scholarly journals Evaluation of the Impact of Codon Optimization and N-Linked Glycosylation on Functional Immunogenicity of Pfs25 DNA Vaccines Delivered byIn VivoElectroporation in Preclinical Studies in Mice

2015 ◽  
Vol 22 (9) ◽  
pp. 1013-1019 ◽  
Author(s):  
Dibyadyuti Datta ◽  
Geetha P. Bansal ◽  
Rajesh Kumar ◽  
Barry Ellefsen ◽  
Drew Hannaman ◽  
...  
Keyword(s):  
Nonhuman Primates ◽  
Vaccine Development ◽  
Dna Vaccines ◽  
Codon Optimization ◽  
Antibody Responses ◽  
Transmission Blocking ◽  
Content Type ◽  
The Impact ◽  
Transmission Blocking Vaccine

ABSTRACTPlasmodium falciparumsexual stage surface antigen Pfs25 is a well-established candidate for malaria transmission-blocking vaccine development. Immunization with DNA vaccines encoding Pfs25 has been shown to elicit potent antibody responses in mice and nonhuman primates. Studies aimed at further optimization have revealed improved immunogenicity through the application ofin vivoelectroporation and by using a heterologous prime-boost approach. The goal of the studies reported here was to systematically evaluate the impact of codon optimization,in vivoelectroporation, and N-linked glycosylation on the immunogenicity of Pfs25 encoded by DNA vaccines. The results from this study demonstrate that while codon optimization andin vivoelectroporation greatly improved functional immunogenicity of Pfs25 DNA vaccines, the presence or absence of N-linked glycosylation did not significantly impact vaccine efficacy. These findings suggest that N-glycosylation of Pfs25 encoded by DNA vaccines is not detrimental to overall transmission-blocking efficacy.

scholarly journals Alga-Produced Malaria Transmission-Blocking Vaccine Candidate Pfs25 Formulated with a Human Use-Compatible Potent Adjuvant Induces High-Affinity Antibodies That Block Plasmodium falciparum Infection of Mosquitoes

2015 ◽  
Vol 83 (5) ◽  
pp. 1799-1808 ◽  
Author(s):  
Kailash P. Patra ◽  
Fengwu Li ◽  
Darrick Carter ◽  
James A. Gregory ◽  
Sheyenne Baga ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Malaria Vaccine ◽  
Vaccine Development ◽  
Vaccine Candidate ◽  
Water Emulsion ◽  
Transmission Blocking ◽  
Content Type ◽  
Oil In Water ◽  
Oil In Water Emulsion ◽  
Transmission Blocking Vaccine

A vaccine to prevent the transmission of malaria parasites from infected humans to mosquitoes is an important component for the elimination of malaria in the 21st century, yet it remains neglected as a priority of malaria vaccine development. The lead candidate forPlasmodium falciparumtransmission-blocking vaccine development, Pfs25, is a sexual stage surface protein that has been produced for vaccine testing in a variety of heterologous expression systems. Any realistic malaria vaccine will need to optimize proper folding balanced against cost of production, yield, and potentially reactogenic contaminants. HereChlamydomonas reinhardtiimicroalga-produced recombinant Pfs25 protein was formulated with four different human-compatible adjuvants (alum, Toll-like receptor 4 [TLR-4] agonist glucopyranosal lipid A [GLA] plus alum, squalene–oil-in-water emulsion, and GLA plus squalene–oil-in-water emulsion) and compared for their ability to induce malaria transmission-blocking antibodies. Alga-produced recombinant Pfs25 plus GLA plus squalene–oil-in-water adjuvant induced the highest titer and avidity in IgG antibodies, measured using alga-produced recombinant Pfs25 as the enzyme-linked immunosorbent assay (ELISA) antigen. These antibodies specifically reacted with the surface ofP. falciparummacrogametes and zygotes and effectively prevented parasites from developing within the mosquito vector in standard membrane feeding assays. Alga-produced Pfs25 in combination with a human-compatible adjuvant composed of a TLR-4 agonist in a squalene–oil-in-water emulsion is an attractive new vaccine candidate that merits head-to-head comparison with other modalities of vaccine production and administration.

scholarly journals Exogenous Alginate Protects Staphylococcus aureus from Killing by Pseudomonas aeruginosa

