Mycobacterium tuberculosis medicament sensitive phenotype detection method and application of method

Abstract

The invention discloses a mycobacterium tuberculosis medicament sensitive phenotype detection method, which comprises the following steps of: 1) adding 2.5 to 6mul of 0.2 percent of resazurin dissolved by using methanol with mass percentage concentration into the inner side of a tube cover of a sterile centrifuge tube, and sterilizing the centrifuge tube after the methanol is volatilized; 2) inoculating 5 to 50mg of mycobacterium tuberculosis to be detected into a 7H9-S culture medium, regulating the concentration of the bacteria solution by adopting turbidimetry, regulating the concentration of the bacteria solution to the concentration of a Mcburney turbidimetric 1 tube by using sterile water or 0.9-percent physiological saline, and diluting the bacteria solution by 20 times by using the 7H9-S culture medium; 3) inoculating 50 to 150mu of diluted bacteria solution into the centrifuge tube of the step 1); 4) adding 100mul of anti-tubercular medicament solution diluted by 7H9-S and to be detected into the centrifuge tube of the step 3), wherein the concentration of the medicament to be detected is 0.01mug / ml to 2.5mg / ml; 5) after the centrifuge tube of the step 4) is covered closely, culturing the solution for 6 to 7 days at the temperature of 37 DEG C; and 6) inverting and oscillating the centrifuge tube of the step 5), and culturing the solution for 12 to 48 hours at the temperature of 37 DEG C after the solution becomes blue. Proved by experiments, in the mycobacterium tuberculosis medicament sensitive phenotype detection method and application of the mycobacterium tuberculosis medicament sensitive phenotype detection method in anti-tubercular medicament screening, the method has low workload and short detection time; compared with a traditional L-J medium medicament sensitive detection method, the accuracy reaches 90 to 99 percent; and the method has good biological safety, does not cause laboratory propagation of tuberculosis, is suitable for large-scale.

Description

technical field

[0001] The invention relates to a method for detecting drug-sensitive phenotype and its application, in particular to a method for detecting drug-sensitive phenotype of Mycobacterium tuberculosis and its application in screening anti-tuberculosis drugs. Background technique

[0002] Tuberculosis is a highly contagious disease worldwide, causing more than one million deaths each year. In recent years, with the emergence of multidrug-resistant Mycobacterium tuberculosis (MDR-TB) and extensively drug-resistant Mycobacterium tuberculosis (XDR-TB), the treatment of tuberculosis has become a great challenge (Espinal M A, Laszlo A , Simonsen L, et al. N Engl J Med. 2001, 344:1294-1303). Due to the emergence of MDR-TB and XDR-TB, tuberculosis drug susceptibility testing and drug resistance testing are particularly important, which can not only better manage and treat tuberculosis patients, reduce the spread of drug-resistant bacteria, but also greatly reduce new tub...

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