To screen novel yeast strains suitable for the low-temperature fermentation of mulberry fruit wine, this study isolated and purified strains from naturally fermented mulberry broth. Yeast strains with excellent low-temperature tolerance and alcohol-producing capacity were then hierarchically screened. The selected yeast strains were further analyzed for tolerance, and their fruit wine fermentation performance and aroma characteristics evaluated. Two Pichia kluyveri strains (PK-1 and PK-2) with excellent aroma producing ability and one Saccharomyces cerevisiae strain (SC-1) with superior fermentation performance were identified. Results showed that the three strains grew well at 16 ℃, exhibiting an ethanol tolerance and high sugar and sulfur dioxide production of 15% (v/v), 400 g.L−1, and 300 mg.L−1, respectively. All strains could also tolerate fruit wine production conditions at pH 3. Fermentation tests showed that the SC-1 group reached an alcohol content of 13.26% vol. A total of 53 volatile SPXaroma compounds were identified, of which 24 were found in the SC-1 group, 32 in the PK-1 group, and 31 in the PK-2 group. Isoamyl alcohol and phenylethanol concentrations (m/v) in the SC-1 group reached 43.09 μg.mL-1 and 143.66 μg.mL-1, respectively. Phenethyl acetate concentrations (m/v) in the PK-1 and PK-2 groups reached 402.96 μg.mL-1 and 421.51 μg.mL-1, respectively, showing significant differences in key aroma components. The SC-1 strains exhibited excellent fermentation performance under lowtemperature conditions, with a notably high isoamyl alcohol content (143.66 μg.mL-1). The two PK strains showed excellent esterproducing ability, which could contribute to the development a novel floral-fruity aroma profile for co-fermented fruit wines. Overall, the results of this study provide a theoretical basis for improving the aroma profile of fruit wine.