The cytotoxicity of the haptophyte Chrysochromulina leadbeateri towards the Atlantic salmon gill cell line ASG-10

Abstract

Fish-killing harmful algal blooms (HABs) threaten fisheries and aquaculture globally. In May and June 2019, the haptophyte Chrysochromulina leadbeateri formed the most devastating fish-killing algal bloom ever recorded in Northern Norway. Despite the severe impact of HABs on farmed fish, our understanding of their toxic mechanisms is limited. Here, we investigated the ichthyotoxicity of C. leadbeateri by using the gill epithelial cell line ASG-10 from Atlantic salmon. Extracts of C. leadbeateri showed potent hemolytic activity towards Atlantic salmon red blood cells, and toxicity to the ASG-10 cells with an EC₅₀ of 3.41 × 10⁶ algal cells mL⁻¹ measured by the Alamar Blue viability assay. Furthermore, lower extract concentrations increased caspase-3/7 activity, suggesting induction of apoptosis, while higher concentrations of the extract showed compromised cell membrane integrity. No intracellular reactive oxygen species (ROS) induction was observed. Exposure to C. leadbeateri extracts cultured under different irradiances or with putrescine supplementation to the algal medium did not increase the toxicity. Exposure of living C. leadbeateri cells to ASG-10 did not show any cytotoxicity from imaging or barrier integrity impairment, but the activity of leaked dead-cell proteases revealed a concentration-dependent effect on the ASG-10 cells. Furthermore, incubation of C. leadbeateri in culture with putrescine for 72 h resulted in increased dead-cell proteases leaked from the ASG-10 cells compared to cultures without putrescine. Our findings suggest that living C. leadbeateri cells exert sublethal effects on Atlantic salmon gill cells. Additionally, putrescine seems to increase the ichthyotoxicity of living C. leadbeateri towards ASG-10 cells.