title: Synergistic killing of NDM-producing MDR Klebsiella pneumoniae by two 'old' antibiotics-polymyxin B and chloramphenicol.
abstract: OBJECTIVE
  Combination therapy is an important option in the fight against Gram-negative 'superbugs'. This study systematically investigated bacterial killing and the emergence of polymyxin resistance with polymyxin B and chloramphenicol combinations used against New Delhi metallo-β-lactamase (NDM)-producing MDR Klebsiella pneumoniae.
  
  
  METHODS
  Four NDM-producing K. pneumoniae strains were employed. The presence of genes conferring resistance to chloramphenicol was examined by PCR. Time-kill studies (inocula ∼10(6) cfu/mL) were conducted using various clinically achievable concentrations of each antibiotic (range: polymyxin B, 0.5-2 mg/L; chloramphenicol, 4-32 mg/L), with real-time population analysis profiles documented at baseline and 24 h. The microbiological response was examined using the log change method and pharmacodynamic modelling in conjunction with scanning electron microscopy (SEM).
  
  
  RESULTS
  Multiple genes coding for efflux pumps involved in chloramphenicol resistance were present in all strains. Polymyxin B monotherapy at all concentrations produced rapid bacterial killing followed by rapid regrowth with the emergence of polymyxin resistance; chloramphenicol monotherapy was largely ineffective. Combination therapy significantly delayed regrowth, with synergy observed in 25 out of 28 cases at both 6 and 24 h; at 24 h, no viable bacterial cells were detected in 15 out of 28 cases with various combinations across all strains. No polymyxin-resistant bacteria were detected with combination therapy. These results were supported by pharmacodynamic modelling. SEM revealed significant morphological changes following treatment with polymyxin B both alone and in combination.
  
  
  CONCLUSIONS
  The combination of polymyxin B and chloramphenicol used against NDM-producing MDR K. pneumoniae substantially enhanced bacterial killing and suppressed the emergence of polymyxin resistance.
authors.affiliation: Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, Melbourne, Victoria, Australia.
authors.forename: Nusaibah
authors.initials: N
authors.lastname: Abdul Rahim
authors.affiliation: Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, Melbourne, Victoria, Australia.
authors.forename: Soon-Ee
authors.initials: SE
authors.lastname: Cheah
authors.affiliation: Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, Melbourne, Victoria, Australia.
authors.forename: Matthew D
authors.initials: MD
authors.lastname: Johnson
authors.affiliation: Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, Melbourne, Victoria, Australia.
authors.forename: Heidi
authors.initials: H
authors.lastname: Yu
authors.affiliation: University of Queensland Centre for Clinical Research, Brisbane, Queensland, Australia.
authors.forename: Hanna E
authors.initials: HE
authors.lastname: Sidjabat
authors.affiliation: Department of Microbiology, Monash University, Melbourne, Victoria, Australia.
authors.forename: John
authors.initials: J
authors.lastname: Boyce
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, Brisbane, Queensland, Australia.
authors.forename: Mark S
authors.initials: MS
authors.lastname: Butler
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, Brisbane, Queensland, Australia.
authors.forename: Matthew A
authors.initials: MA
authors.lastname: Cooper
authors.affiliation: Department of Mechanical and Aerospace Engineering, Faculty of Engineering, Monash University, Clayton, Victoria, Australia.
authors.forename: Jing
authors.initials: J
authors.lastname: Fu
authors.affiliation: University of Queensland Centre for Clinical Research, Brisbane, Queensland, Australia Pathology Queensland, Royal Brisbane and Women's Hospital Campus, Brisbane, Queensland, Australia.
authors.forename: David L
authors.initials: DL
authors.lastname: Paterson
authors.affiliation: Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, Melbourne, Victoria, Australia.
authors.forename: Roger L
authors.initials: RL
authors.lastname: Nation
authors.affiliation: Centre for Medicine Use and Safety, Monash University, Melbourne, Victoria, Australia.
authors.forename: Phillip J
authors.initials: PJ
authors.lastname: Bergen
authors.affiliation: Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, Melbourne, Victoria, Australia.
authors.forename: Tony
authors.initials: T
authors.lastname: Velkov
authors.affiliation: Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, Melbourne, Victoria, Australia colistin.polymyxin@gmail.com.
authors.forename: Jian
authors.initials: J
authors.lastname: Li
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002701:Chloramphenicol / Q000494:pharmacology*; D024901:Drug Resistance, Multiple, Bacterial; D004357:Drug Synergism; D007711:Klebsiella pneumoniae / Q000187:drug effects* / Q000201:enzymology / Q000502:physiology; D050296:Microbial Viability / Q000187:drug effects*; D011112:Polymyxin B / Q000494:pharmacology*; D001618:beta-Lactamases / Q000378:metabolism
chemical_list: D000900:Anti-Bacterial Agents; D002701:Chloramphenicol; D001618:beta-Lactamases; D011112:Polymyxin B
keywords: K. pneumoniae; New Delhi metallo-β-lactamase; synergy