Resistance of Antimicrobial in Pseudomonas aeruginosa

Abstract

This thesis has been written in an article like format for two reasons. First, it is the goal that the results presented in this thesis should be part of a scientific publication about antimicrobial peptide resistance in Pseudomonas aeruginosa (P. aeruginosa). As such I saw this as an opportunity to train myself in communicating research through articles. Secondly, the compact nature of this thesis should reduce the time the readers will need to spend on reading it. I would like to thank my supervisors Anders Folkesson and Søren Molin for giving me the opportunity to work with the fascinating but complex evolution of antimicrobial peptide resistance in P. aeruginosa. I would especially like to express my gratitude to Anders Folkesson for many rewarding discussions and his exceptional guidance. I would also like to thank Lei Yang for excellent advice on DNA microarray experimental procedures, Claus Sternberg for technical support, Rune L. Jensen and Søren Damkiaer for their contributions in the experimental evolution of colistin resistance and the rest of the people from Søren Molin's group for input and ideas and for keeping me company. Abstract Cationic antimicrobial peptides (CAMPs) are attractive candidates for novel drug development due to their activity on bacteria resistant to conventional antibiotics and because it has been suggested that resistance towards CAMPs is less likely to develop compared to conventional antibiotics. Here we investigate the molecular pathways underlying resistance towards the cyclic CAMP colistin in the clinically relevant bacterium Pseudomonas aeruginosa (P. aeruginosa). We show that it is possible to experimentally evolve colistin resistance in P. aeruginosa PAO1 by a continual selection protocol and that resistance is associated with a fitness cost in terms of decreased growth rate. We compare the transcriptional profiles on four strains that evolved resistance during the experiment, two colistin sensitive controls, the ancestral P. aeruginosa PAO1 and two colistin resistant clinical isolates. We show that colistin resistance involves increased expression of the arnB operon and mutations in the genes encoding the regulators of this operon. Furthermore, we present results which show that colistin resistance requires additional mutations indicating that colistin resistance is significantly more complex compared to resistance towards conventional antibiotics.