Taxonomic and functional analysis of bacterial metagenomes and metagenome-assembled genomes from an Alpine Biosphere Reserve

Alpine lakes, small water bodies, ponds and peat bogs are unique and ecologically significant freshwater habitats. These habitats play crucial roles in supporting regional biodiversity and biogeochemical cycles and provide essential ecosystem services, including carbon storage, decomposition of organic matter, and water filtration. Importantly, they support diverse microbial communities and often harbor rare and endangered species. Understanding the composition and functional potential of the bacterial communities residing in freshwater and sediment habitats of highly biodiverse areas is central for assessing and benchmarking ecosystem health and resilience, especially under ongoing climate change. The microbial biodiversity of the freshwater habitats in the ‘Alpi Ledrensi and Judicaria’ Biosphere Reserve has been previously described by Vettorazzo et al. (2024) utilizing metabarcoding analysis based on the V3-V4 regions of the 16S rRNA gene. Here, we employ full-shotgun metagenomics sequencing to i) gain increased taxonomic resolution of the bacterial communities of these habitats, ii) describe their ecological role by looking at their genomic functional potential and iii) compile a catalog of metagenome-assembled genomes (MAGs) to characterize rare bacterial species further. We confirm the high variability of the microbial communities along the physiographic gradient of the water bodies, with a marked difference in bacterial richness and composition between freshwater and sediment habitats. Moreover, by investigating the presence of genes related to the main biogeochemical metabolic pathways, we link community structure to ecosystem functions. A total of 485 high-quality bacterial MAGs were assembled from the sampled sites, of which 72% of species are novel (currently absent in curated bacterial databases such as GTDB, RS220). Most unknown species belonged to the Phyla Cyanobacteria (10 MAGs), Patescibacteria (23 MAGs), Verrucomicrobiota (25 MAGs), Bacteroidota (57 MAGs), Actinomycetota (75 MAGs) and Pseudomonadota (121 MAGs). Amongst the isolated MAGs, 76% were reconstructed from water habitats, while only 23% were from sediment habitats, reflecting the technical difficulties still present when assembling genomes de novo from highly biodiverse soil samples. The MAGs uncovered from these understudied environments contribute to widening the set of environmental reference genomes, thereby increasing the mappability of environmental metagenomes in future studies.