Genetic and Metabolic Diversity of Cyanobacteria on the Rock-Water Interface in Mountainous Ecosystems

Tintenstrich communities are specialized lithic biofilms dominated by free-living cyanobacteria, also occurring in lichen associations, forming a unique ecological interface between rock environments and aquatic habitats in mountainous areas. To better understand their composition and genetic and metabolic potential, we analyzed 207 samples from the Swiss Alps and Jura Mountains. We determined how key environmental factors shaped cyanobacterial abundance, assessed whether these communities harbor genes for toxin biosynthesis, characterized their taxonomic composition at the family and genus level, and evaluated the actual occurrence of cyanotoxins and other bioactive metabolites. Cyanobacterial abundance proved to be influenced by factors such as elevation, exposure, and their interaction with siliceous rock substrata. Targeted PCR and Sanger sequencing revealed the presence of toxin-encoding genes, particularly for ndaF/mcyE fragments, which may encode microcystin and/or nodularin biosynthesis, while specific genes for microcystins, anatoxins, and cylindrospermopsins were rather rare. Metabarcoding analysis identified 11 cyanobacterial families, with Chroococcaceae, Nostocaceae, and Leptolyngbyaceae being the most abundant. Complementary high-resolution mass spectrometry confirmed the occasional presence of nodularins and microcystins, alongside more frequent detection of other bioactive peptides such as anabaenopeptins and cyanopeptolins. Overall, these findings provide the most comprehensive insight to date into Tintenstrich-associated cyanobacteria, underscoring their environmental significance given their genetic and metabolic potential.