Light-enhanced surface film formation in Euglena gracilis is associated with betaine-, phenylalanine-, and arginine-enriched EPS

Euglena blooms in freshwater and brackish habitats often form surface films that pose ecological risks. Although phototaxis is recognized as a key behavioral response, how light-associated physiological changes relate to cell aggregation and surface film development remains poorly understood. Here, Euglena gracilis was used as a model organism to investigate the role of light in surface film formation, with a focus on extracellular polymeric substances (EPS) and associated metabolic profiles. Among the tested irradiance levels under controlled conditions, illumination at 35 μmol·m −2 ·s −1 resulted in the highest degree of surface aggregation, yielding a film-forming rate of 81.51% and a stable film area of 0.075 mm 2 under high cell density (5.23 × 10 8 cells·L −1 ). Combined chemical and spectroscopic analyses revealed significantly increased nitrogen, phosphorus, and protein contents in the bound EPS (BEPS) under illumination. Non-targeted metabolomics identified betaine, L-phenylalanine, and L-arginine as dominant EPS-associated metabolites, with the latter two showing consistent light-associated enrichment. Collectively, these findings suggest that illumination-induced alterations in EPS composition and metabolite profiles contributed to enhanced cell aggregation and surface film development. This study provides insights into the photophysiological mechanisms underlying light-induced surface film formation and EPS-mediated aggregation in Euglena .