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Antimicrobial Agents and Chemotherapy, August 1998, p. 1931-1937, Vol. 42, No. 8
0066-4804/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Molecular Cloning and Functional Analysis of a Novel Tetracycline Resistance Determinant, tet(V), from Mycobacterium smegmatis

Edda De Rossi,1 Marian C. J. Blokpoel,2 Rita Cantoni,1 Manuela Branzoni,1 Giovanna Riccardi,1 Douglas B. Young,2 Koen A. L. De Smet,2 and Orio Ciferri1,*

Department of Genetics and Microbiology, University of Pavia, 27100 Pavia, Italy,1 and Department of Medical Microbiology, Imperial College School of Medicine, London W2 1PG, United Kingdom2

Received 23 October 1997/Returned for modification 17 November 1997/Accepted 1 April 1998

The nucleotide sequence and mechanism of action of a tetracycline resistance gene from Mycobacterium smegmatis were determined. Analysis of a 2.2-kb sequence fragment showed the presence of one open reading frame, designated tet(V), encoding a 419-amino-acid protein (molecular weight, 44,610) with at least 10 transmembrane domains. A database search showed that the gene is homologous to membrane-associated antibiotic efflux pump proteins but not to any known tetracycline efflux pumps. The steady-state accumulation level of tetracycline by M. smegmatis harboring a plasmid carrying the tet(V) gene was about fourfold lower than that of the parental strain. Furthermore, the energy uncoupler carbonyl cyanide m-chlorophenylhydrazone blocked tetracycline efflux in deenergized cells. These results suggest that the tet(V) gene codes for a drug antiporter which uses the proton motive force for the active efflux of tetracycline. By primer-specific amplification the gene appears to be restricted to M. smegmatis and M. fortuitum.

* Corresponding author. Mailing address: Department of Genetics and Microbiology, via Abbiategrasso 207, 27100 Pavia, Italy. Phone: (39-382) 505576. Fax: (39-382) 528496. E-mail: ociferri{at}pillo.unipv.it.

Antimicrobial Agents and Chemotherapy, August 1998, p. 1931-1937, Vol. 42, No. 8
0066-4804/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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