Alewife (Alosa pseudoharengus) recently became established in Lake Champlain and may compete with native rainbow smelt (Osmerus mordax) for food or consume larval rainbow smelt. The strength of this effect depends partly on the spatial and temporal overlap of different age groups of the two species; therefore, we need a better understanding of factors affecting alewife and rainbow smelt distributions in Lake Champlain. We used hydroacoustics, trawls, and gill nets to document vertical fish distribution, and recorded environmental data during 16 day–night surveys over two years. Temperature, temperature change, and light were all predictors of adult and age-0 rainbow smelt distribution, and temperature and light were predictors of age-0 alewives' distribution (based on GAMM models evaluated with AIC). Adult alewives were 5–30 m shallower and age-0 alewives were 2–15 m shallower than their rainbow smelt counterparts. Adult rainbow smelt distribution overlapped with age-0 rainbow smelt and age-0 alewives near the thermocline (10–25 m), whereas adult alewives were shallower (0–6 m) and overlapped with age-0 alewives and rainbow smelt in the epilimnion. Adult rainbow smelt were in water < 10–12 °C, whereas age-0 rainbow smelt were in 10–20 °C, and adult and age-0 alewives were in 15–22 °C water. Predicting these species distributions is necessary for quantifying the strength of predatory and competitive interactions between alewife and rainbow smelt, as well as between alewife and other fish species in Lake Champlain.