Fullerene (C60) Supplementation Stimulated Anammox Granules at Low Doses and Inhibition Occurred at High Doses: Phenomenon and Mechanism

Carbon-based nanomaterials are extensively utilized as additives to enhance microbial activity in pollution remediation. A significant challenge to the widespread adoption of the full-scale anaerobic ammonia oxidation (anammox) process in wastewater treatment is the limited activity of anammox bacteria. Thus, this study investigates the effects of supplementation of fullerene (C60), a carbon-based hollow spherical molecule, on anammox granules, aiming to enhance the efficiency of wastewater treatment by stimulating anammox bacterial activity. The optimum dose of C60 was 25 mg of C60/L of the reactor working volume, which resulted in increasing anammox activity by 22% and the abundance of Candidatus Brocadia from 12 to 20%. The main mechanism for enhanced anammox activity was proposed to be associated with the accumulation of c-di-GMP signal molecules, which was driven by increased c-di-GMP synthesis protein (diguanylate cyclase) and decreased c-di-GMP degradation protein (specific phosphodiesterase) at 25 mg C60/L addition. The protein abundances responsible for the assembly, biogenesis, and stability of pilus were also increased, which might improve extracellular electron transport and accelerate the enzymatic reactions during the anammox process. However, higher C60 dosages (50–100 mg C60/L) inhibited anammox activity and abundance. The economic evaluation revealed a substantial increase in net present value (NPV) from $2,670 to $7,110 with the addition of 25 mg C60/L, emphasizing the economic viability of C60 supplementation for enhanced nitrogen removal in anammox-based wastewater treatment systems. Overall, this study proposes a novel and economically viable approach by supplementing C60 at 25 C60 mg/L (i.e., 3.5 mg C60/gVSS) to stimulate the activity of anammox bacteria, offering promising applications for real wastewater treatment plants.