The damage of chilli, an important spice fruit, comes from fungal diseases caused mainly by Aspergillus niger and Colletotrichum sp. The fungi on chilli fruits would directly harm consumers’ health. Plant extracts containing bio-active compounds with antimicrobial properties could be a good possible solution to deal with the fungi. This study aims to evaluate the in vitro and in vivo efficacy of aqueous extracts from the leaves of Ageratum plants against A. niger and Colletotrichum sp. The results show that the optimal efficacy of the treatment of Ageratum leaf extract is at a concentration of 6%, with the efficacy of Ageratum leaf extract on colonial diameter at approximately 43–44% for the two fungi at 96 hours after inoculation. The 6% Ageratum leaf extract has a high efficacy (~54.23%) on limiting the development of Aspergillus rot lesions on chilli fruits 4 days after inoculation. Meanwhile, the efficacy of the extract on Colletotrichum lesions is 11.34%, lower than that of Aspergillus rot.