Purpose: Reduce the climate cost of plastic mulch is an important issue for the sustainable development of dryland. Previous studies suggest plastic mulch has risk to increase nitrous oxide emission, but how this produced by the complex interaction between soil properties and microbial is still an unsolved problem and need to research to develop targeted practice.Methods: To address this question, molecular ecology tools, including real-time quantitative and high-throughput sequencing were employed to explore how plastic mulch induced change on soil environment and then stimulates denitrification. Three mulching treatments were compared, including no mulching, spring and autumn mulching.Results: Plastic mulch induced significant increase in the richness and diversity of nir-containing denitrifiers. The nirS and nirK-gene abundance was the greatest under the autumn mulching, and a more conspicuous change in the community structure and abundance was observed for nirK gene than for nirS denitrifiers. The most differentially abundant taxa in the autumn mulching were Anaeromyxobacter, Terriglobus, Edwardsiella, Catenulispora and Acidobacterium for nirK genes and Cupriavidus for nirS genes. Mulching increased the potential denitrification enzyme activity by 58% and 29% for autumn and spring mulching, respectively, and this effect was mirrored by the abundance of nirK, but not by nirS denitrifiers. The nirK gene was significantly correlated to variations in soil moisture, temperature, and nitrate nitrogen.Conclusion: Plastic mulch stimulates denitrification by inducing higher moisture, temperature and nitrate nitrogen content and structural change in the community of nirK denitrifying bacteria in dryland agroecosystems.