This work aimed to investigate the activity and diversity of the dissimilatory arsenate-respiring prokaryotes (DARPs) in the paddy soils, and the effects of sulfate on the DARPs-catalyzed dissolution and release of arsenic and iron from paddy soils into aqueous phase. We collected arsenic-rich soils from a farmland region of the Xiantao city, Hubei, China. Microcosm assay was used to detect how DARPs catalyze the reduction, dissolution, and release of arsenic and iron from the soils, and how sulfate affects this microbial reaction. HPLC-ICP-MS technique was used to determine the arsenic and iron species. Quantitative PCR was used to measure the arsenate-respiring reductase gene abundances in the microcosms. We found that there are diverse DARPs in the indigenous microorganisms. Microcosm assays indicated that these DARPs efficiently promoted the mobilization, reduction and release of arsenic from soils under anaerobic condition. Remarkably, when sulfate was added into the reactions, the DARPs-mediated arsenic reduction and release were significantly increased. Agricultural activities may significantly promote arsenic-contamination in groundwater through increasing inputs of sulfate into paddy soils. It should be avoid overuses of sulfate fertilizers.