Rice paddies and rice-aquaculture co-culture systems are important sources of methane (CH₄) and nitrous oxide (N₂O) to the atmosphere. Due to the growing human demand of aquaculture protein and the rapid development of green agriculture, rice-aquaculture co-culture systems have been increasingly developed from rice paddies in southeast China. However, the strength of CH₄ and N₂O emissions from rice-aquaculture co-culture systems remains poorly quantified, in particular the contribution of these emissions from different functional cultivation areas. Here, a two-year parallel field experiment was conducted to examine the changes in CH₄ and N₂O fluxes following the conversion from rice paddies to rice-crayfish co-culture systems in southeast China. Over the two-year period, annual CH₄ and N₂O fluxes from rice-crayfish co-culture systems averaged 4.59 mg m^-2 h^-1 and 39.50 μg m^-2 h^-1, amounting to 391.9 kg ha⁻¹ and 3.46 kg ha⁻¹, respectively. The conversion from rice paddies to rice-crayfish co-culture systems significantly reduced CH₄ and N₂O emissions by 14% and 31%, respectively. The emission factors of N₂O were estimated to be 1.25–1.61% in the rice-crayfish co-culture system. Relative to rice planted areas, CH₄ and N₂O fluxes from crayfish ditched culture areas were three times higher and 50% lower, which contributed 26% and 5% to the total CH₄ and N₂O emissions from rice-crayfish co-culture systems, respectively. Our results highlight that the conversion of paddy rice monoculture to rice-crayfish co-culture systems can benefit low greenhouse gas emissions and high ecosystem economic profits as a potential option to sustain agricultural development and achieve carbon neutrality.