Exogenous and Endogenous Phosphoethanolamine Transferases Differently Affect Colistin Resistance and Fitness in Pseudomonas aeruginosa

Abstract

Colistin represents a last-line treatment option for infections caused by multidrug resistant Gram-negative pathogens, including Pseudomonas aeruginosa . Colistin resistance generally involves the modification of the lipid A moiety of lipopolysaccharide (LPS) with positively charged molecules, namely phosphoethanolamine (PEtN) or 4-amino-4-deoxy- L -arabinose (Ara4N), that reduce colistin affinity for its target. Several lines of evidence highlighted lipid A aminoarabinosylation as the primary colistin resistance mechanism in P. aeruginosa , while the contribution of phosphoethanolamination remains elusive. PEtN modification can be due to either endogenous (chromosomally encoded) PEtN transferase(s) (e.g., EptA in P. aeruginosa ) or plasmid borne MCR enzymes, commonly found in enterobacteria. By individually cloning eptA and mcr-1 into a plasmid for inducible gene expression, we demonstrated that MCR-1 and EptA have comparable PEtN transferase activity in P. aeruginosa and confer colistin resistance levels similar to those provided by lipid A aminoarabinosylation. Notably, EptA, but not MCR-1, negatively affects P. aeruginosa growth and, to a lesser extent, cell envelope integrity when expressed at high levels. Mutagenesis experiments revealed that PEtN transferase activity does not account for the noxious effects of EptA overexpression, that instead requires a C-terminal tail unique to P. aeruginosa EptA, whose function remains unknown. Overall, this study shows that both endogenous and exogenous PEtN transferases can promote colistin resistance in P. aeruginosa , and that PEtN and MCR-1 mediated resistance has no impact on growth and cell envelope homeostasis, suggesting that there may be no fitness barriers to the spread of mcr-1 in P. aeruginosa .

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