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Information on the contribution of various soil nitrogen (N) sources to plant N uptake is often needed for the implementation of sustainable or site-specific management practices in agriculture. Considering the limitations of traditional methods in meeting these needs, this study investigated the potential of leaf δ15N as an early indicator of nutrient deficiency in cotton. The spatial and temporal natural abundance of 15N was measured in the soil and leaves of a fertilized cotton field located near the village of Moschochori (Larissa, Greece). The isotopic signal of the leaves was interpreted in the context of the relative contribution of fertilizer to cotton N uptake,as has been demonstrated in the past for other agricultural crops such as wheat (Triticum aestivam L.) and corn (Zea mays). Spatial variability of leaf δ15N was high early in the growing season (June), reflecting differences in fertilizer N availability and uptake between the east and west side of the field, as well as differences resulting from soil denitrification in depressions. The west side of the field appears to have lost significant amounts of fertilizer N, due to leaching during the rainy period in May, that accumulated in depressions near thewaterway. In the subsequent months, the isotopic signal of the leaves was consistently high and indicated reduced fertilizer N uptake on the west side that resulted in deficiencies of N as well as of phosphorus (P) and potassium (K). The significant correlations of mid-square leaf δ15N with late-season nutrient content and soil elelctrical conductivity(EC) provided evidence that the natural abundance of 15N was a sensitive indicator of soil and plant nutrient status in this fertilized cotton field.