Next Article in Journal
Designing a Waste-Based Culture Medium for the Production of Plant Growth Promoting Microorganisms Based on Cladodes Juice from Opuntia ficus-indica Pruning
Next Article in Special Issue
Integrated Fertilization with Bagasse Vermicompost Changes the Microbiome of Mencía Must and Wine
Previous Article in Journal
Effect of Fermented Camel Milk Containing Pumpkin Seed Milk on the Oxidative Stress Induced by Carbon Tetrachloride in Experimental Rats
Previous Article in Special Issue
New Insights into the Origin of Volatile Sulfur Compounds during Wine Fermentation and Their Evolution during Aging
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Fermentation
Volume 8
Issue 5
10.3390/fermentation8050224
fermentation-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Ecological Distribution and Oenological Characterization of Native Saccharomyces cerevisiae in an Organic Winery

by
Alice Agarbati
,
Laura Canonico
*,
Francesca Comitini
and
Maurizio Ciani
*
Department of Life and Environmental Sciences, Polytechnic University of Marche, Via Brecce Bianche, 60131 Ancona, Italy
*
Authors to whom correspondence should be addressed.
Fermentation 2022, 8(5), 224; https://doi.org/10.3390/fermentation8050224
Submission received: 14 April 2022 / Revised: 2 May 2022 / Accepted: 11 May 2022 / Published: 13 May 2022
(This article belongs to the Special Issue Wine Microbiology)
Browse Figures
Versions Notes

Abstract

:
The relation between regional yeast biota and the organoleptic characteristics of wines has attracted growing attention among winemakers. In this work, the dynamics of a native Saccharomyces cerevisiae population was investigated in an organic winery. In this regard, the occurrence and the persistence of native S. cerevisiae were evaluated in the vineyard and winery and during spontaneous fermentation of two nonconsecutive vintages. From a total of 98 strains, nine different S. cerevisiae biotypes were identified that were distributed through the whole winemaking process, and five of them persisted in both vintages. The results of the oenological characterization of the dominant biotypes (I and II) show a fermentation behavior comparable to that exhibited by three common commercial starter strains, exhibiting specific aromatic profiles. Biotype I was characterized by some fruity aroma compounds, such as isoamyl acetate and ethyl octanoate, while biotype II was differentiated by ethyl hexanoate, nerol, and β-damascenone production also in relation to the fermentation temperature. These results indicate that the specificity of these resident strains should be used as starter cultures to obtain wines with distinctive aromatic profiles.

1. Introduction

For thousands of years, Saccharomycetes have been used by humans to produce fermented foods and beverages. Over the past decades, modern industrial winemaking has been based on the use of starter cultures of specialized and selected wine strains [1,2]. Although the use of selected Saccharomyces cerevisiae strains guarantees the control of fermentation, their widespread utilization can lead to a double consequence: the reduction of indigenous microbiota and the standardization of the wine aroma complexity, which is often linked to the involvement of native yeasts during the fermentation [3,4,5,6,7,8].
To overcome this trend, in recent decades, several winemakers have carried out spontaneous fermentations exploiting indigenous yeasts coming from the vineyard or vinery equipment, with the aim to improve the analytical and aroma complexity and to give peculiar and recognizable flavors to the final wine [5,9]. Indeed, recent studies demonstrated that specific grape varieties and the climate of a specific geographical area seem to influence the yeast community [10,11,12,13,14], indicating a variation of the microbial community of grapes in the function of regional distribution [15,16,17,18,19]. Recently, the correlation between the regional microbial community and the organoleptic characteristics of wine has attracted growing attention [11,20,21]. The understanding of the mechanisms and factors that stabilize a specific epiphytic population in different grape varieties could be useful to establish a correlation with the influence of regional yeast microbiota in the analytical and aromatic characters of the final wine [11,20,21,22]. However, controversies exist regarding the origin of native S. cerevisiae strains. Several studies showed that the direct isolation of S. cerevisiae strains in undamaged grapes and vineyards is very difficult [8,23,24,25]. Mortimer and Polsinelli [26] already showed that damaged grapes represent a depository habitat of Saccharomyces yeasts. Several authors claimed that the detection of S. cerevisiae strains in both vineyards and grapes, is obtained only after grape autoenrichment, due to their deficiency [27,28,29]. On the other hand, it is well known that S. cerevisiae is widely spread in the winery environment and on equipment [30,31,32,33]. The ability of S. cerevisiae to dominate the fermentation process and other competitive advantages allow this species to survive and reproduce in stressful winery conditions.
Knowledge of the occurrence of S. cerevisiae and their colonization on grapes or in the cellar could be of help to preserve the indigenous population and to establish a possible correlation between them and the final wine quality.
In the present study, the occurrence, and the persistence of native S. cerevisiae in the vineyard and winery and in uninoculated wine fermentation was investigated. The organic winery under investigation, which began its activity in 2013 and had never used commercial starter strains, represents an ideal winemaking environment to investigate the dynamics of grape-related and/or winery-related native yeast populations. Within the population of Saccharomyces strains, the dominant biotypes were evaluated and characterized for their oenological aptitude and aromatic profile with the aim to use these native strains as starter cultures.

2. Materials and Methods

2.1. Sampling Campaign

The sampling campaign was carried out in an organic winery (Azienda olivitivinicola Giulia Fiorentini, located in the town of Cupramontana, Ancona, Italy, geographical coordinates: 43°26′48′′ N, 13°6′59′′ E; 505 m altitude) that has never used commercial starter strains.
Thirty-four samples were collected in the 2016 vintage: (i) 10 samples of the Montepulciano grape variety and 10 samples after autoenrichment; (ii) 12 items from the winery environment and pieces of equipment (3 walls, 1 floor, 2 wooden barrels; 4 destemmers, and 2 steel fermentation tanks); and (iii) 2 Verdicchio spontaneous fermentation.
Twenty-four samples were collected in the 2019 vintage: (i) 6 samples of the Verdicchio grape variety and 6 samples after autoenrichment; (ii) 9 items from the winery environment and pieces of equipment (3 walls, 2 floors, 2 destemmers, 2 steel fermentation tanks); and (iii) 2 Verdicchio spontaneous fermentation. Details are reported in Table S1.
The two vineyards (Montepulciano and Verdicchio) each have two hectares and are located close to the winery. The grapes were randomly collected from randomized blocks.
Grape samples (about 1 Kg of undamaged ripe grape bunches) were picked in sterile plastic bags. The related vineyards are located close to each other and close to the cellar (distance of about 300 m). Both vineyards were subjected to the same fungicide treatment used for organic wine production (copper and sulfur compounds). Winery environment and equipment samples were sampled using sterile swabs streaked on random environmental surfaces (10 cm2). Furthermore, samples of spontaneous fermentation were collected through the middle valve of the vat under sterile conditions. All the samples were stored on ice during sampling and immediately transported to the laboratory and processed.

