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Phylogenetic Diversity of Arkansas Vineyard and Wine Microbiota

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Phylogenetic Diversity of Arkansas Vineyard and Wine Microbiota University of Arkansas, Fayetteville ScholarWorks@UARK Theses and Dissertations 5-2020 Phylogenetic Diversity of Arkansas Vineyard and Wine Microbiota Natacha Cureau University of Arkansas, Fayetteville Follow this and additional works at: https://scholarworks.uark.edu/etd Part of the Food Microbiology Commons, Horticulture Commons, and the Viticulture and Oenology Commons Citation Cureau, N. (2020). Phylogenetic Diversity of Arkansas Vineyard and Wine Microbiota. Theses and Dissertations Retrieved from https://scholarworks.uark.edu/etd/3600 This Dissertation is brought to you for free and open access by ScholarWorks@UARK. It has been accepted for inclusion in Theses and Dissertations by an authorized administrator of ScholarWorks@UARK. For more information, please contact [email protected]. Phylogenetic Diversity of Arkansas Vineyard and Wine Microbiota A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Food Science by Natacha Cureau University Pierre and Marie Curie Bachelor of Science in Biology, 2010 University Pierre and Marie Curie Master of Science in Cellular and Molecular Biology, 2012 May 2020 University of Arkansas This dissertation is approved for recommendation to the Graduate Council. Renee T. Threlfall, Ph.D. Dissertation Director Luke R. Howard, Ph.D. Sun-Ok Lee, Ph.D. Committee Member Committee Member Andronikos Mauromoustakos, Ph.D. Mary C. Savin, Ph.D. Committee Member Committee Member Abstract Microorganisms inhabiting the soil, leaves, and grapes in vineyards influence the terroir, a set of environmental factors that impact wine characteristics. Previously, the main method to identify microorganisms was to culture on different media, but a large portion (> 99%) could not be cultured and were consequently unidentified. However, the emergence of new molecular tools has enabled further study and identification of microorganisms such as indigenous vineyard microbiota. The objective of this research was to investigate the phylogenetic diversity of Arkansas vineyard and wine microbiota using high-throughput sequencing. In terms of the Arkansas vineyards, the fungal and bacterial diversity of leaf, soil, and grapes of different varieties from experimental vineyards (one with a high tunnel/covered structure) and commercial vineyards were evaluated. Overall, the fungal diversity of the vineyard grape, leaf, and soil samples was greater in 2017 compared to 2016, while the bacterial diversity was only greater for grapes and leaves. The indigenous grape and leaf microbiota varied depending on the location, grape varieties, and year, but some microorganisms, such as Methylobacterium and Sphingomonas, were identified as a core microbiota and were present in all leaf samples. The covered structure (high tunnel) impacted the grape and leaf microbiota with distinct microbiota compared to the other locations (dominant fungal genera Cladosporium in grapes and leaves). The soil microbiota was similar between the different vineyards with a prevalence of the fungi Mortierella and bacteria from the Acidobacteria phylum. Dissimilarities in soil bacterial communities were observed mainly for the microorganisms present at a small relative abundance (contributing to less than 20% of the total bacterial communities). In terms of wine, the presence and performance of mycobiota during fermentation of two grape varieties with different sulfite levels and yeast inoculations were evaluated. Indigenous juice microbiota of the two grape varieties were similar with the two most abundant fungal genera as Podosphaera and Candida with dissimilarities in fungal communities’ relative abundances. Sulfite levels and yeast impacted the mycobiota and wine composition during fermentation. The fermentation of uninoculated (spontaneous fermentation) juice varieties were dominated by Hanseniaspora and Saccharomyces genera. This research demonstrated the phylogenetic diversity of Arkansas vineyard and wine microbiota as well as the impact that microbiota can have on wine production. Objectives 1. Determine the phylogenetic diversity and taxonomic identity of the indigenous microbiota in Arkansas vineyards using high-throughput sequencing methods 2. Evaluate the phylogenetic diversity and taxonomic identity of microbiota during fermentation of Arkansas grape juice with different sulfite levels and yeast inoculations Acknowledgments I would like to express my sincere gratitude to my advisor Dr. Renee Threlfall for the constant help, advice, and support. Thank you for always answering my concerns quickly, for your edits in pink ink, for always finding something funny to say, and for your overall guidance. I would also like