Anhydrobiosis in yeast: is it possible to reach anhydrobiosis for yeast grown in conditions with severe oxygen limitation?

The yeast Saccharomyces cerevisiae was shown to be extremely sensitive to dehydration-rehydration treatments when stationary phase cells were subjected to conditions of severe oxygen limitation, unlike the same cells grown in aerobic conditions. The viability of dehydrated anaerobically grown yeast cells never exceeded 2 %. It was not possible to increase this viability using gradual rehydration of dry cells in water vapour, which usually strongly reduces damage to intracellular membranes. Specific pre-dehydration treatments significantly increased the resistance of anaerobic yeast to drying. Thus, incubation of cells with trehalose (100 mM), increased the viability of dehydrated cells after slow rehydration in water vapour to 30 %. Similarly, pre-incubation of cells in 1 M xylitol or glycerol enabled up to 50-60 % of cells to successfully enter a viable state of anhydrobiosis after subsequent rehydration. We presume that trehalose and sugar alcohols function mainly according to a water replacement hypothesis, as well as initiating various protective intracellular reactions.

Anhydrobiosis in yeasts: Glutathione synthesis by yeast Ogataea (Hansenula) polymorpha cells after their dehydration-rehydration.

The possibility of using active dry microbial preparations in biotechnological processes is essential for the development of new modern industrial technologies. In this study, we show the possibility of obtaining such preparations of the genetically engineered yeast strain Ogataea (Hansenula) polymorpha with glutathione overproduction. Special pre-treatment involving the gradual rehydration of dry cells in water vapour led to the restoration/reactivation of almost 100% of dehydrated cells. Furthermore, dry cells do not lose their viability during storage at room temperatures. Application of dry cells as the inoculum provides the same levels of glutathione synthesis as that of a native yeast culture.

Anhydrobiosis in yeast: FT-IR spectroscopic studies of yeast grown under conditions of severe oxygen limitation.

Anhydrobiosis is a unique state of living organisms when metabolism is temporarily and reversibly delayed in response to the extreme desiccation of cells. The production of dry active preparations of yeast grown under anaerobic conditions is not currently possible because preparations are extremely sensitive to the dehydration procedure, though they could be very helpful in different biotechnological processes, including bioethanol production. To characterize mechanisms responsible for such sensitivity to the dehydration procedure, Fourier transform infrared spectroscopy was used to study the composition of aerobically grown yeast Saccharomyces cerevisiae resistant to dehydration and grown under conditions of severe oxygen limitation and sensitive to dehydration. Results indicated that significantly lower amounts of lipids in cells, grown under conditions of severe oxygen limitation, may be related to the mechanisms of sensitivity. Dehydration of both resistant and sensitive S. cerevisiae cells was accompanied by similar changes in main cellular compounds. Amounts of nucleic acids and proteins decreased slightly, whereas that of lipids and carbohydrates increased. Artificially reduced sensitivity to dehydration in S. cerevisiae cells, grown under conditions of severe oxygen limitation, led to the increase in the lipid concentration. The chemical composition of S. cerevisiae membranes is proposed to dictate the resistance to dehydration in resistant and sensitive cells.

Immobilisation increases yeast cells' resistance to dehydration-rehydration treatment.

This study was performed with the goal of revealing if the dehydration procedure used in our new immobilisation method noticeably decreases the viability of yeast cells in immobilised preparations. Various yeasts were used in this research: Saccharomyces cerevisiae cells that were rather sensitive to dehydration and had been aerobically grown in an ethanol-containing medium, a recombinant strain of S. cerevisiae grown in aerobic conditions which were completely non-resistant to dehydration and an anaerobically grown bakers' yeast strain S. cerevisiae, as well as a fairly resistant Pichia pastoris strain. Experiments performed showed that immobilisation of all these strains essentially increased their resistance to a dehydration-rehydration treatment. The increase of cells' viability (compared with control cells dehydrated in similar conditions) was from 30 to 60%. It is concluded that a new immobilisation method, which includes a dehydration stage, does not lead to an essential loss of yeast cell viability. Correspondingly, there is no risk of losing the biotechnological activities of immobilised preparations. The possibility of producing dry, active yeast preparations is shown, for those strains that are very sensitive to dehydration and which can be used in biotechnology in an immobilised form. Finally, the immobilisation approach can be used for the development of efficient methods for the storage of recombinant yeast strains.