A physiological characterization in controlled bioreactors reveals a novel survival strategy for Debaryomyces hansenii at high salinity.

Debaryomyces hansenii is traditionally described as a halotolerant non-conventional yeast and has served as a model organism for the study of osmotolerance and salt tolerance mechanisms in eukaryotic systems for the past 30 years. However, unraveling of D. hansenii's biotechnological potential has always been difficult due to the persistent limitations in the availability of efficient molecular tools described for this yeast. Additionally, there is a lack of consensus and contradictory information along the recent years that limits a comprehensive understanding of its central carbon metabolism, mainly due to a lack of physiological studies in controlled and monitored environments. Moreover, there is little consistency in the culture conditions (media composition, temperature, and pH among others) used by different groups, which makes it complicated when trying to get prevalent conclusions on behavioral patterns. In this work, we present for the first time a characterization of D. hansenii in batch cultivations using highly controlled lab-scale bioreactors. Our findings contribute to a more complete picture of the central carbon metabolism and the external pH influence on the yeast's ability to tolerate high Na+ and K+ concentrations, pointing to a differential effect of both salts, as well as a positive effect in cell performance when low environmental pH values are combined with a high sodium concentration in the media. Finally, a novel survival strategy at very high salinity (2 M) is proposed for this yeast, as well as potential outcomes for its use in industrial biotechnology applications. TAKE AWAY: High salt concentrations stimulate respiration in Debaryomyces hansenii. Sodium exerts a stronger positive impact on cell performance than potassium. µmax is higher at a combination of low pH, high salt, and high temperature. Concentrations of 2 M salt result in slower growth but increased biomass yield. The positive effect of salts is enhanced at low glucose concentration.

Phenol degradation by halophilic fungal isolate JS4 and evaluation of its tolerance of heavy metals.

Phenol is one of the most common pollutants in many kinds of industrial wastewater, some of which are in high salinity, resulting in more difficulties of biodegradation. In this work, a halophilic strain capable of utilizing phenol as sole source of carbon and energy in both hypersaline and no-salt media was isolated and identified as genus Debaryomyces. The optimization of environmental parameters including phenol concentration, pH, dissolved oxygen as well as salinity was carried out and tolerance of heavy metals by the strain was evaluated. The strain Debaryomyces sp. was able to grow in culture when initial phenol concentration, pH, agitation and salinity were at wide ranges (0-1200 mg L(-1), 4.0-10.0, 50-200 rpm, 0 %-15 %, respectively). High removal efficiency was hardly affected in the presence of 5 mM of Zn (II) and Mn (II). Under optimal conditions (pH 6.0, 200 rpm, 1 % of salinity without heavy metals), 500 mg L(-1) of phenol could be completely degraded within 32 h. The high removal efficiency of phenol by the strain with significant variations of process parameters might contribute to the bioremediation of phenol-polluted environments under hypersaline or no-salt conditions.

Growth and adaptation of microorganisms on the cheese surface.

Microbial communities living on cheese surfaces are composed of various bacteria, yeasts and molds that interact together, thus generating the typical sensory properties of a cheese. Physiological and genomic investigations have revealed important functions involved in the ability of microorganisms to establish themselves at the cheese surface. These functions include the ability to use the cheese's main energy sources, to acquire iron, to tolerate low pH at the beginning of ripening and to adapt to high salt concentrations and moisture levels. Horizontal gene transfer events involved in the adaptation to the cheese habitat have been described, both for bacteria and fungi. In the future, in situ microbial gene expression profiling and identification of genes that contribute to strain fitness by massive sequencing of transposon libraries will help us to better understand how cheese surface communities function.

Mitochondria from the salt-tolerant yeast Debaryomyces hansenii (halophilic organelles?).

The yeast Debaryomyces hansenii is considered a marine organism. Sea water contains 0.6 M Na(+) and 10 mM K(+); these cations permeate into the cytoplasm of D. hansenii where proteins and organelles have to adapt to high salt concentrations. The effect of high concentrations of monovalent and divalent cations on isolated mitochondria from D. hansenii was explored. As in S. cerevisiae, these mitochondria underwent a phosphate-sensitive permeability transition (PT) which was inhibited by Ca(2+) or Mg(2+). However, D. hansenii mitochondria require higher phosphate concentrations to inhibit PT. In regard to K(+) and Na(+), and at variance with mitochondria from all other sources known, these monovalent cations promoted closure of the putative mitochondrial unspecific channel. This was evidenced by the K(+)/Na(+)-promoted increase in: respiratory control, transmembrane potential and synthesis of ATP. PT was equally sensitive to either Na(+) or K(+). In the presence of propyl-gallate PT was still observed while in the presence of cyanide the alternative pathway was not active enough to generate a Delta Psi due to a low AOX activity. In D. hansenii mitochondria K(+) and Na(+) optimize oxidative phosphorylation, providing an explanation for the higher growth efficiency in saline environments exhibited by this yeast.

Osmotic adaptation in halotolerant yeast, Debaryomyces nepalensis NCYC 3413: role of osmolytes and cation transport.

Debaryomyces nepalensis NCYC 3413, a food spoiling yeast isolated from rotten apple, has been previously demonstrated as halotolerant yeast. In the present study, we assessed its growth, change in cell size, and measured the intracellular polyol and cations (Na(+) or K(+)) accumulated during growth in the absence and presence of different concentrations of salts (NaCl and KCl). Cells could tolerate 2 M NaCl and KCl in defined medium. Scanning electron microscopic results showed linear decrease in mean cell diameter with increase in medium salinity. Cells accumulated high amounts of K(+) during growth at high concentrations of KCl. However, it accumulated low amounts of Na(+) and high amounts of K(+) when grown in the presence of NaCl. Cells grown in the absence of salt showed rapid influx of Na(+)/K(+) on incubation with high salt. On incubation with 2 M KCl, cells grown at 2 M NaCl showed an immediate efflux of Na(+) and rapid uptake of K(+) and vice versa. To withstand the salt stress, osmotic adjustment of intracellular cation was accompanied by intracellular accumulation of polyol (glycerol, arabitol, and sorbitol). Based on our result, we hypothesize that there exists a balanced efflux and synthesis of osmolytes when D. nepalensis was exposed to hypoosmotic and hyperosmotic stress conditions, respectively. Our findings suggest that D. nepalensis is an Na(+) excluder yeast and it has an efficient transport system for sodium extrusion.

Plasma membrane composition of Debaryomyces hansenii adapts to changes in pH and external salinity.

Debaryomyces hansenii is a marine yeast that has to cope with different stress situations. Since changes in membrane properties can play an important function in adaptation, we have examined the fluidity and lipid composition of purified plasma membranes of D. hansenii grown at different external pH values and salt concentrations. Growth at low pH caused an increase in the sterol-to-phospholipid ratio and a decrease in fatty acid unsaturation which was reflected in decreased fluidity of the plasma membrane. High levels of NaCl increased the sterol-to-phospholipid ratio and fatty acid unsaturation, but did not significantly affect fluidity. The sterol-to-phospholipid ratios obtained in D. hansenii grown under any of these conditions were similar to the ratios that have been reported for halophilic/halotolerant black yeasts, but much smaller than those observed in the model yeast Saccharomyces cerevisiae.