Abstract: To reduce the indiscriminate use of pesticides and extend the postharvest shelf life of peach fruit (cv. Baihua) from southeast China, microbial antagonism of indigenous yeasts was mainly studied and applied in construction of composite film. After isolation, purification, cultivation and identification, a total of 14 yeast strains from 9 genera were screened out from the surface of peaches. By experimental analysis of in vitro inhibition zone and in vivo colonizing capacity, Candida oleophila sp-ELPY12B and Cryptococcus laurentii sp-ELPY15A, which have conservative structure of D1/D2 domain sequences and were considered as new species by phylogenetic analysis, were finally chosen as fungicides against the major pathogens. In combination of Na-alginate film (0.4 % glycerin as plasticizer and 0.1 % Tween-80 as emulsifier), the preservative effects of composite-treated groups (1 × 108 CFU mL−1 of Candida oleophila sp-ELPY12B and Cryptococcus laurentii sp-ELPY15A) showed best antifungal effects, which significantly delayed the postharvest preservation period about 6 - 7 d under ambient temperature of 25 ± 3°C and relative humidity of 50 - 70%.