title: Phosphoethanolamine modification of lipid A in colistin-resistant variants of Acinetobacter baumannii mediated by the pmrAB two-component regulatory system.
abstract: Colistin resistance is rare in Acinetobacter baumannii, and little is known about its mechanism. We investigated the role of PmrCAB in this trait, using (i) resistant and susceptible clinical strains, (ii) laboratory-selected mutants of the type strain ATCC 19606 and of the clinical isolate ABRIM, and (iii) a susceptible/resistant pair of isogenic clinical isolates, Ab15/133 and Ab15/132, isolated from the same patient. pmrAB sequences in all the colistin-susceptible isolates were identical to reference sequences, whereas resistant clinical isolates harbored one or two amino acid replacements variously located in PmrB. Single substitutions in PmrB were also found in resistant mutants of strains ATCC 19606 and ABRIM and in the resistant clinical isolate Ab15/132. No mutations in PmrA or PmrC were found. Reverse transcriptase (RT)-PCR identified increased expression of pmrA (4- to 13-fold), pmrB (2- to 7-fold), and pmrC (1- to 3-fold) in resistant versus susceptible organisms. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry showed the addition of phosphoethanolamine to the hepta-acylated form of lipid A in the resistant variants and in strain ATCC 19606 grown under low-Mg(2+) induction conditions. pmrB gene knockout mutants of the colistin-resistant ATCC 19606 derivative showed >100-fold increased susceptibility to colistin and 5-fold decreased expression of pmrC; they also lacked the addition of phosphoethanolamine to lipid A. We conclude that the development of a moderate level of colistin resistance in A. baumannii requires distinct genetic events, including (i) at least one point mutation in pmrB, (ii) upregulation of pmrAB, and (iii) expression of pmrC, which lead to addition of phosphoethanolamine to lipid A.
authors.affiliation: Health Protection Agency, Microbiology Services-Colindale, London, United Kingdom.
authors.forename: Alejandro
authors.initials: A
authors.lastname: Beceiro
authors.forename: Enrique
authors.initials: E
authors.lastname: Llobet
authors.forename: Jesús
authors.initials: J
authors.lastname: Aranda
authors.forename: José Antonio
authors.initials: JA
authors.lastname: Bengoechea
authors.forename: Michel
authors.initials: M
authors.lastname: Doumith
authors.forename: Michael
authors.initials: M
authors.lastname: Hornsey
authors.forename: Hiran
authors.initials: H
authors.lastname: Dhanji
authors.forename: Henrik
authors.initials: H
authors.lastname: Chart
authors.forename: Germán
authors.initials: G
authors.lastname: Bou
authors.forename: David M
authors.initials: DM
authors.lastname: Livermore
authors.forename: Neil
authors.initials: N
authors.lastname: Woodford
mesh_terms: D040981:Acinetobacter baumannii / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D003091:Colistin / Q000494:pharmacology*; D024881:Drug Resistance, Bacterial / Q000235:genetics; D004983:Ethanolamines / Q000378:metabolism*; D008050:Lipid A / Q000378:metabolism*; D008969:Molecular Sequence Data; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D004983:Ethanolamines; D008050:Lipid A; C005448:phosphorylethanolamine; D003091:Colistin