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Journal of Bacteriology, November 2003, p. 6233-6240, Vol. 185, No. 21
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.21.6233-6240.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Identification and Characterization of the emhABC Efflux System for Polycyclic Aromatic Hydrocarbons in Pseudomonas fluorescens cLP6a

Elizabeth M. Hearn,1,2 Jonathan J. Dennis,1 Murray R. Gray,2 and Julia M. Foght1*

Department of Biological Sciences, University of Alberta, Edmonton, Alberta, Canada T6G 2E9,1 Department of Chemical and Materials Engineering, University of Alberta, Edmonton, Alberta, Canada T6G 2G62

Received 1 May 2003/ Accepted 8 August 2003

The hydrocarbon-degrading environmental isolate Pseudomonas fluorescens LP6a possesses an active efflux mechanism for the polycyclic aromatic hydrocarbons phenanthrene, anthracene, and fluoranthene but not for naphthalene or toluene. PCR was used to detect efflux pump genes belonging to the resistance-nodulation-cell division (RND) superfamily in a plasmid-cured derivative, P. fluorescens cLP6a, which is unable to metabolize hydrocarbons. One RND pump, whose gene was identified in P. fluorescens cLP6a and was designated emhB, showed homology to the multidrug and solvent efflux pumps in Pseudomonas aeruginosa and Pseudomonas putida. The emhB gene is located in a gene cluster with the emhA and emhC genes, which encode the membrane fusion protein and outer membrane protein components of the efflux system, respectively. Disruption of emhB by insertion of an antibiotic resistance cassette demonstrated that the corresponding gene product was responsible for the efflux of polycyclic aromatic hydrocarbons. The emhB gene disruption did not affect the resistance of P. fluorescens cLP6a to tetracycline, erythromycin, trimethoprim, or streptomycin, but it did decrease resistance to chloramphenicol and nalidixic acid, indicating that the EmhABC system also functions in the efflux of these compounds and has an unusual selectivity. Phenanthrene efflux was observed in P. aeruginosa, P. putida, and Burkholderia cepacia but not in Azotobacter vinelandii. Polycyclic aromatic hydrocarbons represent a new class of nontoxic, highly hydrophobic compounds that are substrates of RND efflux systems, and the EmhABC system in P. fluorescens cLP6a has a narrow substrate range for these hydrocarbons and certain antibiotics.

* Corresponding author. Mailing address: Department of Biological Sciences, CW405 Biological Sciences Center, University of Alberta, Edmonton, Alberta, Canada T6G 2E9. Phone: (780) 492-3279. Fax: (780) 492-9234. E-mail: julia.foght{at}ualberta.ca.

Journal of Bacteriology, November 2003, p. 6233-6240, Vol. 185, No. 21
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.21.6233-6240.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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