B cell responses to a peptide epitope. VI. The kinetics of antigen recognition modulates B cell-mediated recruitment of T helper subsets

The ability of Ag-primed B cells to recruit distinct Th subsets was examined using two analogous synthetic peptides, G41CT3 and G28CT3, as model Ags. With sequence differences at only two positions, these peptides were identical both with respect to fine specificity of Abs induced and ability to prime T cells. Lymph node cell populations primed with peptide G41CT3, when challenged with the homologous Ag, yielded predominantly Th2 cytokines. In contrast, a challenge with the heterologous Ag, G28CT3, resulted in a markedly increased production of Th1 cytokines. These distinctions derived from altered APC function of Ag-primed B cells due to differential kinetics of recognition of the two Ags by surface Ig receptors, as confirmed by binding studies with a panel of anti-G41CT3 mAbs. A concentration-dependent circular dichroism study revealed differences in the nature of intermolecular associations for these two peptides. Furthermore, the on-rate of peptide G28CT3 binding to Ab also increased with increasing peptide concentration, implying a dependence on intermolecular interactions. This, in turn, correlated well with the ability of peptide G28CT3 to preferentially activate either Th1 or Th2 cells. Thus, the relative proportion of Th1 vs Th2 cells recruited by Ag-primed B cells is governed by the on-rate of Ag binding to surface Ig receptors, with higher on-rates promoting Th1 recruitment. Further, even subtle changes in solution behavior of an Ag can markedly influence the kinetics of recognition by B cells

Mathematical Modeling for Radial Overcut on Electrical Discharge Machining of Incoloy 800 by Response Surface Methodology

AbstractIn the present study, Response surface methodology is applied for prediction of radial overcut in die sinking electrical discharge machining (EDM) process for Incoloy 800 superalloy with copper electrode. The current, pulse-ontime, pulse-off time and voltage are considered as input process parameters to study the ROC. The experiments were planned as per central composite design (CCD) method. After conducting 30 experiments, a mathematical model was developed to correlate the influences of these machining parameters and ROC. The significant coefficients were obtained by performing ANOVA at 5% level of significance. From the obtained results,It was found that current and voltage have significant effect on the radial overcut. The predicted results based on developed models are found to be in good agreement with the The predicted values match the experimental results reasonably well with the coefficient of determination 0.9699 for ROC

B cell responses to a peptide epitope. X. Epitope selection in a primary response is thermodynamically regulated

We examine the etiological basis of hierarchical immunodominance of B cell epitopes on a multideterminant Ag. A model T-dependant immunogen, containing a single immunodominant B cell epitope, was used. The primary IgM response to this peptide included Abs directed against diverse determinants presented by the peptide. Interestingly, affinity of individual monomeric IgM Abs segregated around epitope recognized and was independent of their clonal origins. Furthermore, affinity of Abs directed against the immunodominant epitope were markedly higher than that of the alternate specificities. These studies suggested that the affinity of an epitope-specific primary response, and variations therein, may be determined by the chemical composition of epitope. This inference was supported by thermodynamic analyses of monomer IgM binding to Ag, which revealed that this interaction occurs at the expense of unfavorable entropy changes. Permissible binding required compensation by net enthalpic changes. Finally, the correlation between chemical composition of an epitope, the resultant affinity of the early primary humoral response, and its eventual influence on relative immunogenicity could be experimentally verified. This was achieved by examining the effect of various amino-terminal substitutions on immunogenicity of a, hitherto cryptic, amino-terminal determinant. Such experiments permitted delineation of a hierarchy of individual amino acid residues based on their influence; which correlated well with calculated Gibbs-free energy changes that individual residue side chains were expected to contribute in a binding interaction. Thus, maturation of a T-dependant humoral response is initiated by a step that is under thermodynamic control

B cell responses to a peptide epitope. V. Kinetic regulation of repertoire discrimination and antibody optimization for epitope

The influence of imposing various conformational constraints on immune responses to a model epitope within a synthetic peptide immunogen was examined in mice. Although overall immunogenicity was affected, the model epitope (sequence DPAF) remained the predominant recognition site regardless of the conformation in which it was presented. A comparison of anti-DPAF mAbs obtained in response to two analogue peptides, PS1CT3 and CysCT3, in which the DPAF segment was either unconstrained or held within a cyclic loop, respectively, revealed a significant homology in the paratope composition. At one level a subset of anti-PS1CT3 and anti-CysCT3 mAbs was found to share a common heavy chain variable region. In addition, nucleotide sequence homology comparisons of both heavy and light chain variable regions identified the presence of anti-PS1CT3 and anti-CysCT3 mAbs that collectively appeared to derive from a common progenitor, but with nonidentical somatic mutations. Interestingly, however, no bias toward homologous Ag could be discerned on measurement of relative affinities of the mAbs for the two peptides. In contrast, mAb binding on-rates clearly discriminated between peptides representing the homologous vs the heterologous confomer of the DPAF epitope. Thus, it would appear that the kinetics of Ag recognition dominate over equilibrium binding criteria both in epitope-driven repertoire selection and Ab maturation in a humoral response

Report of dengue outbreak investigation in Jothinagar village, Thiruvallur district, Tamil Nadu, India, 2017: epidemiological, entomological, and geospatial investigations

