H. S. Martinelli, PhD thesis). The fungitoxic activity of Jaburetox was evaluated on germination and growth of Penicillium herguei, Mucor sp. and R solani, as shown in Fig. 6, panels F–H. Mucor sp. showed the highest susceptibility, its growth at 48 h being inhibited at the lowest tested dose (10 μM). For P. herguei, doses www.selleckchem.com/ferroptosis.html of 20 and 40 μM were inhibitory after 72–96 h, affecting also the production

of pigments (data not shown) after hyphae development. In contrast, growth of R. solani was not affected at the highest dose of Jaburetox, 40 μM ( Fig. 6, panel H). Jaburetox at 9 μM inhibited the growth of S. cerevisiae, C. parapsilosis, P. membranisfaciens ( Fig. 7). The other tested yeasts, C. tropicalis, K. marxiannus and C. albicans, were inhibited with 18 μM Jaburetox (not shown). The antifungal effect of Jaburetox did not persist after washing of the treated cells. Additional studies are needed to clarify whether the effect is fungistatic, if the peptide is being hydrolyzed/inactivated, or if the repeated administration of the peptide could lead to the death of the yeasts. Permeabilization of the plasma membrane by Jaburetox was evaluated in S. cerevisiae showing that the treated cells are more permeable to SYTOX Green than controls

( Fig. 3, panels E–F and H–I). As observed for the JBU, the peptide R428 also induces morphological changes in yeasts ( Fig. 3, panel G). The induction of pseudohyphae in C tropicalis and the membrane permeabilization effect in S. cerevisiae occurred at much lower doses (0.36–0.72 μM) than those required to arrest fungi propagation. In this work we have shown that,

besides filamentous fungi, JBU is also toxic against yeasts. The fungitoxic effects consisted in inhibition of proliferation, Chorioepithelioma induction of morphological alterations with formation of pseudohyphae, changes in transport of H+ and in energy metabolism, permeabilization of membranes, eventually leading to cell death. The antifungal effect of the JBU in yeasts or filamentous fungi [7] is independent of its catalytic activity, since the enzymatically inactivated protein, after treatment with the covalent inhibitor p-hydroxy-mercurybenzoate, maintained its fungitoxic properties. The generation of antifungal peptides upon proteolysis of urease further reinforce this fact. On the other hand, the presence of intact urease in the supernatant of cultures after 24 h was observed for most yeasts except for K. marxiannus, which extensively degraded JBU (data not shown). Thus at this point, it is not clear to us whether hydrolysis of JBU by the yeasts is required for expression of its fungicidal effect. Similar to our observation in filamentous fungi [7], the fungicidal activity of JBU in yeasts is also specie-specific, affecting differently in terms of effective dose and type of toxic effects the six yeast species under study.