Environmental and botanical gradients were associated with soil microbiome composition in managed temperate grasslands

Soil microbial communities play a central role in nutrient cycling and the sustainability of pasture-based agricultural systems. The relative influence of grassland management systems, soil pH, soil organic matter (OM), and botanical composition on soil microbiome structure in temperate grazing systems remains unclear. In this study, shotgun metagenomic sequencing was used to characterise microbial communities within the upper 10 cm of soil collected at a single time point in May 2024 from a managed grassland farm systems experiment comprising perennial ryegrass ( Lolium perenne L.; PRG) monocultures receiving 200 kg N ha⁻¹ yr⁻¹ and perennial ryegrass-white clover ( Trifolium repens L.; WC) swards receiving 150 or 100 kg N ha⁻¹ yr⁻¹. Microbial alpha diversity (richness, Shannon and Simpson indices) and beta diversity showed limited differentiation among grassland management systems at both genus and species levels. In contrast, microbial community composition was significantly associated with soil pH, OM, and WC abundance. Genus-level PERMANOVA identified significant relationships with soil organic matter (OM) (R² = 0.085, adjusted p = 0.004), soil pH (R² = 0.070, adjusted p = 0.028), and WC proportion within the plant community (R² = 0.10, adjusted p = 0.026). Several genera, including Saccharomonospora , Thermopolyspora , and Defluviicoccus , were associated with soil OM, whereas fewer genera were strongly associated with soil pH. Nitrogen cycle-associated gene profiles showed limited differentiation among grassland treatments. These findings demonstrate significant associations between soil microbial community composition and environmental and botanical gradients, with limited differentiation among grassland management systems.