Studies aimed at improving the properties of conjugative pCURE plasmids

Abstract

In recent years, the escalating issue of microbial antibiotic resistance has become a global concern. To reduce this challenge and tackle this problem, we need to act quickly by finding and developing new ways of treating infections different from the usual antibiotics. One promising strategy involves addressing multidrug-resistant infections by displacing the resistance plasmid from bacteria exhibiting resistance. This PhD thesis focuses on the concept of antimicrobial resistance plasmid curing (displacement) through incompatibility, aiming to improve the technology for displacing antibiotic resistance plasmids from bacterial strains. The primary objective was to make a derivative of plasmid pUB307, the basis for the curing plasmid pCURE-F-307, that is resistant to suppression of its conjugative transfer (Fertility Inhibition) by competitor plasmids. Initial results showed that when traG, the target gene for the inhibition, was provided on a multi-copy plasmid, Fertility Inhibition was reduced. Based on this finding traG was deleted from pUB307 and reinserted at a new location with its own strong promoter. The new plasmid was able to transfer much more efficiently without suppression by other plasmids. This solution is more general than isolating point mutations in traG because not all Fertility Inhibition proteins interact with TraG in the same way. This progress could help to increase efficiency of plasmid curing strategies and thus help reduce the spread of antimicrobial resistance plasmids.