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. 2023 Feb 2:14:1100188.
doi: 10.3389/fimmu.2023.1100188. eCollection 2023.

Design of a multi-epitope vaccine against six Nocardia species based on reverse vaccinology combined with immunoinformatics

Fei Zhu  1   2   3   4   5 , Caixia Tan  5   6 , Chunhui Li  5   6 , Shiyang Ma  1   2   3   4   5 , Haicheng Wen  5 , Hang Yang  1   2   3   4   5 , Mingjun Rao  1   2   3   4   5 , Peipei Zhang  1   2   3   4   5 , Wenzhong Peng  1   2   3   4   5 , Yanhui Cui  1   2   3   4   5 , Jie Chen  1   2   3   4   5 , Pinhua Pan  1   2   3   4   5
Affiliations

Design of a multi-epitope vaccine against six Nocardia species based on reverse vaccinology combined with immunoinformatics

Fei Zhu et al. Front Immunol. .

Abstract

Background: Nocardia genus, a complex group of species classified to be aerobic actinomycete, can lead to severe concurrent infection as well as disseminated infection, typically in immunocompromised patients. With the expansion of the susceptible population, the incidence of Nocardia has been gradually growing, accompanied by increased resistance of the pathogen to existing therapeutics. However, there is no effective vaccine against this pathogen yet. In this study, a multi-epitope vaccine was designed against the Nocardia infection using reverse vaccinology combined with immunoinformatics approaches.

Methods: First, the proteomes of 6 Nocardia subspecies Nocardia subspecies (Nocardia farcinica, Nocardia cyriacigeorgica, Nocardia abscessus, Nocardia otitidiscaviarum, Nocardia brasiliensis and Nocardia nova) were download NCBI (National Center for Biotechnology Information) database on May 1st, 2022 for the target proteins selection. The essential, virulent-associated or resistant-associated, surface-exposed, antigenic, non-toxic, and non-homologous with the human proteome proteins were selected for epitope identification. The shortlisted T-cell and B-cell epitopes were fused with appropriate adjuvants and linkers to construct vaccines. The physicochemical properties of the designed vaccine were predicted using multiple online servers. The Molecular docking and molecular dynamics (MD) simulation were performed to understand the binding pattern and binding stability between the vaccine candidate and Toll-like receptors (TLRs). The immunogenicity of the designed vaccines was evaluated via immune simulation.

Results: 3 proteins that are essential, virulent-associated or resistant-associated, surface-exposed, antigenic, non-toxic, and non-homologous with the human proteome were selected from 218 complete proteome sequences of the 6 Nocardia subspecies epitope identification. After screening, only 4 cytotoxic T lymphocyte (CTL) epitopes, 6 helper T lymphocyte (HTL) epitopes, and 8 B cell epitopes that were antigenic, non-allergenic, and non-toxic were included in the final vaccine construct. The results of molecular docking and MD simulation showed that the vaccine candidate has a strong affinity for TLR2 and TLR4 of the host and the vaccine-TLR complexes were dynamically stable in the natural environment. The results of the immune simulation indicated that the designed vaccine had the potential to induce strong protective immune responses in the host. The codon optimization and cloned analysis showed that the vaccine was available for mass production.

Conclusion: The designed vaccine has the potential to stimulate long-lasting immunity in the host, but further studies are required to validate its safety and efficacy.

Keywords: Nocardia; immunoinformatics; molecular docking; molecular dynamics (MD) simulation; multi-epitope vaccine; reverse vaccinology (RV).

