A Research On The Effect Of PPS Overexpression Reacts At The Drug Resistance Of MTB And Its Mechanism

Objective: Explore the effect of Mycobacterium tuberculosis Pup, Dop, Paf A and Mpa gene in Pupproteasome system overexpression reacts at the drug resistance of clinical Mycobacterium tuberculosis isolates strains with prevalence in Xinjiang region.Methods: Construction and identification the recombinant shuttle plasmid p MV361/Pup, p MV361/Dop, p MV361/Paf A and p MV361/Mpa, then introduced into drug sensitive strains, Mycobacterium tuberculosis clinical isolates strains of pure resistance to first-line anti-TB drugs witch is Isoniazid, rifampicin, streptomycin and ethambutol, Mycobacterium tuberculosis clinical isolates strains of multidrug-resistant by electroporation. Using real-time quantitative PCR technology to detect and identify them. Determination of minimal inhibitory concentration by using colorimetric method to detect and investigate the effects of drug resistance of over-expression of Pup, Dop, Paf A and Mpa gene in drug sensitive strains, Mycobacterium tuberculosis clinical isolates strains of pure resistance to first-line anti-TB drugs and Mycobacterium tuberculosis clinical isolates strains of multidrug-resistant.Results: Using colorimetric method to determination the MIC value of over-expression of Pup, Dop, Paf A and Mpa gene in Mycobacterium tuberculosis clinical isolates. The result shows that: ①The overexpression of Pup gene can reduce the MIC of drug sensitive strains, Mycobacterium tuberculosis clinical isolates strains of pure resistance to first-line anti-TB drugs and Mycobacterium tuberculosis clinical isolates strains of multidrug-resistant. ②The over-expression of Dop, Paf A and Mpa gene can increased the MIC of drug sensitive strains, Mycobacterium tuberculosis clinical isolates strains of pure resistance to first-line anti-TB drugs and Mycobacterium tuberculosis clinical isolates strains of multidrug-resistant. ③The over-expression of Pup, Dop, Paf A and Mpa gene in drug sensitive strains, Mycobacterium tuberculosis clinical isolates strains of pure resistance to first-line anti-TB drugs and Mycobacterium tuberculosis clinical isolates strains of multidrug-resistant are still sensitive to isoniazid and rifampicin. ④The MIC of drug sensitive strains, Mycobacterium tuberculosis clinical isolates strains of pure resistance to first-line anti-TB drugs and Mycobacterium tuberculosis clinical isolates strains of multidrug-resistant witch were imported in p MV361 does not effect.Conclusions: 1. They successfully constructed the recombinant shuttle plasmid p MV361/Pup, p MV361/Dop, p MV361/Paf A and p MV361/Mpa; 2. Constructed the over-expression of Pup, Dop, Paf A and Mpa gene in drug sensitive strains, Mycobacterium tuberculosis strains of pure resistance to first-line anti-TB drugs witch is Isoniazid, rifampicin, streptomycin and ethambutol,Mycobacterium tuberculosis strains of multidrug-resistant successfully; 3. The over-expression of Pup gene can reduce the drug resistance of Mycobacterium tuberculosis strains of pure resistance to first-line anti-TB drugs and Mycobacterium tuberculosis strains of multidrug-resistant,and the over-expression of Dop, Paf A and Mpa gene can increased their drug resistance.It could be used for the further study of Pup-proteasome system mechanism of drug resistance of Mycobacterium tuberculosis.