The Effect Of Mixed Fermentation Of Torulaspora Delbrueckii And Saccharomyces Cerevisiae On The Aroma Property Of Cabernet Gernischet Wine And The Interactive Mechanism Between Two Yeasts

Torulaspora delbrueckii is a type of aromatic yeast that has attracted much attention in the field of fruit wine making,and its mixed fermentation with Saccharomyces cerevisiae is expected to enhance the quality of wine.In this study,the effects of T.delbrueckii/S.cerevisiae sequential fermentation inoculation ratios on the fermentation process,wine composition and organoleptic properties were investigated at the laboratory level using a Chinese endemic wine grape variety,Cabernet Gernischet,as the raw material,and then the interactions between the two yeasts were investigated at the optimal inoculation ratio,i.e.the transcriptional techniques were used to analyse.The study also investigated the interaction between T.delbrueckii/S.cerevisiae at the optimal inoculation ratio,i.e.using transcriptomics techniques to analyse the differential gene expression between the two yeasts during the fermentation process,and combined with the analysis of the structural composition of the wine to obtain the key genes and synthetic pathways related to the metabolic regulation of the main components in S.cerevisiae,especially the characteristic aromatic substances.The main results of the study include:1.Obtain the optimal inoculation ratios for T.delbrueckii NS-D and S.cerevisiae MERIT in sequential inoculation mode.Three sequential inoculation ratios（T.delbrueckii/S.cerevisiae=1:1,10:1,50:1）and control groups（T.delbrueckii and S.cerevisiae alone:T1,T10,T50 and S1,respectively）were set up for fermentation and the proliferation of each yeast during wine fermentation and the effect on wine composition were examined.The results showed that both yeasts co-existed in the wine in any proportion of the mixed fermentation mode,with T.delbrueckii dominating in the early stages of fermentation and S.cerevisiae in the later stages.The mixed fermentation had a low impact on the physicochemical parameters of the wines,with the lowest volatile acid content（0.41 g/L）in the 10:1 inoculation ratio.The volatile aromatic composition of the wines was measured by GC-MS and the wines from the 50:1 and 10:1 inoculation ratios were found to have the highest total alcohols,esters,aldehydes,ketones and terpenes and the highest sensory scores.Based on these results and the cost of production,the optimal inoculation ratio for T.delbrueckii/S.cerevisiae is 10:1.2.The overall differences in gene transcription levels between the two yeasts in the T.delbrueckii/S.cerevisiae mixed fermentation mode were obtained.A mixed fermentation system of T.delbrueckii and S.cerevisiae was designed,and the RNA-Seq technique was used to examine the changes in differential gene expression between the two yeasts during the whole fermentation process（including the pre-fermentation,mid-fermentation and end-fermentation）at an inoculation ratio of 10:1.S.cerevisiae and T.delbrueckii were detected with 6458 and 5174 genes respectively,while the number of genes affected due to mixed fermentation was 792 and 609 respectively,indicating that mixed inoculation had a greater effect on S.cerevisiae.These differential genes were all localised to biological processes,cellular composition and molecular function in GO enrichment analyses.In the KEGG pathway enrichment analysis,significant differences were found in genes involved in fatty acid degradation,glycolysis,multiple amino acid biosynthesis and metabolism,glyoxylate and dicarboxylic acid metabolism,and vitamin B₆ metabolism in S.cerevisiae under mixed fermentation conditions;most of the differentially expressed genes in T.delbrueckii were involved in amino acid biosynthesis,the catabolism of specific amino acid sugar metabolism,and ribosome biosynthesis,as well as a large number of pathways associated with biological function.3.Preliminary insights into the transcriptional level of interactions between S.cerevisiae and T.delbrueckii were obtained.A total of 68 important genes were discussed in the S.cerevisiae genome that were significantly up-and down-regulated by mixed fermentation,of which 43 genes were involved in the metabolism of volatile esters,alcohols,aldehydes,acids and organic acids.Through in-depth analysis,this study hypothesised that PDCs,ILV5,BATs,ADHs,ALDs,ACS1,ACC1 and ARO10 are key genes in the fermentation process of Cabernet Gernischet wine and are associated with the metabolism of certain amino acids and sugar metabolism in wine,which in turn promotes the synthesis and accumulation of various aromatic substances such as butanol,β-phenylethanol,hexyl acetate,isoamyl acetate,ethyl butyrate and ethyl caprate,thus improving the aroma quality of Cabernet Gernischet wine under mixed fermentation inoculation conditions.In addition,gene-YOL109W,gene-YKR092C and gene-YKL068W-A,unidentified genes that play an important role in the formation of aromatic substances during the metabolism of sugars and amino acids in wine,are also worthy of further study.