The Study Of Bacteriophage-Based Assay Detecting Pyrazinamide Resistance In Mycobacterium Tuberculosis And Mutation In PncA Gene

Tuberculosis is a chronic infectious disease caused by Mycobacteriuni tuberculosis(MTB), which is one of the most epidemic human disease. It is also one of the diseases caused the largest number of death that caused by a single pathogen. Because the drug-resistance Mycobacterium tuberculosis come forth and spread abroad, AIDS prevail and all so, Tuberculosis epidemic situation become serious again in the world. Pyrazinamide(PZA) is one of the most effective one-line drugs which treat tuberculosis, The quick veracious way that detect the PZA resistance in Mycobacterium tuberculosis is very important to Tuberculosis treat and control. The precise molecular mechanism of PZA-resistance is unclear, to dectect the mutation of pncA is very significative in theory and application[Objective] To set up and evaluate the method of Phage Amplified Biologically Assay (PhaB) in the rapid detection of detection of Pyrazinamide (PZA) resistance. To study the mutation of pncA gene.[Methods] To select the optimum detecting conditions and apply developed method detecting PZA resistance in 109 clinical isolates of Mycobacterium tuberculosis and compare it with the results of absolute concentration method. The minimum inhibitory concentration (MIC) was detected for all discrepancy isolates. To amplify the pncA genes of 47 PZA-resistance strains by PCR and sequence pncA genes of PZA-resistance strains.[Results] The results showed that the pH5. 5, PZA 200ug/rnl and 48h was optimum detecting condition. All of 109 strains of Mycobacterium tuberculosis, 34 strains were PZA-susceptible and 75 strains were PZA-resistible detected by PhaB while 35 strains were PZA-suscept ible and 74 strains were PZA-resistible by absolute concentration method. 30 of 108 strains were PZA-susceptibie and 70 strains were PZA-resistible with...