Aquatic viruses have been extensively studied over the past decade, yet aspects of virus communities in cave waters remain poorly described. Our goal was to characterize viromes of cave water sampled in oligotrophic environments where Proteus anguinus, also known as olm or European cave salamander is present. Due to their dependence on in many cases sensitive water habitats, amphibian species are vulnerable to a variety of threats including viral infections. Water samples (5 litres) form 7 different locations of underground cave system in Slovenia were first concentrated using CIM monolithic chromatography, a method that can efficiently concentrate viruses from high-volume water samples. Then, we used shotgun high-throughput sequencing followed by direct similarity search of sequencing reads against comprehensive database on protein level and subsequent taxonomic classification and visualization. Reads classifying as Caudovirales bacteriophages were most abundant in all cave water samples. Nucleocytoplasmic large DNA viruses from Asfarviridae, Iridoviridae, Mimiviridae, Phycodnaviridae families were also abundantly detected together with virophages (Lavidaviridae) that require a coinfection with giant DNA virus. ssDNA phages from Inoviridae and Microviridae family were detected as well as sequences of eukaryotic circular Rep-encoding single-strand (CRESS) DNA viruses. Sequences of ssRNA plant infecting viruses were abundantly present in some cave water samples, part of them possibly reflecting antropogenic contaminations. Targeted detection of ranaviruses (from the family Iridoviridae), the main viral threat to amphibian diversity, showed negative results using qPCR and these viruses were also not detected in metagenomics analysis of cave water samples. Overall, our findings provide insight into cave water viromes describing common virus community of karstic underground caves system and identifying specific differences in pathogens and viral indicators detected in different sampling sites.
