Pichia kluyveri yeast and its role in winemaking

by | Dec 1, 2020 | Oenology research, Winetech Technical

PHOTO: Shutterstock.

The Pichia kluyveri yeast is desired for its ability to enhance volatile thiols and overall fruitiness in wine.

Current and previous names

Pichia kluyveri was previously known as Hansenula kluyveri.1

Where it is found

Pichia kluyveri strains are widely distributed in nature. They have been isolated from grapes, rotting fruit and are commonly encountered in natural fermentations, such as coffee bean and wine fermentations.2,3

What it looks like

Pichia kluyveri is known to form ovoid to elongate cells that may occur singly, in pairs or in short chains, and does not form pseudohyphae. The cells can grow up to 2 – 6 x 3.5 – 10 µm. Sexual reproduction leads to the formation of hat-shaped ascospores 1.5 – 1.75 x 2 – 2.5 µm in size and 2 – 4 per ascus.4 Colonies on Wallerstein Nutrient Agar are cream to mint green in colour, with a rugose texture.

Nitrogen metabolism

Pichia kluyveri grows poorly in synthetic medium and displays low nitrogen consumption in synthetic grape juice.4,5 Strains of this species do not consume asparagine and glycine and show preferential consumption of lysine and arginine when grown in synthetic grape juice.4,5 The presence of ammonium hampers the uptake of valine, isoleucine and phenylalanine.5

Technical characteristics

 

  • Temperature and alcohol tolerance

Pichia kluyveri is a slow grower and a weak fermenter.5 Strains of this species can tolerate 4 – 5% ethanol6 and can grow over a wide temperature range (10 – 28°C), although at lower temperature a longer lag phase is observed.6

  • Glycerol, VA, SO2 and H2S production

Most strains of P. kluyveri produce low levels of H2S, less than 0.7 g/L acetic acid and more than 5 g/L glycerol when fermenting grape juice medium with initial sugars at 21°Brix.7

  • Mixed culture fermentations

Pichia kluyveri is considered a weak fermenter and is unable to produce dry wine when used in monoculture.5,7,8 Consequently, it is used in sequential inoculation with Saccharomyces cerevisiae. An inoculum ratio of 1:9 (P. kluyveri:S. cerevisiae) has been shown to yield desirable wine aroma outputs in Sauvignon blanc.9

Effect on malolactic fermentation

Pichia kluyveri strains have not been shown to consume malic acid or produce lactic acid.

Traits of oenological interest

Generally, P. kluyveri is desired as a co-inoculant in the production of Sauvignon blanc and Riesling due to its ability to enhance thiol production.

  • Extracellular enzymes

Pichia kluyveri produces carbon sulphur lyases. These enzymes are necessary for the release of cysteinylated precursors cys-4MMP and cys-3MH allowing for the release of the volatile thiols 4-mercapto-4-methylpentan-2-one (4MMP), 3-mercaptohexan-1-ol (3MH) and 3-mercaptohexyl acetate (3MHA) which are impact odorants in Sauvignon blanc wines.10 Furthermore, P. kluyveri produces esterases and esterase-lipases involved in the release of esters, as well as pectinases, necessary in grape juice processing.11

  • Contribution to wine aroma

Pichia kluyveri produces considerable levels of 2-phenylethyl acetate (which imparts flowery, rose and fruity aromas in wine) in pure culture and in sequential inoculation with S. cerevisiae.5 It also produces about 10% more ethyl octanoate (which imparts sweet, ripe fruit, pear and pineapple aromas in wine) than S. cerevisiae.12 Sequential inoculation of Riesling with S. cerevisiae increases overall impression of peach/apricot character,8 while the volatile thiols released by this yeast enhance grapefruit and passion fruit aromas in Sauvignon blanc.13

Conclusion

Pichia kluyveri Frootzen® from Chr. Hansen (www.chr-hansen.com) is the only commercial product available. It is supplied as a frozen product used for direct inoculation into grape must without any rehydration. This yeast is especially recommended for Sauvignon blanc, Riesling and Chardonnay wines.8,13

References

  1. Jolly, N.P., Varela, C. & Pretorius, I.S., 2014. Not your ordinary yeast: Non-Saccharomyces yeasts in wine production uncovered. FEMS Yeast Research 14, 215 – 237.
  2. Jolly, N.P., Augustyn, O.P.H. & Pretorius, I.S., 2003. 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. South African Journal of Enology and Viticulture 24, 35 – 42.
  3. Brežná, B., Ženišová, K., Chaovanová, K., Chebeňová, V., Kraková, L., Kuchta, T. & Pangallo, D., 2010. Evaluation of fungal and yeast diversity in Slovakian win-related microbial communities. Antonie van Leeuwenhoek 98, 519 – 529.
  4. Bedford, C.L., 1942. A taxonomic study of the genus Hansenula. Mycologia 34, 628 – 649.
  5. Prior, K.J., Bauer, F.F. & Divol, B., 2019. The utilization of nitrogenous compounds by commercial non-Saccharomyces yeasts associated with wine. Food Microbiology 79, 75 – 84.
  6. Nutt, J., 2018. Multispecies interactions in a simplified wine yeast consortium. MSc Thesis, Stellenbosch University, South Africa. Available on https://scholar.sun.ac.za/handle/10019.1/103538.
  7. Mestre Furlani, M.V., Maturano, Y.P., Combina, M., Mercado, L.A., Toro, M.E. & Vazquez, F., 2017. Selection of non-Saccharomyces yeasts to be used in grape musts with high alcoholic potential: A strategy to obtain wines with reduced ethanol content. FEMS Yeast Research 17, foc010.
  8. Benito, S., Hoffmann, T., Laier, M., Lochbühler, B., Schüttler, A., Ebert, K., Fritsch, S., Röcker, J. & Rauhut, D., 2015. Effect on quality and composition of Riesling wines fermented by sequential inoculation with non-Saccharomyces and Saccharomyces cerevisiae. European Food Research and Technology 24, 707 – 717.
  9. Anfang, N., Brajkovich, M. & Goddard, M.R., 2009. Co-fermentation with Pichia kluyveri increases varietal thiol concentrations in Sauvignon blanc. Australian Journal of Grape and Wine Research 15, 1 – 8.
  10. Padilla, B., Gil, J.V. & Manzanares, P., 2016. Past and future of non-Saccharomyces yeasts: From spoilage microorganisms to biotechnological tools for improving wine aroma complexity. Frontiers in Microbiology 7, 411.
  11. Escribano, R., González-Arenzana, L., Garijo, P., Berlanas, C., López-Alfaro, I., López, R., Gutiérrez, A.R. & Santamaría, P., 2017. Screening of enzymatic activities within different enological non-Saccharomyces yeasts. Journal of Food Science and Technology 54, 1555 – 1564.
  12. Benito, A., Calderón, F. & Benito, S., 2019. The influence of non-Saccharomyces species on wine fermentation quality parameters. Fermentation 5, 54.
  13. Morata, A., Escott, C., Bañuelos, A., Loira, I., De Fresno, J.M., González, C. & Suárez-Lepe, J.A., 2020. Contribution of non-Saccharomyces yeasts to wine freshness – a review. Biomolecules 10, 34.

– For more information, contact Evodia Setati at setati@sun.ac.za.

 

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