Rapid loss of inoculant microorganisms and the slow-growth of autotrophic bacteria pose significant challenges to bio-augmentation technology. Given the low-cost of coal ash and its greater compatibility with nitrifiers, it was utilized as a carrier to maintain bio-augmentation performance and enhance the AAO's nitrogen removal. Results showed the bio-augmented system operated stably, and immobilized coal ash improved the efficiencies of NH-N from 67.6 ± 10.8 % to 94.3 ± 3.0 % and TIN from 63.2 ± 9.2 % to 84.3 ± 4.3 %. An in-depth analysis indicated bio-augmentation altered the microbial community structures and increased the relative abundance of bacterial associated with nitrogen removal, significantly increasing the abundance of Nitrosomonas and Nirospira. In addition, bio-augmentation enhanced the activity of nitrogen removal enzymes, especially amoABC, hao, narGH, nirK, and nosZ transporter genes, according to predicted function profiles. Network analysis between microorganisms and N-metabolic functional genes revealed bio-augmentation improved the complexity of network-structure and the sources of N-metabolic functional genes, thereby enhancing functional stability. Return sludge flow was the most significant factor in regulating the microbial community composition associated with N removal in the bio-augmented AAO system, followed by DO and internal recycle ratio. The aforementioned results obtained from the present study will enable the application of immobilized coal ash in wastewater treatment plants to improve N removal efficiency.