Inhibition of the wine spoilage yeast Dekkera bruxellensis by bovine lactoferrin-derived peptides María Enrique a , Jose F. Marcos b , María Yuste c , Mireia Martínez c , Salvador Vallés a , Paloma Manzanares a, ⁎ a Departamento de Biotecnología de Alimentos, Instituto de Agroquímica y Tecnología de Alimentos, Consejo Superior de Investigaciones Científicas (CSIC) P. O. Box 73, 46100 Burjassot, Valencia, Spain b Departamento de Ciencia de los Alimentos, Instituto de Agroquímica y Tecnología de Alimentos, Consejo Superior de Investigaciones Científicas (CSIC) P. O. Box 73, 46100 Burjassot, Valencia, Spain c Bodegas Miguel Torres S.A., 08720 Villafranca del Penedés, Barcelona, Spain abstract article info Article history: Received 11 March 2008 Received in revised form 25 June 2008 Accepted 8 July 2008 Keywords: Dekkera bruxellensis Wine spoilage Antimicrobial peptide Lactoferrin-derived peptides Binding activity The antimicrobial action of lactoferrin (LF)-derived peptides against Dekkera bruxellensis strains isolated from spoiled wines has been examined. The study included a fifteen-residue peptide (LfcinB 17-31 ) derived from bovine lactoferricin B and a bovine LF pepsin hydrolysate (LFH). In vitro assays showed the inhibitory properties of LfcinB 17-31 on D. bruxellensis growth with IC 50 and MIC values in the micromolar range. Strains tested showed different sensitivity to the peptide. LfcinB 17-31 showed fungicidal properties towards all strains tested in laboratory growth medium. However, the extent of fungicidal activity was strain-dependent in must and wine, confirming the different antimicrobial action of peptides depending on both the food matrix and the target micro-organism. The binding of LfcinB 17-31 to D. bruxellensis cells was visualized by fluorescence microscopy and correlated with the fungicidal activity in the different matrixes. LfcinB 17-31 and LFH showed growth inhibitory properties in wine suggesting their potential use for spoilage control. © 2008 Elsevier B.V. All rights reserved. 1. Introduction Yeasts belonging to the genus Dekkera and its anamorph Bretta- nomyces have been recognized as spoilage yeasts in fermented beverages, and particularly in wine where Dekkera bruxellensis is the most prevalent species (Loureiro and Malfeito-Ferreira, 2003). They can develop in white and red wines at the end of the alcoholic fermentation and mainly during wine aging in wooden barrels where they can grow by fermenting the small amounts of residual sugars in wines producing abnormal amounts of unpleasant compounds such as volatile phenols and tetrahydropyridines (Heresztyn, 1986; Cha- tonnet et al., 1995). Recently, the presence of D. bruxellensis has been established in several vineyards and at different stages of the grape development (Renouf and Lonvaud-Funel, 2007). Several methods including an adequate cellar hygiene, fining and filtration of wines before introducing them into barrels, and aging at low temperatures have been used to prevent wine spoilage by D. bruxellensis (reviewed in Suárez et al., 2007). However, some of these procedures are not preferred by winemakers, and may be detri- mental to wine quality by reducing wine aroma and colour. Chemical preservatives such as sulphur dioxide (SO 2 ) can prevent the growth of Dekkera/Brettanomyces although strains vary considerably in their SO 2 sensitivity (du Toit and Pretorius, 2000) and it is difficult to maintain its effective concentration along the aging process. Moreover dimethyl dicarbonate, a microbial control agent with legal limits of 200 mg/l in wine, cannot completely inhibit their growth at that dose (Delfini et al., 2002) although its effectiveness is dependent on the contami- nation load (Costa et al., 2008). Due to the economic losses and damage to reputation associated with Dekkera/Brettamomyces spoilage, alternative strategies to control their growth are being continuously developed. The use of chitosan with a selective effect against Dekkera/Brettanomyces species in a mixed starter with S. cerevisiae has been described (Gómez-Rivas et al., 2004) as well as the development of thermal treatments for their inactivation both in wine and contaminated equipments (Couto et al., 2005). With regards to biological control, the use of killer toxins produced by Pichia anomala and Kluyveromyces wickerhamii with fungicidal effects against D. bruxellensis in wine has been reported (Comitini et al., 2004). In this context, proteins and peptides exhibiting antimicrobial properties might have a remarkable potential for food preservation and control of spoilage micro-organisms (Cotter et al., 2005; Rydlo et al., 2006) although in the wine making process selectivity is required for not affecting beneficial micro-organisms. Lactoferrin (LF), a major constituent of mammal milk, and derived peptides such as bovine lactoferricin (LfcinB) and derivatives, are known to exhibit a broad spectrum of antimicrobial activity (Orsi, 2004). The antibacter- ial action of either LF-derived peptides or lactoferrin hydrolysates International Journal of Food Microbiology 127 (2008) 229–234 ⁎ Corresponding author. Tel.: +34 963900022; fax: +34 963636301. E-mail address: pmanz@iata.csic.es (P. Manzanares). 0168-1605/$ – see front matter © 2008 Elsevier B.V. All rights reserved. doi:10.1016/j.ijfoodmicro.2008.07.011 Contents lists available at ScienceDirect International Journal of Food Microbiology journal homepage: www.elsevier.com/locate/ijfoodmicro