Prediction,mapping and validation of tick glutathione S-transferase B-cell epitopes

In search of ways to address the increasing incidence of global acaricide resistance, tick control through vaccination is regarded as a sustainable alternative approach. Recently, a novel cocktail antigen tick-vaccine was developed based on the recombinant glutathione S-transferase (rGST) anti-sera cross-reaction to glutathione S-transferases of Rhipicephalus appendiculatus (GST-Ra), Amblyomma variegatum (GST-Av), Haemaphysalis longicornis (GST-Hl), Rhipicephalus decoloratus (GST-Rd) and Rhipicephalus microplus (GST-Rm). Therefore, the current study aimed to predict the shared B-cell epitopes within the GST sequences of these tick species. Prediction of B-cell epitopes and proteasomal cleavage sites were performed using immunoinformatics algorithms. The conserved epitopes predicted within the sequences were mapped on the homodimers of the respective tick GSTs, and the corresponding peptides were independently used for rabbit immunization experiments. Based on the dot blot assay, the immunogenicity of the peptides and their potential to be recognized by corresponding rGST anti-sera raised by rabbit immunization in a previous work were investigated. This study revealed that the predicted conserved B-cell epitopes within the five tick GST sequences were localized on the surface of the respective GST homodimers. The epitopes of GST-Ra, GST-Rd, GST-Av, and GST-Hl were also shown to contain a seven residue-long peptide sequence with no proteasomal cleavage sites, whereas proteasomal digestion of GST-Rm was predicted to yield a 4-residue fragment. Given that a few proteasomal cleavage sites were found within the conserved epitope sequences of the four GSTs, the sequences could also contain a T-cell epitope. Finally, the peptide and rGST anti-sera reacted against the corresponding peptide, confirming their immunogenicity. These data support the claim that the rGSTs, used in the previous study, contain conserved B-cell epitopes, which elucidates why the rGST anti-sera cross-reacted to non-homologous tick GSTs. Taken together, the data suggest that the B-cell epitopes predicted in this study could be useful for constituting epitope-based GST tick vaccines.     

A novel method for enhancement of peptide vaccination utilizing T-cell epitopes from conventional vaccines

Peptide vaccines have two fundamental weak points, namely low antigenicity and MHC-restriction. In our previous study, we proposed the design of vaccine peptide to overcome these weakpoints. The vaccine was constructed in the following order, N-terminal, Arg-Gly-Asp (RGD), T-cell epitope peptide, di-lysine linker (KK) to B-cell epitope peptide. Although the vaccine peptide can basically induce B-cell epitope peptide specific antibodies to the host without immune adjuvants via intraperitoneal, subcutaneous and intranasal administration, some peptide antigens require adjuvants for antibody induction. In this study, we propose a novel protocol to enhance the immunogenicity of the peptide utilizing the host immune response to a conventional toxoid vaccine, which are lymphocyte activities to the T-cell epitope peptide. We selected multiagretope-type T-cell epitopes from diphtheria toxoid, a conventional vaccine antigen, and a part of amyloid-beta peptide (Aβ) as a B-cell epitope. The conventional toxoid vaccine was immunized before the peptide immunization. Using this protocol, we succeeded in the enhancement of the anti-Aβ antibodies induction by intranasal immunization without any immune adjuvants in C57BL/6 and Balb/c mice. Furthermore, the vaccine peptide induced the transformation of peripheral blood lymphocytes collected from healthy volunteers carrying immunities to diphtheria toxoid.     