to thank my committee members; Dr. Luke Howard, Dr. Sun-Ok Lee, Dr. Andy Mauromoustakos, and Dr. Mary Savin for sharing your expertise in your scientific fields, your useful advice, and for your support throughout my studies. I would like to thank Dr. Carbonero for giving me this educational opportunity at the Food Science Department at the University of Arkansas. Thank you to all the vineyard managers and winemakers from Post Vineyards and Winery, Chateau aux Arc Vineyard and Wineries, Keels Creek Winery, the University of Arkansas Fruit Research Station, and the University of Arkansas Agriculture Research Extension for kindly giving me access to their vineyards and letting me collect the valuable samples. Thank you also to Luke Holcombe from Scott Laboratories who generously donated yeast for the fermentation study. Thank you to all my lab mates for the great time we shared in and outside the lab. A special thanks to Laura Lavefve who helped in many ways with my research, from collecting samples in the difficult Arkansas summer weather, to annotating more than 500 tubes, to helping during all the experiments, to fermenting juices, and always with a positive attitude. Thank you to Paulina Sobczak for upgrading my vineyard maps. Thank you to Sarah Mayfield and Andrea Meyers for their help during the final days preceding the submission of this dissertation. I am grateful to each of you for your friendship. Thank you to Daya Marasini for his help with the sequencing and technical support. Thank you to Kevin C. Thompson for his guidance in statistics. Thank you to Cindi Brownmiller who was always available to answer to my many questions with a smile and for her technical help in Biochemistry. Thank you to all the Food Science Department Faculty and Staff members that are always here to help students, especially Cathy S. Hamilton, Eric Friend, and Connie O. Tharel. Thank you to Dr. Ya-Jane Wang for her support and her help in the completion of my degree. Thank you to Dr. Jeyamkondan Subbiah and the Food Science Department, the University of Arkansas Graduate School, the American Society for Enology and Viticulture, the American Society for Enology and Viticulture- Eastern Section, and the Arkansas Association of Grape Growers for their financial support. Thank you to my supportive friends Mary, Eddy, AJ, Jay, and Jola. You became my second family in the United States, and I am so thankful that I have met such welcoming people here. You made this chapter of my life easier. To my long time and supportive friends Wesley (LDG), Bouchra, Mariam, Naoil, Sonia, Nino, Reza, Robin, Meg, Ava, Lindsey, Vishwesh, and Chloë. Thank you for always believing in me even when I had doubts. I am thankful for your support and love. Last but not least, I would like to thank Vitaliy, for his constant support, for being a strong shoulder that I could always lean on, for his advice and for his kindness. Thank you to my family, my mother Joanna and brother Lucas, for all the love you gave me throughout the years and for helping me to become who I am today. I am blessed and thankful to have you all in my life. Dedication This dissertation is dedicated to my mother and brother. Table of Contents Literature Review ......................................................................................................................... 1 Vineyard-Associated Microbiota ............................................................................................. 3 Vineyard soil-associated microbiota ....................................................................................... 3 Grape-associated microbiota ................................................................................................... 7 Winemaking Steps ................................................................................................................... 19 Harvest ................................................................................................................................... 19 Processing grapes .................................................................................................................. 20 Alcoholic fermentation .......................................................................................................... 21 Pressing.................................................................................................................................. 22 Malolactic fermentation......................................................................................................... 22 Racking .................................................................................................................................. 22 Fining ..................................................................................................................................... 23 Filtering
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