The effectiveness of iturin A (IA) and chitosan (CTS) coating, alone or in combination (CTS-IA), against mango anthracnose caused by Colletotrichum gloeosporioides was investigated. In vitro tests, concentrations of IA ranging from 0.03 to 0.12 g/L, and CTS ranging from 10 g/L to 20 g/L, significantly inhibited the growth of C. gloeosporioides. Moreover, the combination of IA with 15 g/L CTS demonstrated an additional inhibitory effect on C. gloeosporioides growth. In vivo experiments, 0.12 g/L IA with 15 g/L CTS had a synergistic effect in preventing lesion expansion in mangoes after 20 days of storage at 12 °C. Compared to using IA or CTS alone, CTS-IA coating showed better results in disease control, respiratory rate, weight loss, total soluble solids, titratable acidity, and fruit firmness maintenance during storage. The combined effect of IA and CTS may be mainly attributed to the enhancement of mango defense enzymes such as GR, CAT, APX, and POD induced by the CTS-IA. In conclusion, CTS-IA may have significant potential in postharvest preservation applications for mangoes.