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@#Rhipicephalus microplus, known as the hard tick, is a vector for the parasites Babesia spp. and Anaplasma marginale, both of which can cause significant financial losses to the livestock industry. There is currently no effective vaccine for R. microplus tick infestations, despite the identification of numerous prospective tick vaccine candidates. As a result, the current research set out to develop an immunoinformatics-based strategy using existing methods for designing a multi-epitope based vaccination that is not only effective but also safe and capable of eliciting cellular and humoral immune responses. First, R. microplus proteins Bm86, Subolesin, and Bm95 were used to anticipate and link B and T-cell epitopes (HTL and CTL) to one another. Antigenicity testing, allergenicity assessment, and toxicity screening were just a few of the many immunoinformatics techniques used to identify potent epitopes. Multi-epitope vaccine design was chosen based on the antigenic score 0.935 that is promising vaccine candidate. Molecular docking was used to determine the nature of the interaction between TLR2 and the vaccine construct. Finally, molecular dynamic simulation was used to assess the stability and compactness of the resulting vaccination based on docking scores. The developed vaccine was shown to be stable, have immunogenic qualities, be soluble, and to have high expression by in silico cloning. These findings suggest that experimental investigation of the multi-epitope based vaccine designed in the current study will produce achievable vaccine candidates against R. microplus ticks, enabling more effective control of infestations.
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Objective:To investigate the effects of different chimerism strategies and different immune ways on the two antigen-dominant regions of Xinjiang hemorrhagic fever virus (XHFV) glycoprotein.Methods:The 5' end was added or not added with interleukin-2 (IL-2) signal peptide and the general-purpose auxiliary T cell epitopes as different design strategies. GcⅠ and GcⅡ and the epitopes previously identified on GcⅠ (Gc 233-248, Gc 241-256 and Gc 281-296) were fused and constructed into the eukaryotic expression vector pVAX1 and the prokaryotic expression vector pET-28a. The recombinant prokaryotic plasmid transformed into E.coli BL21 was induced and purified, and the recombinant eukaryotes were extracted by indirect immunofluorescent assay. BALB/c mice were immunized by protein immunity, gene immunity, and DNA prime-protein boost immunity. The IgG antibody level was measured by ELISA. The immune effect was evaluated by the proliferation of T-lymphocytes and the content of cytokines in the spleen. Results:The results of double enzyme digestion and sequencing showed that eight recombinant plasmids were successfully constructed, and the recombinant eukaryotes were successfully expressed in vitro by fluorescence microscopy. After three times of immunization, the IgG level and the proliferation of T-lymphocytes in the spleen of mice in the experimental group were significantly higher than those in the control group ( P<0.01). The mass concentration test results of Th2 cytokines IL-4 and Th1 cytokines interferon-gamma (IFN-γ) revealed that the response of the DNA prime-protein boost immunity was biased to Th1. Conclusions:The multi-epitope chimeric vaccine of XHFV glycoprotein was successfully constructed, and the target antigen could be expressed effectively in vivo. The immune groups stimulated stronger humoral and cellular immune responses compared with the control group. Among them, the immune effect of pVAX1-ST(GcⅠe+GcⅡ) combined with recombinant protein r(GcⅠe+GcⅡ) was the best, and it is expected to be a new candidate vaccine for XHFV.
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Objective To perform prokaryotic expression and preliminary characterization of the recombinant poly-epitope vaccine EgG1Y162-2 (4) against cystic echinococcosis. Methods The recombinant poly-epitope vaccine EgG1Y162-2 (4) against Echinococcus granulosus based on the linker GSGGSG was subjected to structural three-dimensional (3D) modeling using immunoinformatics to analyze the structural changes and evaluate the antigenicity of the vaccine. The pET30a-EgG1Y162-2 (4) recombinant plasmid was generated using double digestion with EcoR I and Sal I, and then transformed into competent cells. Following protein induction with isopropyl-β-D-thiogalactoside (IPTG), the prokaryotic expression proteins were characterized using Western blotting, and the antigenicity of the recombinant protein was analyzed using sera from cystic echinococcosis patients and health volunteers. Results The four EgG1Y162-2 proteins coupled by the 3D structure of the recombinant vaccine EgG1Y162-2 (4) presented independent and effective expression and good antigenicity. The highest protein expression was detected in the supernatant following induction of the recombinant plasmid pET30a-EgG1Y162-2 (4) by 0.2 mmol/L IPTG at 37 °C for 4 h, and a pure protein component was seen following elution with 60 mmol/L imidazole. Western blotting analysis of the recombinant multiepitope protein HIS-EgG1Y162-2 (4) showed a band at approximately 39 kDa, and this band was recognized by sera from cystic echinococcosis patients. Conclusion A recombinant poly-epitope vaccine EgG1Y162-2 (4) against cystic echinococcosis has been successfully constructed, which provides a preliminary basis for researches on recombinant multi-epitope vaccine against cystic echinococcosis.
