Genome-scale metabolic models reveal determinants of phenotypic differences in non-Saccharomyces yeasts.

BACKGROUND: Use of alternative non-Saccharomyces yeasts in wine and beer brewing has gained more attention the recent years. This is both due to the desire to obtain a wider variety of flavours in the product and to reduce the final alcohol content. Given the metabolic differences between the yeast species, we wanted to account for some of the differences by using in silico models. RESULTS: We created and studied genome-scale metabolic models of five different non-Saccharomyces species using an automated processes. These were: Metschnikowia pulcherrima, Lachancea thermotolerans, Hanseniaspora osmophila, Torulaspora delbrueckii and Kluyveromyces lactis. Using the models, we predicted that M. pulcherrima, when compared to the other species, conducts more respiration and thus produces less fermentation products, a finding which agrees with experimental data. Complex I of the electron transport chain was to be present in M. pulcherrima, but absent in the others. The predicted importance of Complex I was diminished when we incorporated constraints on the amount of enzymatic protein, as this shifts the metabolism towards fermentation. CONCLUSIONS: Our results suggest that Complex I in the electron transport chain is a key differentiator between Metschnikowia pulcherrima and the other yeasts considered. Yet, more annotations and experimental data have the potential to improve model quality in order to increase fidelity and confidence in these results. Further experiments should be conducted to confirm the in vivo effect of Complex I in M. pulcherrima and its respiratory metabolism.

Metschnikowia pulcherrima represses aerobic respiration in Saccharomyces cerevisiae suggesting a direct response to co-cultivation.

The use of non-Saccharomyces species as starter cultures together with Saccharomyces cerevisiae is becoming a common practice in the oenological industry to produce wines that respond to new market demands. In this context, microbial interactions with these non-Saccharomyces species must be considered for a rational design of yeast starter combinations. Previously, transcriptional responses of S. cerevisiae to short-term co-cultivation with Torulaspora delbrueckii, Candida sake, or Hanseniaspora uvarum was compared. An activation of sugar consumption and glycolysis, membrane and cell wall biogenesis, and nitrogen utilization was observed, suggesting a metabolic boost of S. cerevisiae in response to competing yeasts. In the present study, the transcription profile of S. cerevisiae was analyzed after 3 h of cell contact with Metschnikowia pulcherrima. Results show an over-expression of the gluco-fermentative pathway much stronger than with the other species. Moreover, a great repression of the respiration pathway has been found in response to Metschnikowia. Our hypothesis is that there is a direct interaction stress response (DISR) between S. cerevisiae and the other yeast species that, under excess sugar conditions, induces transcription of the hexose transporters, triggering glucose flow to fermentation and inhibiting respiration, leading to an increase in both, metabolic flow and population dynamics.

Novel insights into microbial community dynamics during the fermentation of Central European ice wine.

Culture-dependent and culture-independent strategies were applied to investigate the microbiota of autumn undamaged and damaged berries, winter berries and ice wine must samples of Grüner Veltliner (Veltlínske zelené) from Small Carpathian wine-producing region. One hundred twenty-six yeasts and 242 bacterial strains isolated from several microbiological media (YPD, PDA, R2A, GYC, MRS and MRS-T) were clustered by ITS-PCR and subsequent Qiaxcel electrophoresis. Representatives of each cluster were identified by sequencing. The extracellular hydrolytic properties and intracellular activities of esterase and ß-glucosidase of isolates were assayed. The culture-independent approach permitted the analysis of extracted DNA and RNA coupling DGGE fingerprinting with construction of clone libraries (bacterial and fungal; DGGE-cloning). The combination of the two approaches provided comprehensive data that evidenced the presence of a complex microbiota in each analyzed sample. RNA and DNA analyses facilitated differentiation of living microorganisms from the entire microbiota. Diverse microbial communities colonized the autumn and winter berries. Generally, the combination of results obtained by the methods suggested that the must samples contained mainly Saccharomyces cerevisiae, Metschnikowia spp., Hanseniaspora uvarum, Lactococcus lactis and Leuconostoc spp. The strains exhibited interesting esterase and ß-glucosidase properties, which are important for aroma formation in wine. Fermentation strategies utilising these microorganisms, could be attempted in the future in order to modulate the ice wine characteristics.

Yeast diversity on grapes in two German wine growing regions.

The yeast diversity on wine grapes in Germany, one of the most northern wine growing regions of the world, was investigated by means of a culture dependent approach. All yeast isolates were identified by sequence analysis of the D1/D2 domain of the 26S rDNA and the ITS region. Besides Hanseniaspora uvarum and Metschnikowia pulcherrima, which are well known to be abundant on grapes, Metschnikowia viticola, Rhodosporidium babjevae, and Curvibasidium pallidicorallinum, as well as two potentially new species related to Sporidiobolus pararoseus and Filobasidium floriforme, turned out to be typical members of the grape yeast community. We found M. viticola in about half of the grape samples in high abundance. Our data strongly suggest that M. viticola is one of the most important fermenting yeast species on grapes in the temperate climate of Germany. The frequent occurrence of Cu. pallidicorallinum and strains related to F. floriforme is a new finding. The current investigation provides information on the distribution of recently described yeast species, some of which are known from a very few strains up to now. Interestingly yeasts known for their role in the wine making process, such as Saccharomyces cerevisiae, Saccharomyces bayanus ssp. uvarum, Torulaspora delbrueckii, and Zygosaccharomyces bailii, were not found in the grape samples.