Soil is hosting a tremendous microbial diversity playing a key-role in a number of soil ecosystemic services including nutrient cycling and filtering. ln the European Soil Framework Directive pesticides are clearly marked as one of the major threats for soil biodiversity and functioning. ln orcier to guarantee minimum effects of pesticide application on soil microbes, pesticicleregistration at EU level (Regulation 2006/388) consiclers the toxicity of pesticides ontci non targefsoil microbes by relying on carbon- and nitrcigen-mineralization tests (OECO 216, 217). However tnese tests do not provicle a comprehensive assessment of pesticides onto soil microbes.-ECOFUN-MICROBIODIV was a project cleclicated to cleveloping and evaluating innovativetools toestimate the ecotoxicological impact of sulphonylurea herbicide nicosulfuron on sail micrcibial diversity and functioning. ln orcier to gain full insights into the potential toxiCify of the targeted herbicide a research consortium was established comprised of teams with expertise in agronomy, soif and environmental chemistry, environmental microbiology, soif molecular biology and biochemistry. Two scenario of exposure were considered (i) Tier 1, the worst case scenario, consisting in an experiment conclucted in a greenhouse with up to xl000 of the nicosulfuron dose applied and (ii) Tier Il, the agronomical scenario, consisting in an experiment conducted in the field (1 x, 2x and Sx of nicosulfuron) Agronomical parameters were recorded for bath experiments (yield of crop, weeds development, root and shoot biomass of corn). Sail and corn root sampling was carried out ali along the experiment. The fate of nicosulfuron was monitored in soil by HPLC-UV.The impact of herbicide was estimated using standard methods aimed at studying the abundance (IS014240:2), diversity (ISO/TS29843-1), and the activity (TS29843-1) of the soil microflora. ln addition, new methods based on direct soil DNA extraction (ISO 11 063) were appliecl to determine herbicide impact on structure and abundance of fungal and bacterial communities, structure, abundance, and activity of the functional communities involved in C and N cycling and on the formation of endomycorhizal ; symbiosis. Results suggested that for the good understanding of the impact of low-dose herbicide, such as nicosulfuron, on the abundance, structure, diversity, and activity of sail microbial community it is necessary to use tools of different resolution levels. Application of such multidisciplinary approach relying on biochemistry and molecular biology gives a good tl estimation of the pesticide impact. Results suggested thalif the impact of law-close pesticide wouId beonlyconsidered using the dominant microbial populations it could be underestimated. The consideration of liner approaches allowing to target specifically a functional community or togo deeper in the phylogeny was proven to give a more accurate estimation of the impact of low dose pesticide onto sail microflora.