This Guidance document describes harmonised risk assessment methodologies for combined exposure to multiple chemicals for all relevant areas within EFSA's remit, i.e. human health, animal health and ecological areas. First, a short review of the key terms, scientific basis for combined exposure risk assessment and approaches to assessing (eco)toxicology is given, including existing frameworks for these risk assessments. This background was evaluated, resulting in a harmonised framework for risk assessment of combined exposure to multiple chemicals. The framework is based on the risk assessment steps (problem formulation, exposure assessment, hazard identification and characterisation, and risk characterisation including uncertainty analysis), with tiered and stepwise approaches for both whole mixture approaches and component‐based approaches. Specific considerations are given to component‐based approaches including the grouping of chemicals into common assessment groups, the use of dose addition as a default assumption, approaches to integrate evidence of interactions and the refinement of assessment groups. Case studies are annexed in this guidance document to explore the feasibility and spectrum of applications of the proposed methods and approaches for human and animal health and ecological risk assessment. The Scientific Committee considers that this Guidance is fit for purpose for risk assessments of combined exposure to multiple chemicals and should be applied in all relevant areas of EFSA's work. Future work and research are recommended.

Despite the global importance of forests, it is virtually unknown how their soil microbial communities adapt at the phylogenetic and functional level to long-term metal pollution. Studying 12 sites located along two distinct gradients of metal pollution in Southern Poland revealed that functional potential and diversity (assessed using GeoChip 4.2) were highly similar across the gradients despite drastically diverging metal contamination levels. Metal pollution level did, however, significantly impact bacterial community structure (as shown by MiSeq Illumina sequencing of 16S rRNA genes), but not bacterial taxon richness and community composition. Metal pollution caused changes in the relative abundance of specific bacterial taxa, including Acidobacteria, Actinobacteria, Bacteroidetes, Chloroflexi, Firmicutes, Planctomycetes and Proteobacteria. Also, a group of metal-resistance genes showed significant correlations with metal concentrations in soil. Our study showed that microbial communities are resilient to metal pollution; despite differences in community structure, no clear impact of metal pollution levels on overall functional diversity was observed. While screens of phylogenetic marker genes, such as 16S rRNA genes, provide only limited insight into resilience mechanisms, analysis of specific functional genes, e.g. involved in metal resistance, appears to be a more promising strategy.

Heavy metals can cause a decrease in the taxonomic diversity of soil communities. Because of functional redundancy, it remains unclear to what extent different functions performed by the soil microbial communities may be affected by pollution. We studied the impact of metal contamination on soil bacterial and fungal functional diversity, active microbial biomass, and soil respiration rate. Soil samples were collected from 39 sites along three forest and five meadow pollution transects near an abandoned Pb/Zn smelter in Avonmouth (UK) and Ni smelter in Clydach (UK), in a Cu mining and smelting region near Glogow (Poland), and in a Zn/Pb mining and smelting region near Olkusz (Poland). Biolog® GN2 and SFN2 plates were used to study the bacterial and fungal functional diversity, which subsequently was expressed as Shannon's diversity index (H′). The active microbial biomass was measured as substrate-induced respiration. We found that the bacterial functional diversity significantly decreased, whereas the fungal functional diversity slightly increased, with increasing metal concentration. We also observed a slight negative effect of metal pollution on the active microbial biomass. No relationship was found between metal contamination and total soil respiration rate. This suggests a higher sensitivity of bacterial functional diversity as an indicator for the effects of metal pollution compared with overall soil respiration. All microbial parameters were affected by nutrient concentrations and/or soil pH.