Extracellular carbonic anhydrase from symbiotic bacteria enhances CO2 fixation and microalgal growth in co-culture systems

Microalgae are efficient biological platforms for CO 2 capture and renewable bioresource production; however, carbon assimilation efficiency remains a major bottleneck. In this study, symbiotic bacteria capable of secreting extracellular carbonic anhydrase (CA) were isolated from natural algal-associated environments to enhance CO 2 hydration and utilization in Chlorella sorokiniana . Among six CA-secreting isolates, three strains exhibited high extracellular CA activities and promoted algal growth without inhibitory effects. Co-cultivation with Bacillus safensis YD-1 and B. licheniformis YD-6 increased biomass productivity by up to 74 % and lipid content by 65–70 % compared with axenic controls, while mitigating total inorganic carbon (TIC) depletion during late-stage cultivation. The co-culture systems maintained extracellular CA activities of 3.6–4.8 U/mL and reduced TIC loss to 8.9–16.0 %, compared with 20.3–30.5 % in controls. Immobilization of YD-1 cells using alginate–calcium beads enabled repeated reuse over three cultivation cycles, retaining > 80 % of CA activity and sustaining consistent growth enhancement. These results demonstrate that immobilized CA-secreting symbiotic bacteria provide a self-regenerating and reusable biological strategy to alleviate carbon limitation, offering a practical and scalable approach for improving CO 2 utilization and biomass production in microalgal cultivation systems.