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JAC Advance Access published online on February 13, 2006
Journal of Antimicrobial Chemotherapy, doi:10.1093/jac/dkl033
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© The Author 2006. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org
Received December 6, 2005
Revised January 20, 2006
Accepted January 24, 2006

Original article

Heterologous expression of glycopeptide resistance vanHAX gene clusters from soil bacteria in Enterococcus faecalis

Henrik Hasman 1 *, Frank M. Aarestrup 1, Anders Dalsgaard 2, and Luca Guardabassi 2

1 Danish Institute for Food and Veterinary Research, Bülowsvej 27, 1790 Copenhagen V, Denmark
2 Department of Veterinary Pathobiology, The Royal Veterinary and Agricultural University, Stigbøjlen 4, 1870 Frederiksberg C, Denmark

* To whom correspondence should be addressed.
Henrik Hasman, E-mail: hha{at}dfvf.dk


  Abstract

Objectives: The aim of the study was to determine whether glycopeptide resistance gene clusters from soil bacteria could be heterologously expressed in Enterococcus faecalis and adapt to the new host following exposure to vancomycin.

Methods: The vanHAX clusters from Paenibacillus thiaminolyticus PT-2B1, Paenibacillus apiarius PA-B2B and Amycolatopsis coloradensis DSM 44225 were separately cloned in an appropriately constructed shuttle vector containing the two-component regulatory system (vanRS) of Tn1546. The complete vanAPT operon (vanRSHAXY) from P. thiaminolyticus PT-2B1 was cloned in the same shuttle vector lacking enterococcal vanRS. All plasmid constructs were electroporated into E. faecalis JH2-2 and the MICs of vancomycin and teicoplanin were determined for each recombinant strain before and following exposure to sublethal concentrations of vancomycin.

Results: The vanHAX clusters from P. thiaminolyticus and P. apiarius conferred high-level vancomycin resistance (MIC ≥ 125 mg/L) in E. faecalis JH2-2. In contrast, cloning of the vanHAX cluster from A. coloradensis did not result in a significant increase of vancomycin resistance (MIC = 0.7 mg/L). Resistance to vancomycin was not observed after cloning the complete vanAPT operon from P. thiaminolyticus (MIC = 2 mg/L), but this recombinant rapidly adapted to high concentrations of vancomycin (MIC = 500 mg/L) following exposure to sub-lethal concentrations of this antibiotic.

Conclusion: The results showed that vanAPT in P. thiaminolyticus is a possible ancestor of vanA-mediated glycopeptide resistance in enterococci. Experimental evidence supported the hypothesis that enterococci did not acquire glycopeptide resistance directly from glycopeptide-producing organisms such as A. coloradensis.

Keywords: vanA; Amycolatopsis coloradensis; Paenibacillus.
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