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Antimicrobial Agents and Chemotherapy, November 2008, p. 4001-4009, Vol. 52, No. 11
0066-4804/08/$08.00+0     doi:10.1128/AAC.00308-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

A New Tetracycline Efflux Gene, tet(40), Is Located in Tandem with tet(O/32/O) in a Human Gut Firmicute Bacterium and in Metagenomic Library Clones{triangledown} ,{dagger}

Katarzyna A. Kazimierczak,1 Marco T. Rincon,1 Andrea J. Patterson,1 Jennifer C. Martin,1 Pauline Young,2 Harry J. Flint,1 and Karen P. Scott1*

Microbial Ecology Group,1 Genomics Unit, Rowett Research Institute, Greenburn Road, Bucksburn, Aberdeen AB21 9SB, United Kingdom2

Received 5 March 2008/ Returned for modification 6 May 2008/ Accepted 10 July 2008

The bacterium Clostridium saccharolyticum K10, isolated from a fecal sample obtained from a healthy donor who had received long-term tetracycline therapy, was found to carry three tetracycline resistance genes: tet(W) and the mosaic tet(O/32/O), both conferring ribosome protection-type resistance, and a novel, closely linked efflux-type resistance gene designated tet(40). tet(40) encodes a predicted membrane-associated protein with 42% amino acid identity to tetA(P). Tetracycline did not accumulate in Escherichia coli cells expressing the Tet(40) efflux protein, and resistance to tetracycline was reduced when cells were incubated with an efflux pump inhibitor. E. coli cells carrying tet(40) had a 50% inhibitory concentration of tetracycline of 60 µg/ml. Analysis of a transconjugant from a mating between donor strain C. saccharolyticum K10 and the recipient human gut commensal bacterium Roseburia inulinivorans suggested that tet(O/32/O) and tet(40) were cotransferred on a mobile element. Sequence analysis of a 37-kb insert identified on the basis of tetracycline resistance from a metagenomic fosmid library again revealed a tandem arrangement of tet(O/32/O) and tet(40), flanked by regions with homology to parts of the VanG operon previously identified in Enterococcus faecalis. At least 10 of the metagenomic inserts that carried tet(O/32/O) also carried tet(40), suggesting that tet(40), although previously undetected, may be an abundant efflux gene.

* Corresponding author. Mailing address: Microbial Ecology Group, Gut Health Division, Rowett Research Institute, Greenburn Road, Bucksburn, Aberdeen AB21 9SB, United Kingdom. Phone: 44(0)1224-712751. Fax: 44(0)1224-716687. E-mail: K.Scott{at}rowett.ac.uk

{triangledown} Published ahead of print on 8 September 2008.

{dagger} Supplemental material for this article may be found at http://aac.asm.org/.

Antimicrobial Agents and Chemotherapy, November 2008, p. 4001-4009, Vol. 52, No. 11
0066-4804/08/$08.00+0     doi:10.1128/AAC.00308-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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