Hydrochemical gradients driving extremophile distribution in saline and brine groundwater of southern Poland

Extreme environments, such as highly saline ecosystems, are characterised by a limited presence of microbial communities capable of tolerating and thriving under these conditions. To better understand the limits of life and its chemical and microbiological drivers, highly saline and brine groundwaters of Na-Cl and Na-Ca-Cl types with notably diverse SO₄ contents were sampled in water intakes and springs from sedimentary aquifers located in the Outer Carpathians and the Carpathian Foredeep basin and its basement in Poland. Chemical and microbiological methods were used to identify the composition of groundwaters, determine microbial diversity, and indicate processes controlling their distribution using multivariate statistical analyses. DNA sequencing targeting V3-V4 and V4-V5 gene regions revealed a predominance of Proteobacteriota, Methanobacteria, Methanomicrobia, and Nanoarchaea in most of the water samples, irrespective of their geological context. Despite the sample-size constraint, redundancy analysis employing a compositional approach to hydrochemical predictors identified Cl/SO₄ and Cl/HCO₃ ratios, and specific electrical conductivity, as key gradients shaping microbial communities, depending on the analysed gene regions. Analysis of functional groups revealed that methanogenesis, sulphate oxidation and reduction, and the nitrogen cycle define and distinguish the halotolerant communities in the samples. These communities are characterised by an inverse relationship between methanogens and sulphur-cycling microorganisms.