OXIDATIVE KILLING OF CRYPTOCOCCUS-NEOFORMANS BY HUMAN NEUTROPHILS - EVIDENCE THAT FUNGAL MANNITOL PROTECTS BY SCAVENGING REACTIVE OXYGEN INTERMEDIATES

Polymorphonuclear neutrophils (PMN) kill Cryptococcos neoformans (Cn) by oxidative mechanisms, but the roles of various reactive oxygen inte rmediates (ROIs) are not known. We used a mannitol low-producing Cn mu tant (Cn MLP) and its wild-type parent (Cn H99) to examine the role of ROIs distal to H2O2 in PMN killing and to determine whether mannitol produced by Cn protects the fungus against ROIs. At PMN:Cn cell ratios of 1:1, 10:1, and 100:1, PMN killed significantly more Cn MLP than Cn H99 cells after 2 and 4 h (p < 0.05). Superoxide dismutase and the hy droxyl radical (OH.) scavengers mannitol and DMSO inhibited killing of both strains (p < 0.05), but catalase did not, Cn H99 and Cn MLP stim ulated PMN to produce similar amounts of O-2(-) and H2O2. In contrast, Cn MLP stimulated greater luminol-dependent chemiluminescence than di d Cn H99 (p < 0.05). finally, H2O2 alone killed similar numbers of Cn H99 and Cn MLP cells, but oxidants generated by FeSO4 (1 mu M), H2O2 ( 10 mu M), and iodide (1 to 3 mu M) killed significantly more Cn MLP th an Cn H99 cells in 1 h (17 4 0.05). Mannitol, DMSO, and catalase compl etely inhibited killing of both Cn strains by this cellfree system, bu t superoxide dismutase did not. These results suggest that 1) distal R OIs such as OH. and HOCl are key effector molecules against Cn, acid 2 ) mannitol produced by Cn may protect against oxidative killing by sca venging distal ROIs.