Two B-cell epitope vaccines based on uPA effectively inhibit fertility in male mice

uPA, a trypsin-like serine protease, was found to take active part in male reproduction. Our previous work had demonstrated the antifertility effects of its full length protein immunization, but with immune tolerance and other latent side effects. Here we discovered two effective B-cell epitopes of uPA for male contraception in growth factor-like domain and kringle domain respectively. Together with carrier protein, immunization of these two epitope peptides could induce high titers of specific antibodies in male mice. Significant reduction of fertility was observed in these two groups in mating trial without evident systemic illness or abnormal mating behavior. Epididymal sperms of immunized males exhibited impaired progressive motility and ability to fertilize eggs in vitro. The immunization of another predicted epitope in serine protease domain and the control groups showed no similar positive results. Importantly, T cells were not activated after the challenge of these B-cell epitopes itself, which suggests that these vaccines do not induce cell-mediated autoimmunity. Taken together, our study discovered two uPA B-cell epitopes as novel targets for male immunocontraception with minimum side effects. Considering their high identity with human uPA protein, these two epitope vaccines hold great promise to be developed for man use in the future.     

Specificity of antibodies reactive with hepatitis B surface antigen following immunization with synthetic peptides

The amino acid sequence 139–147 from hepatitis B surface antigen (HBsAg) has previously been shown to represent a B-cell epitope with potential as a component of a synthetic peptide vaccine against hepatitis B. In this paper, two regions of HBsAg which act as T-cell epitopes in inbred mice have been identified (residues 23–34 and residues 160–171). The ability of synthetic peptides representing these epitopes to provide help for the production of antibody against the 139–147 epitope has been assessed following their co-linear synthesis with the B-cell epitope and following co-immunization of the peptides in an uncoupled form. Both these strategies result in the induction of anti-peptide antibodies which specifically react with recombinant HBsAg. The results presented give further support to the concept that synthetic peptides representing appropriately chosen B- and T-cell epitopes from HBsAg could form the basis of a synthetic vaccine against hepatitis B.     

Immunization with phage virus-like particles displaying Zika virus potential B-cell epitopes neutralizes Zika virus infection of monkey kidney cells

Zika virus (ZIKV) is a mosquito-borne flavivirus that has re-emerged and is associated with many debilitating clinical manifestations. Research is currently being conducted to develop a prophylactic vaccine against the virus; however, there has not been any licensed ZIKV vaccine. Recent studies have identified potential B-cell epitopes (amino acids 241–259, 294–315, 317–327, 346–361, 377–388 and 421–437) on the envelope protein of ZIKV, which could be explored to develop peptide vaccines against ZIKV infection. Nevertheless, the immunogenicity of these epitopes has never been assessed. Here, we displayed these epitopes on highly immunogenic bacteriophage virus-like particles (VLPs; MS2, PP7 and Qβ) platforms and assessed their immunogenicity in mice. Mice immunized with a mixture of VLPs displaying ZIKV envelope B-cell epitopes elicited anti-ZIKV antibodies. Although, immunized mice were not protected against a high challenge dose of ZIKV, sera – albeit at low titers – from immunized mice neutralized (in vitro) a low dose of ZIKV. Taken together, these results show that these epitopes are B-cell epitopes and they are immunogenic when displayed on a Qβ VLP platform. Furthermore, the results also show that immunization with VLPs displaying a single B-cell epitope minimally reduces ZIKV infection whereas immunization with a mixture of VLPs displaying a combination of the B-cell epitopes neutralizes ZIKV infection. Thus, immunization with a mixture of VLPs displaying multiple ZIKV B-cell epitopes is a good strategy to enhance ZIKV neutralization.     

Current methods of epitope identification for cancer vaccine design

The importance of the immune system in tumor development and progression has been emerging in many cancers. Previous cancer vaccines have not shown long-term clinical benefit possibly because were not designed to avoid eliciting regulatory T-cell responses that inhibit the anti-tumor immune response. This review will examine different methods of identifying epitopes derived from tumor associated antigens suitable for immunization and the steps used to design and validate peptide epitopes to improve efficacy of anti-tumor peptide-based vaccines. Focusing on in silico prediction algorithms, we survey the advantages and disadvantages of current cancer vaccine prediction tools.     

