Adverse Effects of Glyphosate and Microcystin-LR on Fish Health: Evidence from Structural and Functional Impairments in Zebrafish Gills

Glyphosate (GLY) and microcystin-LR (MC-LR) frequently co-occur in natural water bodies. In this study, a subacute exposure test was conducted on zebrafish treated with 3.5 mg/L GLY and 35 μg/L MC-LR, individually and in combination, for 21 d to determine their effects on the gills of zebrafish and their potential mechanisms. The hematoxylin and eosin staining and scanning electron microscopy examination results showed that GLY and MC-LR exposure caused structural damage to gills. Biochemical analysis revealed Na⁺-K⁺-ATPase activity decreased, and the levels of reactive oxygen species, 8-hydroxy-2'-deoxyguanosine, and malondialdehyde increased, inducing oxidative damage to DNA and lipids of gills. Meanwhile, the inflammatory and immune function of the gill was significantly influenced, as evidenced by the alteration of the expression of tumor necrosis factor-α, interleukin-1β, complement 3, and immunoglobulin M. RNA-seq results revealed that GLY and/or MC-LR treatment induced transcriptional changes in the fish gills, which may affect various biological functions, and the lipid metabolism disruption potentially involved in the aforementioned process. Integrating histopathological, biochemical, and transcriptomic analyses, this study revealed that both individual and combined exposures to GLY and MC-LR had adverse effects on zebrafish gills, with combined exposure appears to result in more pronounced adverse effects, potentially compromising fish health.