3.0 Strategies for Yeast Genetic Improvement in Brewing and Winemaking
Abstract
1. Introduction
2. Non-GMO Genetic Improvement Techniques
2.1. Classical Genetic Improvement Techniques for Non-GMO Yeasts
2.1.1. Random Mutagenesis
2.1.2. Sexual Breeding
2.2. Innovative Genetic Improvement Techniques in Fermentation
2.2.1. Adaptive Laboratory Evolution (ALE)
2.2.2. Big Data, AI, and Omics
2.2.3. Synthetic Microbial Communities
Product | Application | Involved Species | Work |
---|---|---|---|
Wine | Malolactic fermentation for flavor complexity in wine | Oenococcus oeni | [106] |
Adaptation to vineyard microbial terroir | Various LAB and yeast species | [107] | |
Optimization of yeast interactions for improved fermentation | K. apiculate, C. stellata, C. pulcherrima | [108] | |
Influence of nutrient scarcity, oxygen availability, and ethanol on fermentation | Saccharomyces cerevisiae and other yeasts | [109] | |
Ecological interactions driving fermentation outcomes | Multiple yeast species | [110] | |
Persistence of certain non-Saccharomyces yeasts in vineyard ecosystems | Starmerella bacillaris, Lachancea thermotolerans | [111] | |
Yeast ecosystem modulation by S. cerevisiae | Various non-Saccharomyces species | [99] | |
Beer | Bio-acidification and microbial control in beer | Lactic acid bacteria (LAB) | [112] |
Beer | Reduction in final ethanol content | Saccharomyces cerevisiae, Lentilactobacillus brevis | [113] |
3. GMO-Based Genetic Improvement Techniques
3.1. Synthetic Biology and CRISPR/Cas9
Product | Process | Technique | Ref. |
---|---|---|---|
Wine | GPD1 overexpression and ALD6 deletion to reduce alcohol yield in wine yeast | Episomal vector; KanMX deletion cassette | [124] |
Expression of extracellular hydrolytic enzymes to improve juice extraction and release primary aromas | Episomal vector constructed by restriction cloning | [125] | |
Reduction in urea and ethyl carbamate formation | CRISPR/Cas9 | [126] | |
Expression of malolactic enzymes to degrade malate and integrate malolactic fermentation | Episomal vector constructed by restriction cloning | [127] | |
Beer/Wine | Heterologous expression of pediocin to increase resistance to wild yeasts and bacteria | Episomal vector constructed by restriction cloning | [128] |
Beer | Engineering yeast strains to produce methyl anthranilate with grape aroma | CRISPR/Cas9 | [129] |
Expression of acetolactate decarboxylase (ALDC) to reduce diacetyl formation | Episomal vector constructed by restriction cloning | [130] | |
Engineering yeast strains to produce hop monoterpenes | Plasmids obtained by Golden Gate assembly, CRISPR/Cas | [122] |
3.2. Ethical and Commercial Challenges
4. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
- Saerens, S.M.G.; Duong, C.T.; Nevoigt, E. Genetic improvement of brewer’s yeast: Current state, perspectives and limits. Appl. Microbiol. Biotechnol. 2010, 86, 1195–1212. [Google Scholar] [CrossRef] [PubMed]
- Nielsen, J. Production of biopharmaceutical proteins by yeast: Advances through metabolic engineering. Bioengineered 2013, 4, 207–211. [Google Scholar] [CrossRef] [PubMed]
- Botstein, D.; Chervitz, S.A.; Cherry, M. Yeast as a Model Organism. Science 1997, 277, 1259–1260. [Google Scholar] [CrossRef] [PubMed]
- Gobbi, L.; Stanković, M.; Ruggeri, M.; Savastano, M. Craft Beer in Food Science: A Review and Conceptual Framework. Beverages 2024, 10, 91. [Google Scholar] [CrossRef]
- Singh, S. Beer Market Report 2025 (Global Edition). Available online: https://www.cognitivemarketresearch.com/beer-market-report?campaign_source=google_ads&campaign_name=CMR_DPMax_EURO01&gad_source=1&gad_campaignid=21894073608&gclid=Cj0KCQjw0qTCBhCmARIsAAj8C4YL25UW3xkvR4a3hrIno1E3kafRq4dSOkS_NQcDDlEXrgQ4DYVksWoaAnCHEALw_wcB (accessed on 11 June 2025).
- Europe Craft Beer Market Report by Product Type (Ales, Lagers, and Others), Age Group (21–35 Years Old, 40–54 Years Old, 55 Years and Above), Distribution Channel (On-Trade, Off-Trade), and Country 2025–2033. 2025. Available online: https://www.imarcgroup.com/europe-craft-beer-market (accessed on 11 June 2025).