2.2. Samples Processing and S. cerevisiae Isolation

Grape samples were processed as described by Agarbati et al. [23,24,25]. Briefly, ten grape samples from the 2016 vintage and six grape samples from the 2019 vintage were hand-crushed and subjected to shaking at 120 rpm for 30 min to allow the transfer of yeasts from the grape surface to the must. Then, 1 mL of fresh must was used for decimal serial dilution and spread on WL nutrient agar (Wallerstain Laboratories, Oxoid, Hampshire, UK) supplemented with 0.005% chloramphenicol (Sigma-Aldrich, Saint Louis, MI, USA) to suppress bacteria growth. The remaining fresh must was transferred into a 250 mL sterile Erlenmeyer flask closed with Pasteur bung and set up for autoenrichment at room temperature with the aim to promote S. cerevisiae yeast development. The eventual development of this yeast was monitored after 7 and 15 days of autoenrichment through viable cell counts using the same media. Samples were collected by the winery’s equipment, and the cotton swab was placed into 10 mL of sterile water and shaken for 20 min at 120 rpm on a MAXQ 4450 shaker (Thermo Fisher Scientific, Waltham, MA, USA) to allow the transfer of the microorganisms from the swab to the solution. Serial decimal dilutions were carried out and spread on the same media described for grape samples. In the same way, samples collected during must fermentation were diluted and spread on WL nutrient agar (Wallerstain Laboratories, Oxoid, Hampshire, UK) with 0.005% chloramphenicol added. All plates were incubated at 25 °C for 5 days then presumptive S. cerevisiae colonies were isolated on the basis on its typical macro and micromorphology. Each presumptive colony of S. cerevisiae was purified on YPD agar (yeast extract 1%, peptone 2%, D-glucose 2%, and agar 1.8%) medium and then collected for short time at 4 °C using the same medium.

2.3. Identification and Genotyping Characterization of S. cerevisiae Strains

Based on the micro and macromorphology of the colonies, 150 presumptive S. cerevisiae pure cultures were then assayed by genomic DNA analyses. DNA was extracted at 95 °C for 10 min, and then it was amplified by PCR using primers ITS1 (5′-TCCGTAGGTGAACCTCGCG-3′) and ITS4 (5′-TCCTCCGCTTTATTGATATGC-3′) following the procedure reported by Agarbati et al. [23]. A total of 98 strains, the resulting S. cerevisiae strains, were then subjected to genotyping using different primers: firstly, the primer pairs delta12/21 (delta12: 5′-TCAACAATGGAATCCCAAC-3′; delta21: 5′-CATCTTAACACCGTATATGA-3′) were used for interdelta sequence analyses, as described by Legras and Karst [34]. The amplification was performed as per the following program: 3 min at 95 °C, followed by 25 s at 94 °C, 30 s at 45 °C, and 90 s at 72 °C for 9 cycles and 25 s at 94 °C, 30 s at 50 °C, and 90 s at 72 °C for 21 cycles and a final extension at 72 °C for 10 min.
A second genotyped of S. cerevisiae strains was carried out by a PCR reaction using the random amplified microsatellites technique inter-single sequence repeats (ISSR). The primer used was 5′-ARRTYCAGCAGCAGCAG-3′, where R (A or G) and Y (C or T) indicate degenerate sites [35,36]. Amplification was performed as reported: 5 min at 94 °C, followed by 40 cycles of 60 s at 94 °C, 60 s at 55 °C, and 120 s at 72 °C and a final extension at 72 °C for 5 min.
Amplification products from both PCR reactions were separated by electrophoresis on 1.5% (w/v) and 2% (w/v) agarose gels, respectively, submitted to 66 V for 1.5 h in 0.5 × TBE buffer. Before PCR reactions, the DNA of yeasts was extracted at 95 °C for 10 min using a Biorad Thermal Cycler [23].

2.4. Fermentation Trials

Two dominant biotypes (I and II) were used to carry out microfermentation trials to evaluate their fermentative aptitudes at 22 °C and 16 °C fermentation temperatures using fresh Verdicchio must composed as follows: initial sugar content 201 g/L, pH 3.25, total acidity 6.34 g/L, and yeast assimilable nitrogen (YAN) content 98 mgN/L. The YAN was adjusted to 250 mgN/L by the addition of diammonium phosphate and yeast derivative (Genesis Lift® Oenofrance, Bordeaux, France). Before its use, the must was treated with 0.2 mL/L of dimethyl dicarbonate overnight at 4 °C to suppress the wild yeasts, confirmed through viable cell counts using WL nutrient agar (Wallerstain Laboratories, Oxoid, Hampshire, UK).
The strains used were precultured in modified YPD (0.5% w/v yeast extract, 2% w/v glucose, and 0.1% w/v peptone) for 24 h at 25 °C in an orbital shaker (rotation 150 rpm). The cells were collected, washed twice with sterile water, and used to inoculate flasks containing 70 mL of must, at an initial concentration of approximately 1 × 106 cells/mL for each yeast. The flasks were locked with Müller valves containing sulfuric acid to allow CO2 to escape from the system and to avoid contamination. The trials were incubated at 22 °C and 16 °C under static conditions, and the weight loss of the flasks, due to the CO2 evolution, was monitored until a constant weight (for two consecutive days).
The trials were carried out in triplicate, and three commercial starter strains widely used in Verdicchio grape juice fermentation were used as controls: Lalvin EC1118, Lalvin ICV OKAY® (Lallemand Inc., Toulouse, France), and VIN13 (Anchor Wine Yeast, Cape Town, South Africa).

2.5. Analytical Determinations

The resulting wines were analyzed for the main analytical determinations. The fermentation rate was calculated as g CO2/day over the 3rd day of fermentation. Volatile acidity and total SO2 content were obtained following the current analytical methods, according to the Official European Union Methods [37]. Acetaldehyde was determined using a specific enzyme kit (Roche Diagnostics S.p.A., Milano, Italy), while the free α-amino acids were quantified using the o-phthaldialdehyde/N-acetyl-l-cysteine spectrophotometric assay, as described by Dukes and Butzke [38]. Ethanol production evaluated at the end of fermentation was measured according to the Association of Official Analytical Chemists [39]. Specific enzymatic kit (Megazyme, Ireland) was used to determine the concentrations of glucose and fructose, according to the manufacturer instructions.
The volatile compounds were determined by solid-phase microextraction (HS-SPME) method, preparing the sample as follows: 5 mL of wine was placed into a vial; 1 g of NaCl and 3-octanol as the internal standard (1.6 mg/L) were added, and the vial was closed with a septum-type cap and placed on a magnetic stirrer for 10 min at 25 °C. Then the sample was heated to 40 °C and extracted with a fiber Divinylbenzene/Carboxen/Polydimethylsiloxane (DVB/CAR/PDMS) for 30 min by insertion into the vial headspace. The compounds were desorbed by inserting the fiber into Shimadzu gas chromatograph GC injector, in split–splitless modes following the procedure reported by Canonico et al. [40]. The glass capillary column used was 0.25 µm Supelcowax 10, length 60 m, and internal diameter 0.32 mm.

2.6. Data Analyses

The experimental data were analyzed by one-way ANOVA (analysis of variance) using the software STATISTICA 7 (Statsoft inc., Tulsa, OK, USA). The data were considered significantly different with an associated p-value < 0.05 (Duncan tests). The mean data of the main analytical and volatile compounds, normalized to neutralize hidden factors, were used to carry out Principal Component Analysis (PCA) using the package JMP 11® software.

3. Results

3.1. Occurrence of S. cerevisiae on Grapes in Winery Environment and in Uninoculated Fermentations

The source of isolation and number of S. cerevisiae isolated are reported in Table 1.
No S. cerevisiae were found on grape surfaces using the direct isolation procedures. S. cerevisiae were isolated only after the autoenrichment of fresh grape juice (spontaneous fermentation of pressed harvested grapes, under sterile conditions).
A total of 98 S. cerevisiae colonies were isolated in the two years: 37 colonies from grapes after enrichment (10 in the 2016 vintage and 27 in the 2019 vintage), 21 colonies from winery equipment and the environment (13 in the 2016 vintage and 8 in the 2019 vintage) (the specific source are reported in Table S1, Supplementary Material), and 40 strains from spontaneous wine fermentation (half in the 2016 vintage and half in the 2019 vintage). Overall, we isolated a total of 43 and 55 S. cerevisiae in the 2016 and 2019 vintages, respectively.

