title: A nanomechanical study of the effects of colistin on the Klebsiella pneumoniae AJ218 capsule.
abstract: Atomic force microscopy measurements of capsule thickness revealed that that the wild-type Klebsiella pneumoniae AJ218 capsular polysaccharides were rearranged by exposure to colistin. The increase in capsule thickness measured near minimum inhibitory/bactericidal concentration (MIC/MBC) is consistent with the idea that colistin displaces the divalent cations that cross-bridge adjacent lipopolysaccharide (LPS) molecules through the capsule network. Cryo-electron microscopy demonstrated that the measured capsule thickness at near MIC/MBC of 1.2 μM was inflated by the disrupted outer membrane, through which the capsule is excreted and LPS is bound. Since wild-type and capsule-deficient strains of K. pneumoniae AJ218 have equivalent MICs and MBCs, the presence of the capsule appeared to confer no protection against colistin in AJ218. A spontaneously arising colistin mutant showed a tenfold increase in resistance to colistin; genetic analysis identified a single amino acid substitution (Q95P) in the PmrB sensor kinase in this colistin-resistant K. pneumoniae AJ218. Modification of the lipid A component of the LPS could result in a reduction of the net-negative charge of the outer membrane, which could hinder binding of colistin to the outer membrane and displacement of the divalent cations that bridge adjacent LPS molecules throughout the capsular polysaccharide network. Retention of the cross-linking divalent cations may explain why measurements of capsule thickness did not change significantly in the colistin-resistant strain after colistin exposure. These results contrast with those for other K. pneumoniae strains that suggest that the capsule confers colistin resistance.
authors.affiliation: School of Chemistry, University of Melbourne, Melbourne, VIC, 3010, Australia. mularski.a@unimelb.edu.au.
authors.forename: Anna
authors.identifier: http://orcid.org/0000-0003-1140-146X
authors.initials: A
authors.lastname: Mularski
authors.affiliation: Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, VIC, 3010, Australia.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Wilksch
authors.affiliation: Advanced Microscopy Unit, Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Melbourne, VIC, 3010, Australia.
authors.forename: Eric
authors.initials: E
authors.lastname: Hanssen
authors.affiliation: Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, 381 Royal Parade, Parkville, VIC, 3052, Australia.
authors.forename: Jian
authors.initials: J
authors.lastname: Li
authors.affiliation: Microbiological Diagnostic Unit Public Health Laboratory, The Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, VIC, 3010, Australia.
authors.forename: Takehiro
authors.initials: T
authors.lastname: Tomita
authors.affiliation: Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, VIC, 3010, Australia.
authors.forename: Sacha James
authors.initials: SJ
authors.lastname: Pidot
authors.affiliation: Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, VIC, 3010, Australia.
authors.forename: Tim
authors.initials: T
authors.lastname: Stinear
authors.affiliation: School of Chemistry, University of Melbourne, Melbourne, VIC, 3010, Australia.
authors.forename: Frances
authors.initials: F
authors.lastname: Separovic
authors.affiliation: Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, VIC, 3010, Australia.
authors.forename: Dick
authors.initials: D
authors.lastname: Strugnell
mesh_terms: D016667:Bacterial Capsules / Q000187:drug effects* / Q000378:metabolism*; D001696:Biomechanical Phenomena / Q000187:drug effects; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D003091:Colistin / Q000494:pharmacology*; D023281:Genomics; D007711:Klebsiella pneumoniae / Q000166:cytology / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D055595:Mechanical Phenomena / Q000187:drug effects*; D036103:Nanotechnology; D011135:Polysaccharides, Bacterial / Q000378:metabolism
chemical_list: D011135:Polysaccharides, Bacterial; D003091:Colistin
keywords: Antimicrobial peptide; Atomic force microscopy; Capsular polysaccharide; Colistin; Klebsiella pneumoniae; Polymyxin