Development of a specific probe for the aminoglycoside-(3)-N-acetyltransferase resistance gene

Aminoglycoside-(3)-N-acetyltransferase isoenzyme V (AAC(3)V) is produced by > 90% of aminoglycoside-resistant Escherichia coli isolates from Hong Kong. A gentamicin resistance determinant from a conjugative plasmid carried by one of these E. coli strains producing AAC(3)V was cloned as a 6-kb fragment in the PstI site of the Bluescript II KS-phagemid vector. A 0.8-kb SphI-SalI fragment from the cloned insert was used as a probe in an epidemiological study. The specificity of this probe was evaluated by colony hybridization with 17 control strains producing different known aminoglycoside resistance enzymes. Hybridization was observed only with strains producing AAC(3)V. The probe demonstrated the presence of the AAC(3)V gene in 95.5% to 133 gentamicin-resistant E. coli isolates. This result agreed with the earlier data on the distribution of this enzyme that were based on the susceptibility profile.