Combined toxicity of polyethylene microplastics and soil salinization to earthworms is generally antagonistic and additive

Microplastic (MP) accumulation and salinization frequently co-occur in terrestrial ecosystems, posing potential risks to soil health. However, their combined toxicity to soil invertebrates remains unclear. This study investigated combined effects of NaCl and polyethylene MPs (< 35, < 125, < 500 μm) on earthworms. The non-saline avoidance EC50 for < 35 μm MPs was 18 wt%, while avoidance only occurred at 50 wt% for < 125 μm MPs and not at all for < 500 μm MPs. Synergistic effects between NaCl and small MPs increased avoidance. High (10 wt%) MP concentrations by themselves did not increase earthworm mortality; mortality was lower with both MPs and 4000 mg/kg NaCl than with NaCl alone. Co-exposure to MPs and 1000 mg/kg NaCl further reduced growth rates by 16% but increased cocoon production by 259% compared to NaCl exposure alone. NaCl exposure induced earthworm oxidative and osmotic stress; overall stress levels were unchanged by MP co-exposure. 16S rRNA gene sequencing revealed that both NaCl and MPs significantly altered earthworm gut microbiome, suggesting potential impaired gut health. MPs more strongly impacted gut microbial community and functions when no salt was added. Overall, combined effects of NaCl and MPs on earthworms are mainly additive and antagonistic.