2019 ◽  
Vol 202 (8) ◽  
Author(s):  
Courtney E. Price ◽  
Dustin G. Brown ◽  
Dominique H. Limoli ◽  
Vanessa V. Phelan ◽  
George A. O’Toole
Keyword(s):  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Physical Space ◽  
Late Time ◽  
Protective Effects ◽  
Content Type ◽  
Alginate Production ◽  
The Impact

ABSTRACT Cystic fibrosis (CF) patients chronically infected with both Pseudomonas aeruginosa and Staphylococcus aureus have worse health outcomes than patients who are monoinfected with either P. aeruginosa or S. aureus. We showed previously that mucoid strains of P. aeruginosa can coexist with S. aureus in vitro due to the transcriptional downregulation of several toxic exoproducts typically produced by P. aeruginosa, including siderophores, rhamnolipids, and HQNO (2-heptyl-4-hydroxyquinoline N-oxide). Here, we demonstrate that exogenous alginate protects S. aureus from P. aeruginosa in both planktonic and biofilm coculture models under a variety of nutritional conditions. S. aureus protection in the presence of exogenous alginate is due to the transcriptional downregulation of pvdA, a gene required for the production of the iron-scavenging siderophore pyoverdine as well as the downregulation of the PQS (Pseudomonas quinolone signal) (2-heptyl-3,4-dihydroxyquinoline) quorum sensing system. The impact of exogenous alginate is independent of endogenous alginate production. We further demonstrate that coculture of mucoid P. aeruginosa with nonmucoid P. aeruginosa strains can mitigate the killing of S. aureus by the nonmucoid strain of P. aeruginosa, indicating that the mechanism that we describe here may function in vivo in the context of mixed infections. Finally, we investigated a panel of mucoid clinical isolates that retain the ability to kill S. aureus at late time points and show that each strain has a unique expression profile, indicating that mucoid isolates can overcome the S. aureus-protective effects of mucoidy in a strain-specific manner. IMPORTANCE CF patients are chronically infected by polymicrobial communities. The two dominant bacterial pathogens that infect the lungs of CF patients are P. aeruginosa and S. aureus, with ∼30% of patients coinfected by both species. Such coinfected individuals have worse outcomes than monoinfected patients, and both species persist within the same physical space. A variety of host and environmental factors have been demonstrated to promote P. aeruginosa-S. aureus coexistence, despite evidence that P. aeruginosa kills S. aureus when these organisms are cocultured in vitro. Thus, a better understanding of P. aeruginosa-S. aureus interactions, particularly mechanisms by which these microorganisms are able to coexist in proximal physical space, will lead to better-informed treatments for chronic polymicrobial infections.

scholarly journals In VivoEfficacy of Anuran Trypsin Inhibitory Peptides against Staphylococcal Skin Infection and the Impact of Peptide Cyclization

2015 ◽  
Vol 59 (4) ◽  
pp. 2113-2121 ◽  
Author(s):  
U. Malik ◽  
O. N. Silva ◽  
I. C. M. Fensterseifer ◽  
L. Y. Chan ◽  
R. J. Clark ◽  
...  
Keyword(s):  
Antimicrobial Agents ◽  
Cyclic Peptide ◽  
Sequence Similarity ◽  
Infection Model ◽  
Content Type ◽  
Wide Range ◽  
Inhibitory Activities ◽  
The Impact

ABSTRACTStaphylococcus aureusis a virulent pathogen that is responsible for a wide range of superficial and invasive infections. Its resistance to existing antimicrobial drugs is a global problem, and the development of novel antimicrobial agents is crucial. Antimicrobial peptides from natural resources offer potential as new treatments against staphylococcal infections. In the current study, we have examined the antimicrobial properties of peptides isolated from anuran skin secretions and cyclized synthetic analogues of these peptides. The structures of the peptides were elucidated by nuclear magnetic resonance (NMR) spectroscopy, revealing high structural and sequence similarity with each other and with sunflower trypsin inhibitor 1 (SFTI-1). SFTI-1 is an ultrastable cyclic peptide isolated from sunflower seeds that has subnanomolar trypsin inhibitory activity, and this scaffold offers pharmaceutically relevant characteristics. The five anuran peptides were nonhemolytic and noncytotoxic and had trypsin inhibitory activities similar to that of SFTI-1. They demonstrated weakin vitroinhibitory activities againstS. aureus, but several had strong antibacterial activities againstS. aureusin anin vivomurine wound infection model. pYR, an immunomodulatory peptide fromRana sevosa, was the most potent, with complete bacterial clearance at 3 mg · kg−1. Cyclization of the peptides improved their stability but was associated with a concomitant decrease in antimicrobial activity. In summary, these anuran peptides are promising as novel therapeutic agents for treating infections from a clinically resistant pathogen.