Background: During July 2017 to August 2017, five cases of laboratory-confirmed dengue cases were reported from Jothinagar village, Tamil Nadu, India. The episode was investigated to confirm the existence of an outbreak and formulate appropriate recommendations for containment.Methods: The monthly occurrence of dengue cases from 2014 to 2017 was compared to confirm the outbreak. Additional blood specimens from 22 patients were sent for laboratory confirmation. We conducted active case search, eco-entomological survey, and geo-mapping of cases and Aedes breeding spots.Results: The occurrence of 36 cases of dengue in the village, previously free from the disease for the past 3.5 years, confirmed the outbreak. Twelve were laboratory-confirmed while the remaining 24 were probable cases. The attack rate was highest amongst females in the age group 11-15 years (10.8/100 population). Case fatality was zero. The house index, Breteau index, container index (CI) and pupal index was 37.7% (23/61), 54.1% (33/61), 16.7% (33/198) and 32.8% (20/61) respectively. Discarded tyres were the key productive containers (CI=28.36%). Geo-analysis suggested clustering of cases within 70 m of the Aedes breeding spots particularly within the central part of the village.Conclusions: Based on high entomological indices, an intensive vector elimination campaign was implemented with a special focus on managing discarded tyres. Geo-analysis can be incorporated in surveillance to identify clusters early for control measures.

The Pore-Forming Protein Cry5B Elicits the Pathogenicity of Bacillus sp. against Caenorhabditis elegans

The soil bacterium Bacillus thuringiensis is a pathogen of insects and nematodes and is very closely related to, if not the same species as, Bacillus cereus and Bacillus anthracis. The defining characteristic of B. thuringiensis that sets it apart from B. cereus and B. anthracis is the production of crystal (Cry) proteins, which are pore-forming toxins or pore-forming proteins (PFPs). Although it is known that PFPs are important virulence factors since their elimination results in reduced virulence of many pathogenic bacteria, the functions by which PFPs promote virulence are incompletely understood. Here we study the effect of Cry proteins in B. thuringiensis pathogenesis of the nematode Caenorhabditis elegans. We find that whereas B. thuringiensis on its own is not able to infect C. elegans, the addition of the PFP Cry protein, Cry5B, results in a robust lethal infection that consumes the nematode host in 1–2 days, leading to a “Bob” or bag-of-bacteria phenotype. Unlike other infections of C. elegans characterized to date, the infection by B. thuringiensis shows dose-dependency based on bacterial inoculum size and based on PFP concentration. Although the infection process takes 1–2 days, the PFP-instigated infection process is irreversibly established within 15 minutes of initial exposure. Remarkably, treatment of C. elegans with Cry5B PFP is able to instigate many other Bacillus species, including B. anthracis and even “non-pathogenic” Bacillus subtilis, to become lethal and infectious agents to C. elegans. Co-culturing of Cry5B-expressing B. thuringiensis with B. anthracis can result in lethal infection of C. elegans by B. anthracis. Our data demonstrate that one potential property of PFPs is to sensitize the host to bacterial infection and further that C. elegans and probably other roundworms can be common hosts for B. cereus-group bacteria, findings with important ecological and research implications

Interleukin-1 derived synthetic peptide as an added co-adjuvant in vaccine formulations

A synthetic peptide containing the immunostimulatory and receptor binding sequences of human IL-1β was synthesized and tested for its immunoadjuvant properties. Using a commercially available hepatitis B vaccine as model antigen we found that added peptide enhanced both total and protective antibody responses in high and low responder strains of mice but was unable to overcome non-responsiveness in a third strain. Increased antibody response to antigen in the responder strains was not accompanied by any significant alteration in IgG isotype composition. These results suggest that this peptide may prove useful as a co-adjuvant in vaccines

Comparison of immune responses to a native viral antigen and a synthetic peptide derived from it: implications for vaccine development

Murine immune responses to the hepatitis B surface antigen (HBsAg) and a synthetic peptide derived from it were compared at the humoral level. Six of nine strains used responded to either peptide or HBsAg, though restriction profiles were not superimposable. Two of three strains non-responsive to HBsAg produced an antibody response on immunization with peptide which was cross-reactive with both peptide and HBsAg. In in vitro lymphocyte stimulation assays, lymphocyte from all six peptide-immunized mouse strains could be induced to proliferate on challenge with HBsAg. However, of the HBsAg-immunized groups, lymphocytes from only three of six responder strains proliferated on in vitro HBsAg challenge. Cumulatively, these results suggest that a vaccine formulation that includes both protein antigens and synthetic peptides derived from these proteins may be more effective at eliciting an immune response in a broader cross-section of target population

Macromolecular self-association of a synthetic peptide derived from the hepatitis B surface antigen: construction of a quaternary epitope

A major impediment to the development of peptide vaccines has been the inability accurately to mimic conformationally constrained epitopes on the envelope proteins of pathogens. This limitation is further compounded by the fact that several viral envelope proteins exist either as covalently or non-covalently associated homo-oligomers in the native state. Evidence is now accumulating to indicate that, at least in some instances, these homo-oligomers display antigenic determinants that are not present in the dissociated monomer units. Clearly this problem will have to be addressed if peptide-based vaccines are ever to become feasible alternatives. In this report we demonstrate that an oligomerized synthetic peptide that was derived from the hepatitis B surface antigen aggregates in solution to form macromolecular structures. These aggregates appear to represent a 'native' form of the group-specific determinant presented by the hepatitis B surface antigen