title: Antimicrobial synergy between carprofen and doxycycline against methicillin-resistant Staphylococcus pseudintermedius ST71.
abstract: BACKGROUND
  New therapeutic strategies are needed to face the rapid spread of multidrug-resistant staphylococci in veterinary medicine. The objective of this study was to identify synergies between antimicrobial and non-antimicrobial drugs commonly used in companion animals as a possible strategy to restore antimicrobial susceptibility in methicillin-resistant Staphylococcus pseudintermedius (MRSP).
  
  
  RESULTS
  A total of 216 antimicrobial/non-antimicrobial drug combinations were screened by disk diffusion using a clinical MRSP sequence type (ST) 71 strain resistant to all six antimicrobials tested (ampicillin, ciprofloxacin, clindamycin, doxycycline, oxacillin and trimethoprim/sulfamethoxazole). The most promising drug combination (doxycycline-carprofen) was further assessed by checkerboard testing extended to four additional MRSP strains belonging to ST71 or ST68, and by growth inhibition experiments. Seven non-antimicrobial drugs (bromhexine, acepromazine, amitriptyline, clomipramine, carprofen, fluoxetine and ketoconazole) displayed minimum inhibitory concentrations (MICs) ranging between 32 and >4096 mg/L, and enhanced antimicrobial activity of one or more antimicrobials. Secondary screening by checkerboard assay revealed a synergistic antimicrobial effect between carprofen and doxycycline, with the sum of the fractional inhibitory concentration indexes (ΣFICI) ranging between 0.3 and 0.5 depending on drug concentration. Checkerboard testing of multiple MRSP strains revealed a clear association between synergy and carriage of tetK, which is a typical feature of MRSP ST71. An increased growth inhibition was observed when MRSP ST71 cells in exponential phase were exposed to 0.5/32 mg/L of doxycycline/carprofen compared to individual drug exposure.
  
  
  CONCLUSIONS
  Carprofen restores in vitro susceptibility to doxycycline in S. pseudintermedius strains carrying tetK such as MRSP ST71. Further research is warranted to elucidate the molecular mechanism behind the identified synergy and its linkage to tetK.
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Stigbøjlen 4, 1870, Frederiksberg C, Denmark.
authors.forename: Rikke Prejh
authors.initials: RP
authors.lastname: Brochmann
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Stigbøjlen 4, 1870, Frederiksberg C, Denmark.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Helmfrid
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Stigbøjlen 4, 1870, Frederiksberg C, Denmark.
authors.forename: Bimal
authors.initials: B
authors.lastname: Jana
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Stigbøjlen 4, 1870, Frederiksberg C, Denmark.
authors.forename: Zofia
authors.initials: Z
authors.lastname: Magnowska
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Stigbøjlen 4, 1870, Frederiksberg C, Denmark. lguardabassi@rossvet.edu.kn.|Department of Biomedical Sciences, Ross School of Veterinary Medicine, Basseterre, St Kitts, West Indies. lguardabassi@rossvet.edu.kn.
authors.forename: Luca
authors.initials: L
authors.lastname: Guardabassi
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002227:Carbazoles / Q000494:pharmacology*; D052978:Disk Diffusion Antimicrobial Tests; D004318:Doxycycline / Q000494:pharmacology*; D004357:Drug Synergism; D016106:Methicillin Resistance / Q000187:drug effects*; D013210:Staphylococcus / Q000187:drug effects* / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D002227:Carbazoles; C007005:carprofen; D004318:Doxycycline
keywords: Dogs; Multidrug resistance; Non-steroidal anti-inflammatory drugs; Veterinary antimicrobial therapy