fermentation Review Challenges of the Non-Conventional Yeast Wickerhamomyces anomalus in Winemaking Beatriz Padilla 1, Jose V. Gil 2,3 and Paloma Manzanares 2,* 1 INCLIVA Health Research Institute, 46010 Valencia, Spain; [email protected] 2 Department of Biotechnology, Instituto de Agroquímica y Tecnología de Alimentos (IATA), Consejo Superior de Investigaciones Científicas (CSIC), Paterna, 46980 Valencia, Spain; [email protected] 3 Departamento de Medicina Preventiva y Salud Pública, Ciencias de la Alimentación, Toxicología y Medicina Legal, Facultad de Farmacia, Universitat de València, Burjassot, 46100 Valencia, Spain * Correspondence: [email protected]; Tel.: +34-96 390-0022 Received: 27 July 2018; Accepted: 18 August 2018; Published: 20 August 2018 Abstract: Nowadays it is widely accepted that non-Saccharomyces yeasts, which prevail during the early stages of alcoholic fermentation, contribute significantly to the character and quality of the final wine. Among these yeasts, Wickerhamomyces anomalus (formerly Pichia anomala, Hansenula anomala, Candida pelliculosa) has gained considerable importance for the wine industry since it exhibits interesting and potentially exploitable physiological and metabolic characteristics, although its growth along fermentation can still be seen as an uncontrollable risk. This species is widespread in nature and has been isolated from different environments including grapes and wines. Its use together with Saccharomyces cerevisiae in mixed culture fermentations has been proposed to increase wine particular characteristics. Here, we review the ability of W. anomalus to produce enzymes and metabolites of oenological relevance and we discuss its potential as a biocontrol agent in winemaking. Finally, biotechnological applications of W. anomalus beyond wine fermentation are briefly described. Keywords: non-Saccharomyces yeasts; Wickerhamomyces anomalus; Pichia anomala; enzymes; glycosidases; acetate esters; biocontrol; mixed starters; wine 1. Introduction Saccharomyces cerevisiae is the main microorganism involved in the alcoholic fermentation of grape must. Moreover, the use of selected S. cerevisiae strains has provided an improvement in the control and homogeneity of fermentations. However, winemaking is a non-sterile process, and many other species of yeasts belonging to various non-Saccharomyces genera prevail during the early stages of alcoholic fermentation and contribute significantly to the character and quality of the final wine [1]. In the past, non-Saccharomyces yeasts were considered of secondary significance or as undesirable spoilage yeasts. Nowadays, the role of non-Saccharomyces has been re-evaluated, and it is widely accepted that selected strains can positively influence the winemaking process [2]. Beyond the contribution of non-Saccharomyces yeasts to wine aroma complexity [3], these yeasts can help address some of the modern challenges in winemaking, including the reduction of the ethanol content of wine [4–7] or the control of wine spoilage [8,9]. Ecological studies have shown that species of mainly Hanseniaspora (Kloeckera), Candida, and Metschnikowia initiate the fermentation together with species of Pichia, Issatchenkia, and Kluyveromyces. Occasionally, representatives of Brettanomyces, Schizosaccharomyces, Torulaspora, Rhodotorula, Zygosaccharomyces, and Cryptococcus genera are also present. These yeasts decline by mid-fermentation, and then, S. cerevisiae becomes predominant and continues the fermentation [10]. Based on the capability of some of these non-Saccharomyces yeasts to produce flavor-enhancing enzymes or to modify the concentration Fermentation 2018, 4, 68; doi:10.3390/fermentation4030068 www.mdpi.com/journal/fermentation Fermentation 2018, 4, x FOR PEER REVIEW 2 of 13 mid-fermentation, and then, S. cerevisiae becomes predominant and continues the fermentation [10]. FermentationBased on 2018the , capability4, 68 of some of these non-Saccharomyces yeasts to produce flavor-enhancing2 of 14 enzymes or to modify the concentration of secondary metabolites, different mixed starters have been designed and proposed as a tool to enhance wine quality [3,11]. Moreover, several species including Lachanceaof secondary thermotolerans metabolites,, differentMetschnikowia mixed starterspulcherrima have, been Torulaspora designed delbrueckii and proposed, Pichia as a toolkluyvery to enhance, and wineSchizosaccharomyces quality [3,11]. 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