Abstract
Cultures | Thresholds cycles PCR | |
7th- day incubation | 14th-day incubation | |
H37Rv growing culture | 15,2 | 14,3 |
Dormant culture H37Rv after rifampicin processing | 22,4 | 23,0 |
Dormant H37Rv with supernatant from growing culture | 18,6 | 14,5 |
H37Rv+rpfB, 6 mcg | 23,2 | 17,8 |
H37Rv+ rpfB, 12 mcg | 23,7 | 19,1 |
Added mycobacteriophage D29 | 24,0 | 23,1 |
Introduction: The problem of dormant populations of Mycobacterium tuberculosis and their reactivation is one of the key issues in the development of more effective means of treating tuberculosis.
Methods: A dormant culture of MTB of the virulent strain H37Rv with OD 0,5 was created by 3-day incubation in the presence of rifampicin (5 μg/ml) at 370С. The culture was washed from rifampicin and cultured in a 48-well plate in a medium Middlebrook 7H9 with CaCl2 2 mM volume of 600 μl. The experimental wells of the plate were added with growing culture supernatant H37Rv, 6 μg,12 μg of RPF B protein. Cell activation was determined on days 7 and 14 by adding 50 μl of mycobacteriophage D29 (108 pfu/ml) to the test wells. After 24 h incubation, the DNA of the mycobacteriophage was isolated and RT-PCR was performed in comparison with the DNA of the added mycobacteriophage D29. Aims and objectives. Develop a rapid method for determining the reactivation of dormant Mycobacterium tuberculosis. Results are shown in the table. A significant decrease in the threshold fluorescence cycles means a highly significant increase in the multiplication of mycobacteriophage which corresponding increase in the metabolism of mycobacteria. Conclusions. The presented method can be used to accelerate the determination of the specific metabolic activity of Mycobacterium tuberculosis.
Footnotes
Cite this article as: European Respiratory Journal 2021; 58: Suppl. 65, PA671.
This abstract was presented at the 2021 ERS International Congress, in session “Prediction of exacerbations in patients with COPD”.
This is an ERS International Congress abstract. No full-text version is available. Further material to accompany this abstract may be available at www.ers-education.org (ERS member access only).
- Copyright ©the authors 2021