scholarly journals Immunization with a ZmpB-Based Protein Vaccine Could Protect against Pneumococcal Diseases in Mice

2010 ◽  
Vol 79 (2) ◽  
pp. 867-878 ◽  
Author(s):  
Yi Gong ◽  
Wenchun Xu ◽  
Yali Cui ◽  
Xuemei Zhang ◽  
Run Yao ◽  
...  
Keyword(s):  
Polyclonal Antibodies ◽  
Pneumococcal Infection ◽  
A549 Cells ◽  
Passive Immunization ◽  
Vaccine Antigen ◽  
Protein Vaccine ◽  
Survival Times ◽  
Protein Antigens ◽  
Zinc Metalloprotease ◽  
Lung Colonization

ABSTRACTZinc metalloprotease B (ZmpB) is present in all isolated pneumococcal strains and contributes to the pathogenesis of pneumococcal infection. In this study, recombinant ZmpB was cloned and expressed inEscherichia coli. The expression of ZmpB by different pneumococcal strains was detectable by Western blotting with antisera raised to recombinant ZmpB. Flow cytometry analysis demonstrated that anti-ZmpB polyclonal antibodies could bind to the cell surface of the pneumococcal strains analyzed. Both recombinant ZmpB protein and anti-ZmpB polyclonal antibodies significantly inhibited the adhesion ofStreptococcus pneumoniaeD39 to A549 cells. In mouse models, mucosal immunization with recombinant ZmpB could significantly reduce pneumococcal lung colonization caused byS. pneumoniaeserotypes 19F and 14 and significantly increase mice survival times following invasive pneumococcal challenge with different pneumococcal strains, including serotypes 2, 3, 6B, and 14. Furthermore, intraperitoneal immunization with recombinant ZmpB in combination with the recombinant pneumolysin mutant (DeltaA146 Ply) and heat shock protein 40 (DnaJ) could enhance the protection against pneumococcal infection compared to protection provided by single-protein antigens. Passive immunization with hyperimmune antisera against these three antigens also demonstrated that the combination of three hyperimmune antisera could provide better protection than single antisera. Taken together, our results suggest that ZmpB is a good candidate pneumococcal vaccine antigen.

scholarly journals Preclinical Evaluation of the Pht Proteins as Potential Cross-Protective Pneumococcal Vaccine Antigens

2010 ◽  
Vol 79 (1) ◽  
pp. 238-245 ◽  
Author(s):  
Fabrice Godfroid ◽  
Philippe Hermand ◽  
Vincent Verlant ◽  
Philippe Denoël ◽  
Jan T. Poolman
Keyword(s):  
Survival Rates ◽  
Pneumococcal Infection ◽  
Protein Family ◽  
Transfer Model ◽  
Pneumococcal Vaccines ◽  
Protein Antigens ◽  
Human Antibodies ◽  
Lung Colonization ◽  
Vaccine Antigens ◽  
Colonization Model

ABSTRACTCurrent pneumococcal vaccines are composed of capsular polysaccharides (PS) of various serotypes, either as free PS or as protein-PS conjugates. The use of pneumococcus protein antigens that are able to afford protection across the majority of serotypes is envisaged as a relevant alternative and/or complement to the polysaccharides. In this context, based on several studies, the Pht protein family emerged as relevant vaccine candidates. The purpose of the present study was to evaluate the Pht protein family in several preclinical mouse models. Immunization with these antigens was compared with immunization with other pneumococcal antigens, such as CbpA, PspA, and PsaA. In a nasopharyngeal colonization model and in a lung colonization model, the Phts were found to be superior to the other candidates in terms of efficacy of protection and serotype coverage. Likewise, vaccination with PhtD allowed higher animal survival rates after lethal intranasal challenge. Finally, a passive transfer model in which natural anti-PhtD human antibodies were transferred into mice demonstrated significant protection against lethal intranasal challenge. This indicates that natural anti-PhtD human antibodies are able to protect against pneumococcal infection. Our findings, together with the serotype-independent occurrence of the Phts, designate this protein family as valid candidate antigens to be incorporated in protein-based pneumococcal vaccines.

scholarly journals Passive Immunization with a Polyclonal Antiserum to the Hemoglobin Receptor of Haemophilus ducreyi Confers Protection against a Homologous Challenge in the Experimental Swine Model of Chancroid

