify pneumococcal agglutination, which can be used to screen sera from murine or human vaccination studies, in a high-throughput manner.


Subject(s)
Antibodies, Bacterial/immunology , Flow Cytometry/methods , Streptococcus pneumoniae/physiology , Animals , Bacterial Proteins/immunology , Heat-Shock Proteins/immunology , Mice , Streptococcus pneumoniae/immunology
3.
Vaccine ; 34(48): 5968-5974, 2016 11 21.
Article in English | MEDLINE | ID: mdl-27771185

ABSTRACT

Widespread vaccination against Streptococcus pneumoniae (the pneumococcus) has significantly reduced pneumococcal disease caused by vaccine serotypes. Despite vaccination, overall pneumococcal colonization rates in children have not reduced and otitis media (OM) by non-vaccine serotypes remains one of the most common childhood infections. Pneumococcal surface protein A (PspA) has been shown to be a promising protein antigen to induce broad protection against pneumococcal colonization. However, its ability to protect against OM remains unclear. Using our previously established mouse model of influenza-virus induced pneumococcal OM, we here show that intranasal vaccination of mice with PspA together with the mucosal adjuvant CTB results in a decrease in pneumococcal load in the middle ears. This decrease correlated with the induction of PspA-specific IgA, a balanced IgG1:IgG2a antibody response and the induction of a mucosal Th17 response. Our data suggests that the IL-17 response to PspA is more important for protection against OM, whilst the presence of antibodies may be less important, as determined in mice deficient in IL-17 signaling or antibody production. Together, these results suggest that mucosal vaccination with PspA may not only protect against colonization, but also against the development of virus-induced pneumococcal OM.


Subject(s)
Antibodies, Bacterial/immunology , Interleukin-17/immunology , Otitis Media/immunology , Otitis Media/prevention & control , Pneumococcal Infections/immunology , Pneumococcal Vaccines/immunology , Adjuvants, Immunologic/administration & dosage , Administration, Intranasal , Animals , Antibodies, Bacterial/blood , Bacterial Load , Bacterial Proteins/immunology , Disease Models, Animal , Ear, Middle/microbiology , Immunity, Mucosal , Immunoglobulin A/immunology , Interleukin-17/deficiency , Mice, Inbred BALB C , Otitis Media/virology , Pneumococcal Infections/microbiology , Pneumococcal Infections/prevention & control , Pneumococcal Vaccines/administration & dosage , Streptococcus pneumoniae/immunology , Streptococcus pneumoniae/isolation & purification , Th17 Cells/immunology , Vaccination/methods
4.
J Med Microbiol ; 65(2): 129-136, 2016 Feb.
Article in English | MEDLINE | ID: mdl-26628261

ABSTRACT

Streptococcus pneumoniae is responsible for an estimated 1.6 million deaths worldwide every year. While rapid detection and timely treatment with appropriate antibiotics is preferred, this is often difficult due to the amount of time that detection with blood cultures takes. In this study, a novel quantitative PCR assay for the detection of Streptococcus pneumoniae was developed. To identify novel targets, we analysed the pneumococcal genome for unique, repetitive DNA sequences. This approach identified comX, which is conserved and present in duplicate copies in Streptococcus pneumoniae but not in other bacterial species. Comparison with lytA, the current 'gold standard' for detection by quantitative PCR, demonstrated an analytic specificity of 100% for both assays on a panel of 10 pneumococcal and 18 non-pneumococcal isolates, but a reduction of 3.5 quantitation cycle values (± 0.23 sem), resulting in an increased analytical detection rate of comX. We validated our assay on DNA extracted from the serum of 30 bacteraemic patients who were blood culture positive for Streptococcus pneumoniae and 51 serum samples that were culture positive for other bacteria. This resulted in a similar clinical sensitivity between the comX and lytA assays (47%) and in a diagnostic specificity of 98.2 and 100% for the lytA and comX assays, respectively. In conclusion, we have developed a novel quantitative PCR assay with increased analytical sensitivity for the detection of Streptococcus pneumoniae, which may be used to develop a rapid bedside test for the direct detection of Streptococcus pneumoniae in clinical specimens.


