Packing media regulate nitric oxide removal performance by driving community assembly and biofilm evolution in biotrickling filters.

Packing media are essential for optimizing biotrickling filter (BTF) design and pollutant removal efficiency, yet how their integrated physicochemical properties affect the performance by regulating community assembly and biofilm evolution remains unclear. This study compared two parallel BTFs packed with activated carbon (AC) granules (BTFAC) and polyurethane (PU) sponge (BTFPU) for nitric oxide (NO) removal, and tested their resilience to starvation. BTFPU achieved significantly higher and more stable NO removal than BTFAC during both pre-starvation (99% ± 1% over 94% ± 5%) and post-starvation periods (98% ± 1% over 82% ± 9%, P < 0.001). For PU, despite its hydrophobic surface, the large-pore spongy structure (pore sizes of 603 ± 40 μm) and high porosity (97% ± 1%) provided greater protected internal space and lower superficial gas velocity (6 m/h), collectively created a less stressed microenvironment for microbes. This supported the fast evolution of thicker and more viable biofilms with higher microbial diversity, cooperative interactions, and stochastic assembly, facilitating the fast recovery post-starvation. In contrast, for AC, the hydrophobic surface, surface functional groups, higher gas velocity (9 m/h) from lower porosity (62% ± 2%), and small pore size (3.2 ± 0.4 nm) restricted microbial growth to the exposed external surface and imposed stronger selection pressure. These factors collectively drove thinner, less viable, and slower biofilm evolution with lower diversity, reduced proportion of denitrifiers, stronger deterministic assembly, which ultimately compromised the performance stability. These findings reveal how packing media regulate BTF performance by driving community assembly and biofilm evolution.