Investigation of Antifungal Mechanisms of Thymol in the Human Fungal Pathogen, Cryptococcus neoformans.

Kwang-Woo Jung,Moon-Soo Chung,Sungbeom Lee,Hyoung-Woo Bai,Byung-Yeoup Chung

doi:10.3390/molecules26113476

Abstract

Due to lifespan extension and changes in global climate, the increase in mycoses caused by primary and opportunistic fungal pathogens is now a global concern. Despite increasing attention, limited options are available for the treatment of systematic and invasive mycoses, owing to the evolutionary similarity between humans and fungi. Although plants produce a diversity of chemicals to protect themselves from pathogens, the molecular targets and modes of action of these plant-derived chemicals have not been well characterized. Using a reverse genetics approach, the present study revealed that thymol, a monoterpene alcohol from Thymus vulgaris L., (Lamiaceae), exhibits antifungal activity against Cryptococcus neoformans by regulating multiple signaling pathways including calcineurin, unfolded protein response, and HOG (high-osmolarity glycerol) MAPK (mitogen-activated protein kinase) pathways. Thymol treatment reduced the intracellular concentration of Ca2+ by controlling the expression levels of calcium transporter genes in a calcineurin-dependent manner. We demonstrated that thymol decreased N-glycosylation by regulating the expression levels of genes involved in glycan-mediated post-translational modifications. Furthermore, thymol treatment reduced endogenous ergosterol content by decreasing the expression of ergosterol biosynthesis genes in a HOG MAPK pathway-dependent manner. Collectively, this study sheds light on the antifungal mechanisms of thymol against C. neoformans.

Highlights

The incidence rate of systemic and invasive mycoses caused by opportunistic fungal pathogens, including Candida albicans, Aspergillus fumigatus, and Cryptococcus neoformans, has increased over the past decades due to the extension of human lifespan and increase in the number of immunocompromised patients with organ transplantation
We screened loss-of-function strains of C. neoformans, which are mutated in stress-responsive signaling pathways, for their viability post treatment with thyme oil constituents such as thymol, carvacrol, and p-cymene
Spotting assays revealed that the cna1∆, cnb1∆, and ire1∆ mutants were sensitive to treatment with thymol and carvacrol, but not with p-cymene (Figure 1A)

Summary

Introduction

The incidence rate of systemic and invasive mycoses caused by opportunistic fungal pathogens, including Candida albicans, Aspergillus fumigatus, and Cryptococcus neoformans, has increased over the past decades due to the extension of human lifespan and increase in the number of immunocompromised patients with organ transplantation. It has been estimated that there are more than 4,000,000 infections caused by opportunistic fungal pathogens annually [1]. Plants produce a large number of chemicals, some of which are used as primary metabolites, while others are used to protect themselves from pathogens and herbivores. Of the different plant defense compounds, phytoanticipins and phytoalexins have been widely characterized over several decades [5]. Phytoanticipins are pre-existing compounds found in diverse locations that are released from organelles immediately after a pathogen attack

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