Investigating the Effect of Selected Non- Saccharomyces Species on Wine Ecosystem Function and Major Volatiles

Abstract

Natural alcoholic fermentation is initiated by a diverse population of several non-Saccharomyces yeast species. However, most of the species progressively die off, leaving only a few strongly fermentative species, mainly Saccharomyces cerevisiae. The relative performance of each yeast species is dependent on its fermentation capacity, initial cell density, ecological interactions as well as tolerance to environmental factors. However, the fundamental rules underlying the working of the wine ecosystem are not fully understood. Here we use variation in cell density as a tool to evaluate the impact of individual non-Saccharomyces wine yeast species on fermentation kinetics and population dynamics of a multi-species yeast consortium in synthetic grape juice fermentation. Furthermore, the impact of individual species on aromatic properties of wine was investigated, using Gas Chromatography-Flame Ionization Detector. Fermentation kinetics was affected by the inoculation treatment. The results show that some non-Saccharomyces species support or inhibit the growth of other non-Saccharomyces species in the multi-species consortium. Overall, the fermentation inoculated with a high cell density of Starmerella bacillaris displayed the fastest fermentation kinetics while fermentation inoculated with Hanseniaspora vineae showed the slowest kinetics. The production of major volatiles was strongly affected by the treatments, and the aromatic signature could in some cases be linked to specific non-Saccharomyces species. In particular, Wickerhamomyces anomalus at high cell density contributed to elevated levels of 2-Phenylethan-1-ol whereas Starm. bacillaris at high cell density resulted in the high production of 2-methylpropanoic acid and 3-Hydroxybutanone. The data revealed possible direct and indirect influences of individual non-Saccharomyces species within a complex consortium, on wine chemical composition.

Keywords: multi-starter fermentation; population dynamics; wine fermentation; yeast consortium; yeast-yeast interactions.

Figure 1

Outline of the experimental plan showing the dosage treatments with S. cerevisiae included in the inoculum (A) and in the absence of S. cerevisiae (B) Treatments are defined based on the strain codes. The following abbreviations were used in this figure. Mp, M. pulcherrima; Cp, C. parapsilosis; Pt, P. terricola; Wa, W. anomalus; Hv, H. vineae; Lt, L. thermotolerans; Sb, Starm. Bacillaris; NS, Non-Saccharomyces species. In the presence of S. cerevisiae, in each treatment, one non-Saccharomyces species was inoculated at approximately 10⁶ while the rest of yeast species in the consortium (7 NS and Sc) was inoculated at approximately 10⁴. In the absence of S. cerevisiae, the same inoculation strategy was applied. However, S. cerevisiae was not included in the yeast consortium. In control fermentations, in the presence of S. cerevisiae (NS-SC consortium), each NS species was inoculated at approximately 10⁶ and Sc at approximately 10⁴ whereas in the absence of Sc (NS consortium), 7 NS was each inoculated at approximately 10⁶.

Figure 2

Fermentation profiles showing the kinetics of sugar consumption fructose (▴) and glucose (■), in (A) Mp-dose, (B) Cp-dose, (C) Pt-dose, (D), Wa-dose, (E) Hv-dose, (F) Lt-dose, (G) Sb-dose, and (H) NS-SC.

Figure 3

Distribution of yeast species (%) during fermentations at the inoculation time (INC), the beginning (BF), middle (MF), and end of fermentation (EF) in (A) Mp-dose, (B) Cp-dose, (C) Pt-dose, (D), Wa-dose, (E) Hv-dose, (F) Lt-dose, (G) Sb-dose, and (H) NS-SC.

Figure 4

PCA score plot (A) and corresponding loading plot (B) of first and second principal components (PC) derived from PC analysis of the major volatile compounds produced in synthetic must fermentations. The major volatile compounds are represented by light grays whereas the treatments are as follows: Mp-dose (red), Cp-dose (gray), Pt-dose (orange), Wa-dose (brown), Hv-dose (green), Lt-dose (black), Sb-dose (purple), and NS-SC (blue). Some replicates were overlapped in each treatment (e.g. Pt-dose).