Subject(s)
Bacteremia/microbiology , Bacterial Proteins/genetics , Pneumococcal Infections/microbiology , Real-Time Polymerase Chain Reaction/methods , Streptococcus pneumoniae/isolation & purification , Transcription Factors/genetics , Bacteremia/diagnosis , Female , Humans , Male , Pneumococcal Infections/diagnosis , Sensitivity and Specificity , Streptococcus pneumoniae/classification , Streptococcus pneumoniae/genetics
5.
Virol J ; 10: 128, 2013 Apr 23.
Article in English | MEDLINE | ID: mdl-23617940

ABSTRACT

BACKGROUND: Otitis media (OM) affects ≥80% of children before the age of three. OM can arise following co-infection with influenza A virus (IAV) and the bacterium Streptococcus pneumoniae. We have previously shown that H3 IAV strains (such as Udorn/72) induced a higher rate of bacterial OM than H1 strains (such as PR8/34). This was associated with more efficient replication of H3 strains in the middle ear. FINDINGS: Here, we assess if the increased replication of IAV strains such as Udorn/72 in the middle ear is dependent upon the binding of the viral HA to α2,6-linked sialic acid. Using murine and in vitro models, the present study shows that recognition of α2,6-linked sialic acid was not required to facilitate bacterial OM. CONCLUSIONS: Taken together, these data suggest that other features of the HA mediate bacterial OM.


Subject(s)
Influenza A virus/physiology , N-Acetylneuraminic Acid/metabolism , Orthomyxoviridae Infections/complications , Otitis Media/pathology , Pneumococcal Infections/pathology , Viral Tropism , Virus Attachment , Animals , Disease Models, Animal , Mice , Mice, Inbred C57BL , Orthomyxoviridae Infections/virology , Otitis Media/microbiology , Pneumococcal Infections/microbiology , Streptococcus pneumoniae/pathogenicity , Virus Replication
6.
Infect Immun ; 81(3): 645-52, 2013 Mar.
Article in English | MEDLINE | ID: mdl-23319557

ABSTRACT

Influenza A virus (IAV) predisposes individuals to secondary infections with the bacterium Streptococcus pneumoniae (the pneumococcus). Infections may manifest as pneumonia, sepsis, meningitis, or otitis media (OM). It remains controversial as to whether secondary pneumococcal disease is due to the induction of an aberrant immune response or IAV-induced immunosuppression. Moreover, as the majority of studies have been performed in the context of pneumococcal pneumonia, it remains unclear how far these findings can be extrapolated to other pneumococcal disease phenotypes such as OM. Here, we used an infant mouse model, human middle ear epithelial cells, and a series of reverse-engineered influenza viruses to investigate how IAV promotes bacterial OM. Our data suggest that the influenza virus HA facilitates disease by inducing a proinflammatory response in the middle ear cavity in a replication-dependent manner. Importantly, our findings suggest that it is the inflammatory response to IAV infection that mediates pneumococcal replication. This study thus provides the first evidence that inflammation drives pneumococcal replication in the middle ear cavity, which may have important implications for the treatment of pneumococcal OM.


Subject(s)
Inflammation/pathology , Orthomyxoviridae Infections/complications , Otitis Media/pathology , Pneumococcal Infections/pathology , Animals , Influenza A virus/classification , Interleukin-6/genetics , Interleukin-6/metabolism , Interleukin-8/genetics , Interleukin-8/metabolism , Mice , Mice, Inbred C57BL , Orthomyxoviridae Infections/immunology , Orthomyxoviridae Infections/microbiology , Orthomyxoviridae Infections/virology , Otitis Media/immunology , Otitis Media/microbiology , Pneumococcal Infections/immunology , Pneumococcal Infections/microbiology , Viral Load
7.
J Vis Exp ; (82): e50766, 2013 Dec 10.
Article in English | MEDLINE | ID: mdl-24379004

ABSTRACT

Human respiratory syncytial virus (HRSV) infections present a broad spectrum of disease severity, ranging from mild infections to life-threatening bronchiolitis. An important part of the pathogenesis of severe disease is an enhanced immune response leading to immunopathology. Here, we describe a protocol used to investigate the immune response of human immune cells to an HRSV infection. First, we describe methods used for culturing, purification and quantification of HRSV. Subsequently, we describe a human in vitro model in which peripheral blood mononuclear cells (PBMCs) are stimulated with live HRSV. This model system can be used to study multiple parameters that may contribute to disease severity, including the innate and adaptive immune response. These responses can be measured at the transcriptional and translational level. Moreover, viral infection of cells can easily be measured using flow cytometry. Taken together, stimulation of PBMC with live HRSV provides a fast and reproducible model system to examine mechanisms involved in HRSV-induced disease.


