Osmotic adaptation of the halophilic fungus Hortaea werneckii: role of osmolytes and melanization

Abstract

This study was intended to determine the osmoadaptation strategy of Hortaea werneckii, an extremely salt-tolerant melanized ascomycetous fungus that can grow at 0-5.1 M NaCl. It has been shown previously that glycerol is the major compatible solute in actively growing H. werneckii. This study showed that the exponentially growing cells also contained erythritol, arabitol and mannitol at optimal growth salinities, but only glycerol and erythritol at maximal salinities. The latter two were both demonstrated to be major compatible solutes in H. werneckii, as their decrease correlated with the severity of hypoosmotic shock. Besides higher amounts of erythritol and lower amounts of glycerol, stationary-phase cells also contained mycosporineglutaminol-glucoside, which might act as a complementary compatible solute. H. werneckii is constitutively melanized under various salinity conditions. Ultrastructural study showed localization of melanin in the outer parts of the cell wall as a distinct layer at optimal salinity (0.86 M NaCl), whereas cell-wall melanization diminished at higher salinities. The role of melanized cell wall in the effective retention of glycerol is already known, and was also demonstrated in H. werneckii by lower retention of glycerol in cells with blocked melanization compared to melanized cells. However, these non-melanized cells compensated for the lower amounts of glycerol with higher amounts of erythritol and arabitol. We hypothesize that H. werneckii melanization is effective in reducing the permeability of its cell wall to its major compatible solute glycerol, which might be one of the features that helps it tolerate a wider range of salt concentrations than most organisms.

Osmoadaptation Strategy of the Most Halophilic Fungus, Wallemia ichthyophaga, Growing Optimally at Salinities above 15% NaCl

“…The polyols were extracted from 30 mg of freeze-dried cells using the previously described method (17). The cells were suspended in 500 l Bligh and Dyer solution (composed of methanol-chloroform-water [10: 5:4]) and vigorously shaken for approximately 30 min (IKA-Vibrax VXR; Janke & Kunkel).…”

“…The cultures of W. ichthyophaga for the analysis of intracellular compatible solutes after a hypo-osmotic shock were grown in triplicate in liquid YNB medium with 20% (wt/vol) NaCl (14,17). Hypo-osmotic shocks were performed as follows; the cells were grown in YNB with 20% NaCl to the mid-exponential growth phase.…”

“…Therefore, more suitable model organisms for halotolerance studies in eukaryotes were suggested, such as the extremely halotolerant black yeast Hortaea werneckii, the moderately halotolerant yeast Debaryomyces hansenii, and the halophilic W. ichthyophaga (all reviewed in reference 9) and the polyextremotolerant Aureobasidium pullulans (14,15). H. werneckii can grow in vitro at a wide salinity range, from 0% to 32% (saturation) NaCl, and has a broad growth optimum, from 6 to 10% NaCl (16,17), whereas the halotolerant A. pullulans grows best without NaCl but can tolerate up to 17% NaCl and the marine yeast D. hansenii grows better at low sodium and tolerates up to 24% NaCl (18)(19)(20). One of the most important adaptations of fungi to high salinity is the accumulation of a mixture of various compatible solutes, among which glycerol is the most common.…”

Properties of the halophyte microbiome and their implications for plant salt tolerance

“…This has been analysed in the salt sensitive yeast S. cerevisiae and in some halotolerant filamentous fungi; it was shown that mostly glycerol is accumulated upon increasing salt concentrations (Blomberg and Adler 1992). Glycerol is also the compatible solute found in highest concentrations in the halophilic black yeast H. werneckii (Kogej et al 2007). Other modifications upon exposure to high salt concentrations concern the microsomal HMG-CoA reductase (Vaupotic and Plemenitas 2007) and the glycerol-3-phosphate dehydrogenase (Lenassi et al 2011).…”

Aspergillus glaucus Aquaglyceroporin Gene glpF Confers High Osmosis Tolerance in Heterologous Organisms

“…Both the extremely halotolerant yeast Hortaea werneckii and the halophilic fungus Wallemia ichthyophaga accumulate glycerol to adapt to hyperosmotic conditions. The levels of glycerol are increased with increasing salinity and decreased after hy-poosmotic shock (51,52). Several genes known to be involved in glycerol production were found in the genomes of H. werneckii and W. ichthyophaga (53).…”