To isolate environmental bacteria capable of transforming fluoroquinolones to inactive molecules. Bacteria were isolated from the aerobic liquor of a wastewater treatment plant on a medium containing norfloxacin (100 mg l(-1)). Twenty-two isolates were highly resistant (minimal inhibitory concentration: 6.25-200 mu g ml(-1)) to five fluoroquinolones and six of them were positive by PCR amplification for the aminoglycoside resistance gene aac(6')-Ib. Of these, only Escherichia coli strain LR09 had the ciprofloxacin-acetylating variant gene aac(6')-Ib-cr; HPLC and mass spectrometry showed that this strain transformed both ciprofloxacin and norfloxacin by N-acetylation. This bacterium also had mutations in the quinolone-resistance determining regions of the gyrA and parC genes. An E. coli isolate from wastewater, which possessed at least two distinct fluoroquinolone resistance mechanisms, inactivated ciprofloxacin and norfloxacin by N-acetylation. This is the first report of N-acetylation of fluoroquinolones by an aac(6')-Ib-cr-containing bacterium from an environmental source.