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Studies have not fully explained the underlying mechanism of spermidine-mediated heat tolerance. This study investigated the possible role of spermidine (Spd) in regulating citrus heat tolerance. The results showed that exogenous Spd effectively alleviated the limitation of high temperature (HT) on photosynthesis. Exogenous Spd increased the chlorophyll content, net photosynthetic rate, intercellular carbon dioxide concentration, stomatal conductance, maximum and effective quantum yield of PSII photochemistry, nonphotochemical quenching coefficient, and electron transport rate in citrus seedlings under HT stress, but declined the stomatal limitation value. In addition, Spd treatment promoted the dynamic balance of the citrus enzymatic and non-enzymatic antioxidants system. Spd application significantly increased the activity of superoxide dismutase, peroxidase, catalase, ascorbic acid, and glutathione and the expression level of corresponding genes at high temperature, while reducing the content of H 2 O 2 and malondialdehyde. Therefore, our findings suggested exogenous Spd significantly ameliorated citrus physiological and photosynthetic adaptation under HT stress, thereby providing helpful guidance for citrus cultivation in HT events.