Single Species In Vitro Assays with Nitrifying Bacteria and Archaea for Assessing the Toxicity of Pesticides on Soil Microorganisms

Single species tests on surrogate organisms from different trophic levels constitute a cornerstone in aquatic and terrestrial ecotoxicology, representing a major tool for assessing pesticide ecotoxicity. However, there are no assays approved under the EU regulatory framework that specifically target soil microorganisms, despite their pivotal role in ecosystems. This study evaluated the toxicity of pesticides on ammonia-oxidizing microorganisms (AOM), recognized as potential indicators of pesticide toxicity in soil, using a single species in vitro testing system. We assessed the toxicity of ten pesticides of major categories (insecticides, fungicides, herbicides) on a range of ammonia-oxidizing bacteria (AOB), ammonia-oxidizing archaea (AOA) and functionally associated nitrite-oxidizing bacteria (NOB) in liquid cultures, representing all major soil lineages of both groups. Toxicity end points (EC₅₀) were calculated for each strain-pesticide combination. Nitrosotalea sinensis ND2, Nitrosospira briensis and Candidatus Nitrobacter laanbroekii NHB1 were the most sensitive AOA, AOB and NOB, respectively. Fungicides and insecticides were more toxic to AOM than NOB, whereas herbicides had a wide range of toxicity across all groups of nitrifiers. Our study introduces a novel testing approach that includes a range of phylogenetically and ecophysiologically distinct nitrifier strains for assessing pesticides toxicity on soil microorganisms that could potentially, upon standardization, serve as a tier I assay in environmental risk analysis.