Chemosensory stimuli play a crucial role for host selection in insects, including the fruit fly Drosophila melanogaster. Drosophila has been instrumental in unraveling the neurological basis of olfactory processing in insects. Basic knowledge regarding chemical ecology and thorough studies of olfactory preferences are still lacking to a great extent in D. melanogaster, however. We have characterized repeatable variation in olfactory preference between five classical D. melanogaster wild-type strains toward a large array of natural host odors and synthetic compounds. By recording the rate of attraction over up to 24 hr, we could compare stimuli varying in attractiveness and characterize phenotypic parameters on the basis of individual stimuli and the whole stimulus array. Behavioral differences between strains were predominantly due to variation in a single phenotypic parameter: their overall responsiveness toward optimal and suboptimal olfactory stimuli. These differences were not explained by variation in olfactory sensitivity, locomotory activity, or general vigor monitored by survival. Comparisons with three recently established wild-type strains indicated that a high behavioral threshold against accepting suboptimal olfactory stimuli is the characteristic phenotype of wild D. melanogaster.