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@#B7-H3 is an immune checkpoint molecule overexpressed on the surface of a variety of tumors, and is is an ideal target for tumor immunotherapy. In this study, nitrolated T cell epitope designed in the early stage of the laboratory was used to construct an epitope vaccine that can target immune checkpoint B7-H3. The vaccine can significantly inhibit tumor growth in the CT26 colon cancer model, and has a significant synergistic effect with the PD-L1 protein vaccine. B7-H3 vaccine can increase the proportion of CD4+ T cells in splenic T lymphocytes and the proportion of CD8+ T cells in tumor-infiltrating T lymphocytes, while reducing the proportion of suppressor Treg cells in tumor-infiltrating CD4+ T lymphocytes, which effectively improves tumor immunosuppressive microenvironment. Research results suggest that the B7-H3 epitope vaccine can be used as an effective tumor vaccine candidate molecule.
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OBJECTIVE@#To explore RNA dependent RNA polymerase of Chikungunya virus (CHIKV) and develop T cell based epitopes with high antigenicity and good binding affinity for the human leukocyte antigen (HLA) classes as targets for epitopes based CHIKV vaccine.@*METHODS@#In this study we downloaded 371 non-structural protein 4 protein sequences of CHIKV belonging to different regions of the world from the US National Institute of Allergy and Infectious Diseases (NIAID) virus pathogen resource database. All the sequences were aligned by using CLUSTALW software and a consensus sequence was developed by using Uni Pro U Gene Software version 1.2.1. Propred I and Propred software were used to predict HLA I and HLA II binding promiscuous epitopes from the consensus sequence of non-structural protein 4 protein. The predicted epitopes were analyzed to determine their antigenicity through Vaxijen server version 2.0. All the HLA I binding epitopes were scanned to determine their immunogenic potential through the Immune Epitope Database (IEDB). All the predicted epitopes of our study were fed to IEDB database to determine whether they had been tested earlier.@*RESULTS@#Twenty two HLA class II epitopes and eight HLA class I epitopes were predicted. The promiscuous epitopes WMNMEVKII at position 486-494 and VRRLNAVLL at 331-339 were found to bind with 37 and 36 of the 51 HLA class II alleles respectively. Epitope MANRSRYQS at position 58-66 and epitopes YQSRKVENM at positions 64-72 were predicted to bind with 12 and 9 HLA II alleles with antigenicity scores of 0.754 9 and 1.013 0 respectively. Epitope YSPPINVRL was predicted to bind 18 HLA I alleles and its antigenicity score was 1.425 9 and immunogenicity score was 0.173 83. This epitope is very useful in the preparation of a universal vaccine against CHIKV infection.@*CONCLUSIONS@#Epitopes reported in this study showed promiscuity, antigenicity as well as good binding affinity for the HLA classes. These epitopes will provide the baseline for development of efficacious vaccine for CHIKV.
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Objective To explore RNA dependent RNA polymerase of Chikungunya virus (CHIKV) and develop T cell based epitopes with high antigenicity and good binding affinity for the human leukocyte antigen (HLA) classes as targets for epitopes based CHIKV vaccine. Methods In this study we downloaded 371 non-structural protein 4 protein sequences of CHIKV belonging to different regions of the world from the US National Institute of Allergy and Infectious Diseases (NIAID) virus pathogen resource database. All the sequences were aligned by using CLUSTALW software and a consensus sequence was developed by using Uni Pro U Gene Software version 1.2.1. Propred I and Propred software were used to predict HLA I and HLA II binding promiscuous epitopes from the consensus sequence of non-structural protein 4 protein. The predicted epitopes were analyzed to determine their antigenicity through Vaxijen server version 2.0. All the HLA I binding epitopes were scanned to determine their immunogenic potential through the Immune Epitope Database (IEDB). All the predicted epitopes of our study were fed to IEDB database to determine whether they had been tested earlier. Results Twenty two HLA class II epitopes and eight HLA class I epitopes were predicted. The promiscuous epitopes WMNMEVKII at position 486–494 and VRRLNAVLL at 331–339 were found to bind with 37 and 36 of the 51 HLA class II alleles respectively. Epitope MANRSRYQS at position 58–66 and epitopes YQSRKVENM at positions 64–72 were predicted to bind with 12 and 9 HLA II alleles with antigenicity scores of 0.754 9 and 1.013 0 respectively. Epitope YSPPINVRL was predicted to bind 18 HLA I alleles and its antigenicity score was 1.425 9 and immunogenicity score was 0.173 83. This epitope is very useful in the preparation of a universal vaccine against CHIKV infection. Conclusions Epitopes reported in this study showed promiscuity, antigenicity as well as good binding affinity for the HLA classes. These epitopes will provide the baseline for development of efficacious vaccine for CHIKV.
