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Antimicrobial Agents and Chemotherapy, February 2009, p. 603-608, Vol. 53, No. 2
0066-4804/09/$08.00+0     doi:10.1128/AAC.00997-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

qnrD, a Novel Gene Conferring Transferable Quinolone Resistance in Salmonella enterica Serovar Kentucky and Bovismorbificans Strains of Human Origin {triangledown}

L. M. Cavaco,1,2* H. Hasman,1 S. Xia,3 and F. M. Aarestrup1

Research Group for Antimicrobial Resistance and Molecular Epidemiology, National Food Institute, Technical University of Denmark, Copenhagen V, Denmark,1 Department of Veterinary Pathobiology, Faculty of Life Science, University of Copenhagen, 1870 Frederiksberg C, Denmark,2 Branch for Enteric Disease Control and Prevention, Henan, Center for Disease Control and Prevention, Nongye East Road, East New District, Zhengzhou, Henan 450016, People's Republic of China3

Received 25 July 2008/ Returned for modification 10 September 2008/ Accepted 18 November 2008

In a previous study, four Salmonella isolates from humans in the Henan province of China showed reduced susceptibility to ciprofloxacin (MIC, 0.125 to 0.25 µg/ml) but were susceptible to nalidixic acid (MIC, 4 to 8 µg/ml). All isolates were negative for known qnr genes (A, B, and S), aac(6')Ib-cr, and mutations in gyrA and parC. Plasmid DNA was extracted from all four isolates and transformed into Escherichia coli TG1 and DH10B cells by electroporation, and transformants were selected on 0.06 µg/ml ciprofloxacin containing brain heart infusion agar plates. Resistance to ciprofloxacin could be transferred by electroporation, and a similar 4,270-bp plasmid was found in all transformants. By sequence analysis, the plasmid was found to carry an open reading frame that had similarities to other qnr genes and that encoded a 214-amino-acid pentapeptide repeat protein. This gene, designated qnrD, showed 48% similarity to qnrA1, 61% similarity to qnrB1, and 41% similarity to qnrS1. Further subcloning of the qnrD coding region into the constitutively expressed tetA gene of vector pBR322 showed that the gene conferred an increase in the MIC of ciprofloxacin by a factor of 32 (from an MIC of 0.002 to an MIC of 0.06 µg/ml). For comparison, qnrA1 and qnrS1 were also subcloned into pBR322 and transformed into DH10B cells, conferring MICs of 0.125 and 0.5 µg/ml, respectively. A phylogenetic analysis of all known qnr sequences was performed and showed that qnrD was more closely related to the qnrB variants but formed an independent cluster. To our knowledge, this is the first description of this qnrD gene.

* Corresponding author. Mailing address: National Food Institute, Technical University of Denmark, Bülowsvej 27, Room B25, DK-1790 Copenhagen V, Denmark. Phone: 45 72 34 62 69. Fax: 45 7234 63 41. E-mail: licav{at}food.dtu.dk

{triangledown} Published ahead of print on 24 November 2008.

Antimicrobial Agents and Chemotherapy, February 2009, p. 603-608, Vol. 53, No. 2
0066-4804/09/$08.00+0     doi:10.1128/AAC.00997-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.



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