3.2. Biotypes of S. cerevisiae: Frequency and Distribution

The combination of biotype analyses, interdelta sequences, and ISSR polymorphism, revealed eight different biotypes among 98 S. cerevisiae isolates from 13 samples out of the 57 total samples evaluated (Figure S1, Supplementary Materials). The distribution and the frequency of each biotype, expressed as number of isolates and percentage, are reported in Table 2.
Out of the eight biotypes identified, biotypes I and II dominated in both years. Biotype I represented 54%, while biotype II represented 32% of all isolates. In the 2016 vintage, the frequency of biotypes I and II was 68% and 19%, respectively. Biotype I was detected in all samples, mainly during must fermentation and in the winery environment (26% and 28%, respectively). Biotype II was mainly detected during must fermentation (16%), poorly in the winery environment (2%), and not detected on grapes.
In the 2019 vintage, the two biotypes showed a very similar frequency (biotype I: 43%; biotype II: 42%) and were present in all samples, mainly on grapes (both biotypes 24%) then in wine fermentation (biotype I: 11%; biotype II: 13%), while the lowest percentage was found in the winery environment (biotype I: 9%; biotype II: 5%).
The other biotypes (III, IV, V, VI, VII, and VIII) were randomly detected in the different samples of the two vintages. Biotypes III, IV, and V were detected in both vintages, while biotype VIII was only detected in the 2016 vintage and biotypes VI and VII only in the 2019 vintage.

3.3. Oenological Characterization of Predominant Native S. cerevisiae Isolated Strains

3.3.1. Fermentation Kinetics

The oenological characteristics of S. cerevisiae biotypes I and II (the most dominant biotypes) were evaluated and compared with three commercial starter strains, widely used in the Verdicchio winemaking area.
The fermentation kinetics did not show differences among the native S. cerevisiae biotype I, biotype II, and commercial starters at fermentation temperatures of both 16 °C and 22 °C (Figure 1).

3.3.2. Analytical Characteristics

The main oenological characteristics of wines carried out at 16 °C are reported in Table 3. The results obtained show that biotype I exhibited the significant lowest value of all parameters tested apart from volatile acidity that was significantly higher than that shown by the other strains. Regarding the ethanol content, biotype I showed the highest value of 12.31% v/v, comparable with those produced by the starters OKAY and EC1118, while biotype II revealed the lowest ethanol content.
Furthermore, biotype II exhibited a significant increase in acetaldehyde and total SO2 content in comparison with the other strains tested. Among the S. cerevisiae starter strains, VIN13 exhibited a significantly higher fermentation rate (g CO2/day over the 3rd day of fermentation) and free amino acid content in comparison with other fermentation trials.
The results of the main analytical characters of the fermentation carried out at 22 °C are reported in Table 4. The data obtained show a different trend from those obtained at 16 °C (Table 3). Indeed, the fermentation rate did not display significant differences among the strains tested, with the only exception being the S. cerevisiae starter strain OKAY® that showed a significantly lower fermentation rate. About other characteristics, significant differences were found. The commercial starter strains VIN13 and OKAY® showed a significant increase in volatile acidity. Total SO2 was significantly higher in biotype II confirming the behavior shown at 16 °C. All fermentation led to a low acetaldehyde content with the only exception being VIN13 that showed the highest value, but it was below the threshold. The value of free α-amino acids was generally comparable among the strains tested. The same ethanol content was detected at 22 and 16 °C for biotype I with slight variations for the OKAY and VIN13 starters. Conversely, biotype II as well as the starter EC1118 showed an increase in the ethanol content of about 0.4% v/v during fermentation trials carried out at 22 °C.

3.3.3. Volatile Compounds

The results of the main volatile compounds of wines fermented at 16 °C are reported in Table 5. The data show that strains belonging to different biotypes were characterized by the production of ester compounds. Biotype I exhibited a significant increase in isoamyl acetate (banana aroma) and ethyl octanoate (apple, pear, and peach aroma), while biotype II showed a significantly increased amount in ethyl hexanoate (apple aroma). S. cerevisiae starter strains showed different aromatic profiles: OKAY® was characterized by a significant increase in ethyl butyrate content; EC1118 was characterized by relevant amounts of β-phenyl ethanol, and VIN13 showed a significantly high production of phenylethyl acetate. About hexanol, linalool, and geraniol, the results did not show a significant difference among the strains.
The volatile compounds produced at 22 °C are reported in Table 6. The data show a different behavior of the starter strains in comparison with the 16 °C fermentation trials. At 22 °C, biotype I showed a significant increase in isoamyl acetate, while biotype II was characterized by a significantly high production of nerol and β-damascenone. Among the starter strains tested, OKAY® confirmed at 22 °C the high production of ethyl butyrate together with a relevant production of terpens, β-phenyl ethanol, and diethyl succinate. EC1118 was distinguished by the other strains by the production of phenylethyl acetate and ethyl hexanoate, while VIN13 in this fermentation condition did not show any relevant production of a particular compound.
To assess the overall effects of yeast strains under different fermentation temperature, the main analytical characteristics and volatile compounds were analyzed by Principal Component Analyses (PCA) reported in Figure 2. The PCA analysis related to the fermentation carried out at 16 °C (Figure 2a) and showed that all S. cerevisiae starter strains were grouped in the lower left quadrant, while the two dominant biotypes I and II were placed separate with the starter strains between them. Biotype I was in the lower right quadrant, while biotype II was in the upper right quadrant. Under 16° C of fermentation temperature, a greater differentiation of the analytical and volatile compounds was found. Otherwise, data obtained after fermentation carried out at 22 °C reported in Figure 2b showed a different distribution of the strains in comparison with fermentation carried out at 16 °C. Biotype I was located in the lower left quadrant with S. cerevisiae VIN13. The other three strains were each in a different quadrant.

