Enterococcus faecalis is a frequent cause of hospital-acquired infection. Two hundred fifty one E. faecalis isolates from patients with urinary tract infection at Okayama University Hospital over an 8-year period from 1991 through 1998 were collected. The presence of the asaI, cylA, aac (6')-aph (2''), and aph (3')-III genes was analyzed by PCR methods. Of the 251 isolates, 205 (81.7%) were positive for asaI. The 81.5% (167/205) of asaI-positive isolates also possessed either cylA or aminoglycoside resistance genes, compared to only 15.2% of (7/46) asaI-negative isolates (p<0.0001). The incidence of asaI gradually increased from 69.2% in 1991 to 90.7% in 1998. The number of isolates that contain asaI, cylA and aminoglycoside resistance gene (s) also increased. Pulsed-field gel electrophoresis (PFGE) analyses of 28 asaI-positive and hemolysin-producing isolates revealed 22 different banding patterns, including 6 pairs with similar patterns. The plasmid analyses of these isolates showed different patterns except for 1 pair with similar PFGE pattern. These results suggest that E. faecalis possessing the asaI gene may play an important role in the exchange of genetic information among enterococci in the urinary tract.