- Garavaglia, C. Definition, history, and market of craft beer. In Craft Beer; Academic Press: Cambridge, MA, USA, 2025; pp. 1–13. [Google Scholar]
- Pretorius, I.S. Tailoring wine yeast for the new millennium: Novel approaches to the ancient art of winemaking. Yeast 2000, 16, 675–729. [Google Scholar] [CrossRef]
- Bonatto, D. The diversity of commercially available ale and lager yeast strains and the impact of brewer’s preferential yeast choice on the fermentative beer profiles. Int. Food Res. J. 2021, 141, 110125. [Google Scholar] [CrossRef]
- Libkind, D.; Hittinger, C.T.; Valério, E.; Gonçalves, C.; Dover, J.; Johnston, M.; Gonçalves, P.; Sampaio, J.P. Microbe domestication and the identification of the wild genetic stock of lager-brewing yeast. Proc. Natl. Acad. Sci. USA 2011, 108, 14539–14544. [Google Scholar] [CrossRef]
- Gibson, B.; Dahabieh, M.; Krogerus, K.; Jouhten, P.; Magalhães, F.; Pereira, R.; Siewers, V.; Vidgren, V. Adaptive Laboratory Evolution of Ale and Lager Yeasts for Improved Brewing Efficiency and Beer Quality. Annu. Rev. Food Sci. Technol. 2020, 11, 23–44. [Google Scholar] [CrossRef]
- Krogerus, K.; Magalhães, F.; Vidgren, V.; Gibson, B. New lager yeast strains generated by interspecific hybridization. J. Ind. Microbiol. Biotechnol. 2015, 42, 769–778. [Google Scholar] [CrossRef]
- Hutzler, M.; Morrissey, J.P.; Laus, A.; Meussdoerffer, F.; Zarnkow, M. A new hypothesis for the origin of the lager yeast Saccharomyces pastorianus. FEMS Yeast Res. 2023, 23, foad023. [Google Scholar] [CrossRef]
- Steensels, J.; Gallone, B.; Verstrepen, K.J. Interspecific hybridization as a driver of fungal evolution and adaptation. Nat. Rev. Microbiol. 2021, 19, 485–500. [Google Scholar] [CrossRef]
- Zavaleta, V.; Pérez-Través, L.; Saona, L.A.; Villarroel, C.A.; Querol, A.; Cubillos, F.A. Understanding brewing trait inheritance in de novo lager yeast hybrids. mSystems 2024, 9, e00762-24. [Google Scholar] [CrossRef]
- Nasuti, C.; Solieri, L. Yeast Bioflavoring in Beer: Complexity Decoded and Built up Again. Fermentation 2024, 10, 183. [Google Scholar] [CrossRef]
- Li, C.; Zhang, S.; Dong, G.; Bian, M.; Liu, X.; Dong, X.; Xia, T. Multi-omics study revealed the genetic basis of beer flavor quality in yeast. LWT 2022, 168, 113932. [Google Scholar] [CrossRef]
- Saerens, S.; Swiegers, J.H.; Chr Hansen, A.S. Enhancement of Beer Flavor by a Combination of Pichia Yeast and Different Hop Varieties. U.S. Patent US 10,544,385, 28 January 2020. [Google Scholar]
- Giudici, P.; Solieri, L.; Pulvirenti, A.M.; Cassanelli, S. Strategies and perspectives for genetic improvement of wine yeasts. Appl. Microbiol. Biotechnol. 2005, 666, 22–28. [Google Scholar] [CrossRef]
- Pérez-Torrado, R.; Querol, A.; Guillamón, J.M. Genetic improvement of non-GMO wine yeasts: Strategies, advantages and safety. Trends Food Sci. Technol. 2015, 45, 1–11. [Google Scholar] [CrossRef]
- Molinet, J.; Navarrete, J.P.; Villarroel, C.A.; Villarreal, P.; Sandoval, F.I.; Nespolo, R.F.; Stelkens, R.; Cubillos, F.A. Wild Patagonian yeast improve the evolutionary potential of novel interspecific hybrid strains for lager brewing. PLoS Genet. 2024, 20, e1011154. [Google Scholar] [CrossRef]
- Osburn, K.; Amaral, J.; Metcalf, S.R.; Nickens, D.M.; Rogers, C.M.; Sausen, C.; Caputo, R.; Miller, J.; Li, H.; Tennessen, J.M.; et al. Primary souring: A novel bacteria-free method for sour beer production. Food Microbiol. 2018, 70, 76–84. [Google Scholar] [CrossRef]
- Domizio, P.; House, J.F.; Joseph, C.M.; Bisson, L.F.; Bamforth, C.W. Lachancea thermotolerans as an alternative yeast for the production of beer. J. Inst. Brew. 2016, 122, 599–604. [Google Scholar] [CrossRef]
- Varela, C.; Borneman, A.R. Yeasts found in vineyards and wineries. Yeast 2017, 34, 111–128. [Google Scholar] [CrossRef]
- Iattici, F.; Catallo, M.; Solieri, L. Designing new yeasts for craft brewing: When natural biodiversity meets biotechnology. Beverages 2020, 6, 3. [Google Scholar] [CrossRef]
- 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]
- Peter, J.; De Chiara, M.; Friedrich, A.; Yue, J.X.; Pflieger, D.; Bergström, A.; Sigwalt, A.; Barre, B.; Freel, K.; Llored, A.; et al. Genome evolution across 1,011 Saccharomyces cerevisiae isolates. Nature 2018, 556, 339–344. [Google Scholar] [CrossRef] [PubMed]
- Pretorius, I.S.; Boeke, J.D. Yeast 2.0—Connecting the dots in the construction of the world’s first functional synthetic eukaryotic genome. FEMS Yeast Res. 2018, 18, foy032. [Google Scholar] [CrossRef]
- Goold, H.D.; Kroukamp, H.; Erpf, P.E.; Zhao, Y.; Kelso, P.; Calame, J.; Timmins, J.J.; Wightman, E.L.; Peng, K.; Carpenter, A.C.; et al. Construction and iterative redesign of synXVI a 903 kb synthetic Saccharomyces cerevisiae chromosome. Nat. Commun. 2025, 16, 841. [Google Scholar] [CrossRef]
- Rous, C.V.; Snow, R.; Kunkee, R.E. Reduction of higher alcohols by fermentation with a leucine-auxotrophic mutant of wine yeast. J. Inst. Brew. 1983, 89, 274–278. [Google Scholar] [CrossRef]
