Decomposition of organic matter integrates collective activities of organisms within the soil food web. We compared decomposition of museum board (predominantly cellulose) and balsa wood substrates in 18 sites chosen to represent a completely nested design with two disturbance levels nested within three ecosystems (agriculture, wetland, and forest) and ecosystems nested within three land resource regions (LRR) in North Carolina. Percentage mass remaining and daily rate of mass loss of museum board and balsa wood substrates were analyzed using repeated measures analysis of covariance with soil physical and chemical properties as covariates. At the end of the two-year monitoring period, the percentage of museum board and balsa wood substrates remaining was least in agricultural and wetland and greatest in forest ecosystems. Soil pH influenced the percentage of substrate remaining based on days of incubation, and its effects were greater than electrical conductivity, percentage soil organic matter, and total available soil nitrogen (N). Percentage of substrate remaining (museum board or balsa wood) was correlated negatively with pH for all sites, suggesting that pH should be included as a covariate if measures of decomposition are used as environmental indicators. Overall, rate of decomposition of museum board substrates distinguished between relative levels of disturbance in agricultural and wetland but not forest ecosystems. The rate of balsa wood decomposition distinguished between relative levels of disturbance in wetland but not forest or agricultural sites. Forest soils had consistently lower total N and electrical conductivity, and sometimes lower pH, associated with slower decomposition than disturbed wetlands or agricultural lands. We conclude that for short-term monitoring, measures of decomposition of predominantly cellulose substrates can be used to distinguish between relative levels of disturbance in agricultural and wetland but not forest systems. Differences in decomposition may signal either a change in decomposer community or condition of biotic and abiotic resources at a site.