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Objective:To observe the effect of Pidotimod and Thalidomid to enhance the immune response and protective immunity induced by the epitope Vaccines From W2b2a of Toxoplasma gondii in mice.Methods:Mice were immunized intramuscularly with Pidotimod and pcDNA3-W2b2a,thalidomide and pcDNA3-W2b2a,respectively,then the immune response and the survival time of mouse attacked by Toxoplasma gondii were observed.Results:After the immunization,the level of IFN-γin sera of mice inculated with pcDNA3-W2b2a and Pidotimod,pcDNA3-W2b2a and Freund adjuvant,pcDNA3-W2b2a and Thalidomid were significantly higher than pcDNA3-W2b2a (all P<0.05).After the immunization,IgG,CD4+/CD8+T cell ratio ,proliferation of T cell induced by pcDNA3-W2b2a and Pidotimod,pcDNA3-W2b2a and Freund adjuvant were higher than pcDNA3-W2b2a,pcDNA3-W2b2a and thalidomid ( all P<0.05).After challenged with highly virulent tachyzoites,the mean survival time in immunized groups were significantly longer than control group (all P<0.05).Conclusion:Pidotimod ,Thalidomid adjuvant can increase protective immunity of epitope Vaccines From W2b2a of Toxoplasma gondii in mice.
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BACKGROUNDS: Helicobacter pylori colonize the gastric mucosa of half of the world's population. Although it is classified as a definitive type I carcinogen by World Health Organization, there is no effective vaccine against this bacterium. H. pylori evade the host immune response by avoiding toll-like detection, such as detection via toll-like receptor-5 (TLR-5). Thus, a chimeric construct consisting of selected epitopes from virulence factors that is incorporated into a TLR-5 ligand (Pseudomonas flagellin) could result in more potent innate and adaptive immune responses. MATERIALS AND METHODS: Based on the histocompatibility antigens of BALB/c mice, in silico techniques were used to select several fragments from H. pylori virulence factors with a high density of B- and T-cell epitopes. RESULTS: These segments consist of cytotoxin-associated geneA (residue 162-283), neutrophil activating protein (residue 30-135) and outer inflammatory protein A (residue 155-268). The secondary and tertiary structure of the chimeric constructs and other bioinformatics analyses such as stability, solubility, and antigenicity were performed. The chimeric construct containing antigenic segments of H. pylori proteins was fused with the D3 domain of Pseudomonas flagellin. This recombinant chimeric gene was optimized for expression in Escherichia coli. The in silico results showed that the conserved C- and N-terminal domains of flagellin and the antigenicity of selected fragments were retained. DISCUSSION: In silico analysis showed that Pseudomonas flagellin is a suitable platform for incorporation of an antigenic construct from H. pylori. This strategy may be an effective tool for the control of H. pylori and other persistent infections.
Subject(s)
Animals , Antigens, Bacterial/genetics , Bacterial Outer Membrane Proteins/genetics , Bacterial Proteins/genetics , Computer Simulation , Helicobacter pylori/genetics , Mice, Inbred BALB C , Vaccines, DNA/classification , Vaccines, DNA/geneticsABSTRACT
Objective To observe the effect of pidotimod to enhance the immune response and protective immunity induced by the epitope vaccines from new gene wx2b4a of Toxoplasma gondii in mice.Methods Mice were divided into four groups,each group was inoculated intramuscularly three times with pcDNA3-W2b4a,pcDNA3-W2b4a and pidotimod,pcDNA3,natural saline (NS),respectively.The induced immune responses were tested by ELISA detecting IgG,and flow cytometry sorting the subsets of T lymphocyte.All mice were challenged with highly virulent RH tachyzoites to observe the survival time.Results After the immunization,the level of IgG in sera of mice inoculated with wx2b4a and pidotimod,wx2b4a were significantly higher than those control group(all P<0.05).CD4+ and CD8+T cell counts in immunized groups were higher than those control group(P<0.05).After the immunization,CD4+T cell counts and CD4+/CD8+T cell proportion in wx2b4a and pidotimod groups were higher than those in wx2b4a group (all P<0.05).After challenged with highly virulent tachyzoites,the mean survival time of wx2b4a and pidotimod groups was significantly longer than control group and wx2b4a group (all P<0.05).Conclusion Pidotimod can increase protective immunity of epitope vaccines from new gene wx2b4a of Toxoplasma gondii in mice.
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Epitope vaccine is one of the emerging vaccine techniques developing in past decade years.Particularly the advantages of this vaccine on preventing and therapy illness,as cancer and virus,are espacially outstanding.The most importent elements about the vaccine,namely T/B-epitopes obtainment and identification,vehicles for epitope and vaccine construction,are reviewed.In addition,applications of the vaccine technique in some refractory diseases,such as cancer,virus and pathogen infection,are depicted.