sigH-rshA过表达对卡介苗菌株致病性的影响

Effect of sigH-rshA overexpression on the pathogencity of BCG strains

摘要:

卡介苗(Mycobacterium bovis Bacille Calmette-Guérin,BCG)是目前唯一许可的结核病预防用疫苗，但菌株间基因组水平的改变导致疫苗在保护效果、安全性等方面存在诸多差异。【目的】验证DU2 Ⅲ和Ⅳ遗传型中sigH-rshA过表达与BCG菌株毒力变化的相关性。【方法】利用分子克隆技术构建了过表达sigH-rshA的重组BCG China和BCG Japan菌株，通过RT-PCR和Western blotting验证基因表达。利用巨噬细胞和重症联合免疫缺陷(server combined immune-deficiency,SCID)小鼠感染模型评价重组BCG的毒力变化；酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)试验评估肿瘤坏死因子(tumor necrosis factor-α,TNF-α)分泌。【结果】SigH-RshA蛋白过表达能够显著促进BCG的胞内增殖，同时刺激巨噬细胞产生更高水平的TNF-α分泌；SCID小鼠试验结果显示sigH-rshA过表达能提高重组BCG对免疫缺陷小鼠的毒力。【结论】sigH-rshA过表达能够增强BCG菌株对小鼠巨噬细胞和实验小鼠的致病性。

Abstract:

Mycobacterium bovis Bacille Calmette-Guérin (BCG) is the only licensed vaccine for prevention of tuberculosis, whereas the genomic changes lead to differences in the protective effect and safety among strains.[Objective] To study the correlation between sigH-rshA overexpression and pathogenicity of BCG strains in DU2 groups Ⅲ and Ⅳ.[Methods] The recombinant BCG China and BCG Japan strains overexpressing sigH-rshA were established by molecular cloning, and the gene expression was verified by RT-PCR and Western blotting. We used macrophages of mouse origin and server combined immune-deficiency (SCID) mouse infection model to assess the pathogenicity of the recombinant BCG. We employed enzyme-linked immunosorbent assay (ELISA) to assess the secretion of tumor necrosis factor-α (TNF-α).[Results] The overexpression of SigH-RshA protein significantly promoted the intracellular proliferation of BCG and stimulated macrophages to produce more TNF-α. The results of the experiment with SCID mice showed that overexpression of sigH-rshA increased the virulence of recombinant BCG strains to immunodeficient mice.[Conclusion] The overexpression of sigH-rshA can enhance the pathogenicity of BCG strains to macrophages and mice.