2011 ◽  
Vol 79 (8) ◽  
pp. 3168-3177 ◽  
Author(s):  
Isabelle Leduc ◽  
William G. Fusco ◽  
Neelima Choudhary ◽  
Patty A. Routh ◽  
Deborah M. Cholon ◽  
...  
Keyword(s):  
Lipid A ◽  
Polyclonal Antibodies ◽  
Passive Immunization ◽  
Etiologic Agent ◽  
Polyclonal Antiserum ◽  
Haemophilus Ducreyi ◽  
Human Model ◽  
Swine Model ◽  
Content Type ◽  
Monophosphoryl Lipid A

ABSTRACTHaemophilus ducreyi, the etiologic agent of chancroid, has an obligate requirement for heme. Heme is acquired byH. ducreyifrom its human host via TonB-dependent transporters expressed at its bacterial surface. Of 3 TonB-dependent transporters encoded in the genome ofH. ducreyi, only the hemoglobin receptor, HgbA, is required to establish infection during the early stages of the experimental human model of chancroid. Active immunization with a native preparation of HgbA (nHgbA) confers complete protection in the experimental swine model of chancroid, using either Freund's or monophosphoryl lipid A as adjuvants. To determine if transfer of anti-nHgbA serum is sufficient to confer protection, a passive immunization experiment using pooled nHgbA antiserum was conducted in the experimental swine model of chancroid. Pigs receiving this pooled nHgbA antiserum were protected from a homologous, but not a heterologous, challenge. Passively transferred polyclonal antibodies elicited to nHgbA bound the surface ofH. ducreyiand partially blocked hemoglobin binding by nHgbA, but were not bactericidal. Taken together, these data suggest that the humoral immune response to the HgbA vaccine is protective against anH. ducreyiinfection, possibly by preventing acquisition of the essential nutrient heme.

scholarly journals Antibodies against Thrombospondin-Related Anonymous Protein Do Not Inhibit Plasmodium Sporozoite Infectivity In Vivo

2000 ◽  
Vol 68 (6) ◽  
pp. 3667-3673 ◽  
Author(s):  
Soren Gantt ◽  
Cathrine Persson ◽  
Keith Rose ◽  
Ashley J. Birkett ◽  
Ruben Abagyan ◽  
...  
Keyword(s):  
Metal Ion ◽  
Competitive Inhibition ◽  
Polyclonal Antibodies ◽  
Three Dimensional ◽  
Calcium Ionophore ◽  
Plasmodium Yoelii ◽  
Gliding Motility ◽  
Vaccine Antigen ◽  
Dimensional Structure

ABSTRACT Thrombospondin-related anonymous protein (TRAP), a candidate malaria vaccine antigen, is required for Plasmodiumsporozoite gliding motility and cell invasion. For the first time, the ability of antibodies against TRAP to inhibit sporozoite infectivity in vivo is evaluated in detail. TRAP contains an A-domain, a well-characterized adhesive motif found in integrins. We modeled here a three-dimensional structure of the TRAP A-domain of Plasmodium yoelii and located regions surrounding the MIDAS (metal ion-dependent adhesion site), the presumed business end of the domain. Mice were immunized with constructs containing these A-domain regions but were not protected from sporozoite challenge. Furthermore, monoclonal and rabbit polyclonal antibodies against the A-domain, the conserved N terminus, and the repeat region of TRAP had no effect on the gliding motility or sporozoite infectivity to mice. TRAP is located in micronemes, secretory organelles of apicomplexan parasites. Accordingly, the antibodies tested here stained cytoplasmic TRAP brightly by immunofluorescence. However, very little TRAP could be detected on the surface of sporozoites. In contrast, a dramatic relocalization of TRAP onto the parasite surface occurred when sporozoites were treated with calcium ionophore. This likely mimics the release of TRAP from micronemes when a sporozoite contacts its target cell in vivo. Contact with hepatoma cells in culture also appeared to induce the release of TRAP onto the surface of sporozoites. If large amounts of TRAP are released in close proximity to its cellular receptor(s), effective competitive inhibition by antibodies may be difficult to achieve.

scholarly journals Impaired Immunogenicity of a Meningococcal Factor H-Binding Protein Vaccine Engineered To Eliminate Factor H Binding

