Detection of genes encoding aminoglycoside-modifying enzymes in staphylococci by polymerase chain reaction and dot blot hybridization

Abstract

Dot blot hybridization and the polymerase chain reaction (PCR) were used to study aminoglycoside-modifying enzymes in aminoglycoside-resistant staphylococci isolated in hospitals in Kuwait. DNA encoding the acetyltransferase (AAC) (6′)–phosphotransferase (APH) (2′′), nucleotidyltransferase (ANT) (4′) and APH (3′) enzymes were detected in Staphylococcus aureus and coagulase negative staphylococci. ANT (4′) was the most common enzyme detected. The majority of isolates contained genes for all three modifying enzymes, AAC (6′)–APH (2′′), ANT (4′) and APH (3′); only few isolates carried genes for a single modifying enzyme. Genes encoding the AAC (6′)–APH (2′′) were detected in all except two gentamicin-resistant isolates. In these isolates the genes for the AAC (6′)–APH (2′′) enzyme could not be detected by PCR and dot blot hybridization. Whereas antibiotic resistance testing could be used to predict the presence of the AAC (6′)–APH (2′′) enzyme it was not useful in predicting the presence of the ANT (4′) or APH (3′) enzymes in gentamicin-resistant isolates. Results obtained with dot blot hybridization were comparable to those obtained with PCR. However, PCR was fast and results were obtained within the same day. Therefore PCR would be preferred for the detection and confirmation of the presence of aminoglycoside-modifying enzymes in clinical microbiology laboratories.