Wine fermentation was traditionally carried out by indigenous yeasts associated with grapes and cellar equipment. Today majority of wine production is based on use of active dried yeast which ensures rapid and reliable fermentation and reduces the risk of sluggish or stuck fermentation and microbial contamination. Most commercial wine yeast strains available today have been selected in the vineyard for oenological traits. Despite the availability of several Saccharomyces cerevisiae commercial strains intended for wine production, strains isolated from winery regions are usually more adapted to their own climatic conditions, grapes and also partially responsible for particular characteristics that frequently identify specific wines and regions. The selection of yeasts for winemaking consists of identifying specific cultures, mainly Saccharomyces, which can ferment grape juice effectively and can produce good quality wines. Thus, the aim of this is to isolate and characterize S. cerevisiae strains from ”Trifeshti” vineyard that could be used as a tool for improving the wine quality and reproducibility of the wines obtained, allowing the creation of strong identity that could facilitate their market insertion. Isolation and selection of local yeast strains from the vineyard ”Trifeshti” was carried out by means of obtaining pure cultures from fermenting grape must. Thus the microbiota of an important winery region”Trifeshti” was studied in order to isolate and characterize Saccharomyces cerevisiae strains that could be used on wine production. Samples of grape must was characterized by the following initial indices: sugar concentration – 172/218 g/L, acidity titratable-6,2/6,5 g/L (Aligote/Sauvignon); sugar concentration – 220/230 g/L, acidity titratable-7,7/5,4 g/L (Cabernet-Sauvignon/Merlot). During fermentation, before withdrawing the samples, the bottles were shaken to thoroughly mix the contents and get all of microbial cells in suspension. Each of these samples was streaked on 10 Petri dishes containing YPD agar medium (1% yeast extract, 2% peptone, 2% glucose and 1.5% agar) and incubated at 28°C for 3 days to allow colony formation. A total of 47 colonies were selected randomly. Cultures were then stored at 4°C until further analysis. Identification of these isolates was carried out conventionally using morphological and physiological characterization as described by Burian N. (Practicescaia microbiologhia vinodelia, 2003). From 47 yeast isolated, 32 were identified as Saccharomyces cerevisiae, 12 as Saccharomyces pastorianus and 3 as Saccharomyces bayanus. Thirty two strains of Saccharomyces cerevisiae, isolated from the ”Trifeshti” vineyard, were tested for: fermentation vigor, ethanol, SO2, copper and cold resistance, volatile acidity and H2S production, β-glycosidase activity, foam production, killer character and mode of growth in liquid medium. Some of them showed advanced technological characteristics indicating that they could be applied on wine production in order to increase the quality and assure the particular wine characteristics of that region. From the results of this investigation we are able to select ten yeast strains for more detailed fermentation trials and possible use as a starter culture in production of typical wines. In conclusion, this study was useful in obtaining interesting information about the oenological characteristics of Saccharomyces tested, which is a necessary first step in strain selection.