Genomic analysis reveals variation between Mycobacterium tuberculosis H37Rv and the attenuated M. tuberculosis H37Ra strain

Mycobacterium tuberculosis H37Ra is an attenuated tubercle bacillus closely related to the virulent type strain M. tuberculosis H37Rv. Despite extensi ve study, the reason for the decreased virulence of M. tuberculosis H37Ra h as not seen determined, A genomic approach was therefore initiated to ident ify genetic differences between M. tuberculosis H37Rv and M. tuberculosis H 37Ra as a means of pinpointing the attenuating mutation(s). Digestion with the rare-cutting restriction endonuclease DraI revealed two polymorphisms b etween the strains: a 480-kb fragment in M. tuberculosis H37Rv was replaced by two fragments of 220 and 260 kb in M. tuberculosis H37Ra, while there w as a similar to 7.9-kb DraI fragment in M. tuberculosis H37Ra that had no c ounterpart in M. tuberculosis H37Rv, As the M. tuberculosis insertion seque nce IS6110 contains a single DraI restriction site, it was considered possi ble that these polymorphisms were the result of IS6110 transposition events in M. tuberculosis H37Ra, events that may have inactivated virulence genes . The 7.9-kb polymorphism was found to be due to the presence of the previo usly described H37Rv RvD2 deletion in M. tuberculosis H37Ra,,vith sequence analysis suggesting an IS110-mediated deletion mechanism for loss of RvD2. Three other TSdllO-catalyaed deletions from the M. tuberculosis H37Rv chrom osome (RvD3 to RvD5) were also identified, suggesting that this mechanism p lays an important role in genome plasticity in the tubercle bacilli. Compar ative mapping and sequencing revealed that the 480-kb polymorphism was due to an IS6110 insertion in M. tuberculosis H37Ra near oriC, Complementation of M. tuberculosis H37Ra with a 2.9-kb restriction fragment from M. tubercu losis H37Rv that encompassed the IS6110 insertion dig not increase the surv ival of recombinant M. tuberculosis H37Ra in mice. In conclusion, this stud y describes the presence and mechanisms of genomic variation between M. tub erculosis H37Ra and 171. tuberculosis H37Rv, although the role that they pl ay in the attenuation of M, tuberculosis H37Ra is unclear.