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Pesticides are known to affect non-targeted soil microorganisms. Still, studies comparing the effect of multiple pesticides on a wide range of microbial endpoints associated with carbon cycling are scarce. Here, we employed fluorescence enzymatic assay and real-time PCR to evaluate the effect of 20 commercial pesticides, applied at their recommended dose and five times their recommended dose, on soil carbon cycling related enzymatic activities (α-1,4-glucosidase, β-1,4-glucosidase, β-d-cellobiohydrolase and β-xylosidase), and on the absolute abundance of functional genes (cbhl and chiA), in three different South Australian agricultural soils. The effects on cellulolytic and chitinolytic microorganisms, and the total microbial community composition were determined using shotgun metagenomic sequencing in selected pesticide-treated and untreated samples. The application of insecticides significantly increased the cbhl and chiA genes absolute abundance in the acidic soil. At the community level, insecticide fipronil had the greatest stimulating effect on cellulolytic and chitinolytic microorganisms, followed by fungicide metalaxyl-M and insecticide imidacloprid. A shift towards a fungal dominated microbial community was observed in metalaxyl-M treated soil. Overall, our results suggest that the application of pesticides might affect the soil carbon cycle and may disrupt the formation of soil organic matter and structure stabilisation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)