The distribution of phytoplanktonic flagellates in aquatic ecosystems has been widely attributed to a number of driving factors. In this study, we evaluated the influence of behavior on the daytime, seasonal depth distribution and vertical niche separation of five phylogenetically contrasting species of freshwater flagellates. A model predicting distribution was formulated using the dominant behavioral preferences for light, oxygen, and carbon dioxide, previously quantified in laboratory experiments, and was subsequently applied to the physical and chemical conditions measured in a small, strongly stratifying, hypertrophic lake. This model predicted the daytime depth distributions of natural populations of flagellates well, with an average areal fit of above 56% for all species; above 74% for Ceratium furcoides, Chlamydomonas sp., and Dinobryon sertularia; and of up to 93% during stratification. Regression analyses showed no significant variation from a 1 : 1 relationship between the predicted and observed average depths of species in the water column. The model also predicted the constriction into discrete vertical niches upon stratification and delineated the progression from Plagioselmis nannoplanctica in surface waters, through C. furcoides and D. sertularia, to Chlamydomonas sp. and Euglena gracilis deeper in the water column. Changes in observed distributions could not be directly correlated with other members of the plankton community or explained by the segregation of nutrients. The model's wider, transferable applicability suggested that behavioral response to environmental gradients may predict many patterns of distribution, particularly during stratification. In addition to known physiological and biochemical influences, this investigation emphasized the importance of behavioral response in the functional ecology of phytoplanktonic flagellates.Phytoplanktonic flagellates are a diverse group of unicellular or colonial eukaryotic microorganisms characterized by controlled motility combined with an ability to photosynthesize. Commonly occurring in both freshwater and marine habitats, they form an important functional component of the phytoplankton community (Salonen et al. 1984;Stone et al. 1993) and are frequently the principal contributor to harmful algal blooms and red tides (Smayda 1997). The spatial and temporal distributions of flagellates in aquatic ecosystems have been attributed to a large number of driving factors such as physiological response (e.g., Harris et al. 1979), competitive interactions (Riegman et al. 1996), succession (Gasol et al. 1992), the changing distribution of resources (Klausmeier and Litchman 2001), differential predation (Elser and Carpenter 1988), and regimes of turbulent mixing propagated by wind or wave action (Kamykowski 1995). In much the same way, cell motility is also widely considered to be an important influence on distribution; however, to date and due to its complexity, the full ecological significance of active behavioral response still remains uncertai...