It is known that peritoneal murine macrophages activated with interferon-γ exert a fungicidal effect against Paracoccidioides brasiliensis conidia by a nitric oxide (NO)-mediated mechanism. This NO-mediated effect can also be induced by other cytokines such as tumor necrosis factor-α (TNF-α). The aim of this study was to determine if TNF-α-activated peritoneal murine macrophages infected with P. brasiliensis were able to show fungistatic/fungicidal effects mediated by NO. The results indicated that although macrophage activation with TNF-α did not result in NO production, these cells played an important role in inhibiting the conidia from becoming yeast cells. In vivo , the NO-independent inhibitory effect would prove of importance for the establishment of P. brasiliensis in host tissues.