Size-dependent performance and adaptive mechanisms of anammox granular sludge under fluctuating low-temperature conditions

The adaptation mechanisms of anammox granular sludge under fluctuating low-temperature conditions require further elucidation, particularly concerning the interplay between sludge size, microbial community dynamics, and functional gene including nitrogen-removal and cold resistence genes. This study investigated two anammox reactors operated at high-temperature (28.3°C) and fluctuating low-temperature (11–23°C) conditions to elucidate these mechanisms. Results demonstrated that FNA (> 0.5 µg L ^− 1 ), rather than free ammonia (FA), was the primary inhibitor of anammox activity at low temperatures, reducing nitrogen removal efficiency by 16% at 15 ℃. Apparent temperature coefficient ( Q ₁₀ ) ranged from 0.90 to 1.27, indicating typical temperature-dependent bacterial activity. Low temperatures increased EPS structural vulnerability, weakened granular strength and increased small particles (< 0.25 mm) by 32% while decreasing biomass content (VSS/SS to 27.9%). Our findings demonstrated that the system does not merely undergo inhibition but orchestrates a sophisticated adaptive response: larger granules (> 2 mm) serve as stable reservoirs for core anammox bacteria (notably Candidatus Jettenia caeni ), ensuring biomass retention, while smaller granules (< 0.25 mm), generated through low-temperature-induced fragmentation, function as specialized pioneers. These small granules uniquely enrich a suite of cold-adaptation genes, empowering them to directly combat cold stress by modulating membrane fluidity, synthesizing compatible solutes, enhancing antioxidant defense, and fine-tuning cell-to-cell communication, suggesting heightened environmental sensitivity.