- Cordente, A.G.; Solomon, M.; Schulkin, A.; Leigh Francis, I.; Barker, A.; Borneman, A.R.; Curtin, C.D. Novel wine yeast with ARO4 and TYR1 mutations that overproduce ‘floral’aroma compounds 2-phenylethanol and 2-phenylethyl acetate. Appl. Microbiol. Biotechnol. 2018, 102, 5977–5988. [Google Scholar] [CrossRef]
- Diderich, J.A.; Weening, S.M.; Van den Broek, M.; Pronk, J.T.; Daran, J.M. Selection of Pof-Saccharomyces eubayanus variants for the construction of S. cerevisiae × S. eubayanus hybrids with reduced 4-vinyl guaiacol formation. Front. Microbiol. 2018, 9, 1640. [Google Scholar] [CrossRef]
- Jabłoński, S.J.; Mielko-Niziałek, K.A.; Leszczyński, P.; Gasiński, A.; Kawa-Rygielska, J.; Młynarz, P.; Łukaszewicz, M. Examination of internal metabolome and VOCs profile of brewery yeast and their mutants producing beer with improved aroma. Sci. Rep. 2024, 14, 14582. [Google Scholar] [CrossRef]
- Gorter de Vries, A.R.; Knibbe, E.; Van Roosmalen, R.; Van den Broek, M.; de la Torre Cortés, P.; O’Herne, S.F.; Vijverberg, P.A.; El Masoudi, A.; Brouwers, N.; Pronk, J.T.; et al. Improving industrially relevant phenotypic traits by engineering chromosome copy number in Saccharomyces pastorianus. Front. Genet. 2020, 11, 518. [Google Scholar] [CrossRef]
- Kumari, R.; Pramanik, K. Improvement of multiple stress tolerance in yeast strain by sequential mutagenesis for enhanced bioethanol production. J. Biosci. Bioeng. 2012, 114, 622–629. [Google Scholar] [CrossRef] [PubMed]
- Salmon, J.M.; Barre, P. Improvement of Nitrogen Assimilation and Fermentation Kinetics under Enological Conditions by Derepression of Alternative Nitrogen-Assimilatory Pathways in an Industrial Saccharomyces cerevisiae Strain. Appl. Environ. Microbiol. 1998, 64, 3831–3837. [Google Scholar] [CrossRef] [PubMed]
- Quirós, M.; Gonzalez-Ramos, D.; Tabera, L.; Gonzalez, R. A new methodology to obtain wine yeast strains overproducing mannoproteins. Int. J. Food Microbiol. 2010, 139, 9–14. [Google Scholar] [CrossRef]
- Cordente, A.G.; Cordero-Bueso, G.; Pretorius, I.S.; Curtin, C.D. Novel wine yeast with mutations in YAP1 that produce less acetic acid during fermentation. FEMS Yeast Res. 2013, 13, 62–73. [Google Scholar] [CrossRef]
- Wöhrmann, K.; Lance, P. The Polymorphism of Esterases in Yeast (Saccharomyces cerevisiae). J. Inst. Brew. 1980, 86, 174. [Google Scholar] [CrossRef]
- Giudici, P.; Zinnato, A. Influenza dell’uso di mutanti nutrizionali sulla produzione di alcooli superiori. Vignevini 1983, 10, 63–65. [Google Scholar]
- Liu, Z.; Zhang, G.; Sun, Y. Mutagenizing brewing yeast strain for improving fermentation property of beer. J. Biosci. Bioeng. 2008, 106, 33–38. [Google Scholar] [CrossRef]
- Sipiczki, M. Interspecies hybridisation and genome chimerisation in Saccharomyces: Combining of gene pools of species and its biotechnological perspectives. Front. Microbiol. 2018, 9, 3071. [Google Scholar] [CrossRef]
- Eschenbruch, R.; Cresswell, K.J.; Fisher, B.M.; Thornton, R.J. Selective hybridisation of pure culture wine yeasts. Appl. Microbiol. Biotechnol. 1982, 14, 155–158. [Google Scholar] [CrossRef]
- Shinohara, T.; Mamiya, S.; Yanagida, F. Introduction of flocculation property into wine yeasts (Saccharomyces cerevisiae) by hybridization. J. Ferment. Bioeng. 1997, 83, 96–101. [Google Scholar] [CrossRef]
- Rainieri, S.; Zambonelli, C.; Tini, V.; Castellari, L.; Giudici, P. Oenological properties of an interspecific Saccharomyces hybrid. S. Afr. J. Enol. Vitic. 1999, 20, 47–52. [Google Scholar] [CrossRef]
- Origone, A.C.; Rodríguez, M.E.; Oteiza, J.M.; Querol, A.; Lopes, C.A. Saccharomyces cerevisiae × Saccharomyces uvarum hybrids generated under different conditions share similar winemaking features. Yeast 2018, 35, 157–171. [Google Scholar] [CrossRef]
- Pérez-Torrado, R.; González, S.S.; Combina, M.; Barrio, E.; Querol, A. Molecular and enological characterization of a natural Saccharomyces uvarum and Saccharomyces cerevisiae hybrid. Int. J. Food Microbiol. 2015, 204, 101–110. [Google Scholar] [CrossRef]
- Krogerus, K.; Arvas, M.; De Chiara, M.; Magalhães, F.; Mattinen, L.; Oja, M.; Vidgren, V.; Yue, J.X.; Liti, G.; Gibson, B. Ploidy influences the functional attributes of de novo lager yeast hybrids. Appl. Microbiol. Biotechnol. 2016, 100, 7203–7222. [Google Scholar] [CrossRef]
- Krogerus, K.; Holmström, S.; Gibson, B. Enhanced Wort Fermentation with De Novo Lager Hybrids Adapted to High-Ethanol Environments. Appl. Environ. Microbiol. 2018, 84, e02302-17. [Google Scholar] [CrossRef]
- Gyurchev, N.Y.; Coral-Medina, Á.; Weening, S.M.; Almayouf, S.; Kuijpers, N.G.; Nevoigt, E.; Louis, E.J. Beyond Saccharomyces pastorianus for modern lager brews: Exploring non-cerevisiae Saccharomyces hybrids with heterotic maltotriose consumption and novel aroma profile. Front. Microbiol. 2022, 13, 1025132. [Google Scholar] [CrossRef]
- Nikulin, J.; Krogerus, K.; Gibson, B. Alternative Saccharomyces interspecies hybrid combinations and their potential for low-temperature wort fermentation. Yeast 2018, 35, 113–127. [Google Scholar] [CrossRef]
