In the present study, ten yeasts were isolated from fruit juices and subjected to in vitro tests for screening probiotic potentiality. Three isolates exhibited higher tolerance for pH and heat stress. Based on preliminary screening, isolates were subjected to molecular identification and identified as Wickerhamomyces anomalusVIT-ASN01, Saccharomyces cerevisiae VIT-ASN03 and Yarrowia lipolytica VIT-ASN04. The screened yeast isolates survived and grew well at human gastrointestinal conditions in pH 2.5 and 0.3{\%} (w/v) oxgall at 37°C. The antagonistic activity of yeast isolates were tested against clinical pathogens, viz.S. typhii, B. subtilis and E. coli. W. anomalusVIT-ASN01 showed maximum tolerance to high bile salt concentration and acidity, could remove cholesterol 51.28{\%} during 48 h and produced 586.55 mg/L of exopolysaccharide. It can be concluded that the screened novel yeast isolates can serve as promising probiotics in various fields of food industry.