This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Plasmids are mobile genetic elements that play a central role in the dispersion of antibiotic resistance among bacterial species, thereby decreasing the effectiveness of antimicrobial agents used for the treatment of infectious diseases. The objective of our research is to discern patterns of plasmid evolution through experimental evolution studies. In particular, we aim to determine how some plasmids have evolved the ability to transfer and be maintained in a wide range of hosts. The 64.5 kb IncP-1? plasmid pB10 was chosen as a model system because of its high transferability and broad host-range. An examination of the natural host-range of this plasmid in the bacterial community of a sewage treatment plant showed that pB10 transferred to 38 distinct species. These findings also suggested that the host-range of a plasmid was not only determined by intrinsic plasmid characteristics but by the original host and the environmental parameters. From these studies we found two hosts in which plasmid pB10 presents a higher burden or is less stable than in E. coli K12. In an evolution experiment with one of these hosts, P. putida H2 (pB10), the cost of the plasmid to an isogenic H2 strain that had never carried the plasmid before (na ve), was significantly decreased after 1000 generations of plasmid evolution: from 20% to 9% or 16%, depending on whether or not the plasmid was regularly transferred between isogenic hosts. These results indicate that plasmid-encoded adaptive changes were responsible for this drastic plasmid-host adaptation, and the molecular basis is being examined. In parallel, the genome sequences of three naturally occurring IncP-1 plasmids were compared with those of known IncP-1 plasmids. In contrast two all other sequenced IncP-1? plasmids, the oriV region of plasmid pB3 is not interrupted by transposons, shedding new light on the evolutionary history of these promiscuous plasmids.
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