Brine Beneath: The hidden microbial world of Lüneburg's salt

Environments with extremely high salt concentrations serve as analogue sites for astrobiological research. Despite their seemingly inhospitable and harsh conditions, these environments support a diverse array of extremophilic organisms. The saturated Lüneburg brine forms at a depth of 60 m below the surface through dissolution of Zechstein halite. It has been used commercially for salt extraction for over 1000 years until it was closed in 1980 and although it remains easily accessible for sampling, it had not been investigated before. Studying extremophiles from these saline habitats is important, as it offers valuable insights into a hypersaline habitat which cannot benefit from primary production through sunlight and resembles the boundaries of life on Earth and the possibility of life beyond our planet. This work provides a habitat description of the Lüneburg brine, using hydrochemical, cultivation-dependent and independent methods.

The analyses included sequencing of variable regions V1-V2 and V3-V4 of the 16S rRNA gene, isolation of culturable aerobic microorganisms, enrichment of a heterotrophic and an autotrophic anaerobic microbial community as well as chemical profiling of the brine. Selected microbial isolates were tested for their resistance to astrobiological-relevant stressors. The 16S rRNA amplicon analysis identified the presence of halophilic organisms: Desulfohalobiaceae (18%), Thiohalorhabdaceae (11%) and three families from the order Halobacterales (16%). A high abundance of yet-uncultivated Nanosalinaceae (15%) were detected and recognized as promising candidates for pure culture isolation. Anaerobic enrichments demonstrated the presence of Nanosalinaceae, contingent upon substantial co-enrichment of Halobacteriaceae. Microscopic analysis indicated notable morphological diversity, with isolates displaying pleomorphism, encapsulation and biofilm production during growth. The isolates displayed tolerance towards high radiation (>500 Gy) and NaCl concentrations up to 30%, as well as enhanced growth in typically toxic perchlorates, with concentrations that are also present on Mars.

The Lüneburg brine represents an aphotic habitat and thus a representative subsurface analog for other planets in outer space. It hosts a range of halophilic microorganisms capable of tolerating high radiation and salt concentrations. This study highlights the potential of this accessible habitat for astrobiological research.