Abstract
Subsurface Antarctic brines represent highly selective habitats shaped by extreme physicochemical conditions. Here, the microbial communities of three hypersaline brines (B2, BCR, BCM) from Boulder Clay, Northern Victoria Land, which differ in salinity, pH, trace elements, and glaciological context, have been characterized. High-throughput sequencing was used to reveal that each brine hosts distinct bacterial and fungal assemblies, shaped by local environmental filters. Despite their proximity and some connectivity, microbial communities showed strong taxonomic differentiation and habitat-associated structuring, with significant divergence in diversity and indicator taxa. B2, the most saline, hosted halotolerant bacteria and many unclassified fungi, indicating strong selection and the presence of under-sampled lineages. BCM, enriched by lithogenic input, was dominated by cold-adapted yeasts, while BCR, affected by glacial meltwater, had higher microbial diversity and was dominated by terrestrial taxa. This is the first study integrating microbial, geochemical, and geomorphological data across interconnected Antarctic brines, highlighting how fine-scale environmental heterogeneity and filtered connectivity shape microbial differentiation.