猪链球菌2型Dbp缺失株的构建及特性分析

Construction and characterization of Dbp gene knock-out mutant in Streptococcus suis 2

摘要:

【目的】通过构建DNA结合膜蛋白(DNA-binding membrance protein，Dbp)基因的缺失株，探究Dbp基因对猪链球菌2型强毒株毒力的影响。【方法】通过PCR检测Dbp基因的分布。利用同源重组原理构建Dbp基因上下游片段的重组质粒，将构建好的质粒电转入ZY05719感受态细胞中，筛选Dbp缺失突变株，通过PCR及测序分析对其进行验证。生物学特性分析比较缺失株?Dbp和野毒株ZY05719在生长速率、形态特征、毒力等方面的差异。【结果】Dbp基因为猪链球菌2型强毒株中相对保守基因，构建了Dbp基因缺失株?Dbp。体外实验结果显示Dbp基因缺失株的生长速率在对数期减慢，并且缺失株?Dbp的荚膜同野毒株存在显著差异，斑马鱼实验结果表明缺失株?Dbp毒力下降。【结论】Dbp与猪链球菌2型的毒力相关，在猪链球菌2型的致病过程中起一定的作用，这丰富了对该菌致病机理的认识。

Abstract:

[Objective] The contribution of DNA-binding membrane protein (Dbp) to the virulence of Streptococcus suis 2 was evaluated by constructing a gene knock-out mutant of Dbp. [Methods] The distribution of Dbp was analyzed by PCR. An isogenic Streptococcus suis 2 mutant of Dbp, ?Dbp, was constructed based on the principle of homologous recombination, and the biological characteristics, pathogenicity of ?Dbp and wild type ZY05719 was compared. [Results] The Dbp gene may be a relatively conserved in different source of Streptococcus suis 2. Compared with wild type ZY05719, the growth rate of ?Dbp is significantly slower in the logarithmic phase, and the capsule of ?Dbp is different from wild type ZY05719. Zebrafish pathogenicity test showed that the LD50 of ?Dbp had a significant decrease as compared with wild type ZY05719. [Conclusion] Dbp gene is critical for the full virulence of Streptococcus suis 2. All these help us to understand its pathogenic mechanism.