基于鸡毒支原体黏附素蛋白的表位疫苗制备及免疫效果评价

doi:10.11843/j.issn.0366-6964.2023.12.027

Abstract

Abstract: The aim of this study was to design a safer and more effective multi-epitope vaccine candidate using the adhesion proteins of Mycoplasma gallisepticum (MG). In this study, bioinformatics methods were used to predict B and T cell epitopes of several adhesin proteins (CrmA, GapA, Mgc2, PvpA) of MG. The screened antigenic epitopes were synthesized into a new epitope gene peptide (named mEA). The secondary structure, hydrophilicity, antigenicity, and tertiary structure of mEA were analyzed by Biology Online software. The recombinant plasmid pET32a-MG-mEA was constructed, identified by restriction endonucleases (BamH Ⅰ and Hind Ⅲ) and introduced into E. coli, BL21 (DE3) for induction of expression and purification by DNA sequencing.Western blot was used to verify the reactivity of the recombinant protein with chicken negative/positive sera. The purified MG-mEA recombinant protein was mixed 1∶1 with adjuvant and prepared as a multi-epitope vaccine and the immunogenicity of the fusion protein was analysed by SPF chicken immunoassay. The results showed that the secondary structure of the mEA sequence was relatively stable and the antigenicity was good. The recombinant plasmid was amplified by PCR to obtain the target band with a size of 1 680 bp, which was consistent with the prediction. SDS-PAGE detected that the relative molecular mass of the expressed protein was about 48 ku, mainly in soluble form. The multi-epitope vaccine immunised SPF chickens showed the highest antibody levels in all immunised groups at 10 d after the second vaccination, with statistically significant differences between the three recombinant protein immunised groups compared to the PBS group (P<0.05). The results showed that MG infection significantly damaged the tracheal mucosa structure and that the recombinant protein+adjuvant vaccine was effective in providing effective protection. In summary, the results of this study provide a new avenue for the development of a safe and effective MG vaccine candidate.

Key words: Mycoplasma gallisepticum, adhesin proteins, multi-epitope vaccine, prokaryotic expression, immunization

CLC Number:

S852.62

ZHONG Lemiao, LIU Binghui, WU Chunlin, WU Yijian. Preparation and Evaluation of Subunit Vaccine based on Adhesin Protein of Mycoplasma gallisepticum[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(12): 5171-5183.

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Preparation and Evaluation of Subunit Vaccine based on Adhesin Protein of Mycoplasma gallisepticum

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