New Mg2+-dependent oxytetracycline resistance determinant tet 34 in Vibrio isolates from marine fish intestinal contents

Abstract

A new oxytetracycline (OTC) resistance (Otc(r)) determinant, Tet 34, was cloned from chromosomal DNA of Vibrio sp. no. 6 isolated from intestinal contents of cultured yellowtail (Seriola quinqueradiata). The transformant, containing cloned Tet 34, could grow in broth containing 25 microg of drug per ml with 10 mM MgCl2. Tet 34 encoded an open reading frame (ORF) 154 amino acids long. The amino acid sequence of the ORF was homologous to sequences of several bacterial xanthine-guanine phosphoribosyltransferases (XPRTs), which act in purine nucleotide salvage synthesis. Mg2+ binding site residues and the active site were highly conserved in XPRT and the ORF of Tet 34. The results suggest that Tet 34 encodes a new Mg2+-dependent Otc(r) mechanism.

FIG. 1.

Growth of E. coli JM109 transformed with pOV (○, •) or pUC119 (▵, ▴). LB broth contained 25 μg of OTC and 50 μg of ampicillin per ml. Panel A shows results with 10 mM MgCl₂, and panel B shows results without MgCl₂. Open symbols, without OTC; solid symbols, with OTC. O.D., optical density.

FIG. 2.

Alignment of the amino acid sequence of the ORF on Tet 34 with five XPRTs obtained from the database. The PRib-PP motif is boxed, and the conserved amino acid residues related to activity of XRPT are in boldface. Asterisks denote the consensus amino acid residues. See references (noted by no. 1) and (noted by no. 2) for review.