From auto-floating microalgal flocs to granules: Novel approach to cultivating easily harvestable microalgae for...

Microalgae-based wastewater treatment is constrained by the high cost and low efficiency of biomass harvesting. This study validated that a previously developed selective process for auto-floating microalgae is applicable to non-specific inocula and culture media. Using a different inoculum, auto-floating microalgae were successfully enriched while treating aquaculture wastewater under non-sterile conditions. Auto-flotation activity emerged within approximately two months, achieving flotation efficiencies >90% within 5 min at a biomass concentration of ∼0.5 g L^-1. A critical surface hydrophobicity threshold of ∼75%, as measured by the microbial adhesion to hydrocarbons (MATH) assay, was identified for initiating auto-flotation, above which auto-flotation efficiency increased linearly with hydrophobicity. Auto-flotation efficiency was positively correlated with intracellular protein and lipid contents and extracellular polymeric substance (EPS) polysaccharides but negatively correlated with EPS proteins. Subsequently, the loose auto-floating flocs were transformed into stable granules within 23 days under hydrodynamic shear stress ≥ 0.015 Pa. The resulting auto-floating granules exhibited excellent separation performance, with an ultra-low flotation volume index (FVI₃₀) of 12 mL g^-1 and accelerated flotation even at high biomass concentrations (∼2.5 g L^-1). Compared with conventional algal or algal-bacterial granules, the auto-floating microalgal granules formed under substantially lower shear stress and exhibited distinct EPS architecture, characterized by hydrophilic polysaccharides, localized hydrophobic domains, and amphiphilic groups that facilitate bubble-cell adhesion, indicating a fundamentally different granulation mechanism. Granulation also enhanced lipid accumulation to ∼25% of dry biomass. The system consistently achieved >98% nitrogen and >99% phosphorus removal, highlighting the potential of auto-floating microalgal granules for low-cost wastewater treatment and biomass recovery.