- Naseeb, S.; Visinoni, F.; Hu, Y.; Hinks Roberts, A.J.; Maslowska, A.; Walsh, T.; Smart, K.A.; Louis, E.J.; Delneri, D. Restoring fertility in yeast hybrids: Breeding and quantitative genetics of beneficial traits. Proc. Natl. Acad. Sci. USA 2021, 118, e2101242118. [Google Scholar] [CrossRef]
- Mozzachiodi, S.; Krogerus, K.; Gibson, B.; Nicolas, A.; Liti, G. Unlocking the functional potential of polyploid yeasts. Nat. Commun. 2022, 13, 2580. [Google Scholar] [CrossRef]
- Wang, H.; Hou, L. Genome shuffling to improve fermentation properties of top-fermenting yeast by the improvement of stress tolerance. Food Sci. Biotechnol. 2010, 19, 145–150. [Google Scholar] [CrossRef]
- European Parliament and Council. Directive 2001/18/EC of the European Parliament and of the Council of 12 March 2001 on the deliberate release into the environment of genetically modified organisms and repealing Council Directive 90/220/EEC. OJEC 2001, L106, 1–39. [Google Scholar]
- Dragosits, M.; Mattanovich, D. Adaptive laboratory evolution—Principles and applications for biotechnology. Microb. Cell Fact. 2013, 12, 64. [Google Scholar] [CrossRef] [PubMed]
- Lee, S.; Kim, P. Current Status and Applications of Adaptive Laboratory Evolution in Industrial Microorganisms. J. Microbiol. Biotechnol. 2020, 30, 793–803. [Google Scholar] [CrossRef] [PubMed]
- Gallone, B.; Steensels, J.; Prahl, T.; Soriaga, L.; Saels, V.; Herrera-Malaver, B.; Merlevede, A.; Roncoroni, M.; Voordeckers, K.; Miraglia, L.; et al. Domestication and Divergence of Saccharomyces cerevisiae Beer Yeasts. Cell 2016, 166, 1397–1410.e16. [Google Scholar] [CrossRef]
- Gallone, B.; Mertens, S.; Gordon, J.L.; Maere, S.; Verstrepen, K.J.; Steensels, J. Origins, evolution, domestication and diversity of Saccharomyces beer yeasts. Curr. Opin. Biotechnol. 2018, 49, 148–155. [Google Scholar] [CrossRef]
- Whittington, H.D.; Dagher, S.F.; Bruno-Bárcena, J.M. Production and conservation of starter cultures: From “backslopping” to controlled fermentations. In How Fermented Foods Feed a Healthy Gut Microbiota: A Nutrition Continuum; Springer: Cham, Switzerland, 2019; pp. 125–138. [Google Scholar]
- Mavrommati, M.; Daskalaki, A.; Papanikolaou, S.; Aggelis, G. Adaptive laboratory evolution principles and applications in industrial biotechnology. Biotechnol. Adv. 2022, 54, 107795. [Google Scholar] [CrossRef]
- Mattenberger, F.; Fares, M.A.; Toft, C.; Sabater-Muñoz, B. The Role of Ancestral Duplicated Genes in Adaptation to Growth on Lactate, a Non-Fermentable Carbon Source for the Yeast Saccharomyces cerevisiae. Int. J. Mol. Sci. 2021, 22, 12293. [Google Scholar] [CrossRef]
- Brickwedde, A.; van den Broek, M.; Geertman, J.M.; Magalhães, F.; Kuijpers, N.G.; Gibson, B.; Pronk, J.T.; Daran, J.M. Evolutionary engineering in chemostat cultures for improved maltotriose fermentation kinetics in Saccharomyces pastorianus lager brewing yeast. Front. Microbiol. 2017, 8, 1690. [Google Scholar] [CrossRef]
- Tilloy, V.; Ortiz-Julien, A.; Dequin, S. Reduction of ethanol yield and improvement of glycerol formation by adaptive evolution of the wine yeast Saccharomyces cerevisiae under hyperosmotic conditions. Appl. Microbiol. Biotechnol. 2014, 80, 2623–2632. [Google Scholar] [CrossRef]
- Kutyna, D.R.; Varela, C.; Stanley, G.A.; Borneman, A.R.; Henschke, P.A.; Chambers, P.J. Adaptive evolution of Saccharomyces cerevisiae to generate strains with enhanced glycerol production. Appl. Microbiol. Biotechnol. 2012, 93, 1175–1184. [Google Scholar] [CrossRef]
- Mezzetti, F.; De Vero, L.; Giudici, P. Evolved Saccharomyces cerevisiae wine strains with enhanced glutathione production obtained by an evolution-based strategy. FEMS Yeast Res. 2014, 14, 977–987. [Google Scholar] [CrossRef] [PubMed]
- Voordeckers, K.; Kominek, J.; Das, A.; Espinosa-Cantú, A.; De Maeyer, D.; Arslan, A.; Van Pee, M.; van der Zande, E.; Meert, W.; Yang, Y.; et al. Adaptation to high ethanol reveals complex evolutionary pathways. PLoS Genet. 2015, 11, e1005635. [Google Scholar] [CrossRef] [PubMed]
- Bartel, C.; Roach, M.; Onetto, C.; Curtin, C.; Varela, C.; Borneman, A. Adaptive evolution of sulfite tolerance in Brettanomyces bruxellensis. FEMS Yeast Res. 2021, 21, foab036. [Google Scholar] [CrossRef]
- Blieck, L.; Toye, G.; Dumortier, F.; Verstrepen, K.J.; Delvaux, F.R.; Thevelein, J.M.; Van Dijck, P. Isolation and Characterization of Brewer’s Yeast Variants with Improved Fermentation Performance under High-Gravity Conditions. Appl. Environ. Microbiol. 2007, 73, 815–824. [Google Scholar] [CrossRef]
- Avrahami-Moyal, L.; Eelberg, D.; Wenger, J.W.; Sherlock, G.; Braun, S. Turbidostat culture of Saccharomyces cerevisiae W303-1A under selective pressure elicited by ethanol selects for mutations in SSD1 and UTH1. FEMS Yeast Res. 2012, 12, 521–533. [Google Scholar] [CrossRef]
