Habitat Specialization and Airborne Dispersal Shape the Microbiome of a Gypsum Karst Cave

The Gypsum Karst of Sorbas (Almeria, southeastern Spain) is a gypsiferous massif covering approximately 12 km² and constitutes one of the most extensively studied and internationally recognized gypsum karst systems. So far, only a limited number of studies have addressed microbial diversity in gypsum karst caves, and most of them have focused on a reduced number of habitats or specific cave compartments. Here, we characterize the prokaryotic diversity in biofilms, sediments, water and air in the gypsum cave of Covadura to compare the community composition across different habitats, assess connectivity among terrestrial, aquatic and aerial environments, examine associations with environmental and geochemical parameters, and provide baseline information for gypsum cave ecosystems. For this purpose, 28 samples were collected from different compartments distributed within the cave and above it, and analyzed using high-throughput 16 S rRNA gene sequencing. While dominant phyla, classes and genera resembled those of other karst systems, communities differed strongly among habitats. Water samples hosted distinct aquatic assemblages absent from biofilms, sediments, air and soil, indicating a clear separation between aquatic and non-aquatic cave systems. Biofilms were dominated by Crossiella, members of wb1-P19 and of the Euzebyaceae, which were also present in sediments, but scarce in water samples. Interestingly, Crossiella and wb1-P19 members were also detected in the cave air for the first time, suggesting their release from terrestrial communities and subsequent redistribution as bioaerosols and supporting cave air as a key dispersal vector that links stable terrestrial microbial reservoirs in Covadura Cave. This study reveals strong habitat specificity and high spatial heterogeneity in gypsum cave microbiomes, providing new ecological insights relevant for the conservation and management of cave ecosystems.