Urban runoff can reach coastal aquatic environments; however, little is known about the effect of salinity on road runoff toxicity. The objective of this study is to investigate the toxicity of highway road dust over a salinity gradient from 5 to 35‰, in an estuarine benthic amphipod, Grandidierella japonica. Road dust toxicity was evaluated by assessing mortality after 10 days of exposure and short-term microbead ingestion activity of the amphipod. For all road dust samples considered, amphipod mortality increased with increasing salinity, whereas no significant difference in mortality was observed among test salinities in the reference river sediment. Ingestion activity during exposure to road dust decreased with increasing salinity. In fact, none of the individuals ingested any microbeads at salinity of 35‰. If assumed microbead ingestion is a proxy for feeding activity, high mortality at 35‰ could be attributed to aquatic exposure and not to dietary exposure. These findings suggest that road dust may have considerable impact on benthic organisms at high salinity levels.