Understanding of the forces driving the structure of biotic communities has long been an important focus for ecology, with implications for applied and conservation science. To elucidate the factors driving phytoplankton genus richness in the Danish landscape, we analyzed data derived from late-summer samplings in 195 Danish lakes and ponds in a spatially-explicit framework. To account for the uneven sampling of lakes in the monitoring data, we performed 1,000 permutations. A random set of 131 lakes was assembled and a single sample was selected randomly for each lake at each draw and all the analyses were performed on permuted data 1,000 times. The local environment was described by lake water chemistry, lake morphology, land-use in lake catchments, and climate. Analysis of the effects of four groups of environmental factors on the richness of the main groups of phytoplankton revealed contrasting patterns. Lake water chemistry was the strongest predictor of phytoplankton richness for all groups, while lake morphology also had a strong influence on Bacillariophyceae, Cyanobacteria, Dinophyceae, and Euglenophyceae richness. Climate and land-use in catchments contributed only little to the explained variation in phytoplankton richness, although both factors had a significant effect on Bacillariophyceae richness. Notably, total nitrogen played a more important role for phytoplankton richness than total phosphorus. Overall, models accounted for ca. 30% of the variation in genus richness for all phytoplankton combined as well as the main groups separately. Local spatial structure (<30 km) in phytoplankton richness suggested that connectivity among lakes and catchment-scale processes might also influence phytoplankton richness in Danish lakes.