The Preliminary Study Of Constructing The Mycobacterium Tuberculosis Strain Labeled With Bioluminescence Markers And Applying It To The Pharmacodynamics Evaluation Of New Anti-tuberculosis Drugs

Objectives 1.To construct the recombinant MTB strain that expresses stably high levels of firefly luciferase encoded protein and to evaluate the pharmacodynamics of anti-tuberculosis drugs in vitro.2.To infect mice with the recombinant MTB strain that expresses stably high levels of firefly luciferase encoded protein and the recombinant MTB strain that expresses iRFP encoded protein.Then we could estimate the feasibility of evaluating the pharmacodynamics of anti-tuberculosis drugs in vivo.Methods 1.We built the recombinant expression vector plasmid pMV361+FFLuc and used the IVIS Spectrum to identify the expression of FFLuc.We also detected the fluorescence quantity under different concentration of anti-tuberculosis drugs in different time,and then created the method of detecting the MIC on the basic of the recombinant MTB stain that expresses FFLuc.2.The mice were infected with the recombinant MTB stain that expresses FFLuc by the tail vein injection and inhalation of aerosols.The mice were infected with the recombinant MTB stain that expresses iRFP by subcutaneous injection.Then we could detect the luminance of the isolated organs and the infected mice in vivo with the IVIS spectrum Results 1.We applied the recombinant MTB strain that expresses FFLuc to establish MIC detection method and the results were the same with the MABA method.2.We found the bioluminescent signal in belly and chest of the mice that were infected by injecting the recombinant MTB strain that expresses firefly luciferase encoded proteininto the veins of tails.We also found bioluminescent signal in the isolated lung and the spleen.3.We did not find the fluorescence signal in the injection site of the mice that were infected by subcutaneous injection with the recombinant MTB strain that expresses iRFP encoded protein.Conclusions 1.The recombinant MTB strain that expresses FFLuc could be used to evaluate the pharmacodynamics of anti-tuberculosis drugs in vitro rapidly.2.Evaluating the pharmacodynamics of anti-tuberculosis drugs in vivo using the recombinant MTB strain labled with FFLuc is feasible,but still needs further study.