Experimental evolution of ceftazidime resistance in MG1655 with and without TEM-1 on a plasmid or inserted into the chromosome.

Plasmids are thought to play a key role in bacterial evolution by acting as vehicles for horizontal gene transfer, but the role of plasmids as catalysts of gene evolution remains unexplored. We challenged populations of E. coli carrying blaTEM-1 ß-lactamase gene on either the chromosome or a multicopy plasmid (19/cell) with increasing concentrations of ceftazidime. The plasmid accelerated resistance evolution by increasing the rate of appearance of novel TEM-1 mutations conferring resistance to ceftazidime, and then by amplifying the effect of TEM-1 mutations due to increased gene dosage. Crucially, this dual effect was necessary and sufficient for the evolution of clinically relevant levels of resistance. Subsequent evolution occurred by mutations in a regulatory RNA that increased plasmid copy number, resulting in marginal gains in ceftazidime resistance. These results uncover a new role for multicopy plasmids as catalysts for the evolution of antibiotic resistance in bacteria.