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Workflow designed for 6 common Nocardia subspecies vaccines.
Figure 2
Figure 2
The plot of surface positions of all final selected epitopes. (A) The Cytotoxicity T Lymphocytes (CTL) and Helper T lymphocyte (HTL) epitopes of CORE_REP|Org125_Gene1111. (B) The CTL and HTL epitopes of CORE_REP|Org5_Gene928. (C) The CTL and HTL epitopes of CORE_REP|Org97_Gene925; (D) The B-cell epitopes of CORE_REP|Org125_Gene1111. (E) The B-cell epitopes of CORE_REP|Org5_Gene928. (F) The B-cell epitopes of CORE_REP|Org97_Gene925. Each CTL epitope is shown using a combination of C and its starting position. Each HTL epitope is shown using a combination of H and its starting position. Each B-cell epitope is shown using a combination of B and its starting position.
Figure 3
Figure 3
The plot of the binding patterns of CTL epitopes with MHC class I molecules. The MHC I allele molecules were presented in blue and the CTL epitope peptides were presented as magenta. (A) The STDGTGDGY epitope bind with HLA-A0101 (B) The KSGAVRAYY epitope bind with HLA-B5801 (C) The ISPAWFSPY epitope bind with HLA-B1502 (D) The AETASPERI epitope bind with HLA-B4402.
Figure 4
Figure 4
(A) The amino acid sequences of the vaccine construct. The adjuvant is colored blue, the HTL epitopes are colored green, and the CTL epitopes are colored yellow, The B-cell epitopes and the Pan HLA DR-binding epitope (PADRE) are colored red, all linkers are colored black. (B) The secondary structure of the vaccine construct.
Figure 5
Figure 5
(A) Three-dimensional (3D) structure of the vaccine. (B) The Ramachandran plot of the refined 3D model generated by the PROCHECK server, the red-colored regions are the most favored regions, the dark yellow and light yellow regions are the additional allowed and generously allowed regions, the white regions are the disallowed regions. (C) The Z-score plot of the refined 3D model generated by the ProSA-web server. (D) The ERRAT score of the refined 3D model generated by the ERRAT server.
Figure 6
Figure 6
The plot of visualized analysis and the interacting amino acid residues of the docking complex. The TLR2 receptor is colored lightblue, the TLR4 receptor is colored palecyan, and the vaccine is colored limon. (A) The visualization of the vaccine and TLR2 complex and Interacting residues. The chain A refers to TLR2 receptor and the chain B refers to vaccine. (B) The visualization of the vaccine and TLR4 complex and Interacting residues. The chain A refers to TLR4 receptor and the chain B refers to vaccine.
Figure 7
Figure 7
The analysis of molecular dynamics simulation of vaccine-TLRs. (A, B) RMSD (root mean square deviation) plots (C, D) Rg (Radius of gyration) (E, F) SASA(solvent accessible surface area) (G, H) Hydrogen bonds.
Figure 8
Figure 8
Principal component analysis(PCA) and dynamical cross-correlation (DCC) analysis for first MD simulation. (A) The plot of PC1 and PC2 about vaccine-TLR2 complex (B) The plot of PCA analysis about vaccine-TLR2 complex (C) The DCCM plot of vaccine-TLR2 complex (D) The plot of PC1 and PC2 about vaccine-TLR4 complex (E) The plot of PCA analysis about vaccine-TLR4 complex (F) The DCCM plot of vaccine-TLR4 complex.
Figure 9
Figure 9
In silico immune simulation spectrum. (A) Changes in antibody titers after three vaccine injections; (B) Changes in B cell population after three vaccine injections; (C) Changes in plasma B-cell population after three vaccine injections; (D) Changes in helper T cell population after three vaccine injections; (E) Changes in helper T cell population per state after three vaccine injections; (F) Changes in cytotoxic T cell population per state after three vaccine injections; (G) Changes in natural killer (NK) cell population after three vaccine injections; (H) Changes in macrophages (MA) population per state after three vaccine injections (I).
Figure 10
Figure 10
(A) In silico cloning of the multi-epitope vaccine into the pET28a (+) expression vector; the light green part presents the vaccine’s codon. (B) The best secondary structure of the vaccine mRNA. (C) The centroid secondary structure of the vaccine mRNA.
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