Extracellular phospholipase activity has been implicated in the pathogenesis of several bacterial infections. Recently, extracellular phospholipase activity has been proposed as a virulence factor in the opportunistic yeast Candida albicans. Aspergillus fumigatus is the most pathogenic member of its genus, responsible for > 90% of infections. Previously, no specific virulence factors have been determined. We investigated the ability of A. fumigatus to produce extracellular phospholipases at 37 degrees C. Fast atom bombardment was used to compare lipid-containing media before and at 5-h intervals during shaking culture of A. fumigatus. Lipids were extracted and analyzed. Many anions corresponding to phospholipid breakdown products were identified. Specific anion species identified indicated phospholipase A, B, C (PLC), and D activities. PLC activity was further investigated by using the synthetic substrate p-nitrophenylphosphorylcholine. PLC activity was initially observed after 30 h of growth and accumulated in broth cultures up to 50 h. At 55 h, there was a sharp increase in PLC activity which coincided with cultures reaching the stationary phase. Activity of the PLC was measured at different temperatures, with greater activity occurring at 37 degrees C than at lower temperatures. Phospholipases could represent a virulence determinant in A. fumigatus.