4. Discussion

In recent years, several investigations claimed different geographical distributions of wine microbioma [10,11]. This relationship between wine microbiota and regional areas has gained relevance in the wine industry [41]. Indeed, the microbiota from a determined region could be able to confer distinctive characteristics to the wine [20,42].
The concept of native yeasts is linked to the conservation of microorganisms naturally present in a specific viticultural area. The knowledge of distribution and the persistence of native S. cerevisiae strains, together with their fermentation aptitudes and stress resistance, could provide important information that contributes to promoting their use in winemaking.
In this work, the occurrence of S. cerevisiae in different winemaking ecological habitats (grape surfaces, winery environment, and must fermentation) was confirmed. As already demonstrated by several authors, the isolation of S. cerevisiae from the surfaces of grapes was only possible after the autoenrichment procedures [23,28,29], supporting the evidence that grapes represent an unfavorable habitat for S. cerevisiae. The DNA fingerprinting of native S. cerevisiae, following the trend previously reported by other authors [9,43,44,45], showed a high level of DNA polymorphism within the population. This elevated polymorphism in both nonconsecutive vintages indicated a high persistence over the years in this specific organic winery where starter yeasts have never been used. Similar results were reported also by Torija et al. [46] and Granchi et al. [45] that described the persistence of recurrent and predominant S. cerevisiae strains during spontaneous wine fermentations of a specific wine-producing area. In this regard, Granchi et al. [45] found that native S. cerevisiae strains are representative of a specific oenological ecosystem and that some of them termed “dominant” persisted in different fermentations in the same winery from one year to another, and they seemed to be representative of a single winery rather than of an oenological area.
In agreement with the results of Granchi et al. [45], we found that among nine different biotypes, only biotypes I and II were dominant and recurrent in the grapes, winery environment, and uninoculated fermentation. Biotype I was found in both Verdicchio and Montepulciano grapes, suggesting a nonspecific correlation between yeast strains and grape variety, according to Ganucci et al. [47]. On the contrary, other investigations found specific yeast strains for specific grape varieties [48,49,50]. To limit the reproducibility problems of interdelta analyses reported by Franco-Duarte et al. [51], a second genotyped (ISSR) was used. However, further investigations could be necessary to validate the different S. cerevisiae biotypes detected.
Effectively, the concept of the origin of the yeasts is very complex. Alexandre [52] reported that the wine yeast terroir concept is based on the assumption that wines are produced by spontaneous fermentation carried out by the yeast biota naturally present on grape. Actually, the yeast biota present in must is a mix of different yeast species coming from grapes and winery (equipment, tanks…) in which the anthropic management practices (both in vineyard and in the cellar) play a decisive role on the microbiome stability and therefore on its influence in the wine.
In the specific environment considered in this study, it was found that the S. cerevisiae population was stable as almost all biotypes were present throughout the production chain, from the vineyard to the cellar and in the finished product [47,53]. This picture was found in two nonconsecutive years (three-year interval), supporting the thesis that the colonization is strictly dependent on the management of the entire vineyard–cellar “system”. The agronomical and oenological practices used, such as organic open field treatments and spontaneous fermentation, could have determined the stability of the microbiota in the various ecological habitats (grape surface, cellar, equipment, and fermenting must).
The two native dominant biotypes, evaluated for the main oenological aptitude and the aromatic profile and compared with the most used S. cerevisiae starter strains, showed that they can confer a distinctively analytical and volatile profile to wines. Here, the two S. cerevisiae biotypes I and II produced specific ester profiles related to a peculiar aromatic note also depending on the fermentation temperature. The ester production, responsible for the fruity character of wines, is a key characteristic of yeast metabolism [54,55,56]. Different studies reported that the high variability in ester production is strictly related to the yeast strain conferring sensorial divergence to wines [57,58,59]. In this regard, the characterization of the phenometabolomic fermentative profile could be useful to further investigate on these native strains [60].
In addition, these native yeasts resulted in being better adapted to the specific environment condition and substrate composition as previously indicated by other works [61,62].
For these reasons, the use of native S. cerevisiae yeasts could be proposed as starter strains to provide significant diversification to analytical and aroma profiles in line with recent observations that native yeasts may confer peculiar characteristics to wines [11,61,63].
Overall, the results show that the fermentation behavior of these native yeasts was comparable with those exhibited by the commercial starters strains used as the control. Moreover, their occurrence in different habitats of the winemaking process, their temporal persistence in different years, and the fermentation aptitude at different temperatures suggest a suitable application in winemaking.

5. Conclusions

In this specific winemaking environment (vineyard and winery where commercial yeasts have not been used for a long time), a broad and specific S. cerevisiae population was found during two nonconsecutive vintages. Among the native S. cerevisiae, two strains resulted in the dominant biotypes. The evaluation of the oenological characteristics of these two dominant biotypes highlighted specific differences when compared with commercial starter strains. The knowledge of the specific traits of the native strains could be useful to characterize and enhance the distinctive features of a specific winemaking area. This could be a strategy to improve the quality of the fermentation industry maintaining a strict link between products and the environment, as required by consumers.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/fermentation8050224/s1, Figure S1: Electrophoresis gels of native S. cerevisiae strains. (a) represents the different biotypes (I, II, III, IV, V, and VI) obtained by δ12–21 analyses. (b) represents the different biotypes (I, II, III, and IV) obtained by ISSR–PCR. In both agarose gels, the lane indicated as 100 bp represents the molecular weight marker; Table S1: Isolates of S. cerevisiae: source and biotype.

Author Contributions

A.A., L.C., F.C. and M.C., contributed equally to the manuscript. All the authors participated in the design and discussion of the research. A.A. carried out the experimental part of the work. A.A. and L.C. carried out the analysis of the data. All authors have read and agreed to the published version of the manuscript.

Funding

This research received external funding: A.A. was supported by a grant from Polytechnic University of Marche, D.R. No. 1312, 23/10/2019.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

Not applicable.

