A prospective review of all enterococcal isolates for 13 months showe that 9.0% were resistant to ampicillin (MIC ≥ 16 μg/ml; zone diameter < 15 mm), as determined by the Vitek system, disk diffusion, microdilution MIC testing, and macrodilution MIC testing. All were β-lactamase negative. A total of 19 and 3 resistant isolates were from urine and intravascular sites, respectively. Ampicillin-resistant enterococci appear to be a growing clinical problem.