Applications of nanomaterials, including carbon nanotubes (CNTs), are increasing; however, their impact on the environment is still not well understood. A semi-arid soil was treated with multi-walled carbon nanotubes (MWCNTs) at four different concentrations (10-10000 mgMWCNTs kg^-1soil), and incubated with a control (soil only) for 90 d. Soil-basal respiration, enzymatic activities and microbial community structure were evaluated. There was no significant difference in soil basal respiration among the different treatments. Acid phosphatase activities were affected after 28 d, but no significant effects were observed on β-glucosidase and β-glucosaminidase activities. Application of 10000 mgMWCNTs kg^-1soil lowered acid phosphatase and β-glucosidase activity by 25% and 7% after 28 d. However, β-glucosidase and β-glucosamidase activities were increased by 22% and 15 % in the 10000 mgMWCNTs kg^-1soil after 90 d. No significant effects were observed in the acid phosphatase activities in 10000 mgMWCNTs kg^-1soil after 90 d. Dehydrogenase activities were not affected in 10-, 100- and 1000 mgMWCNTs kg^-1soil after both 28 and 90 d. However, the activity was lowered by more than 8 folds in the 10000 mgMWCNTs kg^-1soil. Separate tests demonstrated an interference of the MWCNTs (≥10000 mg kg^-1soil) with the measurement of dehydrogenase activity by affecting absorbance of a color product. Total microbial and fungal biomass, determined with the sum of fatty acids methyl esters (FAMEs), were increased by 27% and 44% in the 10000 mgMWCNTs kg^-1soil after 90 d. Principal component analysis of FAME profiles for both 28 and 90 d samples showed separation of 10000 mgMWCNTs  kg^-1soil from the control along principal component 1 (89% and 90% of variance). This study suggests the presence of MWCNTs may cause shifts in soil microbial community structure and soil enzyme activities.  However, an extensive study is required with evaluation of other components of soil ecosystems to support the preliminary results.