Mineral surfaces in soil are an important interface for organic matter (OM) and nutrient cycling, with associated microorganisms contributing to the formation and turnover of mineral-associated OM (MAOM). However, the relevance of intrinsic (mineral type) versus extrinsic (land-use intensity) factors on the co-development of MAOM and microorganisms under natural conditions remains poorly understood. Mineral containers filled with mixtures of quartz-sand and pristine secondary minerals (goethite or illite) were exposed to 50 grassland topsoils of the Schwäbische Alb (Germany) along a land-use intensity gradient for five years. Mineral samples and soils were analyzed for organic carbon (OC) and nutrients (N and P), abundance and composition of major microbial groups based on phospholipid fatty acid profiles, as well as enzyme activities (β-glucosidase, β-xylosidase, N-acetylglucosaminidase, and acid phosphatase). Microorganisms colonized both mineral samples to the same extent, with goethite samples exhibiting greater MAOM accumulation and higher enzyme activities than illite samples. Both mineral samples differed from the overlying soils with greater relative abundances of fungi and Gram-negative bacteria and greater microbial acquisition of nutrients (N and P) relative to C as indicated by the stoichiometry of enzyme activities. Increasing land-use intensity was associated with decreasing C:N ratios and microbial abundances for goethite samples and increasing β-glucosidase activity for illite samples while the proportion of fungi was reduced in both mineral samples. We conclude that in the studied temperate grasslands the association of OM and microorganisms with secondary minerals is driven more by mineral type and reactivity than by differences in land-use intensity. The different minerals apparently formed distinct microhabitats with unique characteristics that differed in MAOM accumulation and microbial access to OC and nutrients, thus affecting microbial colonization and functionality. • Pristine minerals were buried in grasslands of varying land-use intensity to study co-development of MAOM and microbes. • Mineral type (goethite, illite) affected OC-normalized microbial biomass, fungi to bacteria ratios and enzyme activities. • Minerals were preferentially colonized by fungi and Gram-negative bacteria. • Mineral-associated microbes invested relatively more into nutrient than C acquisition. • Land-use intensity and surrounding soil properties were less important for microbial colonization than mineral type. [ABSTRACT FROM AUTHOR]