Incubation periods of aerobic (AE) and anaerobic (AN) blood-culture bottles with the BacT/Alert system were assessed in our laboratory. We reviewed the results of 6229 blood-culture sets collected at Kyoto University Hospital from January 1999 to December 2000. Of these sets, 731 (11.7%) were positive for bacteria or yeast. Excluding 87 sets with growth evidence on arrival, of the 644 positive blood-culture sets from 341 patients, a total of 691 organisms were isolated. Of the 691 organisms, 413 (59.8%) were recovered from both bottles, 206 (29.8%) were recovered only from the AE bottle, and 72 (10.4%) were recovered only from the AN bottle. The AE bottle was significantly superior to the AN bottle in terms of both recovery rate and detection time for overall organisms, but there was no significant difference in detection time for facultative anaerobic bacteria between the two bottles. Of the 691 organisms, 530 (76.7%) were classified as usual pathogens. Of the 530 usual pathogens, 501 (94.5%) were recovered in at least one bottle of each set within the first 3 days, and 523 (98.7%) within the first 5 days of incubation. Twenty-nine organisms initially isolated on day 4 or later were recovered from 19 patients. Of these, chart reviews indicated that 21 organisms recovered from 11 patients were considered clinically significant bacteria, and the reviews also revealed that no patient had a treatment plan altered based on the results of positive blood culture. Seven organisms initially isolated on day 6 or later were recovered from 7 patients. Chart reviews revealed that 5 of these organisms from 5 patients were considered to be clinically significant. In conclusion, if the incubation period had been less than 3 days, 11 patients with clinically significant bacteremia or fungemia, (3.2% of all patients with bacteremia or fungemia) would have been undiagnosed. Similarly, with an incubation period of 5 days, 5 such patients (1.5%) would have been undiagnosed.