To contribute to a more thorough documentation and understanding of tropical freshwater diatoms, we studied 82 samples of epilithic diatoms, collected on 11 sampling dates in the different seasons from 2015 to 2016 from Rio Cupeyes and Rio Guilarte (southwest Puerto Rico) subjected to different degrees of anthropogenic impacts. On each sampling date, a primary (riffle) and, usually, a secondary habitat (run in Cupeyes and pool in Guilarte) were sampled. The samples were taken in the frame of the National Ecological Observatory Network (NEON). The detailed data on spatial (and temporal) natural variability will be discussed, in particular in relation to the similarity standards imposed to diatom-based assessments by quality control procedures. A clear species replacement in all the main diatom genera was noted in the two streams in relation to the impact of nutrient enrichment. We will also use available NEON data, such us PAR, dissolved oxygen, quantitative measures of periphyton etc. to start outlining the relevance of epilithic diatoms in overall stream metabolism and their contribution to ecosystem functioning. A more in depth understanding of the structure and function of diatom communities in tropical stream ecosystems is important because they are highly vulnerable to changes in land use and vegetation in their catchments.