Optimizing HN-AD bacteria-Chlorella symbiotic system via QS molecules for biogas slurry treatment

Heterotrophic nitrifying-aerobic denitrifying (HN-AD) bacteria- Chlorella symbiotic systems face several limitations, including high energy consumption, weak symbiosis and low pollutant removal efficiency. To address these issues, this study optimized the aeration rate. In addition, three quorum sensing (QS) molecules, namely C4-HSL, C6-HSL and C8-HSL, were introduced based on the structural chain length gradient of signaling molecules. This approach aimed to evaluate their effects on the system's macroscopic performance and to explore the underlying enhancement mechanisms. The results showed that under the optimal aeration rate of 0.6 L·min −1 , C8-HSL significantly enhanced pollutant removal, increasing the average removal efficiencies of NH₄⁺-N, TN, TP and COD by 9.09%, 10.81%, 13.78% and 14.25%, respectively. Highthroughput sequencing revealed that C8-HSL markedly enriched Corynebacterium (48.72%), strengthening bacterial-algal symbiosis and improving nitrogen and phosphorus removal. PICRUSt2 functional prediction indicated that C8-HSL enhanced nitrogen metabolic flux by upregulating key genes involved in denitrification and nitrogen fixation ( nirK and nifDHK ), while also significantly increasing the abundance of the phosphorus metabolism related pat gene. qPCR analysis further confirmed the elevated expression of nirK and nifDHK , validating the reliability of PICRUSt2 predictions. These findings demonstrate that C8-HSL effectively improves the performance of the HN-AD bacteria- Chlorella symbiotic system and provides a theoretical basis for the engineering application of QS molecules.