Novel insights into the connection between peptidoglycan recycling and multidrug resistance in Pseudomonas aeruginosa

Abstract

Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen and a frequent cause of nosocomial infection with multidrug-resistant bacteria. It was classified as one of the three pathogens with the highest priority for the development of novel antibiotics by the World Health Organization. The high resistance of Pseudomonas aeruginosa is mainly caused by three mechanisms: Low permeability of the outer membrane, inactivation of antibiotics by enzymes like the β-lactamase AmpC and the expression of efflux pumps. In two studies, the aim was to identify targets contributing to the first and the second mechanism. In the first study, a transposon library was generated in the multidrug-resistant isolate ID40, which is highly resistant to β-lactam antibiotics due to an overproduction of AmpC. The transposon library was grown in presence of cefepime or meropenem at the breakpoint concentration and then mutants with restored sensitivity were identified by transposon-directed insertion sequencing. Besides a lot of known resistance genes, we identified three genes involved in peptidoglycan recycling as well as a gene with unknown function as most promising candidates, since they were found to be necessary for growth in both the presence of cefepime or meropenem. Deletion of these genes led to strongly reduced ampC expression, β-lactamase activity and consequently to restored sensitivity against several β-antibiotics. All four candidates are promising targets for adjuvants for therapy in combination with β-lactam antibiotics in multidrug-resistant Pseudomonas aeruginosa strains. In the second study, we investigated the impact of proteins promoting the insertion of outer membrane proteins into the outer membrane. Deprivation of the periplasmic chaperone SurA resulted in a drastically altered outer membrane protein composition, impaired virulence and enhanced sensitivity to various antibiotics. SurA could therefore serve as a target to reduce virulence of Pseudomonas aeruginosa and to restore antibiotic sensitivity in multidrug-resistant strains.