Prior clinical trials have demonstrated efficacy and effectiveness of posaconazole in the prophylaxis of invasive fungal diseases in high-risk patients. Controversy exists about the cost-effectiveness of this approach. We performed an analysis comparing the direct costs of posaconazole prophylaxis against polyene mouthwash (thrush) prophylaxis in patients with acute myelogenous leukaemia (AML). Data of AML patients receiving remission-induction chemotherapy were extracted from the CoCoNut (Cologne Cohort of Neutropenic Patients) database to compare hospital costs of patients before (2003–2005) and after (2006–2008) introduction of posaconazole prophylaxis. Treatment on general ward, intensive care unit (ICU), mechanical ventilation, diagnostic procedures, and all anti-infectives were calculated. Patient groups were well matched according to age, gender and duration of neutropenia. The mean costs per patient in the posaconazole group (n = 76) and the polyene mouthwash group (n = 81) were €21 040 (95% confidence interval (CI): €18 204–€23 876) and €23 169 (95% CI: €19 402–€26 937) per patient. Antifungal treatment costs were €4580 (95% CI: €3678–€5482) and €4019 (95% CI: €2825–€5214). Duration on the ICU was 2582 (95% CI: 984.1–4181.7) and 5517 (95% CI: 2206–8827.3) min. In our hospital, primary antifungal prophylaxis by posaconazole was cost-effective. There was a trend towards cost savings, which was primarily caused by a shorter overall length of stay and the less frequent ICU treatment.