scholarly journals The Combinations Chitosan-Pam3CSK4 and Chitosan-Monophosphoryl Lipid A: Promising Immune-Enhancing Adjuvants for Anticaries Vaccine PAc

2019 ◽  
Vol 87 (12) ◽  
Author(s):  
Yongli Bi ◽  
Qingan Xu ◽  
Lingkai Su ◽  
Jiantao Xu ◽  
Zhongfang Liu ◽  
...  
Keyword(s):  
Lipid A ◽  
Vaccine Development ◽  
Protection Effect ◽  
Content Type ◽  
Monophosphoryl Lipid A ◽  
Monophosphoryl Lipid ◽  
Tooth Surfaces ◽  
Antibody Titers

ABSTRACT We previously demonstrated that recombinant protein PAc could be administered as an anticaries vaccine. However, the relatively weak immunogenicity of PAc limits its application. In the present study, we investigated the effect of two adjuvant combinations of chitosan plus Pam3CSK4 (chitosan-Pam3CSK4) and of chitosan plus monophosphoryl lipid A (chitosan-MPL) in the immune responses to the PAc protein in vivo and in vitro. PAc-chitosan-Pam3CSK4 or PAc-chitosan-MPL promoted significantly higher PAc-specific antibody titers in serum and saliva, inhibited Streptococcus mutans colonization onto the tooth surfaces, and endowed better protection effect with significantly less caries activities than PAc alone. Chitosan-Pam3CSK4 and chitosan-MPL showed no statistically significant differences. In conclusion, our study demonstrated that the chitosan-Pam3CSK4 and chitosan-MPL combinations are promising for anticaries vaccine development.

scholarly journals Intestinal IgA Regulates Expression of a Fructan Polysaccharide Utilization Locus in Colonizing Gut Commensal Bacteroides thetaiotaomicron

mBio ◽  
2019 ◽  
Vol 10 (6) ◽  
Author(s):  
Payal Joglekar ◽  
Hua Ding ◽  
Pablo Canales-Herrerias ◽  
Pankaj Jay Pasricha ◽  
Justin L. Sonnenburg ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Ex Vivo ◽  
Microbial Colonization ◽  
Bacteroides Thetaiotaomicron ◽  
Content Type ◽  
Microbial Utilization ◽  
Polysaccharide Utilization Locus ◽  
The Impact

ABSTRACT Gut-derived immunoglobulin A (IgA) is the most abundant antibody secreted in the gut that shapes gut microbiota composition and functionality. However, most of the microbial antigens targeted by gut IgA remain unknown, and the functional effects of IgA targeting these antigens are currently understudied. This study provides a framework for identifying and characterizing gut microbiota antigens targeted by gut IgA. We developed a small intestinal ex vivo culture assay to harvest lamina propria IgA from gnotobiotic mice, with the aim of identifying antigenic targets in a model human gut commensal, Bacteroides thetaiotaomicron VPI-5482. Colonization by B. thetaiotaomicron induced a microbe-specific IgA response that was reactive against diverse antigens, including capsular polysaccharides, lipopolysaccharides, and proteins. IgA against microbial protein antigens targeted membrane and secreted proteins with diverse functionalities, including an IgA specific against proteins of the polysaccharide utilization locus (PUL) that are necessary for utilization of fructan, which is an important dietary polysaccharide. Further analyses demonstrated that the presence of dietary fructan increased the production of fructan PUL-specific IgA, which then downregulated the expression of fructan PUL in B. thetaiotaomicron, both in vivo and in vitro. Since the expression of fructan PUL has been associated with the ability of B. thetaiotaomicron to colonize the gut in the presence of dietary fructans, our work suggests a novel role for gut IgA in regulating microbial colonization by modulating their metabolism. IMPORTANCE Given the significant impact that gut microbes have on our health, it is essential to identify key host and environmental factors that shape this diverse community. While many studies have highlighted the impact of diet on gut microbiota, little is known about how the host regulates this critical diet-microbiota interaction. In our present study, we discovered that gut IgA targeted a protein complex involved in the utilization of an important dietary polysaccharide: fructan. While the presence of dietary fructans was previously thought to allow unrestricted growth of fructan-utilizing bacteria, our work shows that gut IgA, by targeting proteins responsible for fructan utilization, provides the host with tools that can restrict the microbial utilization of such polysaccharides, thereby controlling their growth.

