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Studies on the functional significance of bacteria associated with ectomycorrhizal (ECM) fungi are scarce, as well as information on the metabolism of the host plant when in symbiosis with ECM fungi. Here we intended to evaluate the phenolic profile of seedlings when associated with Bacillus subtilis (B1), Pisolithus tinctorius (Pis) and their combination (PisB1). The interaction between microorganisms was conducted in three stages: (i) in vitro evaluation of fungal/bacterial interaction, (ii) microcosms, (iii) plant transplantation to natural soil. The profile of phenolic compounds was determined at the end of stages (ii) and (iii) and further supplemented with biometric, nutritional and analysis of the ectomycorrhizal community by denaturing gradient gel electrophoresis. In the in vitro compatibility test, B1 inhibited fungal growth at all glucose concentrations tested. In the microcosm, the levels of chlorogenic and p-coumaric acid decreased over time, unlike the protocatechuic acid which tended to increase during 70 days. After transplantation to the soil, the levels of phenolic acids decreased in all treatments, while catechin increased. B. subtilis positively influenced the fungus-plant relationship as was evidenced by higher biomass of seedlings inoculated with the dual inoculum (PisB1), both in the microcosm and soil stages. The presence of the bacteria interfered in the composition of the ECM fungal community installed in Pinus pinea L. in the soil. This leads to infer that B. subtilis may have caused a greater effect on the metabolism of P. pinea, especially in synergy with mycorrhizal fungi, than the action of the isolated fungus.