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Soil enzyme activity and stoichiometry in secondary grasslands along a climatic gradient of subtropical China. The Science of the total environment Soil extracellular enzymes plays key roles in ecosystem carbon (C), nitrogen (N), and phosphorus (P) cycling, and are very sensitive to climatic, plant, and edaphic factors. However, the interactive effects of these factors on soil enzyme activities at large spatial scales remain unclear. Here, we investigated the spatial pattern of the activities of five soil hydrolyzing enzymes [β-D-cellobiohydrolase (CB), β-1,4-glucosidase (BG), β-1,4-N-acetyl-glucosaminidase (NAG), L-leucine aminopeptidase (LAP), and acid phosphatase (AP)], and their C:N:P acquisition ratios in relation to plant inputs and edaphic properties across a 600-km climatic gradient in secondary grasslands of subtropical China. The activities of CB, BG, and NAG decreased while that of LAP increased with the increasing mean annual temperature (MAT). The activities of all enzymes did not significantly vary with the mean annual precipitation (MAP). We found that the activities of BG, NAG, and AP were predominately dependent on plant N contents, while the soil LAP activity was tightly related to soil recalcitrant C and N contents. In contrast, the ecoenzymatic C:nutrient (N and P) acquisition ratios increased with increasing MAP and decreasing MAT, primarily due to the increase in plant input at warmer and wetter sites. In addition to climates, plant C inputs, C use efficiency, soil pH, soil organic C, soil C:P, and N:P ratios explained 79% and 72% of the overall variation in ecoenzymatic C:nutrient and P:N acquisition ratios, respectively. The pattern of ecoenzymatic C:N:P acquisition ratios also revealed unexpected N limitation in subtropical grasslands. Overall, our study highlighted the importance of climate in controlling soil biological C, N, and P acquisition activities through its direct and indirect effects on plant inputs and soil edaphic factors, thereby providing useful information for better understanding and predictions of soil C and nutrient cycling in grassland ecosystems at regional scales. 10.1016/j.scitotenv.2022.154019