In silico CD4+ T-cell epitope prediction and HLA distribution analysis for the potential proteins of Neisseria meningitidis Serogroup B—A clue for vaccine development

Neisseria meningitidis, an exclusive human pathogen, is a major cause of mortality due to meningococcal meningitis and sepsis in many developing countries. Three meningococcal serogroup B proteins, i.e. T-cell stimulating protein A (TspA), autotransporter A (AutA), and IgA-specific serine endopeptidase (IGA1) elicits CD4+ T-cell response and may enhance the effectiveness of meningococcal vaccines by acting as protective immunogens. A very limited data on T-helper cell epitopes in MenB proteins is available. Hence, in silico prediction of peptide sequences which may act as helper T lymphocyte epitopes in MenB proteins was carried out by NetMHCIIpan web server. HLA distribution analysis was done by using the population coverage tool of Immune Epitope Database to determine the fraction of individuals in various populations expected to respond to a given set of predicted T-cell epitopes based on HLA genotype frequencies. Six epitopic core sequences, two from each MenB proteins, i.e. AutA, TspA and IgA1 protease were predicted to associate with a large number of HLA-DR alleles. These six peptides may act as T-cell epitope in more than 95% of populations in 8 out of 12 populations considered. The T-cell stimulation potential of these predicted peptides containing the core epitopic sequences is to be validated by using laboratory experiments for their efficient use as peptide vaccine candidates against N. meningitidis serogroup B.     

Identification and evaluation of an infertility-associated ZP3 epitope from the marsupial brushtail possum (Trichosurus vulpecula)

Immunologically based fertility control vaccines against zona pellucida (ZP) proteins are being developed in New Zealand for biocontrol of the brushtail possum (Trichosurus vulpecula), an introduced Australian marsupial pest. We have shown that immunization of female possums with recombinant possum ZP3 protein (rZP3) reduced fertility by 79%. To enhance the specificity of possum immunocontraceptive vaccines, B-cell epitopes on possum ZP3 protein were mapped using sera of female possums immunized with possum rZP3 and subjected to a fertility trial. The amino acid sequence of the full-length possum ZP3 protein was used to synthesize a complete set of 83 (12-mer) biotinylated peptides each with an overlap of five amino acids with the neighboring peptides. The peptides were used in a modified enzyme-linked immunosorbent assay (ELISA) to identify continuous epitopes recognized by antibodies in the sera of possums immunized with possum rZP3. Sixteen epitopes were identified on the possum ZP3 protein. Comparison of the ELISA binding patterns of these peptides to antibodies in the individual sera with the fertility status of rZP3-immunized possums identified only one epitope (amino acids 156–172) to be associated with infertility. However, female possums immunized with this epitope showed no significant reduction in fertility. The possible reasons for the failure of this potential infertility epitope are discussed.     

Virus and virus-like particle-based immunogens for Alzheimer's disease induce antibody responses against amyloid-beta without concomitant T cell responses

A vaccine targeting the amyloid-beta (Abeta) peptide is a promising potential immunotherapy for Alzheimer's disease patients. However, experience from a recent clinical trial of a candidate Abeta vaccine has suggested that it is important to develop techniques to induce high titer antibodies against Abeta associated with vaccine efficacy while reducing the T cell responses against Abeta that were potentially responsible for serious side effects. We have previously demonstrated that immunization with self- and foreign antigens arrayed in a repetitive fashion on the surface of virus-like particles (VLPs) induces high titer antibody responses at low doses and in the absence of potentially inflammatory adjuvants. In this study, we examined the antibody and T cell responses upon immunization with human papillomavirus VLP- and Qbeta bacteriophage-based Abeta vaccines. Immunization with Abeta conjugated to VLPs or Qbeta elicited anti-Abeta antibody responses at low doses and without the use of adjuvants. The flexibility of these virus-based display systems allowed us to link and induce antibodies against short Abeta-derived peptides from the amino- and carboxyl-termini of the peptide. Immunization of mice with Abeta peptide in combination with Freund's adjuvant elicited predominantly IgG2c antibodies and strong T cell proliferative responses against Abeta. In contrast, VLP-conjugated Abeta peptides elicited more balanced isotype responses, dominated by IgG1. Both VLP and Qbeta-based Abeta vaccines induced weak or negligible T cell responses against Abeta. T cell responses were largely directed against linked viral epitopes. Taken together, virus-based vaccines that allow the presentation of Abeta in a repetitive dense array are new and potentially more effective vaccine candidates for Alzheimer's disease.     

