Complete co-flocculation of microalgae and Penicillium induced by sub-inhibitory erythromycin: Boosting biomass, CO2 capture, and protein yield in organic-free cultivation

Microalgae utilize CO₂ to synthesize bioproducts. However, efficient cell harvesting remains a significant challenge. Bio-flocculation presents a potential solution. We observed that Synechocystis co-flocculated with a naturally contaminating fungus in the presence of the antibiotic erythromycin (EM). The fungus was isolated and identified as Penicillium sp. Co-cultivation in organic compound-free medium showed that Penicillium sp. also co-flocculated with Synechococcus, Oscillatoria, and Chlorella under a sub-inhibitory concentration of EM, but not in the absence of EM. The co-cultivation of Penicillium with each of the four algae under EM resulted in a 2.4- to 14.5-fold increase in biomass production compared to axenic algal cultures. The Synechococcus-Penicillium co-culture with 5 µM EM produced co-floc biomass up to 1.9 g/L, equivalent to 0.3 g CO₂ captured/L/day, representing an 11.2-fold increase in CO₂ capture compared to axenic Synechococcus cultivation. Protein content in Chlorella-Penicillium floc reaches up to 34% of the dry weight. Transcriptomic analysis of the Chlorella-Penicillium co-floc under EM revealed potential Chlorella genes associated with the co-floc process. Up-regulated genes included those involved in lipid synthesis, exopolysaccharide (EPS) production, inhibition of phototactic motility, pilin-like protein synthesis, and transporter/secretion systems. In contrast, genes related to junction proteins, motor proteins, and cytoskeletons were down-regulated. At 160 µM EM, Chlorella cells within the Chlorella-Penicillium floc maintained a normal cell color, while axenic Chlorella suspensions exhibited partial chlorosis. Thus, co-floc between Chlorella and Penicillium protected Chlorella against EM-mediated chlorosis. This co-flocculation offers the production of protein-rich algal-fungal co-floc biomass through organic-free liquid cultivation.