2010 ◽  
Vol 17 (7) ◽  
pp. 1074-1078 ◽  
Author(s):  
Peter T. Beernink ◽  
Jutamas Shaughnessy ◽  
Sanjay Ram ◽  
Dan M. Granoff
Keyword(s):  
Binding Protein ◽  
Factor H ◽  
Vaccine Antigen ◽  
Mouse Strains ◽  
Enzyme Linked Immunosorbent Assay ◽  
Complement Factor ◽  
Protein Vaccine ◽  
Wild Type ◽  
Bacterial Surface ◽  
Cast Doubt

ABSTRACT Meningococcal factor H-binding protein (fHbp) is a promising antigen that is part of two vaccines in clinical development. The protein specifically binds human complement factor H (fH), which downregulates complement activation on the bacterial surface and enables the organism to evade host defenses. In humans, the vaccine antigen forms a complex with fH, which may affect anti-fHbp antibody repertoire and decrease serum bactericidal activity by covering important fHbp epitopes. In a recent study, fHbp residues in contact with fH were identified from a crystal structure. Two fHbp glutamate residues that mediated ion-pair interactions with fH were replaced with alanine, and the resulting E218A/E239A mutant no longer bound the fH fragment. In the present study, we generated the E218A/E239A mutant recombinant protein and confirmed the lack of fH binding. By enzyme-linked immunosorbent assay (ELISA), the mutant fHbp showed similar respective concentration-dependent inhibition of binding of four bactericidal anti-fHbp monoclonal antibodies (MAbs) to fHbp, compared with inhibition by the soluble wild-type protein. In two mouse strains, the mutant fHbp elicited up to 4-fold-lower IgG anti-fHbp antibody titers and up to 20-fold-lower serum bactericidal titers than those elicited by the wild-type fHbp vaccine. Thus, although introduction of the two alanine substitutions to eliminate fH binding did not appear to destabilize the molecule globally, the mutations resulted in decreased immunogenicity in mouse models in which neither the mutant nor the wild-type control vaccine bound fH. These results cast doubt on the vaccine potential in humans of this mutant fHbp.

scholarly journals Two DHH Subfamily 1 Proteins Contribute to Pneumococcal Virulence and Confer Protection against Pneumococcal Disease

2011 ◽  
Vol 79 (9) ◽  
pp. 3697-3710 ◽  
Author(s):  
L. E. Cron ◽  
K. Stol ◽  
P. Burghout ◽  
S. van Selm ◽  
E. R. Simonetti ◽  
...  
Keyword(s):  
Otitis Media ◽  
Pneumococcal Disease ◽  
Capsular Polysaccharide ◽  
Pneumococcal Infection ◽  
Bacterial Pathogen ◽  
Protein Vaccine ◽  
Double Knockout ◽  
Content Type ◽  
Knockout Mutants ◽  
Different Strains

ABSTRACTStreptococcus pneumoniaeis an important human bacterial pathogen, causing such infections as pneumonia, meningitis, septicemia, and otitis media. Current capsular polysaccharide-based conjugate vaccines protect against a fraction of the over 90 serotypes known, whereas vaccines based on conserved pneumococcal proteins are considered promising broad-range alternatives. The pneumococcal genome encodes two conserved proteins of an as yet unknown function, SP1298 and SP2205, classified as DHH (Asp-His-His) subfamily 1 proteins. Here we examined their contribution to pneumococcal pathogenesis using single and double knockout mutants in three different strains: D39, TIGR4, and BHN100. Mutants lacking both SP1298 and SP2205 were severely impaired in adherence to human epithelial Detroit 562 cells. Importantly, the attenuated phenotypes were restored upon genetic complementation of the deleted genes. Single and mixed mouse models of colonization, otitis media, pneumonia, and bacteremia showed that bacterial loads in the nasopharynx, middle ears, lungs, and blood of mice infected with the mutants were significantly reduced from those of wild-type-infected mice, with an apparent additive effect upon deletion of both genes. Minor strain-specific phenotypes were observed, i.e., deletion of SP1298 affected host-cell adherence in BHN100 only, and deletion of SP2205 significantly attenuated virulence in lungs and blood in D39 and BHN100 but not TIGR4. Finally, subcutaneous vaccination with a combination of both DHH subfamily 1 proteins conferred protection to nasopharynx, lungs, and blood of mice infected with TIGR4. We conclude that SP1298 and SP2205 play a significant role at several stages of pneumococcal infection, and importantly, these proteins are potential candidates for a multicomponent protein vaccine.