- Venkataram, S.; Dunn, B.; Li, Y.; Agarwala, A.; Chang, J.; Ebel, E.R.; Geiler-Samerotte, K.; Hérissant, L.; Blundell, J.R.; Levy, S.F.; et al. Development of a comprehensive genotype-to-fitness map of adaptation-driving mutations in yeast. Cell 2016, 166, 1585–1596. [Google Scholar] [CrossRef]
- He, Y.; Yin, H.; Dong, J.; Yu, J.; Zhang, L.; Yan, P.; Wan, X.; Hou, X.; Zhao, Y.; Chen, R.; et al. Reduced sensitivity of lager brewing yeast to premature yeast flocculation via adaptive evolution. Food Microbiol. 2022, 106, 104032. [Google Scholar] [CrossRef]
- Kayacan, Y.; Van Mieghem, T.; Delvaux, F.; Delvaux, F.R.; Willaert, R. Adaptive Evolution of Industrial Brewer’s Yeast Strains towards a Snowflake Phenotype. Fermentation 2020, 6, 20. [Google Scholar] [CrossRef]
- Gonçalves, M.; Pontes, A.; Almeida, P.; Barbosa, R.; Serra, M.; Libkind, D.; Hutzler, M.; Gonçalves, P.; Sampaio, J.P. Distinct Domestication Trajectories in Top-Fermenting Beer Yeasts and Wine Yeasts. Curr. Biol. 2016, 26, 2750–2761. [Google Scholar] [CrossRef]
- Gorter de Vries, A.R.; Voskamp, M.A.; Van Aalst, A.C.; Kristensen, L.H.; Jansen, L.; Van den Broek, M.; Salazar, A.N.; Brouwers, N.; Abeel, T.; Pronk, J.T.; et al. Laboratory evolution of a Saccharomyces cerevisiae × S. eubayanus hybrid under simulated lager-brewing conditions. Front. Genet. 2019, 10, 242. [Google Scholar] [CrossRef]
- Wong, B.G.; Mancuso, C.P.; Kiriakov, S.; Bashor, C.J.; Khalil, A.S. Precise, automated control of conditions for high-throughput growth of yeast and bacteria with eVOLVER. Nat. Biotechnol. 2018, 36, 614–623. [Google Scholar] [CrossRef] [PubMed]
- Winkler, J.; Kao, K.C. Computational identification of adaptive mutants using the VERT system. J. Biol. Eng. 2012, 6, 3. [Google Scholar] [CrossRef]
- LaCroix, R.A.; Palsson, B.O.; Feist, A.M. A model for designing adaptive laboratory evolution experiments. Appl. Environ. Microbiol. 2017, 83, e03115-16. [Google Scholar] [CrossRef]
- Amer, B.; Baidoo, E.E. Omics-driven biotechnology for industrial applications. Front. Bioeng. Biotechnol. 2021, 9, 613307. [Google Scholar] [CrossRef]
- Chen, Y.; Li, F.; Nielsen, J. Genome-scale modeling of yeast metabolism: Retrospectives and perspectives. FEMS Yeast Res. 2022, 22, foac003. [Google Scholar] [CrossRef]
- Zhang, C.; Sánchez, B.J.; Li, F.; Eiden, C.W.; Scott, W.T.; Liebal, U.W.; Blank, L.M.; Mengers, H.G.; Anton, M.; Rangel, A.T.; et al. Yeast9: A consensus genome-scale metabolic model for S. cerevisiae curated by the community. Mol. Syst. Biol. 2024, 20, 1134–1150. [Google Scholar] [CrossRef]
- Gong, X.; Zhang, J.; Gan, Q.; Teng, Y.; Hou, J.; Lyu, Y.; Liu, Z.; Wu, Z.; Dai, R.; Zou, Y.; et al. Advancing microbial production through artificial intelligence-aided biology. Biotechnol. Adv. 2024, 74, 108399. [Google Scholar] [CrossRef]
- Peltier, E.; Friedrich, A.; Schacherer, J.; Marullo, P. Quantitative trait nucleotides impacting the technological performances of industrial Saccharomyces cerevisiae strains. Front. Genet. 2019, 10, 683. [Google Scholar] [CrossRef]
- Krogerus, K.; Rettberg, N. Creating better brewing yeast with the 1011 yeast genomes data sets. Yeast 2025, 42, 5–15. [Google Scholar] [CrossRef]
- Duval, E.H.; Alves, S.L., Jr.; Dunn, B.; Sherlock, G.; Stambuk, B.U. Microarray karyotyping of maltose-fermenting Saccharomyces yeasts with differing maltotriose utilization profiles reveals copy number variation in genes involved in maltose and maltotriose utilization. J. Appl. Microbiol. 2010, 109, 248–259. [Google Scholar] [CrossRef]
- Krogerus, K.; Magalhães, F.; Kuivanen, J.; Gibson, B. A deletion in the STA1 promoter determines maltotriose and starch utilization in STA1+ Saccharomyces cerevisiae strains. Appl. Microbiol. Biotechnol. 2019, 103, 7597–7615. [Google Scholar] [CrossRef] [PubMed]
- Tristão, L.E.; de Sousa, L.I.; de Oliveira Vargas, B.; José, J.; Carazzolle, M.F.; Silva, E.M.; Galhardo, J.P.; Pereira, G.A.; de Mello, F.D. Unveiling genetic anchors in Saccharomyces cerevisiae: QTL mapping identifies IRA2 as a key player in ethanol tolerance and beyond. Mol Genet. Genom. 2024, 299, 103. [Google Scholar] [CrossRef] [PubMed]
- Albillos-Arenal, S.; Alonso-del-Real, J.; Lairón-Peris, M.; Barrio, E.; Querol, A. Identification of a crucial INO2 allele for enhancing ethanol resistance in an industrial fermentation strain of Saccharomyces cerevisiae. bioRxiv 2024. [Google Scholar] [CrossRef]
- Wei, J.; Zhang, Y.; Zhang, X.; Guo, H.; Yuan, Y.; Yue, T. Multi-omics discovery of aroma-active compound formation by Pichia kluyveri during cider production. LWT 2022, 159, 113233. [Google Scholar] [CrossRef]
- van Wyk, N.; Badura, J.; von Wallbrunn, C.; Pretorius, I.S. Exploring future applications of the apiculate yeast Hanseniaspora. Crit. Rev. Biotechnol. 2024, 44, 100–119. [Google Scholar] [CrossRef]
- Zhang, J.; Petersen, S.D.; Radivojevic, T.; Ramirez, A.; Pérez-Manríquez, A.; Abeliuk, E.; Sánchez, B.J.; Costello, Z.; Chen, Y.; Fero, M.J.; et al. Combining mechanistic and machine learning models for predictive engineering and optimization of tryptophan metabolism. Nat. Commun. 2020, 11, 4880. [Google Scholar] [CrossRef]
