Assortative mating is measured as a phenotypic or genotypic correlation between mates. Although biologists typically view assortative mating in terms of mate preference for similar partners, correlations between mates can also arise from phenotypic spatial structure arising from spatial isolation or habitat preferences. Here, we test whether diet-assortative mating within an ecologically variable population of threespine stickleback results from small-scale geographic isolation or microhabitat preference. We find evidence for assortative mating in the form of a positive correlation between mated pairs’ diets (measured using stable isotopes). Stable isotopes reveal diet differences between different nesting areas and among individuals using different nest habitat within a nesting area. This spatial segregation of diet types should generate some assortative mating, but is insufficient to explain the observed assortment strength. Significant male–female isotope correlations remain after controlling for spatial variables. We therefore conclude that sticklebacks’ diet-assortative mating arises from additional behavioral preference. More generally, our results illustrate the point that spatial segregation can only drive appreciable levels of phenotypic assortative mating when environment-phenotype correlations are parallel and strong in both sexes. Consequently, intraspecific assortative mating may typically entail mating preferences rather than just spatial cosegregation of phenotypes.