Identification of B- and T-cell epitopes from glycoprotein B of herpes simplex virus 2 and evaluation of their immunogenicity and protection efficacy

Herpes simplex virus (HSV) infection is a major health concern worldwide. Evidence obtained from animals and humans indicates that B- and T-cell responses contribute to protective immunity against herpes virus infection. Glycoprotein B is a transmembrane envelope component of HSV-1 and HSV-2, which plays an important role in virion morphogenesis and penetration into host cells, and can induce neutralizing antibodies and protective T-cell response when it is used to immunize humans and animals. However, little is known about gB epitopes that are involved in B- and T-cell activities in vitro and in vivo. Thus, the HSV-2 gB sequence was screened using B- and T-cell epitope prediction systems, and the B-cell regions and the HLA-A*0201-restricted epitopes were identified. These B-cell epitopes elicited high IgG antibody titers in Balb/C mice, with a predominantly IgG1 subclass distribution, which indicated a Th2 bias. Specific IgGs induced by these two epitopes were evaluated as the neutralizing antibodies for virus neutralization. The predicted T-cell epitopes stabilized the HLA-A*0201 molecules on T(2) cells, and stimulate interferon-γ-secreting and cytotoxic CD8(+) T cells. Immunization with the predicted peptides reduced virus shedding and protected against lethal viral challenge in mice. The functional epitopes described herein, both B- and T-cell epitopes, are potentially implicated in vaccine development.     

Promiscuous T-cell epitopes of Plasmodium merozoite surface protein 9 (PvMSP9) induces IFN-γ and IL-4 responses in individuals naturally exposed to malaria in the Brazilian Amazon

Plasmodium vivax merozoite surface protein (PvMSP9) stimulates both cellular and humoral immune responses in individuals who are naturally infected by this parasite species. To identify immunodominant human T-cell epitopes in PvMSP9, we used the MHC class II binding peptide prediction algorithm ProPred. Eleven synthetic peptides representing predicted putative promiscuous T-cell epitopes were tested in IFN-γ and IL-4 ELISPOT assays using peripheral blood mononuclear cells (PBMC) derived from 142 individuals from Rondonia State, Brazil who had been naturally exposed to P. vivax infections. To determine whether the predicted epitopes are preferentially recognized in the context of multiple alleles, MHC Class II typing of the cohort was also performed. Five synthetic peptides elicited robust cellular responses, and the overall frequencies of IFN-γ and IL-4 responders to at least one of the promiscuous peptides were 62% and 46%, respectively. The frequencies of IFN-γ and IL-4 responders to each peptide were not associated with a particular HLA-DRB1 allelic group since most of the peptides induced a response in individuals of 12 out of 13 studied allelic groups. The prediction of promiscuous epitopes using ProPred led to the identification of immunodominant epitopes recognized by PBMC from a significant proportion of a genetically heterogeneous population exposed to malaria infections. The combination of several such T-cell epitopes in a vaccine construct may increase the frequency of responders and the overall efficacy of subunit vaccines in genetically distinct populations.     

Characterization of murine T-cell epitopes on mycobacterial DNA-binding protein 1 (MDP1) using DNA vaccination

Mycobacterial DNA-binding protein 1 (MDP1) is a major protein antigen in mycobacteria and induces protective immunity against Mycobacterium tuberculosis infection in mice. In this study we determined murine T-cell epitopes on MDP1 with MDP1 DNA immunization in mice. We analyzed interferon-γ production from the MDP1 DNA-immune splenocytes in response to 20-mer overlapping peptides covering MDP1 protein. We identified several CD4+ T-cell epitopes in three inbred mouse strains and one CD8+ T-cell epitope in C57BL/6 mice. These T-cell epitopes would be feasible for analysis of the role of MDP1-specific T-cells in protective immunity and for future vaccine design against M. tuberculosis infection.     

