Identification of three genes coding for aminoglycoside-modifying enzymes by means of the polymerase chain reaction.

A polymerase chain reaction (PCR) was devised for the simultaneous identification of three widespread aminoglycoside-acetylating enzymes (AACs), namely, AAC(3)-I, AAC(3)-II and AAC(3)-IV. Three sets of 20-mer DNA primers were constructed, based on the nucleotide sequences of the genes coding these enzymes. Each of the three gene regions contained a unique restriction site. This property was used to confirm the specificity of the reactions. Evolutionarily-well-conserved sequences of the 16S rRNA bacterial genes were used to construct two 20-mer primers, which served as positive controls for the amplification reactions. Tests of twenty reference strains, containing ten different aminoglycoside-modifying enzymes, confirmed the specificity and the sensitivity of the method. The PCR method confirmed the presence of the aacC2 gene in 63 clinical strains, in which susceptibility data and colony hybridization had previously suggested the presence of the gene.