Analysis of the expression of <i>hns</i> and Sm<i>qnr25</i>, and copy number of the pBS3.25 plasmid upon experimental evolution.

<p>Panel A, analysis of expression of <i>hns</i> and Sm<i>qnr</i>. Black bars, <i>hns</i> expression; gray bars, Sm<i>qnr25</i> expression. Despite pBS3.25 being stabilized in the <i>hns</i> defective mutant, the expression of this gene did not change in the evolved strains, indicating that plasmid stabilization in these isolates is not due to reduced <i>hns</i> expression. The copy number of values were normalized taking the value for MBS231 as 1. MBS228, MBS229, MBS230 and MBS231 are the evolved strains and MBS25 the original, non-evolved strain. Panel B. Copy number of pBS3.25 after experimental evolution. White bars, copy number of pBS3.25 plasmid in the evolved strains and in the overall MBS25 population. The large deviation observed in the full MBS25 population reflects that it is formed by two subpopulations, one (small colonies) harboring pBS3.25 in high copy numbers, and another (larger colonies) in which the plasmid copy number is low. The copy number of pBS3.25 in the small colonies is shown with a gray bar; the copy number of pBS2.35 in the colonies of normal size is shown with a black bar. As shown, bacteria evolved to reduce the plasmid copy number and hence express low levels of Sm<i>qnr</i>. This reduction is evident even in the original population, which has small colonies with plasmids in high copy number and normal size colonies with low copy numbers of pBS3.25. Values were normalized using the values for the normal-sized colonies of MBS25 (black bar) as 1.