A new high-resolution spectrofluorimetric probe and an automatic water-quality monitoring station (AWQMS) have been used to record seasonal variations in the spatial distribution of three functional groups of phytoplankton in a Mediterranean water-supply reservoir. In comparison with classical methods, the combined use of these innovative techniques enables development of faster and less laborious spatial distribution surveys, thus favouring higher-frequency and spatially more detailed measurements, and, consequently, a better understanding of phytoplankton dynamics. The results show that the observed variations can be explained by the interaction between the buoyancy properties of the phytoplankton and the mixing characteristics of the reservoir. During the winter, when the lake was isothermal and the phytoplankton was dominated by diatoms, there was no significant spatial variation. In the spring, when the phytoplankton was dominated by chlorophytes there was also very little variation but some motile species formed patches when the wind speed was low. The most pronounced nonuniform distributions of phytoplankton were observed during the summer when the phytoplankton community was dominated by positively buoyant cyanobacteria. Then there was a very strong link between the vertical and horizontal gradients which were also related to the prevailing meteorological conditions.