Differential antibody responses to Plasmodium falciparum-derived B-cell epitopes induced by diepitope multiple antigen peptides (MAP) containing different T-cell epitopes

Epitopes of universal character are needed when designing subunit vaccines against infectious diseases such as malaria. We have compared the immunogenicity of B-cell epitopes from the Plasmodium falciparum antigen repeats DPNANPNV (PfCS protein) and VTEEI (Pf332) when assembled with four different universal T-cell epitopes in diepitope multiple antigen peptides (MAP). T-epitopes employed were from P. falciparum antigens (CS.T3, [T(*)]4 and EBP3) or from the Clostridium tetani toxin (P2). In association with either of the T-epitopes, the genetic unresponsiveness to the B-epitopes was successfully bypassed. Our results show that the immunogenicity of a T-epitope alone does not necessarily predict the ability of the T-epitope to provide T-cell help when combined with other epitopes in an immunogen. Further, the nature of the immune responses in terms of total IgG antibodies and their subclass distribution, T-cell proliferation and IFN-gamma production, varied with the T-epitope and mouse strain, which may indicate the need for inclusion of a combination of different universal T-epitopes in a future malaria subunit vaccine.     

Identification of murine T-cell epitopes on low-molecular-mass secretory proteins (CFP11, CFP17, and TB18.5) of Mycobacterium tuberculosis

The low-molecular-mass secretory proteins of Mycobacterium tuberculosis have been shown to be major T-cell antigens during infection with the pathogenic bacterium. In this study, we determined murine T-cell epitopes on three low-molecular-mass proteins, CFP11 (Rv2433c), CFP17 (Rv1827), and TB18.5 (Rv0164) using DNA immunization of inbred mice. We analyzed interferon-γ production from immune splenocytes in response to overlapping peptides covering these proteins. We identified two CD8+ T-cell epitopes on CFP11 and CFP17, one in BALB/c mice and the other in C57BL/6 mice, respectively. On TB18.5, we identified a CD8+ T-cell epitope in BALB/c mice and a CD4+ T-cell epitope in C57BL/6 mice. With the aid of computer algorithms, we could identify the minimal CD8+ T-cell epitopes. These T-cell epitopes are feasible for analysis of the role of antigen-specific T cells during M. tuberculosis infection.     

Cross-reactive immunogenicity of group A streptococcal vaccines designed using a recurrent neural network to identify conserved M protein linear epitopes

The M protein of group A streptococci (Strep A) is a major virulence determinant and protective antigen. The N-terminal sequence of the protein defines the more than 200 M types of Strep A and also contains epitopes that elicit opsonic antibodies, some of which cross-react with heterologous M types. Current efforts to develop broadly protective M protein-based vaccines are directed at identifying potential cross-protective epitopes located in the N-terminal regions of cluster-related M proteins for use as vaccine antigens. In this study, we have used a comprehensive approach using the recurrent neural network ABCpred and IEDB epitope conservancy analysis tools to predict 16 residue linear B-cell epitopes from 117 clinically relevant M types of Strep A (~88% of global Strep A infections). To examine the immunogenicity of these epitope-based vaccines, nine peptides that together shared ≥60% sequence identity with 37 heterologous M proteins were incorporated into two recombinant hybrid protein vaccines, in which the epitopes were repeated 2 or 3 times, respectively. The combined immune responses of immunized rabbits showed that the vaccines elicited significant levels of antibodies against all nine vaccine epitopes present in homologous N-terminal 1–50 amino acid synthetic M peptides, as well as cross-reactive antibodies against 16 of 37 heterologous M peptides predicted to contain similar epitopes. The epitope-specificity of the cross-reactive antibodies was confirmed by ELISA inhibition assays and functional opsonic activity was assayed in HL-60-based bactericidal assays. The results provide important information for the future design of broadly protective M protein-based Strep A vaccines.     

