Assessing realistic soil microbial risks from metal pollution in a copper mining area through a progressive triad...

Ecological risk assessment (ERA) is crucial for soil protection of contaminated areas. A traditional triad ERA integrates chemical, ecological, and toxicological evidence in parallel, potentially obscuring nuanced pollution-effect relationships under complex field conditions. This study developed a progressive triad approach with consecutive tiers across a 20-km radius of a metal-polluted mining area. The chemical tier identified a 10-km decaying risk zone, attributed to mining-derived As, Cu, Pb, and Zn accumulations. The ecological tier, applying in-situ soil phospholipid fatty acids (PLFAs) investigation, demonstrated metal contamination significantly altered PLFA structure, and reduced gram-positive bacteria and actinomycetes abundance by 59.0 % and 63.6 %. Notably, Cr-not a risk driver in chemical tier-emerged as the strongest metal factor on PLFA profiles, although soil total nitrogen and organic carbon exerted greater overall impacts. This discrepancy underscores actual pollution-effect relationships could be obscured in conventional chemical ERAs. Toxicological validation through incubation experiments with field-relevant Cr (33-73 mg/kg) and Cu (17-130 mg/kg) levels, coupled with 16S rRNA sequencing, confirmed Cr induced greater bacterial compositional shifts than Cu, notably promoting Firmicutes dominance. This framework provides a widely applicable and cost-effective pathway to enhance ecological realism of ERAs in contaminated areas, supporting sustainable environmental management and soil health protection worldwide.