Smelt Osmerus eperlanus has two different life history strategies in The Netherlands. The migrating population inhabits the Wadden Sea and spawns in freshwater areas. After the closure of the Afsluitdijk in 1932, part of the smelt population became landlocked. The fresh water smelt population has been in severe decline since 1990, and has strongly negatively impacted the numbers of piscivorous water birds relying on smelt as their main prey. The lakes that were formed after the dike closure, IJsselmeer and Markermeer have been assigned as Natura 2000 sites, based on their importance for (among others) piscivorous water birds. Because of the declining fresh water smelt population, the question arose whether this population is still supported by the diadromous population. Opportunities for exchange between fresh water and the sea are however limited to discharge sluices. The relationship between the diadromous and landlocked smelt population was analysed by means of otolith microchemistry. Our interpretation of otolith strontium ((88)Sr) patterns from smelt specimens collected in the fresh water area of Lake IJsselmeer and Markermeer, compared to those collected in the nearby marine environment, is that there is currently no evidence for a substantial contribution from the diadromous population to the spawning stock of the landlocked population.