- Culley, C.; Vijayakumar, S.; Zampieri, G.; Angione, C. A mechanism-aware and multiomic machine-learning pipeline characterizes yeast cell growth. Proc. Natl. Acad. Sci. USA 2020, 117, 18869–18879. [Google Scholar] [CrossRef]
- Radivojević, T.; Costello, Z.; Workman, K.; Garcia Martin, H. A machine learning Automated Recommendation Tool for synthetic biology. Nat. Commun. 2020, 11, 4879. [Google Scholar] [CrossRef]
- Nguyen, E.; Poli, M.; Durrant, M.G.; Kang, B.; Katrekar, D.; Li, D.B.; Bartie, L.J.; Thomas, A.W.; King, S.H.; Brixi, G.; et al. Sequence modeling and design from molecular to genome scale with Evo. Science 2024, 386, eado9336. [Google Scholar] [CrossRef]
- Bunne, C.; Roohani, Y.; Rosen, Y.; Gupta, A.; Zhang, X.; Roed, M.; Alexandrov, T.; AlQuraishi, M.; Brennan, P.; Burkhardt, D.B.; et al. How to build the virtual cell with artificial intelligence: Priorities and opportunities. Cell 2024, 187, 7045–7063. [Google Scholar] [CrossRef]
- Giri, S.; Shitut, S.; Kost, C. Harnessing ecological and evolutionary principles to guide the design of microbial production consortia. Curr. Opin. Biotechnol. 2020, 62, 228–238. [Google Scholar] [CrossRef] [PubMed]
- Mittermeier, F.; Bäumler, M.; Arulrajah, P.; García Lima, J.D.; Hauke, S.; Stock, A.; Weuster-Botz, D. Artificial microbial consortia for bioproduction processes. Eng. Life Sci. 2023, 23, e2100152. [Google Scholar] [CrossRef]
- Konstantinidis, D.; Pereira, F.; Geissen, E.M.; Grkovska, K.; Kafkia, E.; Jouhten, P.; Kim, Y.; Devendran, S.; Zimmermann, M.; Patil, K.R. Adaptive laboratory evolution of microbial co-cultures for improved metabolite secretion. Mol. Syst. Biol. 2021, 17, e10189. [Google Scholar] [CrossRef]
- Bagheri, B.; Bauer, F.F.; Setati, M.E. The impact of Saccharomyces cerevisiae on a wine yeast consortium in natural and inoculated fermentations. Front. Microbiol. 2017, 8, 1988. [Google Scholar] [CrossRef]
- Conacher, C.G.; Naidoo-Blassoples, R.K.; Rossouw, D.; Bauer, F.F. Real-time monitoring of population dynamics and physical interactions in a synthetic yeast ecosystem by use of multicolour flow cytometry. Appl. Microbiol. Biotechnol. 2020, 104, 5547–5562. [Google Scholar] [CrossRef]
- Canonico, L.; Galli, E.; Ciani, E.; Comitini, F.; Ciani, M. Exploitation of three non-conventional yeast species in the brewing process. Microorganisms 2019, 7, 11. [Google Scholar] [CrossRef]
- Steensels, J.; Gallone, B.; Voordeckers, K.; Verstrepen, K.J. Domestication of industrial microbes. Curr. Biol. 2019, 29, R381–R393. [Google Scholar] [CrossRef]
- Zdaniewicz, M.; Satora, P.; Pater, A.; Bogacz, S. Low lactic acid-producing strain of Lachancea thermotolerans as a new starter for beer production. Biomolecules 2020, 10, 256. [Google Scholar] [CrossRef]
- Klimczak, K.; Cioch-Skoneczny, M.; Ciosek, A.; Poreda, A. Application of Non-Saccharomyces Yeast for the Production of Low-Alcohol Beer. Foods 2024, 13, 3214. [Google Scholar] [CrossRef]
- Zhang, Z.Q.; Bo, L.; Xia, S.Q.; Wang, X.J.; Yang, A.M. Production and application of a novel bioflocculant by multiple-microorganism consortia using brewery wastewater as carbon source. J. Environ. Sci. 2007, 19, 667–673. [Google Scholar] [CrossRef]
- Yang, L.; Zhu, X.; Mao, Y.; Zhang, X.; Xu, B.; Yang, X. Effect of different inoculation strategies of mixed culture Saccharomyces cerevisiae/Oenococcus oeni on the aroma quality of Chardonnay wine. Food Res. Int. 2024, 190, 114636. [Google Scholar] [CrossRef] [PubMed]
- Mas, A.; Portillo, M.C. Strategies for microbiological control of the alcoholic fermentation in wines by exploiting the microbial terroir complexity: A mini-review. Int. J. Food Microbiol. 2022, 367, 109592. [Google Scholar] [CrossRef] [PubMed]
- Jolly, N.P.; Augustyn, O.P.; Pretorius, I.S. The occurrence of non-Saccharomyces cerevisiae yeast species over three vintages in four vineyards and grape musts from four production regions of the Western Cape, South Africa. S. Afr. J. Enol. Vitic. 2003, 24, 35–42. [Google Scholar] [CrossRef]
- Mendoza, L.M.; de Nadra, M.C.M.; Bru, E.; Farías, M.E. Influence of wine-related physicochemical factors on the growth and metabolism of non-Saccharomyces and Saccharomyces yeasts in mixed culture. J. Ind. Microbiol. Biotechnol. 2009, 36, 229–237. [Google Scholar] [CrossRef]
- Wang, C.; Esteve-Zarzoso, B.; Cocolin, L.; Mas, A.; Rantsiou, K. Viable and culturable populations of Saccharomyces cerevisiae, Hanseniaspora uvarum and Starmerella bacillaris (synonym Candida zemplinina) during Barbera must fermentation. Food Res. Int. 2015, 78, 195–200. [Google Scholar] [CrossRef]
- Albergaria, H.; Arneborg, N. Dominance of Saccharomyces cerevisiae in alcoholic fermentation processes: Role of physiological fitness and microbial interactions. Appl. Microbiol. Biotechnol. 2016, 100, 2035–2046. [Google Scholar] [CrossRef]
- Vaughan, A.; O’Sullivan, T.; Van Sinderen, D. Enhancing the Microbiological Stability of Malt and Beer—A Review. J. Inst. Brew. 2005, 111, 355–371. [Google Scholar] [CrossRef]
