We selected two sites in South west China with different levels of Sb and As contamination to study interactions among various Sb and As fractions and the soil microbiota, with a focus on the microbial response to metalloid contamination. Comprehensive geochemical analyses and 16S rRNA gene amplicon sequencing demonstrated distinct soil taxonomic inventories depending on Sb and As contamination levels. In addition, metagenomics revealed the potential metabolic pathways of the soil microbial ecosystems.