Diversity and Identification of Yeasts Isolated from Tumultuous Stage of Spontaneous Table Grape Fermentations in Central China
Abstract
Table grapes are of increasing interest for wine production in China. In this study, 480 yeast isolates were isolated from the tumultuous stage during the spontaneous fermentation of six table grape varieties, which were cultivated in an ecological environment that was not industry-influenced, in Central China. The 26S rDNA D1/D2 domain sequence analysis was more efficient for yeast species identification than the 5.8S-ITS region RFLP analysis in the present study. All the tested strains belonged to nine species from six genera: Hanseniaspora guilliermondii, H. opuntiae, H. uvarum, Pichia terricola, Kazachstania hellenica, K. zonata, P. occidentalis, Saccharomyces cerevisiae and Zygosaccharomyces bailii. The yeast species and populations differed notably among the grape varieties. S. cerevisiae was found in the samples of four grape varieties (Vitis amurensis, Iona, Moldova and V. davidii), but not detected in Cuihong and Alimandeng Rose. Interdelta sequence fingerprinting analysis was used to discriminate between 128 S. cerevisiae isolates. Eight S. cerevisiae genotypes (G1 to G8) were distinguished. Genotypes G1, G2, and G3 were the most dominant strains, accounting for 32.03%, 24.22% and 28.13% of the isolates respectively. This study shows the diversity of yeast species associated with spontaneous fermentations of different table grape varieties grown in an ecological environment without any wine industry effect or footprint.Downloads
References
Bisson, L.F., Kunkee, R.E., 1993. Microbial interactions during wine production. In: Zeikus, J.G., Johnson, E.A. (ed.). Mixed cultures in biotechnology. New York: McGraw-Hill. pp. 37–68.
Clemente-Jimenez, J.M., Mingorance-Cazorla, L., Martı́nez-Rodrı́guez, S., Las Heras-Vázquez, F.J., Rodrı́guez-Vico F., 2004. Molecular characterization and oenological properties of wine yeasts isolated during spontaneous fermentation of six varieties of grape must. Food Microbiol. 21, 149-155.
Di Maro, E., Ercolini, D., Coppola, S., 2007. Yeast dynamics during spontaneous wine fermentation of the Catalanesca grape. Int J Food Microbiol. 117, 201-210.
Drumondeneves, J., Francoduarte, R., Lima, T., Schuller, D., & Pais, C., 2017. Association between grape yeast communities and the vineyard ecosystems. PLoS One, 12(1).
Esteve-Zarzoso, B., Belloch, C., Uruburu, F., Querol, A., 1999. Identification of yeasts by RFLP analysis of the 5.8S rRNA gene and the two ribosomal internal transcribed spacers. Int J Syst Bacteriol. 49, 329-337.
FAO, OIV. 2016. Vitis vinifera Table Grape Cultivar.
Fleet, G.H., 1998. The microbiology of alcoholic beverages. In: Wood BJB (ed.). Microbiology of fermented foods. New York: Blackie Academic & Professional, 217-262.
Goddard, M.R., Greig, D., 2015. Saccharomyces cerevisiae: a nomadic yeast with no niche?. FEMS Yeast Res. 15, 1-6.
Jiang, J.F., Kell, S., Fan, X.C., Zhang, Y,, Wei, W., Kang, D.M., Maxted, N., Ford-Lloyd, B., Liu, CH., 2015. The wild relatives of grape in China: Diversity, conservation gaps and impact of climate change. Agric Ecosyst Environ. 210, 50-58.
Kurtzman, C.P., Robnett, C.J., 1998. Identification and phylogeny of ascomycetous yeasts form analysis of nuclear large subunit 26S ribosomal DNA partial sequences. Antonie Van Leeuwenhoek. 73, 337-371.
Legras, J.L., Karst, F., 2003. Optimisation of interdelta analysis for Saccharomyces cerevisiae strain characterization. FEMS Microbiol Let. 221, 249-255.
Li, E.H., Liu, A.G., Xue, B., Liu, Y.L., 2011. Yeast species associated with spontaneous wine fermentation of Cabernet Sauvignon from Ningxia, China. World J Microbiol Biotechnol. 27, 2475-2482.
Lopandic, K., Tiefenbrunner, W., Gangl, H., Mandl, K., Berger, S., Leitner, G., Abd-Ellah, G.A., Querol, A., Gardner, R.C., Sterflinger, K., Prillinger, H., 2008. Molecular profiling of yeasts isolated during spontaneous fermentations of Austrian wines. FEMS Yeast Res. 8, 1063-1075.
Martini, A., 1993. The origin and domestication of the wine yeast Saccharomyces cerevisiae. J Wine Res. 4, 165-176.
