Soil water repellency (SWR) significantly impacts water infiltration and soil health, influencing ecological processes across various habitats. Although many of the mechanisms behind SWR are still unclear, studies have shown that different soil and biological properties influence SWR. Hydrophobic compounds produced by plants and microorganisms can increase SWR, but microorganisms can also reduce SWR by degrading the compounds. While several studies have examined SWR in agricultural soils, fewer studies have focused on natural habitats. This study investigates the relationship between soil properties, prokaryotic communities, and potential SWR in soil samples collected from natural and semi-natural habitats across Denmark. We analysed 1,153 soil samples covering 33 habitat types to examine how well SWR can be explained by the soil’s prokaryotic communities and other selected soil properties. Furthermore, we assessed the degree of SWR in the habitats and identified prokaryotic genera indicating a specific degree of SWR. Our findings highlight the influence of prokaryotic communities on the degree of SWR while confirming the relationship between SWR and carbon content. Using path model analysis, we show that both biotic and abiotic factors contribute significantly to SWR. A model including pH, electrical conductivity (EC), total carbon content (TC), and prokaryotic community composition (β-diversity) could explain ~50% of the variation in SWR, with β-diversity and TC being the most important properties. Furthermore, we reveal distinct variations in SWR across habitat types, which cover a wide range of hydrophobicity, from hydrophilic to strongly hydrophobic. Prokaryotic α-diversity was negatively correlated to the degree of SWR, and we found a clear gradient in β-diversity from the highest to the lowest degree of SWR. The degree of SWR was divided into classes, and we identified 69 genera indicating one or a combination of the SWR classes, which could potentially be used as indicators of the degree of SWR. This research underscores the importance of including the microbial communities in studies examining SWR. In perspective, the observed relations between SWR and soil prokaryotic diversity and community composition also imply SWR could become a key biophysical indicator of soil health.