scholarly journals Impact of Resolvin E1 on Murine Neutrophil Phagocytosis in Type 2 Diabetes

2014 ◽  
Vol 83 (2) ◽  
pp. 792-801 ◽  
Author(s):  
Bruno S. Herrera ◽  
Hatice Hasturk ◽  
Alpdogan Kantarci ◽  
Marcelo O. Freire ◽  
Olivia Nguyen ◽  
...  
Keyword(s):  
Type 2 Diabetes ◽  
Transgenic Animals ◽  
Genetically Engineered ◽  
Mitogen Activated Protein Kinase ◽  
Air Pouch ◽  
Content Type ◽  
Genetically Engineered Mice ◽  
The Impact

Diabetic complications involve inflammation-mediated microvascular and macrovascular damage, disruption of lipid metabolism, glycosylation of proteins, and abnormalities of neutrophil-mediated events. Resolution of inflamed tissues to health and homeostasis is an active process mediated by endogenous lipid agonists, including lipoxins and resolvins. This proresolution system appears to be compromised in type 2 diabetes (T2D). The goal of this study was to investigate unresolved inflammation in T2D. Wild-type (WT) and genetically engineered mice, including T2D mice (db/db), transgenic mice overexpressing the human resolvin E1 (RvE1) receptor (ERV1), and a newly bred strain ofdb/ERV1mice, were used to determine the impact of RvE1 on the phagocytosis ofPorphyromonas gingivalisin T2D. Neutrophils were isolated and incubated with fluorescein isothiocyanate-labeledP. gingivalis, and phagocytosis was measured in a fluorochrome-based assay by flow cytometry. Mitogen-activated protein kinase (MAPK) (p42 and p44) and Akt (Thr308 and Ser473) phosphorylation was analyzed by Western blotting. The mouse dorsal air pouch model was used to evaluate thein vivoimpact of RvE1. Results revealed that RvE1 increased the neutrophil phagocytosis ofP. gingivalisin WT animals but had no impact indb/dbanimals. InERV1-transgenic andERV1-transgenic diabetic mice, phagocytosis was significantly increased. RvE1 decreased Akt and MAPK phosphorylation in the transgenic animals.In vivodorsal air pouch studies revealed that RvE1 decreases neutrophil influx into the pouch and increases neutrophil phagocytosis ofP. gingivalisin the transgenic animals; cutaneous fat deposition was reduced, as was macrophage infiltration. The results suggest that RvE1 rescues impaired neutrophil phagocytosis in obese T2D mice overexpressingERV1.

scholarly journals Cooperative Role of Antibodies against Heat-Labile Toxin and the EtpA Adhesin in Preventing Toxin Delivery and Intestinal Colonization by Enterotoxigenic Escherichia coli

2012 ◽  
Vol 19 (10) ◽  
pp. 1603-1608 ◽  
Author(s):  
Koushik Roy ◽  
David J. Hamilton ◽  
James M. Fleckenstein
Keyword(s):  
Escherichia Coli ◽  
Diarrheal Disease ◽  
Vaccine Development ◽  
Enterotoxigenic Escherichia Coli ◽  
Intestinal Colonization ◽  
Content Type ◽  
Heat Labile

ABSTRACTEnterotoxigenicEscherichia coli(ETEC) is an important cause of diarrheal disease in developing countries, where it is responsible for hundreds of thousands of deaths each year. Vaccine development for ETEC has been hindered by the heterogeneity of known molecular targets and the lack of broad-based sustained protection afforded by existing vaccine strategies. In an effort to explore the potential role of novel antigens in ETEC vaccines, we examined the ability of antibodies directed against the ETEC heat-labile toxin (LT) and the recently described EtpA adhesin to prevent intestinal colonizationin vivoand toxin delivery to epithelial cellsin vitro. We demonstrate that EtpA is required for the optimal delivery of LT and that antibodies against this adhesin play at least an additive role in preventing delivery of LT to target intestinal cells when combined with antibodies against either the A or B subunits of the toxin. Moreover, vaccination with a combination of LT and EtpA significantly impaired intestinal colonization. Together, these results suggest that the incorporation of recently identified molecules such as EtpA could be used to enhance current approaches to ETEC vaccine development.

scholarly journals Downmodulation of Vaccine-Induced Immunity and Protection against the Intracellular BacteriumFrancisella tularensisby the Inhibitory Receptor FcγRIIB

