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Antimicrobial Agents and Chemotherapy, January 1999, p. 157-160, Vol. 43, No. 1
0066-4804/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Detection of a Streptomycin/Spectinomycin Adenylyltransferase Gene (aadA) in Enterococcus faecalis

Nancye C. Clark,1,* Ørjan Olsvik,2 Jana M. Swenson,1 Carol A. Spiegel,3 and Fred C. Tenover1

Hospital Infections Program, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 303331; Department of Medical Microbiology, School of Medicine, University of Tromsø, Tromsø, Norway2; and Department of Pathology and Laboratory Medicine, University of Wisconsin Hospital and Clinics, Madison, Wisconsin 537923

Received 16 April 1998/Returned for modification 15 July 1998/Accepted 27 October 1998

Genes encoding streptomycin/spectinomycin adenylyltransferases [ANT(3")(9)] have been reported to exist in gram-negative organisms and Staphylococcus aureus. During a study of high-level aminoglycoside resistance in enterococci, we encountered an isolate of Enterococcus faecalis that was streptomycin resistant but did not appear to contain the 6'-adenylyltransferase gene (aadE) when examined by PCR with specific primers. Phosphocellulose paper binding assays indicated the presence of an ANT(3")(9) enzyme. Streptomycin and spectinomycin MICs of 4,000 and 8,000 µg/ml, respectively, were observed for the isolate. PCR primers corresponding to a highly conserved region of the aadA gene were used to amplify a specific 284-bp product. The product hybridized with a digoxigenin-labeled PCR product from E. coli C600(pHP45Omega ) known to contain the aadA gene. The aadA gene was transferred via filter matings from the E. faecalis donor to E. faecalis JH2-2. PCR primers designed for analysis of integrons were used to amplify a 1-kb product containing the aadA gene, which was cloned into the vector pCRII and transformed into Escherichia coli DH5-alpha competent cells. D-Rhodamine dye terminator cycle sequencing was used to determine the gene sequence, which was compared to previously reported sequences of aadA genes. We found the aadA gene in E. faecalis to be identical to the aadA genes reported by Sundström et al. for E. coli plasmid R6-5 (L. Sundström, P. Rådström, G. Swedberg, and O. Sköld, Mol. Gen. Genet. 213:191-201, 1988), by Fling et al. for the aadA within transposon Tn7 (M. E. Fling, J. Kopf, and C. Richards, Nucleic Acids Res. 13:7095-7106, 1985), and by Hollingshead and Vapnek for E. coli R538-1 (S. Hollingshead and D. Vapnek, Plasmid 13:17-30, 1985). Previous reports of the presence of the aadA gene in enterococci appear to be erroneous and probably describe an aadE gene, since the isolates were reported to be susceptible to spectinomycin.

* Corresponding author. Mailing address: Nosocomial Pathogens Laboratory Branch, Centers for Disease Control and Prevention, 1600 Clifton Rd., N.E., Mailstop G08, Atlanta, GA 30333. Phone: (404) 639-0195. Fax: (404) 639-1381. E-mail: ncc1{at}cdc.gov.

Antimicrobial Agents and Chemotherapy, January 1999, p. 157-160, Vol. 43, No. 1
0066-4804/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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