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Effect of residual doxycycline concentrations on resistance selection and transfer in porcine commensal Escherichia coli
Authors:Laura E.J. Peeters  Siska Croubels  Geertrui Rasschaert  Hein Imberechts  Els Daeseleire  Jeroen Dewulf  Marc Heyndrickx  Patrick Butaye  Freddy Haesebrouck  Annemieke Smet
Abstract:Pig feed may contain various levels of antimicrobial residues due to cross-contamination. A previous study showed that a 3% carry-over level of doxycycline (DOX) in the feed results in porcine faecal concentrations of approximately 4?mg/L.The aim of this study was to determine the effect of residual DOX concentrations (1 and 4?mg/L) in vitro on selection of DOX–resistant porcine commensal Escherichia coli and transfer of their resistance plasmids.Three different DOX–resistant porcine commensal E. coli strains and their plasmids were characterised. These strains were each brought in competition with a susceptible strain in a medium containing 0, 1 and 4?mg/L DOX. Resistant bacteria, susceptible bacteria and transconjugants were enumerated after 24?h and 48?h.The tet(A)–carrying plasmids showed genetic backbones that are also present among human E. coli isolates. Ratios of resistant to susceptible bacteria were significantly higher at 1 and 4?mg/L DOX compared with the blank control, but there was no significant difference between 1 and 4?mg/L. Plasmid transfer frequencies were affected by 1 or 4?mg/L DOX in the medium for only one of the resistance plasmids.In conclusion, DOX concentrations of 1 and 4?mg/L can select for resistant E. coli in vitro. Further research is needed to determine the effect of these concentrations in the complex environment of the porcine intestinal microbiota.
Keywords:Doxycycline  Residues  Resistance selection  Plasmid transfer  CFU  colony forming units  DOX  doxycycline  LB  Miller's LB broth  LC-MS/MS  iquid chromatography—tandem mass spectrometry  MC  MacConkey n°3 agar  MIC  minimum inhibitory concentration  RIF  rifampicin  R/S  ratio of resistant to susceptible bacteria  SD  standard deviation
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