This study presents a pilot-scale duckweed-based wastewater treatment system enhanced with CO2 fertilization to improve nutrient removal, CO2 capture, and biomass productivity. Three strains of Landoltia punctata were cultivated continuously for five months. In spring, CO2 supplementation significantly increased the biomass yield and starch and protein productivity (by more than 12.2, 2.4 and 3.5 g/m2/d, respectively), representing increases of 189–259 %, 1,108–2,330 % and 143–166 %, respectively, compared with those of the control group. The removal rates of total nitrogen (TN) and total phosphorus (TP) increased by 28–71 % and 120–148 %, respectively, whereas CO2 capture improved by 196–264 %. Enhanced nutrient recovery was also observed, with N recovery increasing from 0.20–0.25 to 0.54–0.61 g/m2/d and P from approximately 0.05 to 0.11 g/m2/d. Enzyme activity and transcriptome analyses of strain 0202, which presented the highest starch content, revealed that CO2 fertilization upregulated key starch biosynthesis genes (ADP–glucose pyrophosphorylase (AGPase), Granule-bound starch synthase (GBSS), and Soluble Starch Synthase (SSS)), contributing to increased starch accumulation. It also promoted glutamine synthetase expression, increasing nitrogen assimilation and shifting nitrogen removal from microbial nitrification to duckweed uptake. As a result, microbial nitrification decreased significantly, and the proportion of NH4+-N removed via plant assimilation increased to over 69 %. These findings demonstrate that coupling CO2 fertilization with duckweed cultivation is an effective and scalable approach for wastewater treatment, carbon fixation, and renewable bioresource production.