Subject(s)
Leukocytes, Mononuclear/immunology , Leukocytes, Mononuclear/virology , Respiratory Syncytial Virus Infections/immunology , Respiratory Syncytial Viruses/immunology , HeLa Cells , Humans , Respiratory Syncytial Virus Infections/blood
8.
Future Microbiol ; 7(5): 609-24, 2012 May.
Article in English | MEDLINE | ID: mdl-22568716

ABSTRACT

Historically, most research on infectious diseases has focused on infections with single pathogens. However, infections with pathogens often occur in the context of pre-existing viral and bacterial infections. Clinically, this is of particular relevance for coinfections with Streptococcus pneumoniae and influenza virus, which together are an important cause of global morbidity and mortality. In recent years new evidence has emerged regarding the underlying mechanisms of influenza virus-induced susceptibility to secondary pneumococcal infections, in particular regarding the sustained suppression of innate recognition of S. pneumoniae. Conversely, it is also increasingly being recognized that there is not a unidirectional effect of the virus on S. pneumoniae, but that asymptomatic pneumococcal carriage may also affect subsequent influenza virus infection and the clinical outcome. Here, we will review both aspects of pneumococcal influenza virus infection, with a particular focus on the age-related differences in pneumococcal colonization rates and invasive pneumococcal disease.


Subject(s)
Coinfection/microbiology , Coinfection/virology , Influenza, Human/complications , Microbial Interactions , Orthomyxoviridae/pathogenicity , Pneumonia, Pneumococcal/complications , Streptococcus pneumoniae/pathogenicity , Humans , Influenza, Human/virology , Orthomyxoviridae/physiology , Pneumonia, Pneumococcal/microbiology , Streptococcus pneumoniae/physiology
9.
FEMS Microbiol Ecol ; 61(2): 285-94, 2007 Aug.
Article in English | MEDLINE | ID: mdl-17535299

ABSTRACT

The prevalence and distribution of pMEA-like elements in the genus Amycolatopsis was studied. For this purpose, a set of 95 recently isolated Amycolatopsis strains and 16 Amycolatopsis type strains were examined for the presence of two unique pMEA-sequences (repAM and traJ), encoding proteins essential for replication and conjugative transfer. Homologues of repAM and traJ were found in 10 and 26 of 111 investigated strains, respectively, a result which shows that pMEA-like sequences, though not very abundant, can be found in several Amycolatopsis strains. Phylogenetic analysis of the deduced RepAM and TraJ protein sequences revealed clustering with the protein sequences of either pMEA300 or pMEA100. Furthermore, two geographically different populations of pMEA-like elements were distinguished, one originating in Europe and the other in Australia and Asia. Linkage between the distribution of repAM and traJ and the chromosomal identifier, the 16S rRNA gene, indicated that these elements coevolved with their hosts, suggesting that they evolved in an integrated form rather than by horizontal gene transfer of the free replicating form.


Subject(s)
Actinomycetales/genetics , Bacterial Proteins/chemistry , Actinomycetales/isolation & purification , Bacterial Outer Membrane Proteins/chemistry , Bacterial Outer Membrane Proteins/classification , Bacterial Outer Membrane Proteins/genetics , Bacterial Proteins/classification , Bacterial Proteins/genetics , Base Sequence , Chromosome Mapping , DNA Helicases/chemistry , DNA Helicases/classification , DNA Helicases/genetics , DNA-Binding Proteins/chemistry , DNA-Binding Proteins/classification , DNA-Binding Proteins/genetics , Evolution, Molecular , Phylogeny , Sequence Alignment , Sequence Analysis, DNA , Trans-Activators/chemistry , Trans-Activators/classification , Trans-Activators/genetics
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