Immuno-informatics approach for B-cell and T-cell epitope based peptide vaccine design against novel COVID-19 virus

COVID-19 has brought the world to a standstill with a wave of destruction in country after country with tremendous loss of lives and livelihood in advanced to developing nations. Whole world is staring at the prospect of repeated lockdowns with another wave of COVID-19 predicted to hit the world in September of 2020. The second wave is assumed to be even more destructive with severe impact across much of the world. The only way to defeat this pandemic is to quickly develop a safe and effective vaccine against this raging menace and initiate a global vaccination drive. Our study is an attempt to deploy various computational methods to identify B-cell and T-cell epitopes from the spike surface glycoprotein of SARS-COV-2 which have the novel potential for vaccine development against COVID-19. For this we have taken 8 unique strains with one each from India, China, France, USA, Italy, Australia, Iran and Pakistan. The strain data was extracted from NCBI Database. By analyzing the immune parameters like surface accessibility, antigenicity, variability, conservancy, flexibility, hydrophilicity, allergenicity and toxicity of the conserved sequences of spike glycoprotein using various databases and bioinformatics tools, we identified two potential novel linear (SGTNGTKRFDN and ASVYAWNRK) and one structural B-cell epitope as well as two T-cell epitopes (RLFRKSNLK and IPTNFTISV) which can be used as epitope-based peptide vaccines. Docking simulation assay revealed that above T-cell epitopes have minimum free binding energy and showed strong hydrogen bond interaction which strengthened its potential as being a T-cell epitope for the epitope-based novel vaccine against SARS-CoV-2. This study allows us to claim that B-cell and T-cell epitopes mentioned above provide potential pathways for developing an exploratory vaccine against spike surface glycoprotein of SARS-CoV-2 with high confidence for the identified strains. We will need to confirm our findings with biological assays.     

Major CD4 epitopes involved in anti-CD4 T-cell autoimmunity in HIV-1 patients

We studied HIV-positive and -negative subjects for T-cell reactivity to rCD4, and found that 80% of 25 tested HIV-infected patients and 25% of controls manifested T-cell proliferation responses to rCD4. We mapped the major CD4 immunogenic epitopes among the CD4+ responders of both groups by testing T-cell proliferation responses to 31 synthetic overlapping peptides from the human CD4 molecule. Such responses to p1, p4, p14, p21, p28 and p29 were significantly higher in the eight infected patients and, with the exception of p14, these peptides differed from those found in three HIV-negative controls (p11, p14 and p27). Peptides p1, p28 and p29 are major immunogenic epitopes. Our findings suggest: (1) that HIV infection is associated with T-cell reactivity to CD4; and (2) that the use of synthetic CD4 peptides to replace the complete CD4 molecule may therefore lead to a cost-effective T-cell vaccination for HIV-positive patients exhibiting anti-CD4 autoimmunity, as well as to the development of complimentary TCR peptides for future peptide vaccinations.     

Synthetic B- and T-cell epitope peptides of porcine reproductive and respiratory syndrome virus with Gp96 as adjuvant induced humoral and cell-mediated immunity

Highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) has recently caused huge economic losses in the pig industry worldwide. Commercial vaccines, including inactivated vaccines and attenuated live vaccines, are available but fail to provide sustainable protection, especially against genetically heterologous strains. Thus several approaches have been used to develop more effective PRRSV vaccines and/or immune modulators to accelerate and magnify immune responses to PRRSV vaccines. Heat shock protein Gp96 is one such modulator that enhances both the innate and adaptive immune responses. In the present study, two B-cell epitopes and seven T-cell epitopes from PRRSV and a Pan DR T-helper cell epitope were synthesized and mixed with the N-terminal 22–355 aa of Gp96 (Gp96N) as an adjuvant, and immune responses were evaluated. Our results show that Gp96N activated PRRSV-specific humoral immune responses elicited by BCE-peptides and promoted the PRRSV-specific cellular immunity induced by TCE-peptides. Moreover, higher levels of IL-12 and TNF-α and lower levels of IL-4 and IL-10 were observed in the serum of Gp96N-vaccinated piglets compared to piglets immunized with no Gp96N, displaying a predominant Th1 type of immune response induced by Gp96N. Following challenge with the virulent HP-PRRSV isolate JXwn06, piglets vaccinated with the mixture of peptides and Gp96N presented with milder clinical symptoms, lower viremia, and less pathological lesions in their lungs, however, this vaccine could not provide lasting and effective protection against HP-PRRSV infection. These data provide important bases for the development of PRRSV epitope-based synthetic peptide vaccines combined with Gp96N as attractive immunomodulators in swine.     

The design of multiepitope vaccines from plasmids of diarrheagenic Escherichia coli against diarrhoea infection: Immunoinformatics approach

Diarrhoea infection is a major global health public problem and is caused by many organisms including diarrheagenic Escherichia coli pathotypes. The common problem with diarrhoea is the drug resistance of pathogenic bacteria, the most promising alternative means of preventing drug resistance is vaccination. However, there has not been any significant success in the prevention of diarrhoea caused by E. coli through vaccination. Epitope-based vaccine is gaining more attention due to its safety and specificity. Sequence variation of protective antigens of the pathogen has posed a new challenge in the development of epitope-based vaccines against the infection, leading to the necessity of multiepitope based design. In this study, immunoinformatics tools were used to design multiepitope vaccine candidates from plasmid genome sequences of multiple pathotypes of E. coli species involved in diarrhoea infections. The ability of the identified epitopes to be used as a cross-protect multiepitope vaccine was achieved by identifying conserved, immunogenic and antigenic peptides that can elicit CD4⁺ T-cell, CD8⁺ T-cell and B-cell and bind to MHC I and II HLA alleles. The molecular docking results of T-cell epitopes showed their well binding affinity to receptive protein and with a wider population coverage. The different multiepitope-based vaccines (MEVCs) candidates were constructed and based on the types of epitope linker they contained. The MEVCs exhibited very good binding interactions with the human immune receptor. Among multiepitope vaccines constructed, MEVC6, MEVCA and MEVCB are more promising as potential vaccine candidates for cross-protection against gastrointestinal infections according to the computational study. It is also hoped that after validation and testing, the predicted multiepitope-based vaccine candidates will probably resolve the challenge of immunological heterogeneity facing enteric vaccine development.     

Recombinant heat shock protein 65 carrying hepatitis B core antigen induces HBcAg-specific CTL response

Many studies have provided evidence that heat shock protein 65 (Hsp65) can elicit potent specific cellular adaptive immune responses (e.g. CD8(+) cytotoxic T-cell effectors or classic CTLs) based on their ability to chaperone antigenic peptides. Hsp65 is thus an effective carrier for heterologous peptide epitopes for therapeutic vaccines against cancer or chronic infectious diseases. The core antigen of hepatitis B virus (HBcAg) is extremely immunogenic, and functions as both a T-cell-dependent and a T-cell-independent antigen. Therefore, HBcAg may be a promising candidate target for therapeutic vaccine control of chronic HBV infection. Here, a chimeric protein, Hsp65Bc, was created by fusing the HBcAg sequence to the carboxyl terminus of the Hsp65 sequence in E. coli. Analysis of its antigenicity and immunogenicity revealed that HBc epitopes are surface accessible. Hsp65Bc induced moderate anti-HBc immune responses as well as a strong specific T-cell response in BALB/c mice. These results indicate that Hsp65Bc may have potential as a vaccine for treatment of HBV chronic infection.