The objective of this study was to determine the seasonal incidence of ultraviolet A (UVA) and ultraviolet B (UVB) in the water column of the Patos Lagoon estuary and the effects of UV radiation (UVR) at different organizational levels. Field work was conducted to collect UVA and UVB radiance, transparency, salinity, seston and chlorophyll a (Chl a) concentration, and fish larvae at an inner and at an outer sampling site in the Patos Lagoon estuary (32°S). Cell culture techniques combined with tools for fluorimetric detection were used to assess possible damage caused by UVR. GEM-81 erythrophoroma cells and larvae of silverside Odontesthes argentinensis were used as biological indicators. Field results revealed that at the inner station, UVA penetration was deeper in autumn and shallower in spring when water-column was less transparent, fresher, and had the highest concentration of seston and Chl a. At the outer station, UVA penetration in the water-column in spring was significantly different than in summer, when transparency and water salinity were high and Chl a was low. UVB did not present significant variation between seasons at both sampling stations. Experiments with GEM-81 cell lineage revealed that UVA doses similar to the environment (1.35 J cm−2) inhibited cell proliferation, increased the concentration of reactive oxygen species (ROS) and the lipid peroxide content (LPO), and lowered the activity of the antioxidant defense system (total antioxidant capacity against peroxyl radicals (TOSC)). For UVB, 0.075 J cm−2 caused cytotoxicity and 0.015 J cm−2 induced DNA damage in cells. Silverside larvae showed low growth rate when exposed to UVA and low antioxidant capacity when exposed to UVB. These results suggest that planktonic organisms that live close to surface water column in the Patos Lagoon estuary are subject to the effects of continued exposure to UVR. Knowledge of this impact is relevant to understand future changes in the populations dynamics of planktonic organisms and its ecological and economic implications to southern Brazil.