Acknowledgments

The authors wish to thank the winery Azienda olivitivinicola Giulia Fiorentini for the samples and assistance.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Bordet, F.; Roullier-Gall, C.; Ballester, J.; Vichi, S.; Quintanilla-Casas, B.; Gougeon, R.D.; Ortiz, A.J.; Schmitt Kopplin, P.; Alexandre, H. Different wines from different yeasts? “Saccharomyces cerevisiae intraspecies differentiation by metabolomic signature and sensory patterns in wine”. Microorganisms 2021, 9, 2327. [Google Scholar] [CrossRef] [PubMed]
  2. Vaughan-Martini, A.; Martini, A. Saccharomyces meyen ex reess (1870). In The Yeasts, a Taxonomic Study, 5th ed.; Kurtzman, C.P., Fell, J.W., Boekhout, T., Eds.; Elsevier: Amsterdam, The Netherlands, 2011; pp. 733–746. [Google Scholar]
  3. Capece, A.; Pietrafesa, R.; Siesto, G.; Romaniello, R.; Condelli, N.; Romano, P. Selected indigenous Saccharomyces cerevisiae strains as profitable strategy to preserve typical traits of primitivo wine. Fermentation 2019, 5, 87. [Google Scholar] [CrossRef] [Green Version]
  4. Fleet, G.H.; Heard, G.M. Yeasts: Growth during fermentation. In Wine Microbiology and Biotechnology; Fleet, G.M., Ed.; Harwood Academic Publishers: Chur, Switzerland, 1993; pp. 27–54. [Google Scholar]
  5. Le Jeune, C.; Erny, C.; Demuyter, C.; Lollier, M. Evolution of the population of Saccharomyces cerevisiae from grape to wine in a spontaneous fermentation. Food Microbiol. 2006, 23, 709–716. [Google Scholar] [CrossRef] [PubMed]
  6. Romano, P.; Capece, A.; Serafino, V.; Romaniello, R.; Poeta, C. Biodiversity of wild strains of Saccharomyces cerevisiae as tool to complement and optimize wine quality. World J. Microbiol. Biotechnol. 2008, 24, 1797–1802. [Google Scholar] [CrossRef]
  7. Tristezza, M.; Vetrano, C.; Bleve, G.; Spano, G.; Capozzi, V.; Logrieco, A.; Mita, G.; Grieco, F. Biodiversity and safety aspects of yeast strains characterized from vineyards and spontaneous fermentations in the Apulia Region, Italy. Food Microbiol. 2013, 36, 335–342. [Google Scholar] [CrossRef]
  8. Valero, E.; Schuller, D.; Cambon, B.; Casal, M.; Dequin, S. Dissemination and survival of commercial wine yeast in the vineyard: A large-scale, three-years study. FEMS Yeast Res. 2005, 5, 959–969. [Google Scholar] [CrossRef] [Green Version]
  9. Santamaría, P.; López, R.; del Patrocinio Garijo, M.; Escribano, R.; González-Arenzana, L.; López-Alfaro, I.; Gutiérrez, A.R. Biodiversity of Saccharomyces cerevisiae Yeasts in Spontaneous Alcoholic Fermentations: Typical Cellar or Zone Strains? In Advances in Grape and Wine Biotechnology; Morata, A., Loira, I., Eds.; IntechOpen: London, UK, 2019. [Google Scholar]
  10. Bokulich, N.A.; Thorngate, J.H.; Richardson, P.M.; Mills, D.A. Microbial biogeography of wine grapes is conditioned by cultivar, vintage, and climate. Proc. Natl. Acad. Sci. USA 2014, 111, E139–E148. [Google Scholar] [CrossRef] [Green Version]
  11. Knight, S.; Klaere, S.; Fedrizzi, B.; Goddard, M.R. Regional microbial signatures positively correlate with differential wine phenotypes: Evidence for a microbial aspect to terroir. Sci. Rep. 2015, 5, 14233. [Google Scholar] [CrossRef] [Green Version]
  12. Martínez-Romero, D.; Guillén, F.; Valverde, J.M.; Bailén, G.; Zapata, P.; Serrano, M.; Castrillo, S.; Valero, D. Influence of carvacrol on survival of Botrytis cinerea inoculated in table grapes. Int. J. Food Microbiol. 2007, 115, 144–148. [Google Scholar] [CrossRef]
  13. Raspor, P.; Milek, D.M.; Polanc, J.; Možina, S.S.; Čadež, N. Yeasts isolated from three varieties of grapes cultivated in different locations of the Dolenjska vine-growing region, Slovenia. Int. J. Food Microbiol. 2006, 109, 97–102. [Google Scholar] [CrossRef]
  14. Franco-Duarte, R.; Bigey, F.; Carreto, L.; Mendes, I.; Dequin, S.; Santos, M.A.; Pais, C.; Schuller, D. Intrastrain genomic and phenotypic variability of the commercial Saccharomyces cerevisiae strain Zymaflore VL1 reveals microevolutionary adaptation to vineyard environments. FEMS Yeast Res. 2015, 15, fov063. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  15. Belda, I.; Zarraonaindia, I.; Perisin, M.; Palacios, A.; Acedo, A. From vineyard soil to wine fermentation: Microbiome approximations to explain the “terroir” concept. Front. Microbiol. 2017, 8, 821. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  16. Gilbert, J.A.; van der Lelie, D.; Zarraonaindia, I. Microbial terroir for wine grapes. Proc. Natl. Acad. Sci. USA 2014, 111, 5–6. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  17. Morrison-Whittle, P.; Goddard, M.R. Quantifying the relative roles of selective and neutral processes in defining eukaryotic microbial communities. ISME J. 2015, 9, 2003–2011. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  18. Pinto, C.; Pinho, D.; Cardoso, R.; Custódio, V.; Fernandes, J.; Sousa, S.; Pinheiro, M.; Egas, C.; Gomes, A.C. Wine fermentation microbiome: A landscape from different Portuguese wine appellations. Front. Microbiol. 2015, 6, 905. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  19. Taylor, M.W.; Tsai, P.; Anfang, N.; Ross, H.A.; Goddard, M.R. Pyrosequencing reveals regional differences in fruit‐associated fungal communities. Environ. Microbiol. 2014, 16, 2848–2858. [Google Scholar] [CrossRef] [Green Version]
  20. Bokulich, N.A.; Collins, T.S.; Masarweh, C.; Allen, G.; Heymann, H.; Ebeler, S.E.; Mills, D.A. Associations among wine grape microbiome, metabolome, and fermentation behavior suggest microbial contribution to regional wine characteristics. mBio 2016, 7, e00631-16. [Google Scholar] [CrossRef] [Green Version]
  21. Drumonde-Neves, J.; Franco-Duarte, R.; Lima, T.; Schuller, D.; Pais, C. Association between grape yeast communities and the vineyard ecosystems. PLoS ONE 2017, 12, e0169883. [Google Scholar] [CrossRef]
  22. Gao, F.; Chen, J.; Xiao, J.; Cheng, W.; Zheng, X.; Wang, B.; Shi, X. Microbial community composition on grape surface controlled by geographical factors of different wine regions in Xinjiang, China. Food Res. Int. 2019, 122, 348–360. [Google Scholar] [CrossRef]
  23. Agarbati, A.; Canonico, L.; Ciani, M.; Comitini, F. The impact of fungicide treatments on yeast biota of Verdicchio and Montepulciano grape varieties. PLoS ONE 2019, 14, e0217385. [Google Scholar] [CrossRef] [Green Version]
  24. Mercado, L.; Dalcero, A.; Masuelli, R.; Combina, M. Diversity of Saccharomyces strains on grapes and winery surfaces: Analysis of their contribution to fermentative flora of Malbec wine from Mendoza (Argentina) during two consecutive years. Food Microbiol. 2007, 24, 403–412. [Google Scholar] [CrossRef] [PubMed]
