Biology subjects

Marx, F.

Publications and source records attributed to Marx, F..

2 recordsLinked to original sources

bioRxiv2024

The Neosartorya (Aspergillus) fischeri antifungal protein NFAP2 has low potential to trigger resistance development in Candida albicans in vitro

Due to the increase in the number of drug-resistant Candida albicans strains, new antifungal compounds with limited potential for development of resistance are urgently needed. NFAP2, an antifungal protein (AFP) secreted by Neosartorya (Aspergillus) fischeri, is a promising candidate. We investigated the ability of C. albicans to develop resistance to NFAP2 in a microevolution experiment compared with generic fluconazole (FLC). C. albicans adapted to only 1 x minimum inhibitory concentration (MIC) of NFAP2 compared with 32 x MIC of FLC. Genome analysis revealed non-silent mutations in only two genes in NFAP2-resistant strains and in several genes in FLC-resistant strains. Resistance development to NFAP2 did not influence cell morphology. The susceptibility of NFAP2-resistant strains did not change to FLC, amphotericin B, micafungin, terbinafine. These strains did not show altered susceptibility to AFPs from Penicillium chrysogenum, except one which had less susceptibility to P. chrysogenum antifungal protein B. FLC-resistant strains had decreased susceptibility to terbinafine and NFAP2, but not to other drugs and AFPs from P. chrysogenum. NFAP2- and FLC-resistant strains showed decreased and increased NFAP2 binding and uptake, respectively. The development of resistance to NFAP2 decreased tolerance to cell wall, heat, and UV stresses. The development of FLC resistance increased tolerance to cell wall stress and decreased tolerance to heat and UV stresses. Resistance to NFAP2 did not have significant metabolic fitness cost and could not increase virulence, compared with resistance to FLC. ImportanceDue to the increasing number of (multi)drug-resistant strains, only a few effective antifungal drugs are available to treat infections caused by opportunistic Candida species. Therefore, the incidence of hard-to-treat candidiasis has increased dramatically in the past decade, and the demand to identify antifungal compounds with minimal potential to trigger resistance is substantial. The features of NFAP2 make it a promising candidate for the topical treatment of Candida infection. Data on the development of resistance to AFPs in C. albicans are lacking. In this study, we provide evidence that NFAP2 has low potential to trigger resistance in C. albicans in vitro and the developed resistance mechanisms to NFAP2 are not associated with severe phenotypic changes compared with development of resistance to generic FLC. These results suggest the slow emergence of NFAP2-resistant Candida strains and that NFAP2 can reliably be used long-term in the clinic.

microbiology↗

bioRxiv2022

The membrane activity of the amphibian Temporin B peptide analog TB_KKG6K sheds light on the mechanism that kills Candida albicans

Temporin B (TB) is a 13 amino acid long, cationic peptide secreted by the granular glands of the European frog Rana temporaria. We could recently show that the modified TB peptide analog TB_KKG6K rapidly killed planktonic and sessile Candida albicans at low {micro}M concentrations and was neither hemolytic nor cytotoxic to mammalian cells in vitro. The present study aimed to shed light into its mechanism of action, with a focus on its fungal cell membrane activity. We utilized different fluorescent dyes to prove that it rapidly induces membrane depolarization and permeabilization. Studies on model membrane systems revealed that the TB analog undergoes hydrophobic and electrostatic membrane interactions showing a preference for anionic lipids and identified phosphatidylinositol and cardiolipin as possible peptide targets. Fluorescence microscopy using FITC-labelled TB_KKG6K in the presence of the lipophilic dye FM4-64 indicated that the peptide compromises membrane integrity and rapidly enters C. albicans cells in an energy independent manner. Peptide treated cells analyzed by cryo-based electron microscopy exhibited no signs of cell lysis; however, subcellular structures were disintegrated, suggesting that intracellular activity may form part of the killing mechanism of the peptide. Taken together, this study proved that the TB_KKG6K compromises C. albicans membrane function, which explains the previously observed rapid, fungicidal mode of action and promises its great potential as a future anti-Candida therapeutic. ImportanceFungal infections with the opportunistic human pathogen C. albicans are associated with high moratility rates in immunocompromised patients. This is partly due to the yeasts ability to rapidly develop resistance towards currently available antifungals. Small, cationic, membrane-active peptides are promising compounds to fight against resistance development as many of them effectuate rapid fungal cell death. This fast killing is believed to hamper the development of resistance, as the fungi do not have sufficient time to adapt to the antifungal compound. We prevously reported that the synthetic variant of the amphibian Temporin B peptide, TB_KKG6K, rapidly kills C. albicans. In the current study, the mechanism of action of the TB analog was investigated. We show that this TB analog is membrane-active and impairs cell membrane function, highlighting its potential to be developed as an attractive alternative anti-C. albicans therapeutic, which may hinder the development of resistance.

microbiology↗