The implementation of mainstream anaerobic ammonium oxidation (anammox) can facilitate the realization of carbon-neutral wastewater treatment. However, this technology remains challenging owing to the inability to stably provide nitrite. In this study, we developed a novel nitrification and elemental sulfur-based partial autotrophic denitrification/anammox (NS⁰DA) process for mainstream nitrogen removal. The NS⁰DA system consists of a nitrification reactor and a combined elemental sulfur-based denitrification and anammox (S⁰DA) reactor. Each reactor was independently initiated and optimized before being integrated. At mainstream nitrogen levels (48.5 ± 1.7 mg NH₄⁺-N/L) and 25 °C, the NS⁰DA system achieved 89.1 ± 5.7 % total nitrogen (TN) removal efficiency, with an effluent TN concentration of 5.4 ± 2.8 mg N/L. The system exhibited a low N₂O emission factor (0.23 %), significantly lower than other anammox-based systems. The S⁰DA reactor achieved a nitrogen removal rate of 0.53 kg N/(m³·d) with a short hydraulic retention time (2 h). Anammox accounted for 87.3 ± 7.0 % of the TN removal in the S⁰DA reactor. Isotope experiments and kinetic analysis revealed the cooperation between anammox and denitrification for nitrogen removal. Polysulfides formed in the S⁰DA reactor enhanced the utilization rate of elemental sulfur. High-throughput sequencing identified Thiobacillus and Candidatus Brocadia as the dominant genera of sulfur oxidation and anammox, respectively. The nitrogen and sulfur metabolic pathways were further verified through metagenomic analysis. Overall, the NS⁰DA process provides a stable and efficient nitrogen removal process, minimizing oxygen demand, eliminating organic carbon requirements, and reducing N₂O emissions compared to conventional nitrification/denitrification. This approach offers a promising solution for mainstream nitrogen removal in wastewater treatment.