Temporary high-elevation lakes represent vulnerable and unstable environments strongly threatened by tourism, hydrogeological transformations and climate changes. In-depth scientific knowledge on these peculiar habitats is needed, on which to base integrated and sustainable management plans. Freshwater diatoms, thanks to their high diversity and their particular sensitivity to the water chemistry, can be considered powerful ecological indicators, as they are able to reflect environmental changes over time. The aim of the present study was to analyze the diatomic diversity of the Pilato and Palazzo Borghese lakes, two small temporary high-mountain basins, falling in a protected area within the Apennine mountains chain (central Italy). Diatoms data were collected, at the same time as 12 physicochemical parameters, through six microhabitat samplings, from 17 June to 30 August 2019. In both lakes, a total of 111 diatomic species and varieties were identified. The most species-rich genera were Gomphonema, Navicula, and Nitzschia. The Pilato Lake showed a diatomic community dominated by few species, favored by more stable and predictable environmental conditions than the Palazzo Borghese Lake, which hosted a more diversified community, guaranteed by greater spatial and temporal heterogeneity. Both lakes were characterized by the presence of diatomic species typical of good quality waters. The occurrence of numerous aerial species reflected adaptation strategies adopted to colonize environments subjected to extended drought periods. Endangered diatomic species of particular conservational interest were recorded, confirming the need to preserve their habitats.