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To reduce the indiscriminate use of pesticides and extend the postharvest shelf life of peach fruit (Prunus persica, cv. Baihua) from southeast China, mainly the microbial antagonism of indigenous yeasts was studied and applied in the construction of composite film. In this study, 14 yeast strains of 9 genera were screened out from the surface of peaches by isolation, purification, cultivation, and identification. Through an experimental analysis of the in vitro inhibition zone and the in vivo colonizing capacity, 1 × 108 CFU mL−1 of Candida oleophila sp-ELPY12B and Cryptococcus laurentii sp-ELPY15A proved most efficient against the major pathogens and were chosen as candidate fungicides. In combination with Na-alginate film (0.4% glycerin as the plasticizer and 0.1% Tween-80 as the emulsifier), the preservative effects of these composite-treated groups also showed the best antifungal effects, which significantly delayed the postharvest preservation period by about 6–7 d under an ambient temperature of 25 ± 3 °C and a relative humidity of 50–70%. [ABSTRACT FROM AUTHOR]