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Antimicrobial Agents and Chemotherapy, June 1999, p. 1350-1357, Vol. 43, No. 6
0066-4804/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Characterization and Nucleotide Sequence of a Klebsiella oxytoca Cryptic Plasmid Encoding a CMY-Type beta -Lactamase: Confirmation that the Plasmid-Mediated Cephamycinase Originated from the Citrobacter freundii AmpC beta -Lactamase

Shang Wei Wu,1,2,3,* Kathrine Dornbusch,1 Göran Kronvall,1 and Mari Norgren3

Department of Laboratory Medicine, Division of Clinical Microbiology, Karolinska Institute and Karolinska Hospital, 171 76 Stockholm,1 and Department of Clinical Bacteriology, Umeå University, S-901 85 Umeå,3 Sweden, and Laboratory of Microbiology, The Rockefeller University, New York, New York 100212

Received 10 September 1998/Returned for modification 4 January 1999/Accepted 12 March 1999

Plasmid pTKH11, originally obtained by electroporation of a Klebsiella oxytoca plasmid preparation into Escherichia coli XAC, expressed a high level of an AmpC-like beta -lactamase. The enzyme, designated CMY-5, conferred resistance to extended-spectrum beta -lactams in E. coli; nevertheless, the phenotype was cryptic in the K. oxytoca donor. Determination of the complete nucleotide sequence of pTKH11 revealed that the 8,193-bp plasmid encoded seven open reading frames, including that for the CMY-5 beta -lactamase (blaCMY-5). The blaCMY-5 product was similar to the plasmidic CMY-2 beta -lactamase of K. pneumoniae and the chromosomal AmpC of Citrobacter freundii, with 99.7 and 97.0% identities, respectively; there was a substitution of phenylalanine in CMY-5 for isoleucine 105 in CMY-2. blaCMY-5 was followed by the Blc and SugE genes of C. freundii, and this cluster exhibited a genetic organization identical to that of the ampC region on the chromosome of C. freundii; these results confirmed that C. freundii AmpC was the evolutionary origin of the plasmidic cephamycinases. In the K. oxytoca host, the copy number of pTKH11 was very low and the plasmid coexisted with plasmid pNBL63. Analysis of the replication regions of the two plasmids revealed 97% sequence similarity in the RNA I transcripts; this result implied that the two plasmids might be incompatible. Incompatibility of the two plasmids might explain the cryptic phenotype of blaCMY-5 in K. oxytoca through an exclusion effect on pTKH11 by resident plasmid pNBL63.

* Corresponding author. Mailing address: Box 152, The Rockefeller University, 1230 York Ave., New York, NY 10021. Phone: (212) 327-8278. Fax: (212) 327-8688. E-mail: wus{at}rockvax.rockefeller.edu.

Antimicrobial Agents and Chemotherapy, June 1999, p. 1350-1357, Vol. 43, No. 6
0066-4804/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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