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Journal of Bacteriology, September 2006, p. 6506-6514, Vol. 188, No. 18
0021-9193/06/$08.00+0     doi:10.1128/JB.00375-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Functional Characterization of IS1999, an IS4 Family Element Involved in Mobilization and Expression of ß-Lactam Resistance Genes

Daniel Aubert, Thierry Naas,* Claire Héritier, Laurent Poirel, and Patrice Nordmann

Service de Bactériologie-Virologie, Hôpital de Bicêtre, Assistance Publique/Hôpitaux de Paris, Faculté de Médecine Paris-Sud, 94275 K.-Bicêtre, France

Received 16 March 2006/ Accepted 4 July 2006

IS1999 and a point mutant derivative, IS1999.2, have been described inserted upstream of emerging antibiotic resistance genes blaVEB-1 and blaOXA-48. 5' Rapid amplification of cDNA ends experiments revealed that expression of these ß-lactamase genes was driven by the outward-directed promoter, Pout, located in the IS1999 elements. These findings led us to study IS1999-mediated gene mobilization. Thus, the transposition properties of IS1999 and of IS1999-based composite transposons, made of two copies of IS1999 in different orientations, were investigated. IS1999 or IS1999-based composite transposons were capable of transposing onto the conjugative plasmid pOX38-Gen. Sequence analysis of the insertion sites revealed that IS1999 inserted preferentially into DNA targets containing the consensus sequence NGCNNNGCN. Transposition was more efficient when at least one left inverted repeat end was located at an outside end of the transposon. The transposition frequency of IS1999.2 was 10-fold lower than that of IS1999, and transposition frequencies of the putative natural transposon, Tn1999, were below detection limits of our transposition assay. This reduced transposition frequency of IS1999.2-based elements may result from a lower transcription of the transposase gene, as revealed by reverse transcription-PCR analyses.

* Corresponding author. Mailing address: Service de Bactériologie-Virologie, Hôpital de Bicêtre, 78 rue du Général Leclerc, 94275 Le Kremlin-Bicêtre Cedex, France. Phone: 33-1-45-21-29-86. Fax: 33-1-45-21-63-40. E-mail: thierry.naas{at}bct.ap-hop-paris.fr.

Journal of Bacteriology, September 2006, p. 6506-6514, Vol. 188, No. 18
0021-9193/06/$08.00+0     doi:10.1128/JB.00375-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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