- Modzelewska, A.; Jackowski, M.; Trusek, A. Optimization of beer mixed fermentation using Saccharomyces cerevisiae and Lactobacillus brevis. Eur. Food Res. Technol. 2023, 249, 3261–3269. [Google Scholar] [CrossRef]
- Malcı, K.; Walls, L.E.; Rios-Solis, L. Multiplex genome engineering methods for yeast cell factory development. Front. Bioeng. Biotechnol. 2020, 8, 589468. [Google Scholar] [CrossRef]
- DiCarlo, J.E.; Norville, J.E.; Mali, P.; Rios, X.; Aach, J.; Church, G.M. Genome engineering in Saccharomyces cerevisiae using CRISPR-Cas systems. Nucleic Acids Res. 2013, 41, 4336–4343. [Google Scholar] [CrossRef]
- Ren, Q.; Zhong, Z.; Wang, Y.; You, Q.; Li, Q.; Yuan, M.; He, Y.; Qi, C.; Tang, X.; Zheng, X.; et al. Bidirectional promoter-based CRISPR-Cas9 systems for plant genome editing. Front. Plant Sci. 2019, 10, 1173. [Google Scholar] [CrossRef] [PubMed]
- Huang, S.; Geng, A. High-copy genome integration of 2, 3-butanediol biosynthesis pathway in Saccharomyces cerevisiae via in vivo DNA assembly and replicative CRISPR-Cas9 mediated delta integration. J. Biotechnol. 2020, 310, 13–20. [Google Scholar] [CrossRef] [PubMed]
- Zhang, S.; Shen, J.; Li, D.; Cheng, Y. Strategies in the delivery of Cas9 ribonucleoprotein for CRISPR/Cas9 genome editing. Theranostics 2021, 11, 614–648. [Google Scholar] [CrossRef] [PubMed]
- Mertens, S.; Gallone, B.; Steensels, J.; Herrera-Malaver, B.; Cortebeek, J.; Nolmans, R.; Saels, V.; Vyas, V.K.; Verstrepen, K.J. Reducing phenolic off-flavors through CRISPR-based gene editing of the FDC1 gene in Saccharomyces cerevisiae × Saccharomyces eubayanus hybrid lager beer yeasts. PLoS ONE 2019, 14, e0209124. [Google Scholar]
- Dank, A.; Smid, E.J.; Notebaart, R.A. CRISPR-Cas genome engineering of esterase activity in Saccharomyces cerevisiae steers aroma formation. BMC Res. Notes 2018, 11, 682. [Google Scholar] [CrossRef]
- Krogerus, K.; Fletcher, E.; Rettberg, N.; Gibson, B.; Preiss, R. Efficient breeding of industrial brewing yeast strains using CRISPR/Cas9-aided mating-type switching. Appl. Microbiol. Biotechnol. 2021, 105, 8359–8376. [Google Scholar] [CrossRef]
- Denby, C.M.; Li, R.A.; Vu, V.T.; Costello, Z.; Lin, W.; Chan, L.J.; Williams, J.; Donaldson, B.; Bamforth, C.W.; Petzold, C.J.; et al. Industrial brewing yeast engineered for the production of primary flavor determinants in hopped beer. Nat. Commun. 2018, 9, 965. [Google Scholar] [CrossRef]
- Yang, S.; Chen, R.; Cao, X.; Wang, G.; Zhou, Y.J. De novo biosynthesis of the hops bioactive flavonoid xanthohumol in yeast. Nat. Commun. 2024, 15, 253. [Google Scholar] [CrossRef]
- Cambon, B.; Monteil, V.; Remize, F.; Camarasa, C.; Dequin, S. Effects of GPD1 Overexpression in Saccharomyces cerevisiae Commercial Wine Yeast Strains Lacking ALD6 Genes. Appl. Environ. Microbiol. 2006, 72, 4688–4694. [Google Scholar] [CrossRef]
- González, J.P.; González, R.; Querol, A.; Sendra, J.; Ramón, D. Construction of a recombinant wine yeast strain expressing β-(1, 4)-endoglucanase and its use in microvinification process. Appl. Environ. Microbiol. 1993, 59, 2801–2806. [Google Scholar] [CrossRef]
- Vigentini, I.; Gebbia, M.; Belotti, A.; Foschino, R.; Roth, F.P. CRISPR/Cas9 system as a valuable genome editing tool for wine yeasts with application to decrease urea production. Front. Microbiol. 2017, 8, 2194. [Google Scholar] [CrossRef] [PubMed]
- Volschenk, H.; Viljoen, M.; Grobler, J.; Bauer, F.; Lonvaud-Funel, A.; Denayrolles, M.; Subden, R.E.; Van Vuuren, H.J. Malolactic Fermentation in Grape Musts by a Genetically Engineered Strain of Saccharomyces cerevisiae. Am. J. Enol. Vitic. 1997, 48, 193–197. [Google Scholar] [CrossRef]
- Schoeman, H.; Vivier, M.A.; du Toit, M.; Dicks, L.M.T.; Pretorius, I.S. The development of bactericidal yeast strains by expressing the Pediococcus acidilactici pediocin gene (pedA) in Saccharomyces cerevisiae. Yeast 1999, 15, 647–656. [Google Scholar] [CrossRef]
- de Ruijter, J.C.; Aisala, H.; Jokinen, I.; Krogerus, K.; Rischer, H.; Toivari, M. Production and sensory analysis of grape flavoured beer by co-fermentation of an industrial and a genetically modified laboratory yeast strain. Eur. Food Res. Technol. 2023, 249, 1991–2000. [Google Scholar] [CrossRef]
- Sone, H.; Fujii, T.; Kondo, K.; Shimizu, F.; Tanaka, J.; Inoue, T. Nucleotide sequence and expression of the Enterobacter aerogenes alpha-acetolactate decarboxylase gene in brewer’s yeast. Appl. Environ. Microbiol. 1988, 54, 38–42. [Google Scholar] [CrossRef]
- Uzogara, S.G. The impact of genetic modification of human foods in the 21st century: A review. Biotechnol. Adv. 2000, 18, 179–206. [Google Scholar] [CrossRef]
- Davison, J.; Ammann, K. New GMO regulations for old: Determining a new future for EU crop biotechnology. GM Crops Food 2017, 8, 13–34. [Google Scholar] [CrossRef]
- FDA Biotechnology Policy. Available online: https://www.fda.gov/safety/fdas-regulation-plant-and-animal-biotechnology-products (accessed on 11 June 2025).
