In an effort to confer resistance to Cotton Seedling Disease, a fungal complex which results in millions of dollars of revenue loss per year ($10 million in Alabama alone) has had a major affect on farmers and their pockets.  Through genetic modification, a synthetic antimicrobial peptide D4E1, which has been shown in vitro and in-plants to have broad spectrum antimicrobial action against many fungal orders, has been transformed into cotton seeds to examine the efficacy of this peptide on the control of Cotton Seedling Disease Complex.  With the introduction of synthetic antimicrobial peptides as a means of conferring disease resistance, it is important that the potential impact on non-target organisms such as beneficial insects, soil bacteria, and fungi, which play a fundamental role in crop residue degradation and in biogeochemical cycles be investigated (Castaldini et al. 2005).   The objective of this study is to evaluate the effect of D4E1 on phosphatase enzyme activity.  Three 150 x 150 ft test plots, over two field seasons,  were arranged in a completely randomized design and were assigned either one of 3 lines of cotton seed transformed with D4E1 (designated 357, 358, and 373),a control line containing a GUS marker gene, or a non-transgenic control consisting of the parent variety . Soil samples were subjected to Phosphatase enzyme assays and there were no differences between the control and the three isogenic lines; although, there were differences that were associated with time.  Work supported by CREATE- IGERT, USDA and Tuskegee University GWCAES.