<p>Cryoconite holes (water-filled reservoirs) on glacier surfaces are important biodiversity hotspots and biogeochemical factories within terrestrial cryosphere. In this study, we collected cryoconite from the ablation zone of the Forni Glacier (Central Italian Alps) over the whole ablation season. We aimed to describe spatial and temporal patterns in: (i) biomass and community structure of photoautotrophs (cyanobacteria, diatoms, and eukaryotic green algae) and invertebrates; (ii) carbon and nitrogen stable isotopic composition of invertebrates and their potential food; and (iii) the organic matter content and general elemental composition of cryoconite. Structure and biomass of cryoconite biota showed spatio-temporal changes over the season. Dominant cyanobacteria were Oscillatoriaceae and Leptolyngbyaceae, while dominant eukaryotic green algae were Mesotaeniaceae and Chlorellaceae. Eukaryotic green algae dominated in the upper part of the ablation zone, while a seasonal shift from algae- to cyanobacteria-dominated communities was observed in the lower part. Some taxa of photoautotrophs appeared only during specific sampling days. Dominant grazers were tardigrades (<em>Cryobiotus klebelsberg</em>i). The biomass of tardigrades in the upper part was significantly related to the biomass of eukaryotic green algae indicating that algal communities are likely controlled by grazing. The δ<sup>13</sup>C of tardigrades followed fluctuations of δ<sup>13</sup>C in organic matter. We did not observe spatial and temporal changes in the general elemental composition of cryoconite. Thus, changes in community structure and biomass are likely dependent on the interplay between phenology, stochastic events (e.g. rainfall), top-down, or bottom-up controls. Our study shows that understanding the ecology of biota in cryoconite holes requires a spatially explicit and seasonal approach.</p>