  25. Schuller, D.; Casal, M. The use of genetically modified Saccharomyces cerevisiae strains in the wine industry. Appl. Microbiol. Biotechnol. 2005, 68, 292–304. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  26. Mortimer, R.; Polsinelli, M. On the origins of wine yeast. Res. Microbiol. 1999, 150, 199–204. [Google Scholar] [CrossRef]
  27. Agarbati, A.; Canonico, L.; Mancabelli, L.; Milani, C.; Ventura, M.; Ciani, M.; Comitini, F. The influence of fungicide treatments on mycobiota of grapes and its evolution during fermentation evaluated by metagenomic and culture-dependent methods. Microorganisms 2019, 7, 114. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  28. Martini, A.; Ciani, M.; Scorzetti, G. Direct enumeration and isolation of wine yeasts from grape surfaces. Am. J. Enol. Vitic. 1996, 47, 435–440. [Google Scholar]
  29. Mercado, L.; Sturm, M.E.; Rojo, M.C.; Ciklic, I.; Martínez, C.; Combina, M. Biodiversity of Saccharomyces cerevisiae populations in Malbec vineyards from the “Zona Alta del Río Mendoza” region in Argentina. Int. J. Food Microbiol. 2011, 151, 319–326. [Google Scholar] [CrossRef]
  30. Ciani, M.; Maccarelli, F. Oenological properties of non-Saccharomyces yeasts associated with wine-making. World J. Microbiol. Biotechnol. 1997, 14, 199–203. [Google Scholar] [CrossRef]
  31. Ciani, M.; Comitini, F. Yeast ecology of wine production. In Yeasts in the Production of Wine; Romano, P., Ciani, M., Fleet, G.H., Eds.; Springer: NewYork, NY, USA, 2019; pp. 1–42. [Google Scholar]
  32. Ocón, E.; Gutiérrez, A.R.; Garijo, P.; López, R.; Santamaría, P. Presence of non-Saccharomyces yeasts in cellar equipment and grape juice during harvest time. Food Microbiol. 2010, 27, 1023–1027. [Google Scholar] [CrossRef]
  33. Vaughan-Martini, A.; Martini, A. Facts, myths and legends on the prime industrial microorganism. J. Ind. Microbiol. 1995, 14, 514–522. [Google Scholar] [CrossRef]
  34. Legras, J.L.; Karst, F. Optimisation of interdelta analysis for Saccharomyces cerevisiae strain characterization. FEMS Microbiol. Lett. 2003, 221, 249–255. [Google Scholar] [CrossRef] [Green Version]
  35. Gallardo, G.; Ruiz-Moyano, S.; Hernández, A.; Benito, M.J.; Córdoba, M.G.; Pérez-Nevado, F.; Martín, A. Application of ISSR-PCR for rapid strain typing of Debaryomyces hansenii isolated from dry-cured Iberian ham. Food Microbiol. 2014, 42, 205–211. [Google Scholar] [CrossRef] [PubMed]
  36. Larpin, S.; Mondoloni, C.; Goerges, S.; Vernoux, J.P.; Guéguen, M.; Desmasures, N. Geotrichum candidum dominates in yeast population dynamics in Livarot, a French red-smear cheese. FEMS Yeast Res. 2006, 6, 1243–1253. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  37. EC Regulation No. 2870/00; Community reference methods for the analysis of spirit drinks. Official Journal of the European Communities: Luxembourg, 19 December 2000.
  38. Dukes, B.C.; Butzke, C.E. Rapid determination of primary amino acids in grape juice using an o-phthaldialdehyde/Nacetyl-L-cysteine spectrophotometric assay. Am. J. Enol. Vitic. 1998, 49, 125–134. [Google Scholar]
  39. Helrich, K. Official Methods of Analysis of the Association of Official Analytical Chemists; AOAC: Rockville, MD, USA, 1990. [Google Scholar]
  40. Canonico, L.; Comitini, F.; Ciani, M. Influence of vintage and selected starter on Torulaspora delbrueckii/Saccharomyces cerevisiae sequential fermentation. Eur. Food Res. Technol. 2015, 241, 827–833. [Google Scholar] [CrossRef]
  41. Vaudano, E.; Quinterno, G.; Costantini, A.; Pulcini, L.; Pessione, E.; García-Moruno, E. Yeast distribution in Grignolino grapes growing in a new vineyard in Piedmont and the technological characterization of indigenous Saccharomyces spp. strains. Int. J. Food Microbiol. 2019, 289, 154–161. [Google Scholar] [CrossRef] [Green Version]
  42. van Leeuwen, C.; Barbe, J.C.; Darriet, P.; Geffroy, O.; Gomès, E.; Guillaumie, S.; Helwi, P.; Laboyrie, J.; Lytra, G.; Le Menn, N.; et al. Recent advancements in understanding the terroir effect on aromas in grapes and wines. Oeno One 2020, 54, 985–1006. [Google Scholar] [CrossRef]
  43. Bisson, L.F. Geographic origin and diversity of wine strains of Saccharomyces. Am. J. Enol. Vitic. 2012, 63, 165–175. [Google Scholar] [CrossRef] [Green Version]
  44. Börlin, M.; Venet, P.; Claisse, O.; Salin, F.; Legras, J.L.; Masneuf-Pomarede, I. Cellar-associated Saccharomyces cerevisiae population structure revealed high level diversity and perennial persistence at Sauternes wine estates. Appl. Environ. Microbiol. 2016, 82, 2909–2918. [Google Scholar] [CrossRef] [Green Version]
  45. Granchi, L.; Ganucci, D.; Buscioni, G.; Mangani, S.; Guerrini, S. The biodiversity of Saccharomyces cerevisiae in spontaneous wine fermentation: The occurrence and persistence of winery-strains. Fermentation 2019, 5, 86. [Google Scholar] [CrossRef] [Green Version]
  46. Torija, M.J.; Rozes, N.; Poblet, M.; Guillamón, J.M.; Mas, A. Yeast population dynamics in spontaneous fermentations: Comparison between two different wine-producing areas over a period of three years. Antonie Van Leeuwenhoek 2001, 79, 345–352. [Google Scholar] [CrossRef]
  47. Ganucci, D.; Guerrini, S.; Mangani, S.; Vincenzini, M.; Granchi, L. Quantifying the effects of ethanol and temperature on the fitness advantage of predominant Saccharomyces cerevisiae strains occurring in spontaneous wine fermentations. Front. Microbiol. 2018, 9, 1563. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  48. Blanco, P.; Ramilo, A.; Cerdeira, M.; Orriols, I. Genetic diversity of wine Saccharomyces cerevisiae strains in an experimental winery from Galicia (NW Spain). Antonie Van Leeuwenhoek 2006, 89, 351–357. [Google Scholar] [CrossRef] [PubMed]
  49. Li, S.S.; Cheng, C.; Li, Z.; Chen, J.Y.; Yan, B.; Han, B.Z.; Reeves, M. Yeast species associated with wine grapes in China. Int. J. Food Microbiol. 2010, 138, 85–90. [Google Scholar] [CrossRef] [PubMed]
  50. Schuller, D.; Cardoso, F.; Sousa, S.; Gomes, P.; Gomes, A.C.; Santos, M.A.S.; Casal, M. Genetic diversity and population structure of Saccharomyces cerevisiae strains isolated from different grape varieties and winemaking regions. PLoS ONE 2012, 7, e32507. [Google Scholar] [CrossRef] [Green Version]
  51. Franco-Duarte, R.; Mendes, I.; Gomes, A.C.; Santos, M.A.; de Sousa, B.; Schuller, D. Genotyping of Saccharomyces cerevisiae strains by interdelta sequence typing using automated microfluidics. Electrophoresis 2011, 32, 1447–1455. [Google Scholar] [CrossRef] [Green Version]
