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Reservoirs in Hong Kong constitute an essential component of local freshwater habitats, but their biodiversity still needs to be better understood. This study adopted environmental DNA (eDNA) technology to assess faunal biodiversity in local reservoirs. This approach enabled the analysis of metazoan diversity in Hong Kong’s reservoirs, which would have been challenging to achieve through conventional, labourintensive, and invasive ecological surveys. We surveyed eight reservoirs of varying sizes and ages, including their catchment areas, to provide baseline information for establishing a long-term monitoring programme for local reservoir habitats. The results revealed a higher success rate in amplifying eDNA fragments from the reservoirs than from catchment areas. The cytochrome oxidase I (COI) and 18S rRNA markers exhibited differential performance in biodiversity assessment. Notably, eDNA detected known naturalised species, such as Caridina cantonensis (bee shrimp), Cryptopotamon anacoluthon (freshwater crab), Parazacco spilurus (predaceous chub), and Liniparhomaloptera disparis (Hillstream loaches). Moreover, potential exotic species, Cherax quadricarinatus (the red-clawed crayfish), Oreochromis niloticus (Nile tilapia), and Paramisgurnus dabryanus (large-scaled loach), were discovered through both eDNA metabarcoding and ecological surveys. The findings suggest that reservoir-associated human activities, such as the release of aquarium pets, are potential sources of species introduction in these freshwater habitats. Regular biodiversity monitoring using eDNA technology can be an effective tool in freshwater ecosystem management and invasive species detection.