A simple method for the identification of aminoglycoside-modifying enzymes.

In order to study the correlation between the presence of aminoglycoside-modifying enzymes in bacteria and the susceptibility of these bacteria to aminoglycosides, 133 resistant strains were collected, representing the most frequently occurring modifying enzymes in clinical isolates today. Enzymes in these resistant strains were identified by the determination of substrate profiles for eight different aminoglycosides in vitro. Thirteen different enzymes or combinations of enzymes appeared to be present in this collection, whereas in seven cases the resistance appeared to be non-enzyme-mediated. The enzyme activities were not reflected in the bacterial susceptibility data for each antibiotic. Cluster analysis of the combined sensitivity data, with biochemical enzyme analysis as a guideline, proved to be unsatisfactory for the identification of aminoglycoside-modifying enzymes in clinical isolates. Much better results were obtained with a stepwise determination scheme. This method relies upon inhibition zone diameters around commercially available sensitivity discs for six aminoglycoside antibiotics. These zone diameters are compared with empirically established critical values, which are characteristic of an enzyme or group of enzymes. By this procedure proper identification of the enzyme(s) proved to be possible in 84% of the 133 resistant strains. For the evaluation of the method 100 consecutively isolated aminoglycoside resistant clinical isolates were analysed by means of the stepwise scheme and the biochemical method. Results were identical for 97 of the 100 strains.