scholarly journals Prevotella intermedia Induces Severe Bacteremic Pneumococcal Pneumonia in Mice with Upregulated Platelet-Activating Factor Receptor Expression

2013 ◽  
Vol 82 (2) ◽  
pp. 587-593 ◽  
Author(s):  
Kentaro Nagaoka ◽  
Katsunori Yanagihara ◽  
Yoshitomo Morinaga ◽  
Shigeki Nakamura ◽  
Tatsuhiko Harada ◽  
...  
Keyword(s):  
Pneumococcal Pneumonia ◽  
Pneumococcal Infection ◽  
Platelet Activating Factor ◽  
A549 Cells ◽  
Receptor Expression ◽  
Prevotella Intermedia ◽  
Transcript Levels ◽  
Content Type ◽  
Bacteremic Pneumococcal Pneumonia ◽  
Platelet Activating Factor Receptor

ABSTRACTStreptococcus pneumoniaeis the leading cause of respiratory infection worldwide. Although oral hygiene has been considered a risk factor for developing pneumonia, the relationship between oral bacteria and pneumococcal infection is unknown. In this study, we examined the synergic effects ofPrevotella intermedia, a major periodontopathic bacterium, on pneumococcal pneumonia. The synergic effects of the supernatant ofP. intermedia(PiSup) on pneumococcal pneumonia were investigated in mice, and the stimulation of pneumococcal adhesion to human alveolar (A549) cells by PiSup was assessed. The effects of PiSup on platelet-activating factor receptor (PAFR) transcript levelsin vitroandin vivowere analyzed by quantitative real-time PCR, and the differences between the effects of pneumococcal infection induced by various periodontopathic bacterial species were verified in mice. Mice inoculated withS. pneumoniaeplus PiSup exhibited a significantly lower survival rate, higher bacterial loads in the lungs, spleen, and blood, and higher inflammatory cytokine levels in the bronchoalveolar lavage fluid (macrophage inflammatory protein 2 and tumor necrosis factor alpha) than those infected without PiSup. In A549 cells, PiSup increased pneumococcal adhesion and PAFR transcript levels. PiSup also increased lung PAFR transcript levels in mice. Similar effects were not observed in the supernatants ofPorphyromonas gingivalisorFusobacterium nucleatum. Thus,P. intermediahas the potential to induce severe bacteremic pneumococcal pneumonia with enhanced pneumococcal adhesion to lower airway cells.

scholarly journals Expression of Heterologous Antigens in Commensal Neisseria spp.: Preservation of Conformational Epitopes with Vaccine Potential

2004 ◽  
Vol 72 (11) ◽  
pp. 6511-6518 ◽  
Author(s):  
Clíona A. O'Dwyer ◽  
Karen Reddin ◽  
Denis Martin ◽  
Stephen C. Taylor ◽  
Andrew R. Gorringe ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Meningococcal Disease ◽  
Protective Efficacy ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Heterologous Proteins ◽  
Protein Vaccine ◽  
Native Conformation ◽  
Protein Antigens

ABSTRACT Commensal neisseriae share with Neisseria meningitidis (meningococcus) a tendency towards overproduction of the bacterial outer envelope, leading to the formation and release during growth of outer membrane vesicles (OMVs). OMVs from both meningococci and commensal neisseriae have shown promise as vaccines to protect against meningococcal disease. We report here the successful expression at high levels of heterologous proteins in commensal neisseriae and the display, in its native conformation, of one meningococcal outer membrane protein vaccine candidate, NspA, in OMVs prepared from such a recombinant Neisseria flavescens strain. These NspA-containing OMVs conferred protection against otherwise lethal intraperitoneal challenge of mice with N. meningitidis serogroup B, and sera raised against them mediated opsonophagocytosis of meningococcal strains expressing this antigen. This development promises to facilitate the design of novel vaccines containing membrane protein antigens that are otherwise difficult to present in native conformation that provide cross-protective efficacy in the prevention of meningococcal disease.

scholarly journals Evaluation of a toxoid fusion protein vaccine produced in plants to protect poultry against necrotic enteritis

2018 ◽  
Author(s):  
Joseph G Hunter ◽  
Shyra Wilde ◽  
Amanda M Tafoya ◽  
Jamie Horsman ◽  
Miranda Yousif ◽  
...  
Keyword(s):  
Fusion Protein ◽  
Expression System ◽  
Vaccine Antigen ◽  
Economic Losses ◽  
Mutant Form ◽  
Necrotic Enteritis ◽  
Protein Vaccine ◽  
Human Pathogens ◽  
Metal Affinity ◽  
The Impact

