Lately, agriculture, livestock, forestry, and aquaculture activities have been greatly developed in Chilean North Patagonia, negatively impacting the balance of the environmental conditions in lakes and affecting the development and survival of several native species. The aim of this study was to assess the anthropic impact on a zooplankton community in two North Patagonian lakes. We collected samples from four sites belonging to Lake Icalma and Lake Llanquihue, including four replicates per site. Water samples were analyzed for physicochemical characteristics and zooplankton communities. We focused on the presence of Daphnia pulex, a species of zooplanktonic crustacean that performs a key role in capturing energy from primary producers to deliver it to final consumers such as fish. We found that Llanquihue showed higher total phosphorus, nitrogen, copper, iron, manganese, total dissolved solids (TDS), and conductivity (EC) than Icalma. Furthermore, ecological variables were greatly decreased due to total P, total N, manganese, copper, total dissolved solids, and conductivity, which changed the species dominance of the zooplankton community in Llanquihue, indicating some degree of anthropization. This study provides fundamental information on the anthropogenic impact on water quality, as well as on zooplankton diversity, highlighting the importance of monitoring the health of these North Patagonia freshwater ecosystems.