2015 ◽  
Vol 2015 ◽  
pp. 1-13 ◽  
Author(s):  
Brian J. Franz ◽  
Ying Li ◽  
Constantine Bitsaktsis ◽  
Bibiana V. Iglesias ◽  
Giang Pham ◽  
...  
Keyword(s):  
Vaccine Development ◽  
Immune Protection ◽  
Fc Gamma Receptor ◽  
Gamma Receptor ◽  
Significant Difference ◽  
Cytokine Levels ◽  
Iga Production ◽  
Protection Against Infection ◽  
The Impact

Fc gamma receptor IIB (FcγRIIB) is the only Fc gamma receptor (FcγR) which negatively regulates the immune response, when engaged by antigen- (Ag-) antibody (Ab) complexes. Thus, the generation of Ag-specific IgG in response to infection or immunization has the potential to downmodulate immune protection against infection. Therefore, we sought to determine the impact of FcγRIIB on immune protection againstFrancisella tularensis(Ft), a Category A biothreat agent. We utilized inactivatedFt(iFt) as an immunogen. Naïve and iFt-immunized FcγRIIB knockout (KO) or wildtype (WT) mice were challenged withFt-live vaccine strain (LVS). While no significant difference in survival between naïve FcγRIIB KO versus WT mice was observed, iFt-immunized FcγRIIB KO mice were significantly better protected than iFt-immunized WT mice.Ft-specific IgA in serum and bronchial alveolar lavage, as well as IFN-γ, IL-10, and TNF-αproduction by splenocytes harvested from iFt-immunized FcγRIIB KO, were also significantly elevated. In addition, iFt-immunized FcγRIIB KO mice exhibited a reduction in proinflammatory cytokine levelsin vivoat 5 days after challenge, which correlates with increased survival followingFt-LVS challenge in published studies. Thus, these studies demonstrate for the first time the ability of FcγRIIB to regulate vaccine-induced IgA production and downmodulate immunity and protection. The immune mechanisms behind the above observations and their potential impact on vaccine development are discussed.

scholarly journals Generic Vancomycin Enriches Resistant Subpopulations of Staphylococcus aureus after Exposure in a Neutropenic Mouse Thigh Infection Model

2011 ◽  
Vol 56 (1) ◽  
pp. 243-247 ◽  
Author(s):  
Carlos A. Rodriguez ◽  
Maria Agudelo ◽  
Andres F. Zuluaga ◽  
Omar Vesga
Keyword(s):  
Staphylococcus Aureus ◽  
Population Analysis ◽  
Recovery Process ◽  
Control Group ◽  
Infection Model ◽  
Content Type ◽  
Sterile Saline ◽  
Generic Products ◽  
The Impact

ABSTRACTPrevious studies have shown that “bioequivalent” generic products of vancomycin are less effectivein vivoagainstStaphylococcus aureusthan the innovator compound. Considering that suboptimal bactericidal effect has been associated with emergence of resistance, we aimed to assessin vivothe impact of exposure to innovator and generic products of vancomycin onS. aureussusceptibility. A clinical methicillin-resistantS. aureus(MRSA) strain from a liver transplant patient with persistent bacteremia was used for which MIC, minimum bactericidal concentration (MBC), and autolytic properties were determined. Susceptibility was also assessed by determining a population analysis profile (PAP) with vancomycin concentrations from 0 to 5 mg/liter. ICR neutropenic mice were inoculated in each thigh with ∼7.0 log10CFU. Treatment with the different vancomycin products (innovator and three generics; 1,200 mg/kg of body weight/day every 3 h) started 2 h later while the control group received sterile saline. After 24 h, mice were euthanized, and the thigh homogenates were plated. Recovered colonies were reinoculated to new groups of animals, and the exposure-recovery process was repeated until 12 cycles were completed. The evolution of resistance was assessed by PAP after cycles 5, 10, 11, and 12. The initial isolate displayed reduced autolysis and higher resistance frequencies thanS. aureusATCC 29213 but without vancomycin-intermediateS. aureus(VISA) subpopulations. After 12 cycles, innovator vancomycin had significantly reduced resistant subpopulations at 1, 2, and 3 mg/liter, while the generic products had enriched them progressively by orders of magnitude. The great capacity of generic vancomycin to select for less susceptible organisms raises concerns about the role of therapeutic inequivalence of any antimicrobial on the epidemiology of resistance worldwide.

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