OBJECTIVE: To determine the prevalence and impact on patient outcome of active human cytomegalovirus infections in patients with prolonged treatment in an intensive care unit. DESIGN: Retrospective analysis of stored plasma samples. SETTING: Anesthesiological intensive care unit of a university hospital. PATIENTS: All 138 patients treated for at least 14 days (of a total of 4940 patients admitted during the study period). Immunocompromised patients and patients with inconclusive results for cytomegalovirus DNA were excluded. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Stored plasma samples of patients with prolonged intensive care unit stay were tested for cytomegalovirus DNA. Sixty-four of 255 evaluable samples from 99 immunocompetent patients tested cytomegalovirus DNA-positive with a mean DNA concentration of 8,600 genome equivalents per milliliter. Active cytomegalovirus infection was diagnosed by reproducibly positive results in 35 patients (35%). Only one case had been diagnosed clinically. Patients with and without active cytomegalovirus infection were not significantly different in parameters, such as age, sex, admission category, source of admission, or comorbidities. Even review of specific surgical procedures or the use of a heart-lung-machine showed no significant differences between the groups. The mortality rate in patients with cytomegalovirus infection was significantly increased (28.6% vs. 10.9%, p = 0.048), and surviving patients had a longer intensive care unit stay (32.6 vs. 22.1 days, p <0.001). CONCLUSIONS: Active cytomegalovirus infection is a frequent but seldom diagnosed finding in surgical patients with prolonged intensive care unit stay, which is associated with increased mortality and prolonged intensive care unit stay of surviving patients.