Study On Mechanisms Of Overexpression PhoPR Two-component System Effects On Mycobacterium Tuberculosis Pathogenicity

Objective:To study the effect and its mechanism.of Mycobacterium tuberculosis Pho PR two-component system on the pathogenicity of Mycobacterium tuberculosis.Methods :Construction and identification of recombinant shuttle plasmid p MV361-Pho P and p MV361-Pho R,and the recombinant shuttle plasmids were introduced to H37 Rv and H37 Ra competent cells by using the electricity transformation technology to build gene expression strains. To detect and identify the overexpression strains by using real-time fluorescence quantitative PCR technology, the over expression of macrophages were infected with different strains and control strains, CFU colony counting method was used to detect the strain in the infected macrophage proliferation and the infected macrophage apoptosis rate by flow cytometry.Results:1.Construction and identification of recombinant shuttle plasmid p MV361-Pho R and p MV361-Pho P;2.Construction and identification Pho P/Pho R: H37Rv:: Pho P strain, H37Rv:: Pho R strain,: H37Ra:: Pho P strain, H37Ra:: Pho R strain;3. The CFU colony count test results: compared with group H37 Ra strains colony count, H37Ra: : Pho P strains group in 0.5 days there was no significant difference, the infection of the 1 and 3 days colony number significantly higher, statistically significant difference(P < 0.05); H37Ra: : Pho R strains group in infection of 0.5, the 1day, 3day had no significant difference. Compared with H37 Rv strain group of colony count, H37Rv: : Pho P strains group in 0.5 days there was no significant change of infection, the infection of the 1 day, 3 day colony number significantly higher, statistically significant difference(P < 0.05); H37Rv: :Pho R strains group in infection of 0.5, the 1 day, 3day had no significant difference;4.Cells streaming technology to detect:compared with H37 Ra strains, H37Ra: : Pho P strain infection in 0 h,12 h, 24 h had no significant change; higher in infected 6 h, the difference was statistically significant(P <0.05); H37Ra: : Pho R strain infection in 0 h, 6 h, 12 h, 24 h had no significant change. compared with H37 Rv strain, H37Rv: : Pho P strain infection in 0 h, 6 h, 12 h, 24 h had no significant change; H37Rv: :Pho R strain infection in 0 h, 6 h, 12 h, 24 h had no significant change.Conclusion:1.The recombinant plasmid p MV361-Pho P and p MV361-Pho R were constructed successfully;2.H37Rv:: Pho P strain, H37Rv:: Pho R strain,: Pho PR strain, H37Ra:: Pho P strain, H37Ra:: Pho R strain;were constructed successfully;3.The overexpression of Pho P in the Pho PR MTB two-component system might enhance the pathogenicity of MTB, and the overexpression of Pho R had no significant effect on the pathogenicity of MTB.