The mode of action of Epicoccum derived antimicrobials

Current antifungal drugs possess two major limitations: i) emergence of fungal strains that are resistant to their antifungal modes of action, and ii) toxic side effects of the drugs caused by lack of selectivity towards fungal targets. One option to overcome these limitations is thought to be the discovery of antifungal compounds with novel modes of action. Fungi are of great interest as a possible source due to their ability to produce a diverse range of secondary metabolites that are tailored to their specific niche for survival. In addition, there are a vast number of fungi, or their products that are yet undiscovered or studied thoroughly. Epicoccum species are fungi that are already known to produce secondary metabolites with promising antifungal properties. However, no studies have been carried out to identify their modes of action. This research focused on the purification and identification of an antifungal compound produced by Epicoccum italicum ICMP 19927 through agar diffusion assays, minimal inhibitory concentration (MIC) assays, time-to-kill assays, flash column chromatography, and nuclear magnetic resonance (NMR) spectroscopy. In addition, attempts were made to elucidate its mode/s of action by the combination of chemical-genetic profiling, haploinsufficiency profiling, green fluorescent protein (GFP)-tagged protein library screening, and gas-chromatography massspectrometry (GC-MS) based metabolomics approach to observe the responses of different Saccharomyces cerevisiae strains to sub-lethal dosage of the antifungal compound. Antifungal compound was purified from the crude extract of E. italicum, with activity against both moulds and yeasts. Its activity was as low as 30 µg/mL against S. cerevisiae. It was identified as a disalt (presumably of sodium) of epipyrone A (DEA). Results suggest DEA affects sphingolipids and their biosynthesis in yeast, while very long chain fatty acids (VLCFAs) are possibly contributing to resistance. Furthermore, additional antifungal modes of action were indicated including oxidative stress, disruption of the membrane protein endocytosis and multivesicular body (MVB) sorting pathways, and β-oxidation of fatty acids. To our knowledge, this was the first time to report a polyene class of antifungal compound directly affecting sphingolipids – possibly not ergosterol, which is a well-known target of other drugs in the same class. These results support that discovery of an antifungal compound with novel modes of action from fungi is possible, and we should not underestimate fungi and the compounds they produce that remain understudied.

Name thesis:degree_name

PhD

Level thesis:degree_level

Doctoral

Discipline thesis:degree_discipline

Biological Sciences

Grantor dc:publisher

ResearchSpace@Auckland

Year dc:date.issued

2019

Author dc:creator

• Lee, Alex Jungwook

Advisors dc:contributor.advisor

• Lewis, Gillian
• Swift, Simon
• Villas-Boas, Silas

Statement dc:rights

• Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher.

Licence dc:rights.uri

• https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htm

Handle dc:identifier.uri

https://hdl.handle.net/2292/50174

OAI identifier oai:identifier

oai:researchspace.auckland.ac.nz:2292/50174