  52. Alexandre, H. Wine yeast terroir: Separating the wheat from the chaff—for an open debate. Microorganisms 2020, 8, 787. [Google Scholar] [CrossRef]
  53. Pérez-Torrado, R.; Rantsiou, K.; Perrone, B.; Navarro-Tapia, E.; Querol, A.; Cocolin, L. Saccharomyces cerevisiae strains: Insight into the dominance phenomenon. Sci. Rep. 2017, 7, 43603. [Google Scholar] [CrossRef]
  54. Ugliano, M.; Henschke, P.A. Yeasts and wine flavour. In Wine Chemistry and Biochemistry; Moreno-Arribas, M.V., Polo, M.C., Eds.; Springer: New York, NY, USA, 2009; pp. 251–274. [Google Scholar]
  55. Mendes, I.; Sanchez, I.; Franco-Duarte, R.; Camarasa, C.; Schuller, D.; Dequin, S.; Sousa, M.J. Integrating transcriptomics and metabolomics for the analysis of the aroma profiles of Saccharomyces cerevisiae strains from diverse origins. BMC Genom. 2017, 18, 1–13. [Google Scholar] [CrossRef] [Green Version]
  56. Nikolaou, E.; Soufleros, E.H.; Bouloumpasi, E.; Tzanetakis, N. Selection of indigenous Saccharomyces cerevisiae strains according to their oenological characteristics and vinification results. Food Microbiol. 2006, 23, 205–211. [Google Scholar] [CrossRef]
  57. Englezos, V.; Rantsiou, K.; Cravero, F.; Torchio, F.; Pollon, M.; Fracassetti, D.; Ortiz-Julien, A.; Gerbi, V.; Rolle, L.; Cocolin, L. Volatile profile of white wines fermented with sequential inoculation of Starmerella bacillaris and Saccharomyces cerevisiae. Food Chem. 2018, 257, 350–360. [Google Scholar] [CrossRef]
  58. Sadoudi, M.; Tourdot-Maréchal, R.; Rousseaux, S.; Steyer, D.; Gallardo-Chacón, J.J.; Ballester, J.; Vichi, S.; Guérin-Schneider, R.; Caixach, J.; Alexandre, H. Yeast–yeast interactions revealed by aromatic profile analysis of Sauvignon Blanc wine fermented by single or co-culture of non-Saccharomyces and Saccharomyces yeasts. Food Microbiol. 2012, 32, 243–253. [Google Scholar] [CrossRef] [PubMed]
  59. Soles, R.M.; Ough, C.S.; Kunkee, R.E. Ester concentration differences in wine fermented by various species and strains of yeasts. Am. J. Enol. Vitic. 1982, 33, 94–98. [Google Scholar]
  60. Franco-Duarte, R.; Umek, L.; Mendes, I.; Castro, C.C.; Fonseca, N.; Martins, R.; Silva-Ferreira, A.C.; Sampaio, P.; Pais, C.; Schuller, D. New integrative computational approaches unveil the Saccharomyces cerevisiae pheno-metabolomic fermentative profile and allow strain selection for winemaking. Food Chem. 2016, 211, 509–520. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  61. Nisiotou, A.; Sgouros, G.; Mallouchos, A.; Nisiotis, C.S.; Michaelidis, C.; Tassou, C.; Banilas, G. The use of indigenous Saccharomyces cerevisiae and Starmerella bacillaris strains as a tool to create chemical complexity in local wines. Food Res. Int. 2018, 111, 498–508. [Google Scholar] [CrossRef] [PubMed]
  62. Rantsiou, K.; Dolci, P.; Giacosa, S.; Torchio, F.; Tofalo, R.; Torriani, S.; Suzzi, G.; Rolle, L.; Cocolin, L. Candida zemplinina can reduce acetic acid produced by Saccharomyces cerevisiae in sweet wine fermentations. Appl. Environ. Microbiol. 2012, 78, 1987–1994. [Google Scholar] [CrossRef] [Green Version]
  63. Banilas, G.; Sgouros, G.; Nisiotou, A. Development of microsatellite markers for Lachancea thermotolerans typing and population structure of wine-associated isolates. Microbiol. Res. 2016, 193, 1–10. [Google Scholar] [CrossRef]
Fermentation 08 00224 g001 550
Figure 1. Fermentation kinetic of S. cerevisiae biotypes I and II in comparison with three commercial starters. (A) Fermentation carried out at 16 °C. (B) Fermentation carried out at 22 °C.
Figure 1. Fermentation kinetic of S. cerevisiae biotypes I and II in comparison with three commercial starters. (A) Fermentation carried out at 16 °C. (B) Fermentation carried out at 22 °C.
Fermentation 08 00224 g001
Fermentation 08 00224 g002 550
Figure 2. Principal Component Analyses (PCA) for the main analytical characteristics and volatile compounds of wines obtained by two biotypes at 16 °C (A) and 22 °C (B) Fermentation temperatures.
Figure 2. Principal Component Analyses (PCA) for the main analytical characteristics and volatile compounds of wines obtained by two biotypes at 16 °C (A) and 22 °C (B) Fermentation temperatures.
Fermentation 08 00224 g002
Table
Table 1. Source and number of S. cerevisiae isolates during 2016 and 2019 vintages.
Table 1. Source and number of S. cerevisiae isolates during 2016 and 2019 vintages.
Sampling YearSource of S. cerevisiaeNumber of SamplesNumber of Samples with S. cerevisiaeNumber
of S. cerevisiae
Isolated
2016Grapes (direct isolation)1000
Grapes (after autoenrichment)10210
Winery equipment and environment12413
Spontaneous fermentation2220
2019Grapes (direct isolation)600
Grapes (after autoenrichment)6227
Winery equipment and environment918
Spontaneous fermentation2220
Table
Table 2. Occurrence of S. cerevisiae biotypes in different years and samples. Percentage values were calculated on the basis of the number of isolates of the same year. For both vintages (2016/2019), the percentage values were calculated considering the sum of the isolates of both sampling years. * No presence of S. cerevisiae.
Table 2. Occurrence of S. cerevisiae biotypes in different years and samples. Percentage values were calculated on the basis of the number of isolates of the same year. For both vintages (2016/2019), the percentage values were calculated considering the sum of the isolates of both sampling years. * No presence of S. cerevisiae.
Sampling YearNumber of IsolatesBiotypeOrigin of SamplingTotal of Yeast for Each Biotype
GrapeWinery EnvironmentWine Fermentation
201643I6 (14%)12 (28%)11 (26%)29 (68%)
II/ *1 (2%)7 (16%)8 (19%)
III2 (5%)//2 (5%)
IV1 (2%)//1 (2%)
V//1 (2%)1 (2%)
VI1 (2%)//1 (2%)
VII//1 (2%)1 (2%)
VIII////
IX////
201955I13 (24%)5 (9%)6 (11%)24 (43%)
II13 (24%)3 (5%)7 (13%)23 (42%)
III1 (2%)/2 (4%)3 (5%)
IV//1 (2%)1 (2%)
V//2 (4%)2 (4%)
VI////
VII////
VIII//1 (2%)1 (2%)
IX//1 (2%)1 (2%)
2016/201998I19 (19%)17 (17%)17 (17%)53 (54%)
II13 (13%)4 (4%)14 (14%)31 (32%)
III3 (3%)/2 (2%)5 (5%)
IV1 (1%)/1 (1%)2 (2%)
V//3 (3%)3 (3%)
VI1 (1%)//1 (1%)
VII//1 (1%)1 (1%)
VIII//1 (1%)1 (1%)
IX//1 (1%)1 (1%)
Table
Table 3. Main analytical characteristics of wines obtained by the two dominant biotypes compared with commercial starter strains, fermented at 16 °C. Data are means ± standard deviations, and those with different superscript letters (a–e) within each column are significantly different, according to Duncan’s test (p-value < 0.05).
Table 3. Main analytical characteristics of wines obtained by the two dominant biotypes compared with commercial starter strains, fermented at 16 °C. Data are means ± standard deviations, and those with different superscript letters (a–e) within each column are significantly different, according to Duncan’s test (p-value < 0.05).
