Road construction for mineral exploration and extraction in the Little Missouri National Grasslands (LMNG) in southwestern North Dakota has produced thousands of miles of roads and utility corridors. Such roads are required to be restored to pre-construction vegetative biodiversity as described by the US Forest Service as part of the Federal Multiple Use Sustained Yield Act of 1960. Previous research has shown that 25 years after restoration neither vegetation nor soil physical/chemical properties of the roads have returned to conditions of adjacent native prairie. Several studies have found that reductions in SOM and changes in plant community may lead to differences in microbial ecology and related ecosystem services of reclaimed roads. The objective of this study was to characterize the soil microbial communities of roads reclaimed during three different time periods. Soil biological properties were evaluated among 9 roads from 3 different reclamation periods to assess the effect of time since reclamation on microbial community structure and function. Soil microbial community structure, microbial biomass, using phospholipid fatty acidy analysis, and enzyme profiles were measured on the roads and compared to values of the adjacent native prairie. Soil biological properties were evaluated on samples collected in close proximity to sites from previous studies to permit inclusion of vegetation and soil physical and chemical data in the analysis. Initial enzyme data indicates that roads’ enzyme activity have not returned to that of the adjacent native prairie substantiating findings of previous studies on these sites.