JJo, a recombinant dimer of conformationally restricted peptide elicits protective response against Group A Streptococcus (GAS) isolates from a GAS-endemic region

A peptide (J14) containing conformationally restricted epitopes from the M protein of Group A Streptococcus (GAS) is capable of eliciting protective immune response against GAS infection. However, the protective response may be lost possibly due to its weak secondary-structure when the antigen is fused with other antigens in a recombinant polyepitope vaccine construct. We previously showed that JJo, a conformationally stabilized derivative of dimeric J14, overcomes this problem. We now show that anti JJo antibodies react with diverse GAS isolates found in the Indian sub-continent and that these antibodies are opsonic for GAS. The GAS strains used in this study were isolated from throat and skin swabs from Mumbai, Chennai and Vellore. Sera from mice immunized with recombinant JJo peptide were tested by ELISA, immunofluorescence, flow-cytometry, indirect bactericidal assay and mouse challenge assays to determine specific immunogenicity, opsonic functions and protection against an Indian isolate. We propose that JJo is a robust antigen suitable for inclusion in recombinant multi-epitope vaccines which are potentially affordable option for the pediatric population of developing countries

An efficient, chemically-defined semisynthetic lipid-adjuvanted nanoparticulate vaccine development system

A novel vaccine development platform that enables the site-specific conjugation of synthetic lipid adjuvants to recombinant proteins was produced. This technology facilitates the simple and efficient production of homogeneous, chemically-defined, semisynthetic lipoprotein vaccines. Using a polytope 'string-of-beads' approach, a synthetic gene incorporating seven Streptococcus pyogenes M protein strain-specific antigens, and a conserved M protein antigen (J14) was produced, expressed, and attached to a lipoamino acid based adjuvant (lipid core peptide; LCP). Nanoparticles (40 nm diameter) of an optimal size for stimulating antibody-mediated immunity were formed upon the addition of these lipoproteins to aqueous buffer (PBS). Systemic antigen-specific IgG antibodies were raised against all eight antigens in C57BL/6 J mice, without the need to formulate with additional adjuvant. These antibodies bound cell surface M proteins of S. pyogenes strains represented within the polytope sequence, with higher antibody levels observed where a dendritic cell targeting peptide (DCpep) was incorporated within the LCP adjuvant. Crown Copyright (C) 2013 Published by Elsevier Inc. All rights reserved

Immunogenicity of recombinant polytope constructs containing protective group A streptococcal antigens, J14 and H12

A number of polytope recombinant proteins were constructed that contained two group A streptococcal (GAS) protective epitopes, J14 and H12, in various combinations and molar ratios. The ability of each of these polytopes to stimulate an immune response to both of the antigens was tested in a murine model. Antigen-specific serum IgG varied between polytope constructs, indicating that the relative position of J14 and H12 influences the immune response and may need to be considered in multi-epitope vaccine design containing these antigens

Gone with the weed: Population growth of Sitophilus oryzae and Rhyzopertha dominica in wheat and barley containing seeds of Silybum marianum

In the present study we examined the effect of seeds of the weed Silybum marianum (L.) Gaertn. (Asterales: Asteraceae) in different combinations with wheat or barley in the population growth of two major stored product insects, Rhyzopertha dominica (F.) (Coleoptera: Bostryhidae) and Sitophilus oryzae (L.) (Coleoptera: Curculionidae). The evaluation took place on 7 different quantitative combinations of the weed, i.e. 0, 1, 5, 10, 20, 50 and 100% of the total amount of the grain mass. All combinations were kept under constant conditions of 25 οC and 65% relative humidity (r.h.) for 65 days. After this interval, adult progeny production was counted, and classified as dead or alive. In general, progeny production was higher on wheat than on barley for R. dominica, but the reverse was recorded for S. oryzae. No progeny production was recorded for either species when S. marianum containment was 100%. Moreover, for both species, the decrease of the percentage of S. marianum caused an increase in progeny production, but this decrease was not linear. The study concludes that R. dominica and S. oryzae cannot develop on S. marianum seeds, but they can develop in mixtures of these seeds with grains. © 2020 Elsevier Lt

Evaluation of novel Streptococcus pyogenes vaccine candidates incorporating multiple conserved sequences from the C-repeat region of the M-protein

