New insights into the transposition mechanisms of IS6110 and its dynamic distribution between Mycobacterium tuberculosis Complex lineages
Fig 5
Transposition in the laboratory and in a mouse infection model using reference M. bovis and M. tuberculosis strains.
(a) Experimental model to measure transposition rates in M. tuberculosis H37Rv (15 IS6110 copies) and M. bovis AF2122/97 (1 IS6110 copy). Both strains are transformed with pIR-Km and used to inoculate liquid media or to intranasally infect C57BL/6 mice. After the indicated time points aliquots are plated in kanamycin and sucrose containing plates to ensure pIR-km loss and to recover colonies resulting from transposition. (b) Transposition frequencies in laboratory medium in M. bovis and M. tuberculosis are indicated by red and blue columns respectively. Error bars indicate the standard deviation of the mean value from three independent cultures. Transposition preferentially occurs in M. tuberculosis after the stationary phase reaching it maximum in the starvation period. (c) Expression of M. tuberculosis IS6110 during exponential, stationary and starvation periods in vitro. Expression of ORF1 and ORF2 are indicated by dark and light blue columns respectively. Results are from three independent cultures. (d) Transposition frequencies during mouse infection with M. bovis or M. tuberculosis are indicated by red and blue columns respectively. Data from lung and spleen are shown and error bars indicate the standard deviation of the mean value from three independent mice. M. bovis does not exhibit increased transposition rates in vivo relative to liquid culture. Conversely M. tuberculosis show 10-fold higher transposition rates compared to exponential growth in vitro. In all cases, transposition frequencies were calculated relative to the total number of CFU in either cultures or mouse organs.