Soil translocation is an ecological habitat restoration technique, which consists of moving the entire topsoil from a donor site to a chosen receptor site. We investigated soil chemistry and microbiology changes three growing seasons after the salvage of semi‐ancient woodland soil (0.94 ha) to a nearby receptor pasture due to road widening works (Kent, United Kingdom). We sampled (1) Intact woodland soils adjacent to the area of soils that was translocated to represent the lost donor site; (2) The soil from three growing seasons after it had been translocated; and (3) Grassland soils adjacent to the translocated soil to represent the original receptor site. The intention was to ascertain if shifts in soil chemistry and microbial community composition (phospholipid fatty acid analysis, PLFA) occurred due to soil translocation. PLFA signature biomarkers demonstrated the overall microbial community profile of the translocated and woodland soils were similar; however, salvaged soils had a 40% increase in the arbuscular mycorrhizal fungi bioindicator fatty acid 16:1ω5, a 10% decrease in the Gram‐positive bacterial fatty acids, and increased pH (5.01–5.77) compared to the original donor woodland soil. Microbial community composition (PLFA) correlated well with soil pH across all sites. Considering that pH increases with soil depth in this location, it is likely that mixing of soil horizons during translocation increased the topsoil pH causing changes in the soil microbial communities. We concluded that after three growing seasons, the chemical and microbial properties of the salvaged soil were characteristic of a woodland soil but showed signs of disturbance.