We assessed microbial community composition as a function of altered above- and belowground inputs to soil in forest ecosystems of Oregon, Pennsylvania, and Hungary as part of a larger Detritus Input and Removal Treatment (DIRT) experiment. DIRT plots, which include root trenching, aboveground litter exclusion, and doubling of litter inputs, have been established in forested ecosystems in the US and Europe that vary with respect to dominant tree species, soil C content, N deposition rate, and soil type. This study used phospholipid fatty-acid (PLFA) analysis to examine changes in the soil microbial community size and composition in the mineral soil (0-10 cm) as a result of the DIRT treatments. At all sites, the PLFA profiles from the plots without roots were significantly different from all other treatments. PLFA analysis showed that the rootless plots generally contained larger quantities of actinomycete biomarkers and lower amounts of fungal biomarkers. At one of the sites in an old-growth coniferous forest, seasonal changes in PLFA profiles were also examined. Seasonal differences in soil microbial community composition were greater than treatment differences. Throughout the year, treatments without roots continued to have a different microbial community composition than the treatments with roots, although the specific PLFA biomarkers responsible for these differences varied by season. These data provide direct evidence that root C inputs exert a large control on microbial community composition in the three forested ecosystems studied.