title: The metabolically active subpopulation in Pseudomonas aeruginosa biofilms survives exposure to membrane-targeting antimicrobials via distinct molecular mechanisms.
abstract: Biofilms are reported to be inherently refractory toward antimicrobial attack and, therefore, cause problems in industrial and medical settings. Pseudomonas aeruginosa biofilms contain subpopulations that exhibit high metabolic activity and subpopulations that exhibit low metabolic activity. We have found that membrane-targeting antimicrobials such as colistin, EDTA, SDS, and chlorhexidine specifically kill the inactive subpopulation in P. aeruginosa biofilms, whereas the active subpopulation survives exposure to these compounds. Because treatment of P. aeruginosa biofilms with the membrane-targeting compounds colistin, EDTA, SDS, and chlorhexidine resulted in the same spatial distribution of live and dead bacteria, we investigated whether tolerance to these compounds originated from the same molecular mechanisms. Development of colistin-tolerant subpopulations was found to depend on the pmr genes encoding lipopolysaccharide modification enzymes, as well as on the mexAB-oprM, mexCD-oprJ, and muxABC-opmB genes encoding antimicrobial efflux pumps, but does not depend on the mexPQ-opmE efflux pump genes. Development of chlorhexidine-tolerant subpopulations was found to depend on the mexCD-oprJ genes, but does not depend on the pmr, mexAB-oprM, mexPQ-opmE, or muxABC-opmB genes. Tolerance to SDS and EDTA in P. aeruginosa biofilms is linked to metabolically active cells, but does not depend on the pmr, mexAB, mexCD, mexPQ, or muxABC genes. Our data suggest that the active subpopulation in P. aeruginosa biofilms is able to adapt to exposure to membrane-targeting agents through the use of different genetic determinants, dependent on the specific membrane-targeting compound.
authors.affiliation: Department of International Health, Immunology and Microbiology, Faculty of Health Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Wen-Chi
authors.initials: WC
authors.lastname: Chiang
authors.forename: Sünje Johanna
authors.initials: SJ
authors.lastname: Pamp
authors.forename: Martin
authors.initials: M
authors.lastname: Nilsson
authors.forename: Michael
authors.initials: M
authors.lastname: Givskov
authors.forename: Tim
authors.initials: T
authors.lastname: Tolker-Nielsen
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D018441:Biofilms / Q000187:drug effects*; D002462:Cell Membrane / Q000187:drug effects*; D006801:Humans; D050296:Microbial Viability / Q000187:drug effects*; D011550:Pseudomonas aeruginosa / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism* / Q000502:physiology
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins