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Gray mold is a main disease of strawberry fruit (<i>Fragaria</i> × <i>xananassa</i> cv. Camino Real) caused by <i>Botrytis cinerea</i>, which leads to marketable value losses in the supply chain. The purpose of this study was to investigate the effects of exogenous melatonin (MT) on the physicochemical quality, antioxidant defense system, and disease resistance of strawberry fruit to <i>B. cinerea</i> infection. The results revealed that strawberry fruit immersed in 100 µM MT for 15 min effectively maintained its brightness and delayed the change in fruit color. MT also maintained the level of titratable acidity and slowed down the increase of total soluble solids in strawberry fruit. Moreover, strawberries immersed in MT maintained a fresh weight and fruit firmness, as well as reduced <i>B. cinerea</i> infection when compared to the untreated control fruit and fruit treated with 5% NaOCl. In addition, MT increased the accumulation of DPPH scavenging capacity and the activity of antioxidant enzymes (SOD, POD, and APX) with the exception of CAT. The same effect was also observed in strawberry fruit after immersion in MT and followed by <i>B. cinerea</i> inoculation. These findings demonstrated that exogenous MT could effectively maintain the postharvest quality of strawberries, even when the fruit was inoculated with <i>B. cinerea</i>.