Mixture matters: a probabilistic assessment of cumulative exposure and health risks from multiple toxic elements in Italian mussels and clams

Bivalve mollusks are cornerstone products of European aquaculture, yet their filter-feeding biology promotes the bioaccumulation of potentially toxic elements. While regulated contaminants such as Cd, Pb, and inorganic As (iAs) are subject to routine monitoring, numerous toxicologically relevant elements remain unaddressed by surveillance programs, and traditional single-element risk assessments fail to account for the health implications of dietary co-exposure to elemental mixtures. This study addresses both limitations through the first comprehensive, population-specific probabilistic cumulative risk assessment for multiple toxic elements in mussels (Mytilus galloprovincialis) and clams (Chamelea gallina). A total of 102 mussel and 26 clam batches were collected from the Italian retail network and their multi-elemental profiles were analyzed by inductively coupled plasma mass spectrometry (Ag, B, Ba, Be, Cd, Cr, As, Mn, Mo, Ni, Sb, Se, Sr, U, V, and Zn) and thermal decomposition atomic absorption spectrometry (Hg). Since total As, Cr, and Hg were measured, the species iAs, Cr(III), Cr(VI), and MeHg were estimated to account for 3.5%, 100%, 1%, and 80% of their respective total concentrations. Dietary exposure was assessed for Italian children, adolescents, and adults using probabilistic methods based on Monte Carlo simulations, integrating measured element concentrations with food the Hazard Index (HI) [1], obtained by summing individual Target Hazard Quotients for 18 elements. Cumulative cancer risk was expressed as Total Cancer Risk (TCR) [1], derived from the sum of Incremental Lifetime Cancer Risks for iAs and Cr(VI). The two bivalve species exhibited low contamination levels but distinct elemental profiles: mussels showed higher concentrations of Zn, B, Se, Cd, Pb, and Hg, whereas clams were enriched in Sr, Mn, V, Ni, Ba, and Cr (p < 0.05). Estimated non-cancer risk, mainly driven by iAs, Cd, Se, and Zn (> 96% contribution), was negligible, as HI values remained well below the threshold of 1 across all groups and exposure percentiles. In contrast, TCR exceeded the acceptable threshold of 1×10⁻⁴ in every simulated scenario, with over 99% of the total risk attributable to Cr(VI). The extent of exceedance was pronounced, reaching up to two orders of magnitude in adults at high-exposure levels and one order of magnitude in adolescents at median exposure levels. Sensitivity analysis identified consumption frequency as the predominant determinant of risk (57–85% contribution), consistently surpassing the influence of contamination levels. These findings suggest that long-term, regular consumption of bivalves may pose a significant cancer risk and underscore the need for expanded research on elemental speciation and harmonized European methodologies for mixture risk assessment of potentially toxic elements in seafood to safeguard public health.