Yeast biodiversity in patagonian ciders: effect of temperature, apple variety and fermentation scale

GONZÁLEZ FLORES M.; DI NICCOLO R.; VERA MACAYA D.; RODRÍGUEZ M.E.; LOPES C.A.

Jornada; V Jornadas Sudamericanas de Biología y Biotecnología de Levaduras; 2015

Cider is the product of alcoholic fermentation of juice obtained from healthy and clean apples, andNorth Patagonia is the major apple and cider production region of Argentina. Most ciders areelaborated using commercial yeast starters traditionally used for winemaking; however, a fewartisanal producers make still use of the natural biota to produce differential beverages. The aim ofthis work was to evaluate the diversity of yeasts present in ciders elaborated by means of naturalfermentation and the effect of substrate, temperature and fermentation scale on this biodiversity.These yeasts will be used as a genetic bank to develop a regional cidermaking starter.Yeast samples were obtained from fermentations carried out in flasks containing 1L of Red Delicious(RD) or Granny Smith (GS) musts (substrate effect) at 13ºC and 26ºC (temperature effect).Additionally, the same musts were fermented in cellar conditions (5000L, scale effect). Yeasts wereisolated in complete medium (GPY) and identified by sequencing the D1/D2 domains of 26S DNAr.The 100% of the isolates recovered to the initial stages of both RD and GS musts belonged to thegenera Pichia spp., Hanseniaspora spp., Rhodotorula spp. and Cryptococcus spp, among these,Hanseniaspora guillermondii (50%) and Cryptococcus tephrensis (38%) were the majority speciesin RD and GS respectively. Contrarily, yeasts belonging to Saccharomyces genus dominated middleand end stages. This behaviour was observed at both 1L and 5000L scales and the two temperatures.Among Saccharomyces isolates, the species S. cerevisiae (Sc) was majority in most conditions withthe exception of RD must fermented at 13ºC where Saccharomyces uvarum (Su) predominated (75%and 62% of total biota at middle and end stages, respectively).The intraspecific variability of Su (56 isolates) was then evaluated by mtDNA-RFLP and RAPD-PCR(Primers p24 and p28) methods. Genetic profiles obtained for these isolates were compared withthose present in Su isolates from natural habitats (Araucaria araucana, 9 strains) and apple chichas(14 strains) from Andean North Patagonia. UPGMA analysis demonstrated that most Su cider isolateswere closely related to Su strains from chichas, while those strains from natural habitats formed adifferent cluster. Intraspecific variability (IV) was higher in Su isolates from natural habitats (50%IV) than in those from fermentations (chicha 40%, cider 8.92%), which could indicate a selectivepressure of fermentative conditions on this species. Summarizing, a strong effect of fermentationtemperature and apple variety on yeast diversity was evidenced. S. uvarum was detected for the firsttime in ciders from South America; these strains were genetically related to strains isolated fromapple chicha, a traditional fermented beverage from Andean North Patagonia.