Papadopoulos Christos, Roshanfekrrad Marjan, Tsikou Daniela, Papadopoulou Kalliope K, Calonne-Salmon Maryline, Declerck Stephan, Karpouzas Dimitrios G
Laboratory of Plant and Environmental Biotechnology, Department of Biochemistry and Biotechnology, University of Thessaly, Biopolis Campus, Larissa 41500, Greece.
Universite Catholique de Louvain, Earth and Life Institute, Mycology, Croix du Sud 2, Box L7.05.06, Louvain‑La‑Neuve 1348, Belgium.
Ecotoxicol Environ Saf. 2025 Sep 15;303:118892. doi: 10.1016/j.ecoenv.2025.118892. Epub 2025 Aug 18.
Soil microorganisms are a key protection goal in the European Union (EU) pesticide regulatory framework. Arbuscular mycorrhizal fungi (AMF) were identified as good proxies for assessing pesticides toxicity on the soil microbiota. This could involve ecotoxicity testing at the different life stages of AMF. We evaluated the effects of five pesticides (pyraclostrobin, fludioxonil, hymexazol, etridiazole, glyphosate) and a transformation product (AMPA), with distinct mode of action, on the development and functionality of Rhizophagus irregularis at the asymbiotic and symbiotic phase using a spore germination assay and a gnotobiotic AMF-host plant system (AMF-sandwich test), respectively. Based on arbuscular colonization in the AMF-sandwich test, fludioxonil was the most toxic (EC 0.085 mg/L) followed by glyphosate (EC 2.58 mg/L) and pyraclostrobin (EC 9.22 mg/L), while etridiazole, hymexazol, and AMPA showed EC values higher than the highest tested concentration. However, for glyphosate and pyraclostrobin negative effects on symbiosis functioning were observed at lower concentrations than for colonization, as depicted by the expression of plant marker genes and/or P-uptake, suggesting the establishment of non-functional arbuscular symbiosis. The high toxicity of fludioxonil (EC 0.03 mg/L) and the low toxicity of AMPA (EC > 432 mg/L) on R. irregularis was verified also for the asymbiotic phase via spore germination assay. Comparative tests showed differences in the toxicity of pure active substances and commercial formulations of fludioxonil and pyraclostrobin on the AMF-sandwich test. We propose that the AMF-sandwich system together with the spore germination test could be used as a toolbox for Tier-I assessment of pesticides toxicity on AMF.
土壤微生物是欧盟农药监管框架中的一个关键保护目标。丛枝菌根真菌（AMF）被确定为评估农药对土壤微生物群毒性的良好替代指标。这可能涉及在AMF的不同生命阶段进行生态毒性测试。我们分别使用孢子萌发试验和无菌AMF-宿主植物系统（AMF三明治试验），评估了五种作用方式不同的农药（吡唑醚菌酯、咯菌腈、恶霉灵、土菌灵、草甘膦）和一种转化产物（氨甲基膦酸）对不规则球囊霉在非共生和共生阶段的发育及功能的影响。基于AMF三明治试验中的丛枝定殖情况，咯菌腈毒性最大（EC为0.085毫克/升），其次是草甘膦（EC为2.58毫克/升）和吡唑醚菌酯（EC为9.22毫克/升），而土菌灵、恶霉灵和氨甲基膦酸的EC值高于最高测试浓度。然而，对于草甘膦和吡唑醚菌酯，在低于定殖浓度时就观察到了对共生功能的负面影响，如植物标记基因的表达和/或磷吸收所示，这表明建立了无功能的丛枝共生。通过孢子萌发试验也证实了咯菌腈（EC为0.03毫克/升）对不规则球囊霉的高毒性和氨甲基膦酸（EC>432毫克/升）的低毒性在非共生阶段同样存在。比较试验表明，咯菌腈和吡唑醚菌酯的纯活性物质和商业制剂在AMF三明治试验中的毒性存在差异。我们建议，AMF三明治系统与孢子萌发试验可作为一级评估农药对AMF毒性的工具箱。