SamplesFermentation Rate (gCO2/Day) *Volatile Acidity (g/L)Total SO2 (mg/L)Acetaldehyde (mg/L)Free α-Amino Acids (mgN/L)Ethanol
(% v/v)
Biotype I0.9 ± 0.1 c0.7 ± 0.1 a4.0 ± 0.0 e9.5 ± 0.7 d35.8 ± 0.7 d12.3 ± 0.1 a
Biotype II1.1 ± 0.0 b0.3 ± 0.0 bc41.0 ± 1.4 a62.5 ± 0.76 a32.2 ± 0.6 e11.7 ± 0.4 b
OKAY1.1 ± 0.1 bc0.3 ± 0.0 c11.0 ± 0.7 d40.1 ± 0.3 c43.6 ± 0.9 b12.2 ± 0.2 a
EC11181.2 ± 0.0 b0.4 ± 0.1 b23.0 ± 0.7 b48.4 ± 0.9 b38.8 ± 0.6 c12.3 ± 0.2 a
VIN131.4 ± 0.1 a0.2 ± 0.0 d16.0 ± 0.7 c47.3 ± 0.5 b46.0 ± 0.7 a11.9 ± 0.1 ab
* Fermentation rate calculated over the 3rd day of fermentation. Sugars (glucose/fructose) at the end of the fermentation were completely consumed.
Table
Table 4. Main analytical characteristics of wines obtained by the two dominant biotypes compared with commercial starter strains, fermented at 22 °C. Data are means ± standard deviations, and those with different superscript letters (a–e) within each column are significantly different, according to Duncan’s test (p-value < 0.05).
Table 4. Main analytical characteristics of wines obtained by the two dominant biotypes compared with commercial starter strains, fermented at 22 °C. Data are means ± standard deviations, and those with different superscript letters (a–e) within each column are significantly different, according to Duncan’s test (p-value < 0.05).
SamplesFermentation Rate (gCO2/Day) *Volatile Acidity (g/L)Total SO2 (mg/L)Acetaldehyde (mg/L)Free α-Amino Acids (mgN/L)Ethanol
(% v/v)
Biotype I1.2 ± 0.1 a0.3 ± 0.1 bc2.0 ± 0.0 c45.7 ± 0.6 b40.3 ± 0.5 b12.3 ± 0.1 b
Biotype II1.1 ± 0.1 ab0.2 ± 0.0 c24.0 ± 0.7 a19.2 ± 1.0 e45.5 ± 0.2 a12.1 ± 0.3 b
OKAY1.0 ± 0.1 b0.4 ± 0.1 ab3.0 ± 0.0 c25.3 ± 0.5 c45.9 ± 0.3 a12.3 ± 0.0 b
EC11181.2 ± 0.0 a0.3 ± 0.0 bc10.0 ± 0.7 b21.3 ± 0.9 d39.6 ± 0.9 b12.7 ± 0.1 a
VIN131.2 ± 0.0 a0.5 ± 0.1 a2.0 ± 0.0 c74.6 ± 0.6 a45.9 ± 0.9 a12.2 ± 0.0 b
* Fermentation rate calculated over the 3rd day of fermentation. The residual sugars (glucose/fructose) at the end of the fermentation were totally consumed.
Table
Table 5. Main volatile compounds of wines obtained by the two native S. cerevisiae strains compared with commercial starters, fermented at 16 °C. Data are means ± standard deviations, and those with different superscript letters (a–d) within each row are significantly different, according to Duncan’s tests (p-value < 0.05).
Table 5. Main volatile compounds of wines obtained by the two native S. cerevisiae strains compared with commercial starters, fermented at 16 °C. Data are means ± standard deviations, and those with different superscript letters (a–d) within each row are significantly different, according to Duncan’s tests (p-value < 0.05).
Volatile CompoundsBiotype IBiotype IIOKAYEC1118VIN13
Alcohols
Hexanol (µg/L)10.0 ± 0.0 b20.0 ± 0.0 a20.0 ± 0.0 a20.0 ± 0.0 a20.0 ± 0.0 a b
β-Phenyl ethanol (mg/L)29.1 ± 0.4 b24.8 ± 0.4 d20.2 ± 0.9 e50.5 ± 0.1 a27.2 ± 1.1 c
Esters
Isoamyl acetate (mg/L)2.5 ± 0.1 a2.2 ± 0.0 b1.2 ± 0.1 e1.6 ± 0.1 c1.3 ± 0.1 d
Phenylethyl acetate (mg/L)0.3 ± 0.0 c0.3 ± 0.0 c0.2 ± 0.0 d0.6 ± 0.0 b0.7 ± 0.0 a
Ethyl hexanoate (mg/L)0.2 ± 0.0 d0.4 ± 0.0 a0.3 ± 0.0 b0.2 ± 0.0 cd0.2 ± 0.0 c
Ethyl butyrate (mg/L)0.2 ± 0.0 d0.3 ± 0.0 c1.4 ± 0.0 a0.7 ± 0.0 b0.7 ± 0.0 b
Ethyl octanoate (µg/L)50.0 ± 0.1 a30.0 ± 0.1 b10.0 ± 0.0 b20.0 ± 0.0 b10.0 ± 0.0 b
Diethyl succinate (µg/L)10.0 ± 0.1 a20.0 ± 0.0 a10.0 ± 0.1 a10.0 ± 0.0 a10.0 ± 0.0 a
Terpenes
Linalool (µg/L)40.0 ± 0.1 a50.0 ± 0.1 a60.0 ± 0.1 a50.0 ± 0.1 a10.0 ± 0.0 b
Nerol (µg/L)30.0 ± 0.1 a20.0 ± 0.1 a20.0 ± 0.1 a20.0 ± 0.0 a20.0 ± 0.0 a
Geraniol (µg/L)30.0 ± 0.0 a40.0 ± 0.1 a10.0 ± 0.0 a0.0 ± 0.0 a10.0 ± 0.0 a
Enones
β-Damascenone (µg/L)10.0 ± 0.0 ab30.0 ± 0.1 a10.0 ± 0.0 b10.0 ± 0.0 b10.0 ± 0.1 b
Table
Table 6. Main volatile compounds of wines carried out at 22 °C. Data are means ± standard deviations, and those with different superscript letters (a–d) within each column are significantly different, according to Duncan’s tests (p-value < 0.05).
Table 6. Main volatile compounds of wines carried out at 22 °C. Data are means ± standard deviations, and those with different superscript letters (a–d) within each column are significantly different, according to Duncan’s tests (p-value < 0.05).
Volatile CompoundsBiotype IBiotype IIOKAYEC1118VIN13
Alcohols
Hexanol (µg/L)20.0 ± 0.0 a10.0 ± 0.0 a20.0 ± 0.1 a10.0 ± 0.0 a20.0 ± 0.1 a
β-Phenyl ethanol (mg/L)30.7 ± 3.1 c12.9 ± 2.9 e35.2 ± 0.7 a32.7 ± 2.9 b16.3 ± 2.1 d
Esters
Isoamyl acetate (mg/L)3.8 ± 0.3 a1.8 ± 0.1 b1.3 ± 0.0 c1.1 ± 0.1 c1.5 ± 0.0 bc
Phenylethyl acetate (mg/L)0.2 ± 0.0 b0.1 ± 0.0 c0.1 ± 0.0 bc0.4 ± 0.1 a0.1 ± 0.0 c
Ethyl hexanoate (mg/L)0.2 ± 0.1 c0.4 ± 0.2 b0.36 ± 0.04 b0.43 ± 0.04 a0.21 ± 0.03 c
Ethyl butyrate (mg/L)0.3 ± 0.0 c0.4 ± 0.0 c1.3 ± 0.2 a0.5 ± 0.1 bc0.5 ± 0.0 b
Ethyl octanoate (µg/L)40.0 ± 0.1 a40.0 ± 0.1 a40.0 ± 0.2 a10.0 ± 0.0 b30.0 ± 0.1 a
Diethyl succinate µg/L)20.0 ± 0.0 b10.0 ± 0.0 c60.0 ± 0.1 a10.0 ± 0.0 c10.0 ± 0.0 c
Terpenes
Linalool (µg/L)40.0 ± 0.01 b30.0 ± 0.1 b60.0 ± 0.2 a40.0 ± 0.1 b20.0 ± 0.1 b
Nerol (µg/L)10.0 ± 0.01 b30.0 ± 0.0 a30.0 ± 0.1 a20.0 ± 0.1 b10.0 ± 0.0 b
Geraniol (µg/L)10.0 ± 0.0 ab0.0 ± 0.0 b30.0 ± 0.1 a20.0 ± 0.1 ab10.0 ± 0.0 a b
Enones
β-Damascenone (µg/L)10.0 ± 0.0 b20.0 ± 0.1 a10.0 ± 0.0 b10.0 ± 0.0 b10.0 ± 0.0 b
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Share and Cite

MDPI and ACS Style

Agarbati, A.; Canonico, L.; Comitini, F.; Ciani, M. Ecological Distribution and Oenological Characterization of Native Saccharomyces cerevisiae in an Organic Winery. Fermentation 2022, 8, 224. https://doi.org/10.3390/fermentation8050224

AMA Style

Agarbati A, Canonico L, Comitini F, Ciani M. Ecological Distribution and Oenological Characterization of Native Saccharomyces cerevisiae in an Organic Winery. Fermentation. 2022; 8(5):224. https://doi.org/10.3390/fermentation8050224

Chicago/Turabian Style

Agarbati, Alice, Laura Canonico, Francesca Comitini, and Maurizio Ciani. 2022. "Ecological Distribution and Oenological Characterization of Native Saccharomyces cerevisiae in an Organic Winery" Fermentation 8, no. 5: 224. https://doi.org/10.3390/fermentation8050224

APA Style

Agarbati, A., Canonico, L., Comitini, F., & Ciani, M. (2022). Ecological Distribution and Oenological Characterization of Native Saccharomyces cerevisiae in an Organic Winery. Fermentation, 8(5), 224. https://doi.org/10.3390/fermentation8050224

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop