Effect of growth rate and selection pressure on rates of transfer of an antibiotic resistance plasmid between E. coli strains Jasper M. Schuurmans a , Sacha A.F.T. van Hijum b,c , Jurgen R. Piet d , Nadine Händel a , Jan Smelt a , Stanley Brul a , Benno H. ter Kuile a,e,⇑ a Department of Molecular Biology & Microbial Food Safety, Swammerdam Institute of Life Sciences, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands b NIZO Food Research B.V., Kernhemseweg 2, 6718 ZB Ede, The Netherlands c Centre for Molecular and Biomolecular Informatics (CMBI), NCMLS, Radboud University Medical Centre, Nijmegen, The Netherlands d Department of Medical Microbiology, Center of Infection and Immunity Amsterdam (CINIMA), Academic Medical Center, P.O. Box 22660, 1100 DD Amsterdam, The Netherlands e Office for Risk Assessment and Research, Netherlands Food and Consumer Product Safety Authority, Catharijnesingel 59, 3511 GG Utrecht, The Netherlands article info Article history: Received 16 October 2013 Accepted 30 January 2014 Available online 10 February 2014 Communicated by Dr. J. Casadesus Keywords: Plasmid transfer Selection regimen Tetracycline abstract Antibiotic resistance increases costs for health care and causes therapy failure. An impor- tant mechanism for spreading resistance is transfer of plasmids containing resistance genes and subsequent selection. Yet the factors that influence the rate of transfer are poorly known. Rates of plasmid transfer were measured in co-cultures in chemostats of a donor and a acceptor strain under various selective pressures. To document whether specific mutations in either plasmid or acceptor genome are associated with the plasmid transfer, whole genome sequencing was performed. The DM0133 TetR tetracycline resistance plas- mid was transferred between Escherichia coli K-12 strains during co-culture at frequencies that seemed higher at increased growth rate. Modeling of the take-over of the culture by the transformed strain suggests that in reality more transfer events occurred at low growth rates. At moderate selection pressure due to an antibiotic concentration that still allowed growth, a maximum transfer frequency was determined of once per 10 11 cell divisions. In the absence of tetracycline or in the presence of high concentrations the frequency of transfer was sometimes zero, but otherwise reduced by at least a factor of 5. Whole gen- ome sequencing showed that the plasmid was transferred without mutations, but two functional mutations in the genome of the recipient strain accompanied this transfer. Exposure to concentrations of antibiotics that fall within the mutant selection window stimulated transfer of the resistance plasmid most. Ó 2014 Elsevier Inc. All rights reserved. 1. Introduction Because of therapy failure, the costs for human health- care of antibiotic resistance of pathogens are immense (Kock et al., 2010; Maragakis et al., 2008). Antibiotic resis- tance can be brought about by three main mechanisms: physiological adaptation, genetic mutations and transfer of genes coding for resistance from resistant bacteria to sensitive ones. The latter one is responsible for most of the resistance found in human pathogens. In the ultra-long time scale resistance genes mostly originate from the envi- ronment (Martinez, 2009) and may be spread especially by exposure to sublethal concentrations of antibiotics http://dx.doi.org/10.1016/j.plasmid.2014.01.002 0147-619X/Ó 2014 Elsevier Inc. All rights reserved. ⇑ Corresponding author at: Department of Molecular Biology & Micro- bial Food Safety, Swammerdam Institute of Life Sciences, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands. E-mail addresses: B.H.terKuile@uva.nl, benno.ter.kuile@vwa.nl (B.H. ter Kuile). Plasmid 72 (2014) 1–8 Contents lists available at ScienceDirect Plasmid journal homepage: www.elsevier.com/locate/yplas