Glitter particles are increasingly recognized as a potential source of microplastic contamination in aquatic environments, yet their effects on freshwater invertebrates remain poorly investigated. This study assessed the accumulation and biological effects of pristine, non-conditioned glitter particles (NCG) and environmentally conditioned glitter particles (CG) in the red swamp crayfish Procambarus clarkii. Crayfish were exposed under laboratory conditions to three glitter concentrations, 0.067, 0.134, and 0.268 g/L, alongside a control treatment. Since no significant concentration-dependent responses were detected, subsequent analyses focused on glitter type and individual glitter burden in the gastrointestinal tract.

Both NCG- and CG-exposed groups showed markedly higher gastrointestinal burdens than controls. Oxidative stress biomarkers and elemental analyses in the hepatopancreas revealed significant treatment-related differences in glutathione peroxidase (GPx) activity and in several elements, including Cr, Cs, Ga, Si, Tl, and Zn. Moreover, glitter burden was positively correlated with glutathione reductase (GR), GPx, carboxylesterase activity measured using pNPB as substrate (CE-pNPB), and N-acetyl-β-D-glucosaminidase (NAG), indicating activation of antioxidant, detoxification, and lysosomal pathways with increasing particle accumulation.

Multivariate analyses provided stronger evidence of treatment effects than individual biomarkers alone. Principal component analysis highlighted treatment-related shifts in integrated biochemical and elemental profiles, while PERMANOVA demonstrated a significant effect of treatment on the overall physiological status of crayfish. Integrated Biomarker Response analysis using the IBR-T approach further supported these findings, revealing a higher biological response in crayfish exposed to non-conditioned glitter (IBR-T = 1.88; 5/13 biomarkers modulated) than in those exposed to conditioned glitter (IBR-T = 1.73; 4/13 biomarkers modulated).

Overall, these findings demonstrate that glitter particles can induce measurable physiological alterations in freshwater crayfish and suggest that environmental conditioning may influence both the magnitude and nature of biological responses. Glitter should therefore be considered an emerging contaminant of ecotoxicological concern.