Mercado, L., Jubany, S., Gaggero, C., Masuelli, R.W., Combina, M., 2010. Molecular Relationships Between Saccharomyces cerevisiae Strains Involved in Winemaking from Mendoza, Argentina. Curr Microbiol. 61, 506-514.
Nisiotou, A.A., Nychas, G.E., 2008. Kazachstania hellenica sp. nov., a novel ascomycetous yeast from a Botrytis-affected grape must fermentation. Int J Syst Bacteriol. 58, 1263-1267.
OIV. 2017. Statistical Report on World Vitiviniculture.
Oliveira, M., Arenas, M., Lage, O., Cunha, M., & Amorim, M. I., 2017. Epiphytic fungal community in vitis vinifera of the portuguese wine regions. Lett Appl Microbiol, 66(1).
Pallmann, C.L., Brown, J.A., Olineka, T.L., Cocolin, L., Mills, D.A., Bisson, L.F., 2001. Use of WL medium to profile native flora fermentations. Am J Enol Vitic. 52, 198-203.
Pei, Y.F., Wang, G.P., Liu, Y.L., 2009. Strain Typing of Saccharomyces cerevisiae During Spontaneous Fermentation of Cabernet Sauvigon Wine. Microbiol China. 10, 1532-1535.
Pretorius, I.S., van der Westhuizen T.J., Augustyn O.P.H., 1999. Yeast biodiversity in vineyards and wineries and its importance to the South African Wine Industry. S Afr J Enol Viti. 20, 61-74.
Renouf, V., Claisse, O., Lonvaud-Funel, A., 2007. Inventory and monitoring of wine microbial consortia. Appl Microbiol Biotechnol. 75, 149-164.
Rojas, V., Gil, J., Piñaga, F., Manzanares, P., 2003. Acetate ester formation in wine by mixed cultures in laboratory fermentations. Int J Food Microbiol. 86, 181-188.
Sun, H.H., Ma, H.Q., Hao, M.L., Pretorius, I.S., Chen, S.W., 2009. Identification of yeast population dynamics of spontaneous fermentation in Beijing wine region, China. Ann Microbiol. 59, 69-76.
Sun, Y., Liu, Y.L., 2014. Investigating of yeast species in wine fermentation using terminal restriction fragment length polymorphism method. Food Microbiol. 38, 201-207.
Sun, Y., Guo, J.J., Liu, F.B., Liu, Y.L., 2014. Identification of indigenous yeast flora isolated from the five winegrape varieties harvested in Xiangning, China. Antonie Van Leeuwenhoek. 105, 533-540.
Sun, Y., Li, E.H., Qi, X.T., Liu, Y.L., 2015. Changes of diversity and population of yeasts during the fermentations by pure and mixed inoculation of Saccharomyces cerevisiae strains. Ann Microbiol. 2, 911-919.
Torija, M.J., Rozeès, N., Poblet, M., Guillamón, J.M., Mas, A., 2001. Yeast population dynamics in spontaneous fermentations: Comparison between two different wine-producing areas over a period of three years. Antonie van Leeuwenhoek. 79, 345-352.
Tofalo, R., Perpetuini, G., Fasoli, G., Schirone, M., Corsetti, A., Suzzi, G., 2014. Biodiversity study of wine yeasts belonging to the “terroir” of Montepulciano d’Abruzzo “Colline Teramane” revealed Saccharomyces cerevisiae strains exhibiting atypical and unique 5.8S-ITS restriction patterns. Food Microbiol. 39, 7-12.
Tofalo, R., Perpetuini, G., Schirone, M., Fasoli, G., Aguzzi, I., Corsetti, A., Suzzi, G., 2013. Biogeographical characterization of Saccharomyces cerevisiae wine yeast by molecular methods. Front Microbiol. 4, 166.
Valero, E., Cambon, B., Schuller, D., Casal, M., Dequin, S., 2007. Biodiversity of Saccharomyces yeast strains from grape berries of wine-producing areas using starter commercial yeasts. FEMS Yeast Res. 7, 317-329.
Wang, C.X., Liu, Y.L., 2013. Dynamic study of yeast species and Saccharomyces cerevisiae strains during the spontaneous fermentations of Muscat blanc in Jingyang, China. Food Microbiol. 33, 172-177.
White, T.J., Burns, T., Lee, S., Tayler, J., 1990. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis, M.A., Gelfand, D.H., Sninsdy, J.J., White, T.J. (eds), In PCR Protocols: a guide to methods and applications. Academics Press, San Diego. 315-322.
Zhang, H.Y., Lee, S.A., Bradbury, J.E., Warren, R.N., Sheth, H., Hooks, D.O., Richards, K.D., Gardnery, R.C., 2010. Yeasts isolated from New Zealand vineyards and wineries. Aust J Grape Wine Res. 16, 491-496.
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