Abstract
Karst freshwater systems represent unique ecological niches where physicochemical and biological interactions promote intensive calcium carbonate precipitation and the formation of tufa barriers. Here, we studied the composition, diversity, and functional potential of prokaryotic and microeukaryotic communities involved in early-stage tufa formation at two sites within the Plitvice Lakes National Park, Croatia. Over a 5-day period in two different seasons, tufa and water samples were collected at Prošćansko Lake and Novakovića Brod to examine temporal and spatial microbial dynamics. High-throughput sequencing revealed early stabilization of prokaryotic communities within tufa biofilms, dominated by genera, such as Bacillus, Delftia, Hyphomicrobium, and Methylobacterium-Methylorubrum, which are linked to carbonate precipitation processes. With biofilm maturation, shifts toward Acinetobacter and Rhodoferax indicated increasing heterotrophic activity and organic matter degradation. In contrast, microeukaryotic communities were more variable, with diatoms and Zygnemophyceae contributing to extracellular polymeric substance production, crucial for carbonate entrapment. Site-specific patterns reflected environmental influences, such as hydrodynamics and terrestrial organic input. The results underscore the importance of microbial succession and community specialization in the formation and stability of tufa barriers, offering new insights into microbial contributions to biogeochemical processes in karst freshwater systems.