A major challenge for Streptococcus pyogenes vaccine development is the identification of epitopes that confer protection from infection by multiple S. pyogenes M-types. Here we have identified and characterised the distribution of common variant sequences from individual repeat units of the C-repeat region (CRR) of M-proteins representing 77 different M-types. Three polyvalent fusion vaccine candidates (SV1, SV2 and SV3) incorporating the most common variants were subsequently expressed and purified, and demonstrated to be alpha-helical by Circular Dichroism (CD), a secondary conformational characteristic of the CRR in the M-protein. Antibodies raised against each of these constructs recognise M-proteins that vary in their CRR, and bind to the surface of multiple S. pyogenes isolates. Antibodies raised against SV1, containing five variant sequences, also kill heterologous S. pyogenes isolates in in vitro bactericidal assays. Further structural characterisation of this construct demonstrated the conformation of SV1 was stable at different pHs, and thermal unfolding of SV1 a reversible process. Our findings demonstrate that linkage of multiple variant sequences into a single recombinant construct overcomes the need to embed the variant sequences in foreign helix promoting flanking sequences for conformational stability, and demonstrates the viability of the polyvalent candidates as global S. pyogenes vaccine candidates

Geostatistical Analysis of permeability data

This dissertation is submitted for the degree of Master of Science.Summarization: Properties such as porosity and permeability are of utmost significance in petroleum industry. They are important parameters for both reservoir engineering and reservoir modelling, since they constitute the basis upon which hydrocarbon reserves, as well as hydrocarbon flow characteristics are determined. For that reason, accurate estimation of these properties has been the subject of continuous studies. For the purposes of this thesis, the porosity and permeability data provided by the 10th SPE benchmark reservoir model were used. This model is part of the PUNQ Complex Model and is a highly heterogeneous model, consisting of two parts: A relatively permeable Tarbert formation on top and an Upper Ness formation at the bottom. The latter comprises of permeable anastomosed channels laid on a non–permeable background. The data were analysed using visualization, statistical and geostatistical techniques, in order to investigate the statistical properties and to subsequently quantify and evaluate their spatial correlation and variability. The exploratory analysis was carried out using classical statistics (e.g., histograms, statistical moments, distribution fitting and scatter plots). However, the main results were derived using geostatistical methods. Geostatistical analysis included the calculation of various variograms (i.e., directional, anisotropic, omnidirectional and 3D variograms), and their subsequent fitting with appropriate theoretical variogram models. In addition to this analysis, an upscaling of the reservoir model was performed by implementing the Simplified Renormalization method to both porosity and permeability data. Various conclusions were drawn from this project concerning the behaviour and spatial correlation of the reservoir, as well as the general implementation of the geostatistical methods. The most important outcome was the confirmation of the high heterogeneity and anisotropy characterizing the entire reservoir model. An equally significant observation was that results depend greatly on the number and locations of data included. More specifically, considering more data across the horizontal plane increases the interpretable information of variogram analysis, while considering more data along the vertical direction increases the variability. Finally, upscaling leads to coarse–grained versions of the reservoir model; such reduced dimensionality models should be further evaluated by means of subsequent flow simulation

Differences among group A streptococcus epidemiological landscapes: consequences for M protein-based vaccines?

Group A streptococcus (GAS) is a bacterial pathogen responsible for a wide array of disease pathologies in humans. GAS surface M protein plays multiple key roles in pathogenesis, and serves as a target for typing and vaccine development. In this review, we have compiled GAS epidemiological studies from several countries around the world to highlight the consequences on the theoretical efficacy of two different M protein-based vaccine strategies.Journal ArticleReviewSCOPUS: re.jinfo:eu-repo/semantics/publishe

Structural optimisation of a conformational epitope improves antigenicity when expressed as a recombinant fusion protein

A conformationally restricted B cell epitope has been identified as a potential safe vaccine candidate from the major group A streptococcal virulence factor, the M protein. To maintain α-helical secondary structure, the minimal epitope is flanked with heterologous sequences to produce the chimeric vaccine candidate called J14. As a strategy toward developing an affordable multivalent GAS vaccine, we have expressed J14 recombinantly with a second GAS protective antigen H12 (rJ14H12). When administered to mice sub-cutaneously, the fusion protein stimulated a strong serum IgG response to the H12 component, but J14 was poorly immunogenic. To increase the immunogenicity of J14 when expressed with the model fusion partner, amino acid modifications were made to the initial recombinant J14 construct to produce rJJo. These changes stabilised the α-helical conformation of the recombinant antigen as assessed by circular dichroism. Mice immunised with rJJoH12, the fusion protein incorporating JJo, effectively stimulated a humoral response to both of the included antigens. These data support the feasibility of developing a multivalent vaccine incorporating the conformationally restricted protective antigen J14