- Government of Canada Novel Foods: Overview. Available online: https://www.canada.ca/en/health-canada/services/food-nutrition/genetically-modified-foods-other-novel-foods.html (accessed on 11 June 2025).
Technique | Pros | Cons |
---|---|---|
Mutagenesis Non-GMO | Generates a wide variety of potential phenotypes; relatively inexpensive and straightforward; suitable for monogenic phenotypes. | Random outcomes make it difficult to predict results; can introduce harmful mutations or undesired traits; not suitable for polygenic or complex phenotypes. |
Hybridization Non-GMO | Simple method to combine beneficial traits from different yeast strains; heterosis compared with parents; well-established and cost-effective. | May result in sterility or instability of the hybrid offspring; difficult to obtain for poorly sporulating strains; undesirable characteristics may emerge. |
Adaptive Laboratory Evolution (ALE) Non-GMO | Mimics natural selection, leading to improved fitness-related traits over time; suitable for phenotypes that can be directly selected under controlled environments. | Time-consuming and labor-intensive; Unintended side effects may occur, as it is difficult to control specific outcomes; May not be suitable for traits without clear selection markers. |
Multi-Omics and AI Integration Non-GMO | Provides a comprehensive view of yeast metabolism and gene expression; unintended side effects may occur, as it is difficult to control specific outcomes. | Requires large datasets and significant computational resources; interpreting the data can be complex and requires expert knowledge. |
Synthetic Microbial Communities (SMC) GMO or non-GMO (depending on strains used) | Enables creation of yeast strains consortia with complementary traits that work together; Can improve metabolic networks. | Requires compatibility between strains, including nutrient requirements; If GMO strains are used, there are ethical concerns around genetic modifications. |
CRISPR-Cas9 GMO | Precise and efficient genome editing; Allows for modifications of multiple genes at once; highly versatile across different yeast strains. | Ethical concerns around gene editing; Requires optimization for different yeast species; Requires advanced understanding of yeast genetics. |
Product | Objective | Work |
---|---|---|
Wine | Increased ethanol tolerance | [35] |
Enhanced nitrogen source utilization | [36] | |
Increased mannoprotein release | [37] | |
Reduced volatile acidity | [38] | |
Increased aroma compound concentration (esters) | [39] | |
Increased higher alcohols via leucine auxotrophic mutants | [40] | |
Beer | Elimination of phenolic off-flavors (POFs) in S. eubayanus | [32] |
Enhancement of ethanol tolerance in lager yeast | [34] | |
Increase in maltotriose utilization | [41] |
Product | Cross | Objectives | Work |
---|---|---|---|
Wine | S. cerevisiae × S. cerevisiae | Elimination of undesirable traits (e.g., SO2 formation, excessive foam production) | [43] |
S. cerevisiae × S. cerevisiae | Increase in flocculation | [44] | |
S. cerevisiae × S. uvarum | Increase optimal temperature range; modulation of by-products | [45,46,47] | |
Beer | S. cerevisiae × S. eubayanus | Improved fermentation and aroma production in lager hybrids | [12,48,49,50] |
S. mikatae × S. eubayanus and S. jurei × S. eubayanus | Improved fermentation and aroma production in lager hybrids | [50] | |
S. cerevisiae × S. mikatae | Enhancement of beer flavor under cold temperatures | [51] |
Product | Objective | Ref. |
---|---|---|
Wine | Increased glycerol production for low-ethanol wines | [64,65] |
Development of yeast strains producing higher levels of glutathione (GSH) | [66] | |
Increased ethanol tolerance | [67] | |
Enhanced sulfite resistance in B. bruxellensis | [68] | |
Beer | Improved beer yeast performance via UV mutagenesis and high-gravity wort fermentations | [69] |
Enhanced ethanol tolerance in evolved de novo lager yeasts | [12] | |
Genomic adaptations linked to chromosomal duplications and mutations in IRA2 and UTH1 | [70,71] | |
Adaptation of Saccharomyces variants to overcome premature yeast flocculation (PYF) | [72] | |
Increased flocculation to favor yeast removal | [73] | |
Reduction in phenolic off-flavors via PAD1 and FDC1 mutations | [74] | |
Improvement of fermentation efficiency | [75] |
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content. |
© 2025 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
Share and Cite
Nasuti, C.; Solieri, L.; Krogerus, K. 3.0 Strategies for Yeast Genetic Improvement in Brewing and Winemaking. Beverages 2025, 11, 100. https://doi.org/10.3390/beverages11040100
Nasuti C, Solieri L, Krogerus K. 3.0 Strategies for Yeast Genetic Improvement in Brewing and Winemaking. Beverages. 2025; 11(4):100. https://doi.org/10.3390/beverages11040100
Chicago/Turabian StyleNasuti, Chiara, Lisa Solieri, and Kristoffer Krogerus. 2025. "3.0 Strategies for Yeast Genetic Improvement in Brewing and Winemaking" Beverages 11, no. 4: 100. https://doi.org/10.3390/beverages11040100
APA StyleNasuti, C., Solieri, L., & Krogerus, K. (2025). 3.0 Strategies for Yeast Genetic Improvement in Brewing and Winemaking. Beverages, 11(4), 100. https://doi.org/10.3390/beverages11040100