Taxonomy and functional profile of microbial communities across the depths of the Alpine Cenote Abyss ice cave

Investigating the geomicrobiology of the cryosphere offers insights into past climate dynamics and the potential impacts of ongoing climate change. Here, we present the characterization of the microbial communities inhabiting ice sediments collected at various depths within the Cenote Abyss cave, located in the Italian Alps. First explored in 1994 following the drainage of an overlying lake, this site harbours one of the largest cave glaciers in the Dolomites. Metabarcoding and metagenomic analyses revealed a dominance of cold-adapted bacterial taxa, primarily Actinomycetota, Bacteroidota, and Pseudomonadota, with functional genes linked to distinct steps of the nitrogen cycle varying by cave depth. From the shallowest to the deepest ice cave zones, microbial communities shifted from nitrogen-fixing bacterial genera, including Parafrigobacterium, Polaromonas, and Pedobacter, to a higher prevalence of nitrifying bacteria such as Nitrospira. Functional metagenomic analyses revealed that genes involved in nitrogen and carbon cycling are broadly distributed across the cave depth zones, with the inner samples displaying the highest potential for nitrogen transformations, including complete denitrification pathways. CO₂ fixation pathways, including the Calvin–Benson–Bassham and Wood–Ljungdahl cycles, were partially represented and taxonomically diverse across the cave depths. Culturable bacterial strains from all depths demonstrated enzymatic activities relevant to organic matter degradation, while phenotype microarray analysis highlighted the metabolic versatility of the inner microbial community in utilizing organic nitrogen substrates, supporting the higher diversity of the inner cave zone compared to the outer cave zone. These findings underscore the ecological complexity and functional potential of microbial life in subterranean ice, offering insights into biogeochemical processes in cold and nutrient-poor environments with implications for climate change studies.