Abstract: This study utilized glucose as the carbon source, melamine as the nitrogen source, sodium chloride (NaCl) as the porogen, and manganese acetylacetonate as the manganese source to prepare a porous biochar-based manganese single-atom catalyst (MnCN) through a "ball milling-calcination" process. The research focused on optimizing the preparation process, evaluating the efficiency of the catalyst in activating peroxymonosulfate (PMS) for inactivating antibiotic-resistant bacteria (ARB), and elucidating the underlying mechanisms. The results show that the MnCN catalyst achieved optimal performance with a NaCl dosage of 15 g and a manganese acetylacetonate dosage of 75 mg. The prepared MnCN catalyst exhibited a porous structure with a BET specific surface area of 369.13 m²·g^-1, and Mn elements were anchored as single atoms on the carbon matrix. In terms of application performance, the MnCN/PMS system achieved a logarithmic removal efficiency for ARB of -6.45 within 30 minutes of reaction and demonstrated stable and efficient catalytic performance across a pH range of 3 to 9. Moreover, the system exhibited strong tolerance to coexisting anions, highlighting its outstanding anti-interference capability in complex water matrices. Mechanistic analysis reveal that the MnCN/PMS system primarily operates via a non-radical pathway mediated by catalyst-facilitated electron transfer. This pathway not only achieves efficient inactivation of ARB but also significantly suppresses the horizontal transfer of antibiotic resistance genes (ARGs), thereby reducing the risk of resistance gene dissemination. The findings of this study provide a theoretical basis and technical reference for the precise prevention and control of antibiotic resistance pollution in aquatic environments.