Background. Necrotic enteritis (NE) is caused by type A strains of the bacterium Clostridium perfringens. Total global economic losses to the poultry industry due to NE is estimated to be over 2 billion dollars annually. Traditionally, NE has been effectively controlled by inclusion of antibiotics in the diet of poultry. However, recent concerns regarding the impact of this practice on increasing antibiotic resistance in human pathogens have led us to consider alternative approaches, such as vaccination, for controlling this disease. NE strains of C. perfringensproduce two major toxins, a-toxin and NetB. Immune responses against either toxin can provide partial protection against NE. Methods. We have developed a fusion protein combining a non-toxic carboxyl-terminal domain of a-toxin (PlcC) and an attenuated, mutant form of NetB (NetB-W262A) for use as a vaccine antigen to immunize poultry against NE. We utilized a DNA sequence that was codon-optimized for Nicotiana benthamianato enable high levels of expression. The 6-His tagged PlcC-NetB fusion protein was synthesized in N. benthamianausing a geminiviral replicon transient expression system, purified by metal affinity chromatography, and used to immunize broiler birds. Results. Immunized birds produced a strong serum IgY response against both the plant produced PlcC-NetB protein and against bacterially produced His-PlcC and His-NetB. Immunized birds were significantly protected against a subsequent in-feed challenge with virulent C. perfringenswhen treated with the fusion protein. These results indicate that a plant-produced PlcC-NetB toxoid is a promising vaccine candidate for controlling NE in poultry.

scholarly journals An Engineered Receptor-Binding Domain Improves the Immunogenicity of Multivalent SARS-CoV-2 Vaccines

mBio ◽  
2021 ◽  
Vol 12 (3) ◽  
Author(s):  
Yan Guo ◽  
Wenhui He ◽  
Huihui Mou ◽  
Lizhou Zhang ◽  
Jing Chang ◽  
...  
Keyword(s):  
Amino Acid ◽  
Receptor Binding ◽  
Binding Domain ◽  
Full Length ◽  
Vaccine Antigen ◽  
Receptor Binding Domain ◽  
Protein Vaccine ◽  
Wild Type ◽  
S Protein ◽  
Glycosylation Sites

ABSTRACT The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) protein mediates viral entry into cells expressing angiotensin-converting enzyme 2 (ACE2). The S protein engages ACE2 through its receptor-binding domain (RBD), an independently folded 197-amino-acid fragment of the 1,273-amino-acid S-protein protomer. The RBD is the primary SARS-CoV-2 neutralizing epitope and a critical target of any SARS-CoV-2 vaccine. Here, we show that this RBD conjugated to each of two carrier proteins elicited more potent neutralizing responses in immunized rodents than did a similarly conjugated proline-stabilized S-protein ectodomain. Nonetheless, the native RBD is expressed inefficiently, limiting its usefulness as a vaccine antigen. However, we show that an RBD engineered with four novel glycosylation sites (gRBD) is expressed markedly more efficiently and generates a more potent neutralizing responses as a DNA vaccine antigen than the wild-type RBD or the full-length S protein, especially when fused to multivalent carriers, such as a Helicobacter pylori ferritin 24-mer. Further, gRBD is more immunogenic than the wild-type RBD when administered as a subunit protein vaccine. Our data suggest that multivalent gRBD antigens can reduce costs and doses, and improve the immunogenicity, of all major classes of SARS-CoV-2 vaccines. IMPORTANCE All available vaccines for coronavirus disease 2019 (COVID-19) express or deliver the full-length SARS-CoV-2 spike (S) protein. We show that this antigen is not optimal, consistent with observations that the vast majority of the neutralizing response to the virus is focused on the S-protein receptor-binding domain (RBD). However, this RBD is not expressed well as an independent domain, especially when expressed as a fusion protein with a multivalent scaffold. We therefore engineered a more highly expressed form of the SARS-CoV-2 RBD by introducing four glycosylation sites into a face of the RBD normally occluded in the full S protein. We show that this engineered protein, gRBD, is more immunogenic than the wild-type RBD or the full-length S protein in both genetic and protein-delivered vaccines.

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