Seed harvesting ants (Pogonomyrmex rugosus) concentrate organic matter and nutrients near their nests and create biogeochemical hotspots in desert soil. We examined factors regulating denitrification and soil respiration in a Mojave Desert ecosystem to determine the role harvester ant colonies play in nitrogen loss and carbon mineralization. Organic matter and nutrient storage were significantly greater in colonies than under the dominant vegetation (i.e., Pleuraphis rigida, a bunch grass) and in bare soil, with standing stocks of inorganic nitrogen in colonies nearly 4-fold greater than in the other microhabitats. Soil respiration, measured with laboratory incubations, was below detection limits under ambient soil conditions. Respiration rate in soil from bare patches and under grass was limited by water and labile organic carbon, with secondary nitrate limitation evident only once carbon limitation was alleviated. Soil respiration in ant nest soil was limited by water and labile organic carbon only. Denitrification, measured by the acetylene block technique, was elevated in bare soil and under grass with the addition of nitrate. Ant nest soil also responded to a nitrate addition; however, denitrification rate was greatest with addition of glucose. Ordinarily desert soil has low rates of respiration and denitrification due to dry ambient conditions. However, ant colonies likely function as important sites for nitrogen loss and carbon mineralization following rain storms, especially if storms coincide with seed set and the temporally pulsed input of organic matter into colonies.