Co-exposure to zinc pyrithione and polystyrene nanoplastics alters developmental processes, DNA methylation, and immunogenic pathways in zebrafish larvae.

Marine aquaculture uses a wide variety of chemicals that go directly out into waterbodies, but little is known about the combined impact of these substances. As much of salmon aquaculture is situated in fjords where early development occurs for fish and other aquatic organisms, we have used zebrafish larvae as a model organism to assess the impacts of the combined exposure of a common antifoulant (ZnPT) and nanoplastics (NPs). Embryos were exposed to a control, 100 nm NPs (50 mg/L), 0.002 μM ZnPT (635.4 ng/L), and their combination. Zebrafish were studied from fertilization to 116 hpf, and developmental processes, early embryonic movement, heart rate, oxygen saturation, and swimming behavior, as well as DNA methylation and RNA transcriptome, were included as endpoints. We observed higher toxicity in larvae exposed to a combined treatment of ZnPT+NPs, with significant decreases in eye size and body length at 72 hpf, significantly increased burst activity at 24 hpf, significantly decreased oxygen saturation, hypomethylation of 5mC%, and 3054 significantly differentially expressed genes. These differentially expressed genes were mainly associated with immunogenic pathways, such as leukocyte (white blood cell) activation and proliferation, interleukin-1 production, neutrophil migration, interleukin-17 (antimicrobial proteins), cytokine and chemokine-mediated pathways, and C-type lectin receptors. At 72 hpf, co-exposure resulted in significantly stronger effects on developmental parameters (body length and eye size) and behavioral responses (burst activity). By 116 hpf, immunotoxic outcomes were substantially more pronounced in the co-exposure group, while impacts on global methylation levels had decreased in the co-exposure. Overall, these findings demonstrate that simultaneous exposure to ZnPT and NPs produce significantly stronger biological impacts than exposure to either compound alone.